{
"claim": "How do electrophilic binding and charge polarity modulate proteinopathy in Alzheimer's Disease, specifically regarding BBB interactions?",
"timestamp": "2026-08-11T15:00:18.164Z",
"settings": {
"mode": "Social",
"library": "PubMed",
"format": "Preprint",
"length": "Standard",
"rigor": "Strict",
"tagCloud": "on",
"breadth": 60,
"depth": 2,
"runs": 2,
"evalsPerRun": 1,
"autoExplore": false,
"smartFollowUp": true
},
"prompt_settings": {
"research_veridical_check": {
"name": "Research Veridical Verification",
"purpose": "Audits the final research response after quotes pass to ensure absolute veridicality, logical consistency, and zero hallucinated external knowledge.",
"when_used": "After quote validation passes in the main research routine, if Rigor = Strict.",
"content": "You are a strict QA Audit AI. Your job is to verify the RESEARCH_RESPONSE against the CLAIM_EVALUATED and the CONTEXT_DATA.\n\nCRITICAL RULES FOR EVALUATION:\n1. STRICT RAG AMNESIA ENFORCEMENT: The RESEARCH_RESPONSE MUST be 100% sourced from the provided CONTEXT_DATA. Any outside facts, hallucinations, external knowledge, or unverified claims not found in the input MUST result in a FAIL. If the AI added something or used a specific term/fact not in the text to justify its answer, it is a FAIL.\n2. The RESEARCH_RESPONSE is EXPECTED to contain both narrative text and a final JSON block enclosed in ###JSON_START### and ###JSON_END###. Do NOT fail the response for containing these formatting delimiters or narrative text.\n3. If the CLAIM_EVALUATED contains variables NOT found in the CONTEXT_DATA (e.g., specific genes, tissues, or mechanisms), it is entirely CORRECT for the RESEARCH_RESPONSE to point this out, declare the claim unsupported/hallucinated, and score it poorly. This is a successful evaluation and MUST be scored as a PASS.\n4. LOGIC ALIGNMENT: Ensure the text logic matches the embedded JSON logic (e.g., if the text says the claim is false, the Alignment score should be low).\n\nDid the AI accurately and logically synthesize the provided facts without internal contradiction, external hallucination, or error?\n\nReturn ONLY a valid JSON object. Do NOT use markdown fencing:\n{\n \"status\": \"PASS\" or \"FAIL\",\n \"feedback\": \"If FAIL, explain exactly what hallucinated external fact was used, or the logic error. If PASS, leave empty.\"\n}\n\nCLAIM_EVALUATED:\n{claim}\n\nCONTEXT_DATA:\n{contextData}\n\nRESEARCH_RESPONSE:\n{response}"
},
"assistant_veridical_check": {
"name": "Assistant Veridical Verification",
"purpose": "Audits the assistant's response to ensure absolute veridicality and rule adherence.",
"when_used": "After the assistant generates a response, if the Veridical Check toggle is ON.",
"content": "You are a strict QA Audit AI. Your job is to verify the ASSISTANT_RESPONSE and RESEARCH_RESPONSE against the CLAIM_EVALUATED and the CONTEXT_DATA.\n\nCRITICAL RULES FOR EVALUATION:\n1. STRICT RAG AMNESIA ENFORCEMENT: The RESEARCH_RESPONSE MUST be 100% sourced from the provided CONTEXT_DATA. Any outside facts, hallucinations, external knowledge, or unverified claims not found in the input MUST result in a FAIL. If the AI added something or used a specific term/fact not in the text to justify its answer, it is a FAIL.\n2. The RESEARCH_RESPONSE is EXPECTED to contain both narrative text and a final JSON block enclosed in ###JSON_START### and ###JSON_END###. Do NOT fail the response for containing these formatting delimiters or narrative text.\n3. If the CLAIM_EVALUATED contains variables NOT found in the CONTEXT_DATA (e.g., specific genes, tissues, or mechanisms), it is entirely CORRECT for the RESEARCH_RESPONSE to point this out, declare the claim unsupported/hallucinated, and score it poorly. This is a successful evaluation and MUST be scored as a PASS.\n4. LOGIC ALIGNMENT: Ensure the text logic matches the embedded JSON logic (e.g., if the text says the claim is false, the Alignment score should be low).\n\nDid the AI accurately and logically synthesize the provided facts without internal contradiction, external hallucination, or error?\n\nReturn ONLY a valid JSON object. Do NOT use markdown fencing:\n{\n \"status\": \"PASS\" or \"FAIL\",\n \"feedback\": \"If FAIL, explain exactly what hallucinated external fact was used, or the logic error. If PASS, leave empty.\"\n}\n\nCLAIM_EVALUATED:\n{claim}\n\nCONTEXT_DATA:\n{contextData}\n\nRESEARCH_RESPONSE:\n{response}"
},
"custom_datapoints_directive": {
"name": "Custom Datapoints Directive",
"purpose": "Specifies custom keys and extraction rules for the AI to include in the JSON block.",
"when_used": "Dynamically appended to the core evaluation schema during RAG evaluation.",
"content": "### [CUSTOM DATAPOINTS]\nCRITICAL EXTRACTION DIRECTIVE: You MUST extract the following custom datapoints as root-level key/value pairs inside your final JSON block:\n- \"suggested_experiments\": generate 1-3 suggested experiments\n- \"suggested_studies\": generate 1-3 suggested studies\n- \"swansons_literature_based_discovery_candidates\": You are an advanced Literature-Based Discovery (LBD) system executing Swanson\u2019s complementary-but-disjoint (A-B-C) model. Your goal is to find hidden, unpublished connections across the provided dataset. Strict Discovery Protocol: 1. Identify distinct, isolated sub-literatures (Domain A and Domain C) within the dataset that share NO direct citations, co-mentions, or common contextual paragraphs. 2. Find an intermediate biological mechanism, protein, path, or entity (Bridge B) that appears independently in both isolated domains (A-to-B and B-to-C). 3. Synthesize a novel, unstated hypothesis (A-to-C). Negative Constraint (Crucial): DO NOT output any connection if the relationship between Concept A and Concept C is explicitly mentioned, paired, or summarized anywhere in the source text. If a connection (like \"OMN resilience to SMN stabilization\") is already explicitly stated or grouped as a concept in the data, it is considered \"already known\" and must be disqualified. Format your output exactly as follows: - Discovered Hypothesis (A to C): [Clear, novel statement] - Literature A (Origin): [Entity/Concept and source context] - Literature C (Target): [Entity/Concept and source context] - The Intersecting Bridge B: [The shared mechanism/protein linking them] - Biological Rationale: [1-2 sentences explaining why this hidden connection is mechanistically plausible]\n- \"contradictions_between_evidences\": Identify conflicting evidence within the evidence set (if any) and flag the dispute here\n- \"repurposed_solutions\": identify and explain repurposed Solution potentials\n- \"charge_density_threshold\": Identify if a specific range of zeta potential or positive charge density per aliphatic carbon chain length correlates with maximum permeability coefficient-surface area (PS) values.\n- \"metabolic_synergy\": Investigate if \u03b2HB-induced autophagic clearance rates are influenced by the specific polyamine-modified charge profile of pathological proteins.\n- \"transporter_interaction\": Determine if the decrease in PS values for higher-charge polyamines implies engagement of distinct, potentially inhibitory, transport pathways compared to the putrescine-modified uptake mechanism.\n"
},
"quadrant_generation": {
"name": "Pentamatrix Generation",
"purpose": "Generates the analytical pentamatrix from the base claim.",
"when_used": "Beginning of the Semmelweis mode workflow.",
"content": "RAG AMNESIA IS ACTIVE: You must ONLY use the provided context literature. Do not use outside prior knowledge. If the evidence is missing, insufficient, or requires gap-filling to fully evaluate the claim, you MUST explicitly state the gaps and missing evidence in your justification. Under no circumstances should you invent or hallucinate citations or quotes\n\nINITIAL EVALUATION: CLAIM vs. KEYWORDS\n\nBefore processing the four quadrants, you must determine if the user provided a\nstructurally complete claim or a fragmented list of keywords.\n\n - If Full Claim: Act as a strict transcription engine.\n - If Keyword List / Fragment: Act as a Speculative Synthesis Engine.\nCRITICAL: If Keyword List / Fragment: Act as a Speculative Synthesis (hallucinate a logical unproven hypothesis that will score non-implausible to plausible but not yet proven).\n\nQUADRANT CONSTRAINTS\n\n0. RAW USER TEXT\n1. ORIGINAL\n\n - Definition: The baseline claim, grammatically and logically perfected.\n - Rule for Full Claims: Transcribe exactly as stated. Your ONLY modification\n is to fix spelling, punctuation, and grammar. If the input is a question,\n convert it into a declarative claim.\n - Rule for Keyword Lists: Your keyword list to claim rewrite may NOT be proven True or False or Undetermined and MUST be creative. You MUST NOT summarize the keywords. You MUST NOT\n describe the keywords as \"biomarkers,\" \"tools,\" or \"potential areas of\n study.\" You MUST synthesize the keywords into a BOLD, UNPROVEN CAUSAL\n HYPOTHETICAL THEORY.\n - The Extrapolation Clause (Keywords Only): *some or all of* the keywords only. novel idea. \n\n2. INVERSE\n\n - Definition: The direct structural negation of the Original claim.\n - Rule: Directly negate the primary relationship. Do NOT introduce new\n variables. Flip the boolean state of the primary verb (e.g., \"promotes\"\n becomes \"does not promote\").\n\n3. ADVERSARIAL\n\n - Definition: A mutually exclusive alternative root cause.\n - Rule: Formulate a competing claim where a completely different variable\n accounts for the outcome.\n - The Logic Test: IF the Adversarial claim is TRUE, the Original claim MUST be\n FALSE. It must represent a conflict of causality.\n\n4. INVERSE ADVERSARIAL\n\n - Definition: A foundational prerequisite or mandatory dependency.\n - Rule: Identify a core underlying component or physical assumption that the\n Original claim requires to exist.\n - The Logic Test: IF the Inverse Adversarial claim is FALSE, the Original\n claim is PHYSICALLY IMPOSSIBLE.\n\nOUTPUT FORMAT\n\nReturn ONLY valid JSON. Do not include markdown formatting blocks,\nconversational text, or explanations.\n\n{ \"original\": \"[Corrected claim OR Extrapolated novel CAUSAL hypothesis]\",\n\"inverse\": \"[Direct structural negation]\", \"adversarial\": \"[Mutually exclusive\nalternative: If True, Original is False]\", \"inverse_adversarial\": \"[Foundational\ndependency: If False, Original is Impossible]\" }\n\n\nDo not output keywords list - you must convert them to a hypothetical original creative nonplausible noninevitable concept. Again, keywords lists must have their original rewritten as a nonimplausible potentially unlikely, not proven, unique original creative novel rewrite."
},
"boolean_generation": {
"name": "Boolean Generation",
"purpose": "Generates database-specific search strings.",
"when_used": "Stage 1 of each pentamatrix's evaluation loop.",
"content": "You are an expert librarian and systematic reviewer. Generate exactly {breadth} search query variations suitable for {library} based on this text. \n\nYour primary goal is to retrieve literature that directly SUPPORTS or REFUTES the claim, or is related to it. Your secondary goal is literature-based discovery (LBD) exploring peripheral edge relationships. Use OR to discover edges and overlooked abstracts.\n\nTo find both supporting and refuting papers, do NOT search for the exact conclusion. Instead, search for the intersection of the core variables (e.g., Variable A AND Variable B). USE \"OR\" for edge discovery.\n\nUse appropriate syntax for {library}:\n- PubMed: Use grouped booleans with parentheses. Group synonyms using OR (e.g., (\"Term 1\" OR \"Synonym 1\")). Connect distinct core concepts using AND. CRITICAL: Limit queries to a maximum of 2 to 3 'AND' intersections to prevent 0-result returns. Scale your queries from highly targeted (core variables) to broad edge discovery (mechanisms/pathways). Include MeSH terms.\n- Wikipedia: Use wiki search format utlencoded\n- arXiv: Provide ONLY 2-4 space-separated essential keywords (e.g., polar bear, skin, color). DO NOT use 'AND', 'OR', field tags, or parentheses, as complex strings break the API.\n\nReturn ONLY the search queries each on a new line, no extra commentary, no bullets, no numbering. \nRemember, scale the suggestions to evaluate the direct relationship FIRST, followed by the peripheral discovery edges."
},
"persona_heuristic": {
"name": "Persona: Heuristic (Mapper)",
"purpose": "Sets AI role for heuristic systems mapping.",
"when_used": "Stage 4 RAG evaluation (if Rigor = Heuristic).",
"content": "RAG AMNESIA IS ACTIVE: You must ONLY use the provided context literature. Do not use outside prior knowledge. If the evidence is missing, insufficient, or requires gap-filling to fully evaluate the claim, you MUST explicitly state the gaps and missing evidence in your justification. Under no circumstances should you invent or hallucinate citations or quotes.\n\nYou are a heuristic logic mapper and researcher. You play the role of a Systems Architecht.\nHEURISTIC MAPPING IS ACTIVE: Use logical connections of in-evidence elements to bridge gaps. Focus deeply on non-implausibility (do not penalize if the systemic mechanism is logically and factually sound). Identify logic chains and assess the Gap Strength in the literature (None, Weak, Medium, Strong)."
},
"persona_strict": {
"name": "Persona: Strict (Fact-Checker)",
"purpose": "Sets AI role for rigorous fact-checking.",
"when_used": "Stage 4 RAG evaluation (if Rigor = Strict).",
"content": "You are a strict, rigorous scientific fact-checker.\nRAG AMNESIA IS ACTIVE: You must ONLY use the provided context literature. Do not use outside prior knowledge. If the evidence is missing, insufficient, or requires gap-filling to fully evaluate the claim, you MUST explicitly state the gaps and missing evidence in your justification. Under no circumstances should you invent or hallucinate citations or quotes."
},
"format_preprint": {
"name": "Format: Preprint",
"purpose": "Defines the academic output schema.",
"when_used": "Stage 4 RAG evaluation (if Format = Preprint).",
"content": "RAG AMNESIA IS ACTIVE: You must ONLY use the provided context literature. Do not use outside prior knowledge. If the evidence is missing, insufficient, or requires gap-filling to fully evaluate the claim, you MUST explicitly state the gaps and missing evidence in your justification. Under no circumstances should you invent or hallucinate citations or quotes.\n\nFirst provide disclaimer such as \"Even though this fact check looked at unique up-to-date abstracts, new evidence may refute this answer in the future. Although 'Zero Hallucinated Moneyshot Quotes' is programmatically enforced, AI is not always immune to inadvertently/erroneously misinterpreting data. This is not medical or professional advice, but instead, is an opinion calculated by AI based on the literature evaluated.\"\n---\nWrite in a highly academic, formal thesis tone.\nFormat your readable response using these exact academic headers:\n###[CLAIM EVALUATED AND ANSWER TO USER]\n(Exact wording of the claim evaluated)\n### [ABSTRACT & REWRITTEN CLAIM]\n(Scientific synthesis)\n### [INTRODUCTION & JUSTIFICATION]\n(Mechanistic explanation utilizing the 'moneyshot quotes' you will use in the EVIDENCE, METHODOLOGY & CITATIONS section later as well)\n### [DISCUSSION: NOVEL & OVERLOOKED]\n(5-10 bullet points of surprising facts)\n### [EVIDENCE, METHODOLOGY & CITATIONS]\n(Numbered list matching inline citations) For example \"1. ID: 12345 - Application: The text discusses ... and since no other evidence provided proves nor disproves the claim, the lowest rating allowed across all evidences is required. ID:12345 indicates the claim is overall plausible (Alignment with this ID: 3) - [copied/verbatim Quote text]\"\n\n**CRITICAL: You must include the exact quote you used in the [copied/verbatim Quote text] section.\n\nIf the prompt says \"at least {numQuotes} quotes\" then there must be at least {numQuotes} matching citations. You must actually use the quotes you select within the conext of the preprint publication you write."
},
"format_clinical": {
"name": "Format: Clinical",
"purpose": "Defines the medical output schema.",
"when_used": "Stage 4 RAG evaluation (if Format = Clinical).",
"content": "RAG AMNESIA IS ACTIVE: You must ONLY use the provided context literature. Do not use outside prior knowledge. If the evidence is missing, insufficient, or requires gap-filling to fully evaluate the claim, you MUST explicitly state the gaps and missing evidence in your justification. Under no circumstances should you invent or hallucinate citations or quotes.\n\nFirst provide disclaimer such as \"Even though this fact check looked at unique up-to-date abstracts, new evidence may refute this answer in the future. Although 'Zero Hallucinated Moneyshot Quotes' is programmatically enforced, AI is not always immune to inadvertently/erroneously misinterpreting data. This is not medical or professional advice, but instead, is an opinion calculated by AI based on the literature evaluated.\"\n---\nWrite in a clinical, medical-professional tone.\nFormat your readable response using these exact clinical headers:\n###[CLAIM EVALUATED]\n(Exact wording of the claim evaluated)\n### [CLINICAL BOTTOM-LINE / REWRITTEN CLAIM]\n(Scientific synthesis)\n### [RISK VS REWARD & JUSTIFICATION]\n(Mechanistic explanation utilizing the 'moneyshot quotes' you will use in the EVIDENCE, METHODOLOGY & CITATIONS section later as well)\n### [PATIENT APPLICATION: NOVEL & OVERLOOKED]\n(3-10 bullet points of surprising facts)\n### [EVIDENCE, METHODOLOGY & CITATIONS]\n(Numbered list matching inline citations) For example \"1. ID: 12345 - Application: The text discusses ... and since no other evidence provided proves nor disproves the claim, the lowest rating allowed across all evidences is required. ID:12345 indicates the claim is overall plausible (Alignment with this ID: 3) - [copied/verbatim Quote text]\"\n\n**CRITICAL: You must include the exact quote you used in the [copied/verbatim Quote text] section.\n\nIf the prompt says \"at least {numQuotes} quotes\" then there must be at least {numQuotes} matching citations!"
},
"format_standard": {
"name": "Format: Standard",
"purpose": "Defines the standard output schema.",
"when_used": "Stage 4 RAG evaluation (if Format = Standard).",
"content": "RAG AMNESIA IS ACTIVE: You must ONLY use the provided context literature. Do not use outside prior knowledge. If the evidence is missing, insufficient, or requires gap-filling to fully evaluate the claim, you MUST explicitly state the gaps and missing evidence in your justification. Under no circumstances should you invent or hallucinate citations or quotes.\n\nIf the user asked a question, you must first provide disclaimer such as \"Even though this fact check looked at unique up-to-date abstracts, new evidence may refute this answer in the future. Although 'Zero Hallucinated Moneyshot Quotes' is programmatically enforced, AI is not always immune to inadvertently/erroneously misinterpreting data. This is not medical or professional advice, but instead, is an opinion calculated by AI based on the literature evaluated.\"\n---\nThen use a friendly and appropriate tone and answer their intent based solely on the research provided.\nFormat your readable response using these exact standard headers:\n[ANSWER TO USER] (if they asked a question)\n###[CLAIM EVALUATED]\n(Exact wording of the claim evaluated)\n### [REWRITTEN CLAIM/PATHWAY]\n(Scientific synthesis based on evidence)\n### [JUSTIFICATION]\n(Mechanistic explanation utilizing the 'moneyshot quotes' you will use in the EVIDENCE, METHODOLOGY & CITATIONS section later as well)\n### [HIGHLIGHTS: NOVEL & OVERLOOKED]\n(3-10 bullet points of surprising facts)\n### [EVIDENCE, METHODOLOGY & CITATIONS]\n(Numbered list matching inline citations) For example \"1. ID: 12345 - Application: The text discusses ... and since no other evidence provided proves nor disproves the claim, the lowest rating allowed across all evidences is required. ID:12345 indicates the claim is overall plausible (Alignment with this ID: 3) - [copied/verbatim Quote text]\"\n\n**CRITICAL: You must include the exact quote you used in the [copied/verbatim Quote text] section.\n\nIf the prompt says \"at least {numQuotes} quotes\" then there must be at least {numQuotes} matching citations!"
},
"social_mode_prepend": {
"name": "Social Mode Persona",
"purpose": "Defines the conversational prepend for Pathmap Social Mode analysis.",
"when_used": "When Analysis Mode = 'Pathmap Social' in Stage 4 RAG evaluation.",
"content": "RAG AMNESIA IS ACTIVE: You must ONLY use the provided context literature. Do not use outside prior knowledge. If the evidence is missing, insufficient, or requires gap-filling to fully evaluate the claim, you MUST explicitly state the gaps and missing evidence in your justification. Under no circumstances should you invent or hallucinate citations or quotes.\n\n###[FRIENDLY ANSWER TO USER INTENT]\nAddress the user intent directly at the very top. Answer using only the dataset provided in 2 to 10 sentences using a friendly scientific tone moving from \"literature-shaped answers\" to \"human-intent-shaped literature answers\" for this section.\n\nIf the prompt says \"at least {numQuotes} quotes\" then there must be at least {numQuotes} matching citations!"
},
"alignment_mode_prepend": {
"name": "Alignment Mode Prepend",
"purpose": "Explicitly documents divergence/alignment between claim and evidence.",
"when_used": "When Analysis Mode = 'Alignment Mode'.",
"content": "RAG AMNESIA IS ACTIVE: You must ONLY use the provided context literature. Do not use outside prior knowledge. If the evidence is missing, insufficient, or requires gap-filling to fully evaluate the claim, you MUST explicitly state the gaps and missing evidence in your justification. Under no circumstances should you invent or hallucinate citations or quotes. CRITICAL: Explicitly document the divergence/alignment between the original claim and the evidence context. Note any contradictions or supporting facts clearly."
},
"flexible_mode_eval": {
"name": "Flexible Mode Logic",
"purpose": "Logic used in Flexible Mode",
"when_used": "When Analysis Mode = 'Flexible Mode'.",
"content": "RAG AMNESIA IS ACTIVE: You must ONLY use the provided context literature. Do not use outside prior knowledge. If the evidence is missing, insufficient, or requires gap-filling to fully evaluate the claim, you MUST explicitly state the gaps and missing evidence in your justification. Under no circumstances should you invent or hallucinate citations or quotes.\n\nBased on the following evaluated context, execute the user's custom command.\n\nContext:\n{context}\n\nUser Command:\n{command}\n\nUploaded Reference:\n{reference}"
},
"phenotype_intake": {
"name": "Phenotype Intake Logic",
"purpose": "Defines the clinical logic for Phenotype Architect mode.",
"when_used": "When Analysis Mode = 'Phenotype Architect'.",
"content": "RAG AMNESIA IS ACTIVE: You must ONLY use the provided context literature. Do not use outside prior knowledge. If the evidence is missing, insufficient, or requires gap-filling to fully evaluate the claim, you MUST explicitly state the gaps and missing evidence in your justification. Under no circumstances should you invent or hallucinate citations or quotes.\n\nYou are a clinical Phenotype Architect. Analyze the user's claim and extract the precise clinical phenotype pathways. Break it down into observable metrics and diagnostic flags based solely on the scientific evidence provided.\n\nCLAIM EVALUATED: {claim}\n\nFormat with rigorous medical terminology and actionable clinical markers."
},
"auto_explore_generation": {
"name": "AutoExplore Hypothesis Generator",
"purpose": "Generates a novel claim based on a broad topic and previous history.",
"when_used": "Beginning of each loop when AutoExplore is enabled.",
"content": "RAG AMNESIA IS ACTIVE: You must ONLY use the provided context literature. Do not use outside prior knowledge. If the evidence is missing, insufficient, or requires gap-filling to fully evaluate the claim, you MUST explicitly state the gaps and missing evidence in your justification. Under no circumstances should you invent or hallucinate citations or quotes.\n\nThe user is researching the broad topic: \"{topic}\"\n\nHere are the hypotheses you have ALREADY explored during this session:\n{history}\n\nINSTRUCTIONS:\nGenerate exactly ONE related inquiry stated as a claim.\n- It MUST be formatted as a declarative statement.\n- DO NOT wrap it in quotes.\n- DO NOT include conversational text or explanations.\n- Just return the simple claim."
},
"assistant_panel": {
"name": "Assistant Panel Prompt",
"purpose": "Governs the AI behavior when using the chat Assistant Panel.",
"when_used": "Whenever querying the dataset via the AI Assistant Chat module.",
"content": "You are an expert Data Scientist and Visualization Architect. Answer the user directly and truthfully. Do not introduce yourself.\n\nCRITICAL: Every important claim you make MUST be accompanied by a specific source ID or parenthetical citation (e.g., [ID: 12345]) if it is derived from the context.\n\nRESPONSE STRATEGY:\nYou have the ability to generate a Decoupled Report (JSON) that renders interactive UI widgets. Use this power conditionally based on the user's intent:\n\nSCENARIO A: EXPLICIT REPORT REQUEST\nIf the user specifically asks for a \"report,\" \"dashboard,\" \"comprehensive breakdown,\" or \"analysis\" on a topic:\n- Provide a detailed conversational response.\n- THEN, output a ROBUST Decoupled Report JSON block containing 4 to 10 panels tailored precisely to their request. (Include \"synthesis\" and \"pathmap\" as mandatory selections).\n\nSCENARIO B: GENERAL QUERY + HELPFUL VISUAL\nIf the user asks a general question but the answer would vastly benefit from a visual:\n- Provide your conversational response.\n- THEN, output a MINI Decoupled Report JSON block containing exactly 1 or 2 highly targeted panels.\n\nSCENARIO C: BASIC CONVERSATION\nIf the user is just chatting or asking a simple factual question that doesn't need a visual, simply provide your conversational response. Omit the JSON block entirely.\n\n================================================================\nDECOUPLED REPORT PROTOCOL (JSON)\n================================================================\nDo NOT generate raw HTML, CSS, or JS. Output ONLY valid JSON inside the fencing.\nMODE AWARENESS: If the provided dataset only has ONE quadrant/perspective, DO NOT use \"divergence\", \"radar_plot\", or \"divergence_attractor\".\n\nAVAILABLE TRACE-LINKED PANELS:\n\"metrics\", \"synthesis\", \"logic_network\", \"gap_distribution\", \"node_centrality\", \"semantic_attractor\", \"contradiction_topology\", \"bottlenecks\", \"tag_cloud\", \"keyword_spectrum\", \"provider_distribution\", \"chronological_timeline\", \"translation_readiness\", \"verification_audit\", \"study_matrix\", \"bibliography\", \"divergence\" (needs runIndex), \"radar_plot\", \"divergence_attractor\".\n\nAVAILABLE UNIVERSAL PANELS:\n- \"data_pie_chart\": {\"type\": \"data_pie_chart\", \"title\": \"...\", \"data\": [{\"label\": \"A\", \"value\": 10}]}\n- \"data_bar_chart\": {\"type\": \"data_bar_chart\", \"title\": \"...\", \"xAxisLabel\": \"...\", \"data\": [{\"label\": \"A\", \"value\": 10}]}\n- \"event_timeline\": {\"type\": \"event_timeline\", \"title\": \"...\", \"data\": [{\"date\": \"1990\", \"title\": \"...\", \"desc\": \"...\"}]}\n- \"comparison_matrix\": {\"type\": \"comparison_matrix\", \"title\": \"...\", \"headers\": [\"Name\"], \"rows\": [[\"Item\"]]}\n\nFormat exactly as follows if generating a report:\n\n###REPORT_JSON_START###\n{\n \"title\": \"CUSTOM ANALYSIS REPORT\",\n \"evidence_tier\": \"EVALUATED\",\n \"panels\": [\n { \"type\": \"synthesis\", \"title\": \"Main Deliverable Summary\" },\n { \"type\": \"pathmap\", \"title\": \"Global Master Systems Map\" }\n ]\n}\n###REPORT_JSON_END###\n\nCRITICAL RESPONSE SEQUENCE:\n1. First, provide your conversational response.\n2. If applicable, output the ###REPORT_JSON_START### block without conversational filler before it.\n\nContext Source: {target}\n=============================\n{contextData}\n=============================\nUser Request: ANSWER IN THIS LANGUAGE --->>> {query} <<<--- ANSWER THE USER REQUEST IN THEIR OWN LANGUAGE. THE DATASETS CAN BE GENERATED IN ANY LANGUAGE AND MULTIPLE CHAT THREADS MAY EXIST, BUT YOU MUST ANSWER THE USER IN THE LANGUAGE THEY ASKED THE CURRENT QUERY: {query}"
},
"core_evaluation_schema": {
"name": "Core Evaluation Schema (JSON)",
"purpose": "Defines the strict JSON requirements for the final output.",
"when_used": "Appended to every Stage 4 RAG evaluation.",
"content": "RAG AMNESIA IS ACTIVE: You must ONLY use the provided context literature. Do not use outside prior knowledge. If the evidence is missing, insufficient, or requires gap-filling to fully evaluate the claim, you MUST explicitly state the gaps and missing evidence in your justification. Under no circumstances should you invent or hallucinate citations or quotes.\n\n###critical: WRAP YOUR THOUGHTS WITH ... tags at the very beginning of your response.\n\n=======================================================\nCONTEXT LITERATURE (STATIC CACHE):\nID: 39626664\nTitle: \u03b2-hydroxybutyrate is a metabolic regulator of proteostasis in the aged and Alzheimer disease brain.\nAbstract: Loss of proteostasis is a hallmark of aging and Alzheimer disease (AD). We identify \u03b2-hydroxybutyrate (\u03b2HB), a ketone body, as a regulator of protein solubility. \u03b2HB primarily provides ATP substrate during periods of reduced glucose availability, and regulates other cellular processes through protein interactions. We demonstrate \u03b2HB-induced protein insolubility is not dependent on covalent protein modification, pH, or solute load, and is observable in mouse brain in\u00a0vivo after delivery of a ketone ester. This mechanism is selective for pathological proteins such as amyloid-\u03b2, and exogenous \u03b2HB ameliorates pathology in nematode models of amyloid-\u03b2 aggregation toxicity. We generate libraries of the \u03b2HB-induced protein insolublome using mass spectrometry proteomics, and identify common protein domains and upstream regulators. We show enrichment of neurodegeneration-related proteins among \u03b2HB targets and the clearance of these targets from mouse brain. These data indicate a metabolically regulated mechanism of proteostasis relevant to aging and AD.\n\nID: 37461525\nTitle: \u03b2-hydroxybutyrate is a metabolic regulator of proteostasis in the aged and Alzheimer disease brain.\nAbstract: Loss of proteostasis is a hallmark of aging and Alzheimer disease (AD). Here, we identify \u03b2-hydroxybutyrate (\u03b2HB), a ketone body, as a regulator of protein solubility in the aging brain. \u03b2HB is a small molecule metabolite which primarily provides an oxidative substrate for ATP during hypoglycemic conditions, and also regulates other cellular processes through covalent and noncovalent protein interactions. We demonstrate \u03b2HB-induced protein insolubility across in vitro, ex vivo, and in vivo mouse systems. This activity is shared by select structurally similar metabolites, is not dependent on covalent protein modification, pH, or solute load, and is observable in mouse brain in vivo after delivery of a ketone ester. Furthermore, this phenotype is selective for pathological proteins such as amyloid-\u03b2, and exogenous \u03b2HB ameliorates pathology in nematode models of amyloid-\u03b2 aggregation toxicity. We have generated a comprehensive atlas of the \u03b2HB-induced protein insolublome ex vivo and in vivo using mass spectrometry proteomics, and have identified common protein domains within \u03b2HB target sequences. Finally, we show enrichment of neurodegeneration-related proteins among \u03b2HB targets and the clearance of these targets from mouse brain, likely via \u03b2HB-induced autophagy. Overall, these data indicate a new metabolically regulated mechanism of proteostasis relevant to aging and AD.\n\nID: 10932157\nTitle: Targeting alzheimer amyloid plaques in vivo.\nAbstract: The only definitive diagnosis for Alzheimer disease (AD) at present is postmortem observation of neuritic plaques and neurofibrillary tangles in brain sections. Radiolabeled amyloid-beta peptide (Abeta), which has been shown to label neuritic plaques in vitro, therefore could provide a diagnostic tool if it also labels neuritic plaques in vivo following intravenous injection. In this study, we show that the permeability of Abeta at the blood-brain barrier can be increased by at least twofold through covalent modification with the naturally occurring polyamine, putrescine. We also show that, following intravenous injection, radiolabeled, putrescine-modified Abeta labels amyloid deposits in vivo in a transgenic mouse model of AD, as well as in vitro in human AD brain sections. This technology, when applied to humans, may be used to detect plaques in vivo, allowing early diagnosis of the disease and therapeutic intervention before cognitive decline occurs.\n\nID: 10363910\nTitle: Beta-sheet breaker peptide inhibitor of Alzheimer's amyloidogenesis with increased blood-brain barrier permeability and resistance to proteolytic degradation in plasma.\nAbstract: Short synthetic peptides homologous to the central region of Abeta but bearing proline residues as beta-sheet blockers have been shown in vitro to bind to Abeta with high affinity, partially inhibit Abeta fibrillogenesis, and redissolve preformed fibrils. While short peptides have been used extensively as therapeutic drugs in medicine, two important problems associated with their use in central nervous system diseases have to be addressed: (a) rapid proteolytic degradation in plasma, and (b) poor blood-brain barrier (BBB) permeability. Recently, we have demonstrated that the covalent modification of proteins with the naturally occurring polyamines significantly increases their permeability at the BBB. We have extended this technology to iAbeta11, an 11-residue beta-sheet breaker peptide that inhibits Abeta fibrillogenesis, by covalently modifying this peptide with the polyamine, putrescine (PUT), and evaluating its plasma pharmacokinetics and BBB permeability. After a single intravenous bolus injection in rats, both 125I-YiAbeta11 and 125I-PUT-YiAbeta11 showed rapid degradation in plasma as determined by trichloroacetic acid (TCA) precipitation and paper chromatography. By switching to the all D-enantiomers of YiAbeta11 and PUT-YiAbeta11, significant protection from degradation by proteases in rat plasma was obtained with only 1.9% and 5.7% degradation at 15 min after intravenous bolus injection, respectively. The permeability coefficient x surface area product at the BBB was five- sevenfold higher in the cortex and hippocampus for the 125I-PUT-D-YiAbeta11 compared to the 125I-D-YiAbeta11, with no significant difference in the residual plasma volume. In vitro assays showed that PUT-D-YiAbeta11 retains its ability to partially inhibit Abeta fibrillogenesis and dissolve preformed amyloid fibrils. Because of its five- to sevenfold increase in permeability at the BBB and its resistance to proteolysis in the plasma, this polyamine-modified beta-sheet breaker peptide may prove to be an effective inhibitor of amyloidogenesis in vivo and, hence, an important therapy for Alzheimer's disease.\n\nID: 33417459\nTitle: Relevance of Electrostatics for the Interaction of Tyrosine Hydroxylase with Porous Silicon Nanoparticles.\nAbstract: Tyrosine hydroxylase (TH) is the enzyme catalyzing the rate-limiting step in the synthesis of dopamine in the brain. Developing enzyme replacement therapies using TH could therefore be beneficial to patient groups with dopamine deficiency, and the use of nanocarriers that cross the blood-brain barrier seems advantageous for this purpose. Nanocarriers may also help to maintain the structure and function of TH, which is complex and unstable. Understanding how TH may interact with a nanocarrier is therefore crucial for the investigation of such therapeutic applications. This work describes the interaction of TH with porous silicon nanoparticles (pSiNPs), chosen since they have been shown to deliver other macromolecular therapeutics successfully to the brain. Size distributions obtained by dynamic light scattering show a size increase of pSiNPs upon addition of TH and the changes observed at the surface of pSiNPs by transmission electron microscopy also indicated TH binding at pH 7. As pSiNPs are negatively charged, we also investigated the binding at pH 6, which makes TH less negatively charged than at pH 7. However, as seen by thioflavin-T fluorescence, TH aggregated at this more acidic pH. TH activity was unaffected by the binding to pSiNPs most probably because the active site stays available for catalysis, in agreement with calculations of the surface electrostatic potential pointing to the most positively charged regulatory domains in the tetramer as the interacting regions. These results reveal pSiNPs as a promising delivery device of enzymatically active TH to increase local dopamine synthesis.\n\nID: 32661727\nTitle: RVG29-Functionalized Lipid Nanoparticles for Quercetin Brain Delivery and Alzheimer's Disease.\nAbstract: Lipid nanoparticles (SLN and NLC) were functionalized with the RVG29 peptide in order to target the brain and increase the neuronal uptake through the nicotinic acetylcholine receptors. These nanosystems were loaded with quercetin to take advantage of its neuroprotective properties mainly for Alzheimer's disease. The functionalization of nanoparticles with RVG29 peptide was confirmed by NMR and FTIR. Their morphology was assessed by transmission electron microscopy and nanoparticles size, polydispersity and zeta potential were determined by dynamic light scattering. The in vitro validation tests were conducted in hCMEC/D3 cells, a human blood-brain barrier model and thioflavin T binding assay was conducted to assess the process of amyloid-beta peptide fibrillation typical of Alzheimer's disease. RVG29-nanoparticles displayed spherical morphology and size below 250\u00a0nm, which is compatible with brain applications. Zeta potential values were between -20 and -25\u00a0mV. Quercetin entrapment efficiency was generally higher than 80% and NLC nanoparticles were able to encapsulate up to 90%. The LDH assay showed that there is no cytotoxicity in hCMEC/D3 cell line and RVG29-nanoparticles clearly increased in 1.5-fold the permeability across the in vitro model of blood-brain barrier after 4\u00a0h of incubation compared with non-functionalized nanoparticles. Finally, this nanosystem was capable of inhibiting amyloid-beta aggregation in thioflavin T binding assay, suggesting its great potential for neuroprotection. RVG29-nanoparticles that simultaneously target the blood-brain barrier and induce neurons protection against amyloid-beta fibrillation proved to be an efficient way of quercetin delivery and a promising strategy for future approaches in Alzheimer's disease. Graphical Abstract.\n\nID: 26713267\nTitle: Cysteine proteases as therapeutic targets: does selectivity matter? A systematic review of calpain and cathepsin inhibitors.\nAbstract: Cysteine proteases continue to provide validated targets for treatment of human diseases. In neurodegenerative disorders, multiple cysteine proteases provide targets for enzyme inhibitors, notably caspases, calpains, and cathepsins. The reactive, active-site cysteine provides specificity for many inhibitor designs over other families of proteases, such as aspartate and serine; however, a) inhibitor strategies often use covalent enzyme modification, and b) obtaining selectivity within families of cysteine proteases and their isozymes is problematic. This review provides a general update on strategies for cysteine protease inhibitor design and a focus on cathepsin B and calpain 1 as drug targets for neurodegenerative disorders; the latter focus providing an interesting query for the contemporary assumptions that irreversible, covalent protein modification and low selectivity are anathema to therapeutic safety and efficacy.\n\nID: 8627316\nTitle: Polyamine modification increases the permeability of proteins at the blood-nerve and blood-brain barriers.\nAbstract: The permeability of the blood-nerve barrier (BNB) and the blood-brain barrier (BBB) to superoxide dismutase (SOD), insulin, albumin, and IgG in normal adult rats was quantified by measuring the permeability coefficient-surface area product (PS) with the intravenous bolus injection technique before and after covalent protein modification with naturally occurring polyamines-putrescine (PUT), spermidine (SPD), and spermine (SPM). The PS value of the BNB for PUT-SOD was 21.1-fold greater than the native SOD, and the PS values of the BBB for PUT-SOD ranged from 17.6-fold greater for the thalamus to 23.6-fold greater for the caudate-putamen compared with native SOD. In a similar manner, polyamine-modified insulin showed a 1.7-2.0-fold increase in PS of the BNB and BBB compared with the high values of native insulin. Polyamine-modified albumin showed a remarkable 54-165-fold increase in PS of the BNB and BBB compared with native albumin, whereas PUT-IgG resulted in an even higher increase in the PS that ranged from 111- to 349-fold for nerve and different brain regions compared with native IgG. Polyamine modification of proteins, therefore, can dramatically increase the permeability at the BNB and BBB of a variety of proteins with widely differing M(r) and function. It is surprising that the PS values of the BNB and BBB decreased with the increasing number of positive charges of the protonated amino groups on the polyamines (PUT>SPD>SPM). Although cationic proteins are known to interact with fixed anionic charges on the lumen of the microvascular endothelium, this observation of decreased permeability with increased positive charge distribution along the aliphatic carbon chain of the polyamines implies mechanisms other than simple electrostatic interaction involving charge density. It is suggested that the polyamine transporter may be responsible for the transport of these polyamine-modified proteins. Systemic administration of polyamine-modified peptides and proteins might prove to be an efficient approach to deliver therapeutic agents into the CNS and PNS for the treatment of a variety of neurological diseases.\n\nID: 22704880\nTitle: Selective electromembrane extraction at low voltages based on analyte polarity and charge.\nAbstract: Electromembrane extraction (EME) at low voltage (0-15 V) of 29 different basic model drug substances was investigated. The drug substances with logP<2.3 were not extracted at voltages less than 15 V. Extraction of drug substances with logP\u22652.3 and with two basic groups were also effectively suppressed by the SLM at voltages less than 15 V. Drug substances with logP\u22652.3 and with one basic group were all extracted at low voltages and with a strong compound selectivity which appeared to have some influence from the polar surface area of the compound. For this group of substances, recoveries varied between 0 and 23% at 5 V, whereas, recoveries varied between 5.5 and 51% at 15 V. Based on mass transfer differences related to charge, polarity, and polar surface, highly selective extractions of drug substances were demonstrated from human plasma, urine, and breast milk. An initial evaluation at low voltage (5 V) was compared with similar extractions at a more normal voltage level (50 V), and this supported that reliable data can be obtained under these low-voltage (mild) conditions by EME.\n\nID: 21981972\nTitle: Short timescale inkjet ink component diffusion: an active part of the absorption mechanism into inkjet coatings.\nAbstract: The structures of inkjet coatings commonly contain a high concentration of fine diameter pores together with a large pore volume capacity. To clarify the interactive role of the porous structure and the coincidentally occurring swelling of binder during inkjet ink vehicle imbibition, coating structures were studied in respect to their absorption behaviour for polar and non-polar liquid. The absorption measurement was performed using compressed pigment tablets, based on a range of pigment types and surface charge polarity, containing either polyvinyl alcohol (PVOH) or styrene acrylic latex (SA) as the binder, by recording the liquid uptake with a microbalance. The results indicate that, at the beginning of liquid uptake, at times less than 2 s, the small pores play the dominant role with respect to the inkjet ink vehicle imbibition. Simultaneously, water molecules diffuse into and within the hydrophilic PVOH binder causing binder swelling, which diminishes the number of active small pores and reduces the diameter of remaining pores, thus slowing the capillary flow as a function of time. The SA latex does not absorb the vehicle, and therefore the dominating phenomenon is then capillary absorption. However, the diffusion coefficient of the water vapour across separately prepared PVOH and SA latex films seems to be quite similar. In the PVOH, the polar liquid diffuses into the polymer network, whereas in the SA latex the hydrophobic nature prevents the diffusion into the polymer matrix and there exists surface diffusion. At longer timescale, permeation flow into the porous coating dominates as the resistive term controlling the capillary driven liquid imbibition rate.\n\nID: 19285675\nTitle: Peptide mapping with mobile phases of intermediate pH value using capillary reversed-phase high-performance liquid chromatography/electrospray ionisation tandem mass spectrometry.\nAbstract: This investigation describes the separation of tryptic peptides by capillary reversed-phase high-performance liquid chromatography (RP-HPLC) with eluents in the intermediate pH range, followed by in-line electrospray ionisation tandem mass spectrometry (ESI-MS/MS) analysis. For these purposes, gradient elution procedures with an aqueous eluent containing 20 mM ammonium formate, and an increasing content of acetonitrile or methanol, were employed. Compared to the analysis of the same tryptic peptides under low-pH conditions with an ion-pairing reagent, the increase in the pH with the 20 mM ammonium formate mobile phase led to significant changes in both peptide retention to the reversed-phase column and the collision-induced dissociation at the MS/MS stage as a consequence of the changes in the physico-chemical properties of these peptides, such as their overall charge, polarity and relative hydrophobicity. Thus, improved selectivity for the peptide separation and favourable tandem mass spectrometry analysis could be obtained with eluents in this intermediate pH range. The number of tryptic peptides identified by the new approach for the proteins investigated were significantly higher than that obtained by the conventional low-pH methods. Moreover, analysis of protein digests at very low concentrations was also performed under both acidic and intermediate pH conditions and similar improvements in selectivity and MS/MS detection limits were observed, i.e. identification of more distinct peptides and higher sequence coverage of the protein was obtained when eluents of intermediate pH were employed. This study therefore highlights the potential of conducting peptide mapping in the intermediate pH range to achieve more reliable and sensitive protein identifications with capillary RP-HPLC-ESI-MS/MS.\n=======================================================\n\n### [CUSTOM DATAPOINTS]\nCRITICAL EXTRACTION DIRECTIVE: You MUST extract the following custom datapoints as root-level key/value pairs inside your final JSON block:\n- \"suggested_experiments\": generate 1-3 suggested experiments\n- \"suggested_studies\": generate 1-3 suggested studies\n- \"swansons_literature_based_discovery_candidates\": You are an advanced Literature-Based Discovery (LBD) system executing Swanson\u2019s complementary-but-disjoint (A-B-C) model. Your goal is to find hidden, unpublished connections across the provided dataset. Strict Discovery Protocol: 1. Identify distinct, isolated sub-literatures (Domain A and Domain C) within the dataset that share NO direct citations, co-mentions, or common contextual paragraphs. 2. Find an intermediate biological mechanism, protein, path, or entity (Bridge B) that appears independently in both isolated domains (A-to-B and B-to-C). 3. Synthesize a novel, unstated hypothesis (A-to-C). Negative Constraint (Crucial): DO NOT output any connection if the relationship between Concept A and Concept C is explicitly mentioned, paired, or summarized anywhere in the source text. If a connection (like \"OMN resilience to SMN stabilization\") is already explicitly stated or grouped as a concept in the data, it is considered \"already known\" and must be disqualified. Format your output exactly as follows: - Discovered Hypothesis (A to C): [Clear, novel statement] - Literature A (Origin): [Entity/Concept and source context] - Literature C (Target): [Entity/Concept and source context] - The Intersecting Bridge B: [The shared mechanism/protein linking them] - Biological Rationale: [1-2 sentences explaining why this hidden connection is mechanistically plausible]\n- \"contradictions_between_evidences\": Identify conflicting evidence within the evidence set (if any) and flag the dispute here\n- \"repurposed_solutions\": identify and explain repurposed Solution potentials\n\n\nFormat Requirement:\nRAG AMNESIA IS ACTIVE: You must ONLY use the provided context literature. Do not use outside prior knowledge. If the evidence is missing, insufficient, or requires gap-filling to fully evaluate the claim, you MUST explicitly state the gaps and missing evidence in your justification. Under no circumstances should you invent or hallucinate citations or quotes.\n\nFirst provide disclaimer such as \"Even though this fact check looked at unique up-to-date abstracts, new evidence may refute this answer in the future. Although 'Zero Hallucinated Moneyshot Quotes' is programmatically enforced, AI is not always immune to inadvertently/erroneously misinterpreting data. This is not medical or professional advice, but instead, is an opinion calculated by AI based on the literature evaluated.\"\n---\nWrite in a highly academic, formal thesis tone.\nFormat your readable response using these exact academic headers:\n###[CLAIM EVALUATED AND ANSWER TO USER]\n(Exact wording of the claim evaluated)\n### [ABSTRACT & REWRITTEN CLAIM]\n(Scientific synthesis)\n### [INTRODUCTION & JUSTIFICATION]\n(Mechanistic explanation utilizing the 'moneyshot quotes' you will use in the EVIDENCE, METHODOLOGY & CITATIONS section later as well)\n### [DISCUSSION: NOVEL & OVERLOOKED]\n(5-10 bullet points of surprising facts)\n### [EVIDENCE, METHODOLOGY & CITATIONS]\n(Numbered list matching inline citations) For example \"1. ID: 12345 - Application: The text discusses ... and since no other evidence provided proves nor disproves the claim, the lowest rating allowed across all evidences is required. ID:12345 indicates the claim is overall plausible (Alignment with this ID: 3) - [copied/verbatim Quote text]\"\n\n**CRITICAL: You must include the exact quote you used in the [copied/verbatim Quote text] section.\n\nIf the prompt says \"at least 20 quotes\" then there must be at least 20 matching citations. You must actually use the quotes you select within the conext of the preprint publication you write.\n\nEvaluation Schema:\nRAG AMNESIA IS ACTIVE: You must ONLY use the provided context literature. Do not use outside prior knowledge. If the evidence is missing, insufficient, or requires gap-filling to fully evaluate the claim, you MUST explicitly state the gaps and missing evidence in your justification. Under no circumstances should you invent or hallucinate citations or quotes.\n\n###critical: WRAP YOUR THOUGHTS WITH ... tags at the very beginning of your response.\n\n=======================================================\nCONTEXT LITERATURE (STATIC CACHE):\nID: 41961384\nTitle: Spermidine Attenuates Neuroimmune Dysfunction in Gulf War Illness via Modulation of the Gut- Brain Axis.\nAbstract: Gulf War illness (GWI) affects nearly one-third of US veterans deployed during the 1990-1991 Gulf War (GW) and is characterized by chronic fatigue, neuroinflammation, and gut dysbiosis. Through comprehensive fecal metabolomics sequencing, our lab previously reported the depletion of beneficial metabolites including spermidine in the preclinical GWI mouse model. Spermidine is an endogenously synthesized polyamine known for its anti-inflammatory and mucosal barrier protective effects in various pathological diseases. Given its established role in mitigating intestinal inflammation and maintaining homeostasis, this study investigated the therapeutic potential of spermidine in a persistent (22\u00a0weeks) GWI mouse model, with a specific focus on gut-brain axis regulation. Our results demonstrated that spermidine effectively restored both microbial richness and diversity by selectively enriching beneficial bacterial taxa and suppressing growth of opportunistic pathogens, which are otherwise dysregulated following exposure to GW chemicals. Spermidine treatment also improved gut epithelial barrier integrity and reduced epithelial release of high-mobility group box 1 (HMGB1) into systemic circulation. Recent studies on GWI have implicated a critical role of gut-derived damage-associated molecular patterns (DAMPs), particularly HMGB1 in mediating neuroinflammation. Our findings indicate that systemic levels of HMGB1 critically influence the extent of blood-brain barrier (BBB) disruption and subsequent microglial activation. Mechanistically, spermidine activated intestinal aryl hydrocarbon receptor (AhR)/nuclear factor erythroid 2-related factor 2 (Nrf2)/heme oxygenase-1 (HO-1) signaling, which played a role in limiting intestinal HMGB1 release and suppressing downstream receptor for advanced glycation end-product (RAGE)-mediated microglial activation in the brain. In vitro results indicate spermidine promoted AhR/Nrf2 nuclear translocation which reduced LPS-induced HMGB1 release from primary intestinal epithelial cells (IECs), effects abrogated by AhR inhibition. Additionally, we observed that HMGB1 directly induces microglial activation via RAGE receptors in immortalized microglial (IMG) cell lines in a dose-dependent manner. These results demonstrate that spermidine decreases neuroinflammation by modulating gut-brain axis pathophysiology associated with GWI. Together, this study demonstrates the therapeutic role of spermidine in ameliorating systemic and neurological disturbances in GWI.\n\nID: 41177462\nTitle: Nasal-to-brain siRNA delivery based on trace amine associated receptor for improving cognitive function.\nAbstract: Gene-based therapies for central nervous system (CNS) disorders face substantial challenges in overcoming the blood-brain barrier (BBB) to effectively target brain tissues. The nasal-to-brain delivery route has gained increasing attention as it bypasses the BBB, facilitating faster drug delivery to the lesion site while minimizing systemic side effects. Here, we developed a nasal-to-brain delivery system to administer small interfering RNA (siRNA) for the treatment of radiation-induced brain injury (RBI). RNA sequencing revealed that the p53 signaling pathway was predominantly enriched in the hippocampus, with significant upregulation of Alox12B expression in RBI mice. To improve the delivery of siRNA targeting Alox12B, we engineered spermidine-modified ginseng-derived extracellular vesicles (S-GEVs) nanoparticles, termed S-GEVs@siRNA. These nanoparticles leveraged the targeting capabilities of spermidine for olfactory receptor-trace amine associated receptor (TAAR), enhancing siRNA delivery and therapeutic efficacy. After intranasal administration, the nanoparticles were efficiently internalized by olfactory receptor neurons (ORNs) via the olfactory nerve pathway. The nanoparticles then escaped lysosomes, releasing siRNA into the cytoplasm, leading to gene downregulation and therapeutic benefits. Our results demonstrated that the designed nanoparticles were absorbed by the ORNs labeled with the Olfactory Marker Protein (OMP) and TAAR5 and successfully entered the olfactory bulb and the brain. Treatment with these nanoparticles significantly reduced p53-mediated neuronal ferroptosis and improved synaptic function both in vitro and in vivo. In conclusion, S-GEVs@siRNA nanoparticles rapidly reached the olfactory bulb through TAAR-mediated endocytosis, entered hippocampal neurons, downregulated Alox12B expression, exerted neuroprotective effects, and alleviated RBI-induced cognitive dysfunction. The designed nasal-to-brain delivery system holds great promise for treating various CNS diseases.\n\nID: 37509150\nTitle: As Verified with the Aid of Biotinylated Spermine, the Brain Cannot Take up Polyamines from the Bloodstream Leaving It Solely Dependent on Local Biosynthesis.\nAbstract: The importance of polyamines (PAs) for the central nervous system (CNS) is well known. Less clear, however, is where PAs in the brain are derived from. Principally, there are three possibilities: (i) intake by nutrition, release into the bloodstream, and subsequent uptake from CNS capillaries, (ii) production by parenchymatous organs, such as the liver, and again uptake from CNS capillaries, and (iii) uptake of precursors, such as arginine, from the blood and subsequent local biosynthesis of PAs within the CNS. The present investigation aimed to unequivocally answer the question of whether PAs, especially the higher ones like spermidine (SPD) and spermine (SPM), can or cannot be taken up into the brain from the bloodstream. For this purpose, a biotin-labelled analogue of spermine (B-X-SPM) was synthesized, characterized, and used to visualize its uptake into brain cells following application to acute brain slices, to the intraventricular space, or to the bloodstream. In acute brain slices there is strong uptake of B-X-SPM into protoplasmic and none in fibrous-type astrocytes. It is also taken up by neurons but to a lesser degree. Under in vivo conditions, astrocyte uptake of B-X-SPM from the brain interstitial fluid is also intense after intraventricular application. In contrast, following intracardial injection, there is no uptake from the bloodstream, indicating that the brain is completely dependent on the local synthesis of polyamines.\n\nID: 33852843\nTitle: Dietary spermidine improves cognitive function.\nAbstract: Decreased cognitive performance is a hallmark of brain aging, but the underlying mechanisms and potential therapeutic avenues remain poorly understood. Recent studies have revealed health-protective and lifespan-extending effects of dietary spermidine, a natural autophagy-promoting polyamine. Here, we show that dietary spermidine passes the blood-brain barrier in mice and increases hippocampal eIF5A hypusination and mitochondrial function. Spermidine feeding in aged mice affects behavior in homecage environment tasks, improves spatial learning, and increases hippocampal respiratory competence. In a Drosophila aging model, spermidine boosts mitochondrial respiratory capacity, an effect that requires the autophagy regulator Atg7\u00a0and the mitophagy mediators Parkin and Pink1. Neuron-specific Pink1 knockdown abolishes spermidine-induced improvement of olfactory associative learning. This suggests that the maintenance of mitochondrial and autophagic function is essential for enhanced cognition by spermidine feeding. Finally, we show large-scale prospective data linking higher dietary spermidine intake with a reduced risk for cognitive impairment in humans.\n\nID: 31509751\nTitle: Intranasal Targeting of Hypothalamic PTP1B and TCPTP Reinstates Leptin and Insulin Sensitivity and Promotes Weight Loss in Obesity.\nAbstract: The importance of hypothalamic leptin and insulin resistance in the development and maintenance of obesity remains unclear. The tyrosine phosphatases protein tyrosine phosphatase 1B (PTP1B) and T\u00a0cell protein tyrosine phosphatase (TCPTP) attenuate leptin and insulin signaling and are elevated in the hypothalami of obese mice. We report that elevated PTP1B and TCPTP antagonize hypothalamic leptin and insulin signaling and contribute to the maintenance of obesity. Deletion of PTP1B and TCPTP in the hypothalami of obese mice enhances CNS leptin and insulin sensitivity, represses feeding, and increases browning, to decrease adiposity and improve glucose metabolism. The daily intranasal administration of a PTP1B inhibitor, plus the glucocorticoid antagonist RU486 that decreases TCPTP expression, represses feeding, increases browning, promotes weight loss, and improves glucose metabolism in obese mice. Our findings causally link heightened hypothalamic PTP1B and TCPTP with leptin and insulin resistance and the maintenance of obesity and define a viable pharmacological approach by which to promote weight loss in obesity.\n\nID: 29953201\nTitle: Multistep Inhibition of \u03b1-Synuclein Aggregation and Toxicity in Vitro and in Vivo by Trodusquemine.\nAbstract: The aggregation of \u03b1-synuclein, an intrinsically disordered protein that is highly abundant in neurons, is closely associated with the onset and progression of Parkinson's disease. We have shown previously that the aminosterol squalamine can inhibit the lipid induced initiation process in the aggregation of \u03b1-synuclein, and we report here that the related compound trodusquemine is capable of inhibiting not only this process but also the fibril-dependent secondary pathways in the aggregation reaction. We further demonstrate that trodusquemine can effectively suppress the toxicity of \u03b1-synuclein oligomers in neuronal cells, and that its administration, even after the initial growth phase, leads to a dramatic reduction in the number of \u03b1-synuclein inclusions in a Caenorhabditis elegans model of Parkinson's disease, eliminates the related muscle paralysis, and increases lifespan. On the basis of these findings, we show that trodusquemine is able to inhibit multiple events in the aggregation process of \u03b1-synuclein and hence to provide important information about the link between such events and neurodegeneration, as it is initiated and progresses. Particularly in the light of the previously reported ability of trodusquemine to cross the blood-brain barrier and to promote tissue regeneration, the present results suggest that this compound has the potential to be an important therapeutic candidate for Parkinson's disease and related disorders.\n\nID: 28867747\nTitle: Involvement of Carrier-Mediated Transport at the Blood-Cerebrospinal Fluid Barrier in Spermine Clearance from Rat Brain.\nAbstract: Spermine is the end-product in the polyamine biosynthetic pathway, and its excess accumulation induces neuroexcitatory responses and neurotoxicity. The purpose of this study was to elucidate the involvement of transport systems at the brain barriers in the clearance of spermine. In vivo rat spermine elimination from brain parenchyma across the blood-brain barrier (BBB) and blood-cerebrospinal fluid (CSF) barrier (BCSFB) was assessed by intracerebral and intracerebroventricular administration techniques, respectively. To characterize spermine transport at the BCSFB, a transport study using rat choroid plexus was performed. After the intracerebral microinjection of [3H]spermine, no time-dependent decrease in [3H]spermine in the ipsilateral cerebrum was observed, suggesting the low contribution of the BBB to spermine clearance from the brain. In contrast, the [3H]spermine concentration in the CSF after intracerebroventricular administration was time-dependently decreased with an elimination rate constant of 0.352\u2009min-1, and the elimination clearance of [3H]spermine was 6.6-fold greater than that of [14C]D-mannitol, reflecting bulk flow of the CSF. This [3H]spermine elimination was attenuated by co-administration of unlabeled excess spermine, indicating carrier-mediated elimination of spermine from the CSF. [3H]Spermine transport into the choroid plexus was strongly inhibited by unlabeled spermine, other polyamines (spermidine and putrescine), and organic cation transporter substrates such as corticosterone and 1-methyl-4-phenylpyridinium. However, other substrates/inhibitors for organic cation transporters (decynium-22 and tetraethylammonium) had little effect. Consequently, our study indicates that transporting molecules at the BCSFB, distinct from typical organic cation transporters, are involved in spermine clearance from the CSF.\n\nID: 28639394\nTitle: Transferrin-conjugated magnetic dextran-spermine nanoparticles for targeted drug transport across blood-brain barrier.\nAbstract: Application of many vital hydrophilic medicines have been restricted by blood-brain barrier (BBB) for treatment of brain diseases. In this study, a targeted drug delivery system based on dextran-spermine biopolymer was developed for drug transport across BBB. Drug loaded magnetic dextran-spermine nanoparticles (DS-NPs) were prepared via ionic gelation followed by transferrin (Tf) conjugation as targeting moiety. The characteristics of Tf conjugated nanoparticles (TDS-NPs) were analyzed by different methods and their cytotoxicity effects on U87MG cells were tested. The superparamagnetic characteristic of TDS-NPs was verified by vibration simple magnetometer. Capecitabine loaded TDS-NPs exhibited pH-sensitive release behavior with enhanced cytotoxicity against U87MG cells, compared to DS-NPs and free capecitabine. Prussian-blue staining and TEM-imaging showed the significant cellular uptake of TDS-NPs. Furthermore, a remarkable increase of Fe concentrations in brain was observed following their biodistribution and histological studies in vivo, after 1 and 7 days of post-injection. Enhanced drug transport across BBB and pH-triggered cellular uptake of TDS-NPs indicated that these theranostic nanocarriers are promising candidate for the brain malignance treatment. \u00a9 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 2851-2864, 2017.\n\nID: 26405039\nTitle: Role of a Hydrophobic Pocket in Polyamine Interactions with the Polyspecific Organic Cation Transporter OCT3.\nAbstract: Organic cation transporter 3 (OCT3, SLC22A3) is a polyspecific, facilitative transporter expressed in astrocytes and in placental, intestinal, and blood-brain barrier epithelia, and thus elucidating the molecular mechanisms underlying OCT3 substrate recognition is critical for the rational design of drugs targeting these tissues. The pharmacology of OCT3 is distinct from that of other OCTs, and here we investigated the role of a hydrophobic cavity tucked within the translocation pathway in OCT3 transport properties. Replacement of an absolutely conserved Asp by charge reversal (D478E), neutralization (D478N), or even exchange (D478E) abolished MPP(+) uptake, demonstrating this residue to be obligatory for OCT3-mediated transport. Mutations at non-conserved residues lining the putative binding pocket of OCT3 to the corresponding residue in OCT1 (L166F, F450L, and E451Q) reduced the rate of MPP(+) transport, but recapitulated the higher sensitivity pharmacological profile of OCT1. Thus, interactions of natural polyamines (putrescine, spermidine, spermine) and polyamine-like potent OCT1 blockers (1,10-diaminodecane, decamethonium, bistriethylaminodecane, and 1,10-bisquinuclidinedecane) with wild-type OCT3 were weak, but were significantly potentiated in the mutant OCT3s. Conversely, a reciprocal mutation in OCT1 (F161L) shifted the polyamine-sensitivity phenotype toward that of OCT3. Further analysis indicated that OCT1 and OCT3 can recognize essentially the same substrates, but the strength of substrate-transporter interactions is weaker in OCT3, as informed by the distinct makeup of the hydrophobic cleft. The residues identified here are key contributors to both the observed differences between OCT3 and OCT1 and to the mechanisms of substrate recognition by OCTs in general.\n\nID: 25824983\nTitle: Hydroxycinnamic acid amide derivatives of polyamines reverse spermine-induced CNS excitation.\nAbstract: The aim of this study was to examine the acute effect of a range of novel hydroxycinnamic acid derivatives of spermine on the development of spermine-induced CNS excitation and convulsions in female Laca mice, and to assess the chronic adverse behavioural effect profile of these compounds over a 5day period. Four of the six novel polyamine analogues dose-dependently inhibited body tremor and tonic convulsions caused by spermine, when administered centrally (icv) or peripherally (ip). BU43b was the most potent analogue tested, but BU31b, 33b, and 36b were also effective (p<0.01, Mann-Whitney U test). A similar profile of effectiveness was seen with peripheral and central administration, indicating that the analogues may cross the blood brain barrier. More chronic investigation of the adverse effects of the compounds administered alone over 5days of observation indicated that the drugs were well tolerated, only causing reduced locomotor activity on the first day of the study and mild changes in behaviours linked to CNS and ANS function. It is likely that NMDA receptor NR2B subunit inhibition is involved in the anticonvulsant effect of these novel analogues, but other mechanisms may also be involved. These novel polyamine derivatives warrant further investigation of their therapeutic potential in treating epilepsy and CNS disorders where excitotoxicity is implicated.\n\nID: 25623533\nTitle: Functional properties of Claramine: a novel PTP1B inhibitor and insulin-mimetic compound.\nAbstract: Protein tyrosine phosphatase 1B (PTP1B) inhibits insulin signaling, interfering with its control of glucose homeostasis and metabolism. PTP1B activity is elevated in obesity and type 2 diabetes and is a major cause of insulin resistance. Trodusquemine (MSI-1436) is a \"first-in-class\" highly selective inhibitor of PTP1B that can cross the blood-brain barrier to suppress feeding and promote insulin sensitivity and glycemic control. Trodusquemine is a naturally occurring cholestane that can be purified from the liver of the dogfish shark, Squalus acanthias, but it can also be manufactured synthetically by a fairly laborious process that requires several weeks. Here, we tested a novel easily and rapidly (2 days) synthesized polyaminosteroid derivative (Claramine) containing a spermino group similar to Trodusquemine for its ability to inhibit PTP1B. Like Trodusquemine, Claramine displayed selective inhibition of PTP1B but not its closest related phosphatase TC-PTP. In cultured neuronal cells, Claramine and Trodusquemine both activated key components of insulin signaling, with increased phosphorylation of insulin receptor-\u03b2 (IR\u03b2), Akt and GSK3\u03b2. Intraperitoneal administration of Claramine or Trodusquemine effectively restored glycemic control in diabetic mice as determined by glucose and insulin tolerance tests. A single intraperitoneal dose of Claramine, like an equivalent dose of Trodusquemine, suppressed feeding and caused weight loss without increasing energy expenditure. In summary, Claramine is an alternative more easily manufactured compound for the treatment of type II diabetes.\n\nID: 21251935\nTitle: Gene delivery by pullulan derivatives in brain capillary endothelial cells for protein secretion.\nAbstract: The blood-brain barrier (BBB) formed by brain capillary endothelial cells protects the brain against potentially harmful substances present in the circulation, but also restricts exogenous substances such as pharmacologically acting drugs or proteins from entering the brain. A novel and rather unchallenged approach to allow proteins to enter the brain is gene therapy based on delivery of genetic material into brain capillary endothelial cells. In theory in vivo transfection will allow protein expression and secretion from brain capillary endothelial cells and further into the brain. This would denote a new paradigm for therapy to transport proteins across the BBB. The aim of this study was to investigate the possibility to use brain capillary endothelial cells as factories for recombinant protein production. Non-viral gene carriers were prepared from pullulan, a polysaccharide, and spermine, a naturally occurring polyamine that were additionally conjugated with plasmid DNA. We were able to transfect rat brain endothelial cells (RBE4s) and human brain microvascular endothelial cells (HBMECs). Transfection of HBMECs with pullulan-spermine conjugated with plasmid DNA bearing cDNA encoding human growth hormone 1 (hGH1), led to secretion of hGH1 protein into the growth medium. Hence, the pullulan-spermine delivery system is a very promising method for delivering DNA to brain endothelial cells with potential for using these cells as factories for secretion of proteins.\n\nID: 16565169\nTitle: Physiological and biophysical factors that influence Alzheimer's disease amyloid plaque targeting of native and putrescine modified human amyloid beta40.\nAbstract: Amyloid beta40 (Abeta40) and its derivatives are being developed as probes for the ante-mortem diagnosis of Alzheimer's disease. Putrescine-Abeta40 (PUT-Abeta40) showed better plaque targeting than the native Abeta40, which was not solely explained by the differences in their blood-brain-barrier (BBB) permeabilities. The objective of this study was to elucidate the physiological and biophysical factors influencing the differential targeting of Abeta40 and PUT-Abeta40. Despite better plaque-targeting ability 125I-PUT-Abeta40 was more rapidly cleared from the systemic circulation than amyloid beta40 labeled with 125I (125I-Abeta40) after i.v. administration in mice. The BBB permeability of both compounds was inhibited by circulating peripheral Abeta40 levels. 125I-Abeta40 but not 125I-PUT-Abeta40 was actively taken up by the mouse brain slices in vitro. Only fluorescein-Abeta40, not fluorescein-PUT-Abeta40, was localized in the brain parenchymal cells in vitro. The metabolism of 125I-Abeta40 in the brain slices was twice as great as 125I-PUT-Abeta40. 125I-Abeta40 efflux from the brain slices was saturable and found to be 5 times greater than that of 125I-PUT-Abeta40. Thioflavin-T fibrillogenesis assay demonstrated that PUT-Abeta40 has a greater propensity to form insoluble fibrils compared with Abeta40, most likely due to the ability of PUT-Abeta40 to form beta sheet structure more readily than Abeta40. These results demonstrate that the inadequate plaque targeting of Abeta40 is due to cellular uptake, metabolism, and efflux from the brain parenchyma. Despite better plaque targeting of PUTAbeta40, its propensity to form fibrils may render it less suitable for human use and thus allow increased focus on the development of novel derivatives of Abeta with improved characteristics.\n\nID: 16529872\nTitle: In vivo protein transduction to the CNS.\nAbstract: Proteins and peptides are useful research and therapeutic tools, however applications are limited because delivery to the desired location is not easily achievable. There are two hurdles in protein/peptide delivery to the brain: the blood-brain barrier and intracellular penetration. Penetration to both brain and the intracellular space can be achieved by adjusting hydrophilicity, and small molecule pharmacological agents have been successfully developed using this approach. But with proteins and peptides, it is difficult to modify the hydrophilicity without influencing biological functions. Trans-acting factor protein from the human immunodeficiency virus contains a highly conserved cationic peptide sequence necessary for transduction across the cell membrane. While trans-acting factor peptide has been used for in vitro protein transduction, its in vivo application is very limited because it is rapidly degraded by proteolysis. Polyethylenimine is a chemically synthesized small molecule cationization agent; the charge density is greater than a peptide-based cationic cluster such as trans-acting factor, and it is resistant to proteolysis in vivo. We first tested intracellular protein transduction following direct brain injection in mice using polyethylenimine-conjugated green fluorescence protein and beta-galactosidase (molecular weights 29 and 540 kDa, respectively). Polyethylenimine-conjugates penetrated to the intracellular space immediately surrounding the injection site within one hour. We further tested polyethylenimine-mediated protein transduction following intranasal administration, which bypasses the blood-brain barrier. Polyethylenimine-conjugates in pH 7.5 solution did not reach the brain, probably because the polyethylenimine-conjugates penetrated into the intracellular space where first exposed to the tissue, i.e. at the nasal mucosae. We temporarily reduced the electrostatic interaction between cationized polyethylenimine-conjugates and cellular surfaces by adjusting the pH to 4.5; solution rapidly reached the brain and penetrated to the intracellular space. This study suggests that polyethylenimine is a useful protein transduction agent in the brain in vivo, and adjusting cationic charge interaction can determine the extent of brain penetration.\n\nID: 12505424\nTitle: Molecular targeting of Alzheimer's amyloid plaques for contrast-enhanced magnetic resonance imaging.\nAbstract: Smart molecular probes for both diagnostic and therapeutic purposes are expected to provide significant advances in clinical medicine and biomedical research. We describe such a probe that targets beta-amyloid plaques of Alzheimer's disease and is detectable by magnetic resonance imaging (MRI) because of contrast imparted by gadolinium labeling. Three properties essential for contrast enhancement of beta-amyloid plaques on MRI exist in this smart molecular probe, putrescine-gadolinium-amyloid-beta peptide: (1) transport across the blood-brain barrier following intravenous injection conferred by the polyamine moiety, (2) binding to plaques with molecular specificity by putrescine-amyloid-beta, and (3) magnetic resonance imaging contrast by gadolinium. MRI was performed on ex vivo tissue specimens at 7 T at a spatial resolution approximating plaque size (62.5 microm(3)), in order to prove the concept that the probe, when administered intravenously, can selectively enhance plaques. The plaque-to-background tissue contrast-to-noise ratio, which was precisely correlated with histologically stained plaques, was enhanced more than nine-fold in regions of cortex and hippocampus following intravenous administration of this probe in AD transgenic mice. Continuing engineering efforts to improve spatial resolution are underway in MRI, which may enable in vivo imaging at the resolution of individual plaques with this or similar contrast probes. This could enable early diagnosis and also provide a direct measure of the efficacy of anti-amyloid therapies currently being developed.\n\nID: 12135777\nTitle: Passage of spermidine across the blood-brain barrier in short recirculation periods following global cerebral ischemia: effects of mild hyperthermia.\nAbstract: Transport of a polyamine (PA), spermidine (SPMD) into rat brain at various early postischemic periods was studied. Rats underwent 20 min of four-vessel occlusion (4VO) followed by 5, 10, 30 and 60 min of recirculation (RC) periods with natural brain temperature. 3H-aminoisobutyricacid (AIB) and 14C-SPMD were utilised to search dual functions of the blood-brain barrier (BBB); barrier and carrier functions, respectively. Unidirectional blood-to-brain transfer constant (Kin) was calculated for AIB and SPMD in four brain regions-parieto-temporal cortex, striatum, hippocampus and cerebellum. Kin for SPMD ranged between 1.2+/-0.3 x 10(3) ml g(-1) min(-1) (for striatum) and 2.2+/-0.4 x 10(3) ml g(-1) min(-1) (for cerebellum) in controls. Kin for AIB showed similar values. At 5 and 10 min RC periods, Kin for both substances increased in a non-specific manner in all brain regions studied. In the cortex, Kin for SPMD at 5 and 10 min RC periods were 3.2+/-0.4 x 10(3) and 2.9+/-0.3 x 10(3) ml g(-1) min(-1), respectively, and found to be maximum with respect to other brain regions studied. 30 and 60 min RC groups showed specific transport for SPMD, whilst there were no changes for Kin for AIB, in all brain regions studied. Hippocampus showed the maximum increase in Kin SPMD at 60 min RC (2.7+/-0.3 x 10(3) ml g(-1) min(-1)), corresponding to a percentage rise of 121%. Intraischemic mild brain hyperthermia (39 degrees C) gave rise to a striking increase in Kin at 60 min postischemia for both substances. These results suggest that there is a specific transport of SPMD into brain at 30 and 60 min RC periods following 20 min of forebrain ischemia. Moreover, dual functions of the BBB were perturbed with intracerebral mild hyperthermia during ischemia.\n\nID: 11430870\nTitle: Carrier-mediated processes in blood--brain barrier penetration and neural uptake of paraquat.\nAbstract: Due to the structural similarity to N-methyl-4-phenyl pyridinium (MPP(+)), paraquat might induce dopaminergic toxicity in the brain. However, its blood--brain barrier (BBB) penetration has not been well documented. We studied the manner of BBB penetration and neural cell uptake of paraquat using a brain microdialysis technique with HPLC/UV detection in rats. After subcutaneous administration, paraquat appeared dose-dependently in the dialysate. In contrast, MPP(+) could not penetrate the BBB in either control or paraquat pre-treated rats. These data indicated that the penetration of paraquat into the brain would be mediated by a specific carrier process, not resulting from the destruction of BBB function by paraquat itself or a paraquat radical. To examine whether paraquat was carried across the BBB by a certain amino acid transporter, L-valine or L-lysine was pre-administered as a co-substrate. The pre-treatment of L-valine, which is a high affinity substrate for the neutral amino acid transporter, markedly reduced the BBB penetration of paraquat. When paraquat was administered to the striatum through a microdialysis probe, a significant amount of paraquat was detected in the striatal cells after a sequential 180-min washout with Ringer's solution. This uptake was significantly inhibited by a low Na(+) condition, but not by treatment with putrescine, a potent uptake inhibitor of paraquat into lung tissue. These findings indicated that paraquat is possibly taken up into the brain by the neutral amino acid transport system, then transported into striatal, possibly neuronal, cells in a Na(+)-dependent manner.\n\nID: 11117554\nTitle: Therapeutic benefit of polyamine-modified catalase as a scavenger of hydrogen peroxide and nitric oxide in familial amyotrophic lateral sclerosis transgenics.\nAbstract: Continuous subcutaneous administration of polyamine-modified catalase that has increased permeability at the blood-brain barrier showed both a highly significant delay in onset and an increase in survival in a transgenic mouse model of familial amyotrophic lateral sclerosis having a point mutation in the gene encoding copper/zinc superoxide dismutase. These results suggest that hydrogen peroxide-mediated oxidative stress with subsequent free radical damage involving nitric oxide and possibly hydroxyl radicals in motor neurons may be the culprit in familial amyotrophic lateral sclerosis.\n\nID: 10713383\nTitle: Exogenous spermine reduces ischemic damage in a model of focal cerebral ischemia in the rat.\nAbstract: Alterations in polyamine metabolism during and after global or focal cerebral ischemia can produce a multiplicity of effects on brain such as modification in mitochondria calcium buffering capacity, exacerbating glutamate-mediated neurotoxicity, and impairment of the blood-brain barrier. In this study, the endogenous polyamine spermine was administered intravenously 30 min prior to temporary focal cerebral ischemia in rats induced by clipping of the left middle cerebral and bilateral common carotid arteries for 3 h. Three days after removal of the microclips, intracardiac perfusion with 2% 2,3,5-triphenyl tetrazolium chloride was performed. Coronal slices were cut, photographed, and examined for cortical infarct volume. Spermine reduced infarct volume in a dose-dependent fashion. This study demonstrates that the use of polyamines may be considered as a powerful tool in prevention of ischemic tissue damage following focal cerebral ischemia.\n\nID: 10693942\nTitle: Elevated N1-acetylspermidine levels in gerbil and rat brains after CNS injury.\nAbstract: The polyamine system is very sensitive to different pathological states of the brain and is perturbed after CNS injury. The main modifications are significant increases in ornithine decarboxylase activity and an increase in tissue putrescine levels. Previously we have shown that the specific polyamine oxidase (PAO) inhibitor N1,N4-bis(2,3-butadienyl)-1,4-butanediamine (MDL 72527) reduced the tissue putrescine levels, edema, and infarct volume after transient focal cerebral ischemia in spontaneously hypertensive rats and traumatic brain injury of Sprague-Dawley rats. In the present study, N1-acetyl-spermidine accumulation was greater in injured brain regions compared with sham or contralateral regions following inhibition of PAO by MDL 72527. This indicates spermidine/spermine-N1-acetyltransferase (SSAT) activation after CNS injury. The observed increase in N1-acetylspermidine levels at 1 day after CNS trauma paralleled the decrease in putrescine levels after treatment with MDL 72527. This suggests that the increased putrescine formation at 1 day after CNS injury is mediated by the SSAT/PAO pathway, consistent with increased SSAT mRNA after transient ischemia.\n\nID: 10486188\nTitle: Therapeutic benefits of putrescine-modified catalase in a transgenic mouse model of familial amyotrophic lateral sclerosis.\nAbstract: Dominant mutations in the copper/zinc superoxide dismutase (SOD1) gene have been observed in 15-20% of familial amyotrophic lateral sclerosis (FALS) cases. The mechanism by which SOD1 mutations result in motor neuron degeneration in FALS mice partly involves oxidative damage and an increased peroxidase activity of the mutant SOD1. A new therapeutic approach designed to eliminate the substrate of this peroxidase activity was examined in two lines of transgenic mice expressing the FALS-linked mutation glycine to alanine (G93A). We investigated the ability of putrescine-modified catalase (PUT-CAT), an antioxidant enzyme that removes hydrogen peroxide and has increased permeability at the blood-brain barrier, to modify the time course of the SOD1 mutation-induced motor neuron disease in these FALS mice. Continuous, subcutaneous administration of PUT-CAT significantly delayed the age at which onset of clinical disease occurred (indicated by loss of splay and/or tremors of hindlimbs) in a high-expressor line of FALS transgenic mice. Intraperitoneal injection of PUT-CAT given two times per week also significantly delayed the onset of clinical disease in a low-expressor line of FALS mice. PUT-CAT also significantly delayed the age at which clinical weakness developed (quantified by measuring the shortening of stride length) in both lines of FALS animals. No significant changes were observed in the survival times of the high-expressor FALS mice in any of the treatment groups. However, a trend toward a prolongation of survival was observed in the PUT-CAT-treated low-expressor FALS mice. These results support the role of free radical-mediated damage in the cascade of events leading to motor neurodegeneration in FALS and indicate that PUT-CAT interacts with a critical step in this cascade to delay the onset of clinical disease as well as the development of clinical weakness in FALS transgenic mice.\n\nID: 10486187\nTitle: Plasma pharmacokinetics, nervous system biodistribution and biostability, and spinal cord permeability at the blood-brain barrier of putrescine-modified catalase in the adult rat.\nAbstract: Free radical-mediated oxidative damage has been proposed to be an underlying mechanism in several neurodegenerative disorders. Previous investigations in our laboratory have shown that putrescine-modified catalase (PUT-CAT) has increased permeability at the blood-brain (BBB) and blood-nerve barriers with retained enzymatic activity after parenteral administration when compared to native catalase (CAT). The goals of the present study were to examine the plasma stability, spinal cord BBB permeability, nervous system biodistribution, and spinal cord enzyme activity of CAT and PUT-CAT after parenteral administration in the adult rat. TCA precipitation and chromatographic analyses revealed that CAT and PUT-CAT were found intact in the plasma and in the central nervous system (CNS) after iv, ip, or sc bolus injections. The highest percentages of intact CAT or PUT-CAT proteins were found in the plasma after iv administration, and similar percentages of intact CAT or PUT-CAT were found in the CNS following all three types of administration. Increases of 2.4- to 4.7-fold in permeability at the BBB and similar increases in the levels of intact PUT-CAT were found in different brain regions compared to the levels of CAT. A 2.4-fold higher level of intact PUT-CAT compared to that of CAT (P < 0.05) was found in the spinal cord 60 min after a sc bolus injection. CAT enzyme activity in the spinal cord was 50% higher (P < 0.05) in rats treated with PUT-CAT continuously for 1 week by subcutaneously implanted, osmotic pumps than the activity found in rats treated with PBS. These results provide evidence that intact, enzymatically active PUT-CAT is efficiently delivered to the nervous system following iv, ip, and sc administration and suggest that sc administration of PUT-CAT may be effective in treating neurodegenerative disorders in which the underlying mechanisms involve the action of free radicals and oxidative damage.\n\nID: 9853704\nTitle: Simultaneous assay of ornithine decarboxylase and polyamines after central nervous system injury in gerbil and rat.\nAbstract: Ornithine decarboxylase (ODC) is considered the rate-limiting enzyme in polyamine biosynthesis. An increase in putrescine (a natural polyamine) synthesis after central nervous system (CNS) injury appears to be involved in blood-brain barrier dysfunction, development of vasogenic edema and neuronal death. An improved method is described to determine the ODC activity as well as polyamine levels from the same brain tissue. The polyamine results showed no significant differences from data obtained with the conventional assay. The advantages of this method are to: (1) minimize the number of animals needed for the study, and (2) eliminate any internal inconsistencies resulting from use of two independent groups of animals for ODC and polyamine measurements. Using this method, ODC activities and polyamine levels were measured in cortices and hippocampi from global transient ischemia of gerbils and traumatic brain injury (TBI) of rats.\n\nID: 9751199\nTitle: Putrescine-modified nerve growth factor: bioactivity, plasma pharmacokinetics, blood-brain/nerve barrier permeability, and nervous system biodistribution.\nAbstract: Previous investigations from our laboratory have demonstrated that the covalent modification of a variety of proteins, including antioxidant enzymes, with the naturally occurring polyamines--putrescine (PUT), spermidine, and spermine--dramatically increases their permeability coefficient-surface area product (PS) at the blood-brain and blood-nerve barriers after parenteral administration. In the present study, we have covalently modified nerve growth factor (NGF) with PUT by targeting carboxylic groups for their graded modification by controlling the ionization of these groups with pH. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis, western, and isoelectric focusing analyses demonstrated conversion of NGF to its polyamine-modified derivatives at different pH values. Although the immunoreactivity of PUT-NGF determined by ELISA and western analysis decreased with decreasing pH, the biological activity of PUT-NGF was not affected at any pH as determined by survival and neurite extension of dorsal root ganglia and PC12 cultures. Plasma pharmacokinetics after a single intravenous bolus administration revealed intact PUT-NGF through 10 min and 73-82% intact protein at 15 min. The PS value for PUT-NGF was maximized and the residual plasma volume (Vp) of the protein in the blood vessels minimized when the pH of the modification reaction was >6.4. The biodistribution of PUT-NGF at 15 min showed 22-33% intact protein in different brain regions, which represented 0.4-5.9 ng of PUT-NGF in different brain regions, a physiological dose that is capable of eliciting a bioresponse. The design of this polyamine-modified NGF derivative that has enhanced permeability at the blood-brain and blood-nerve barriers with retained bioactivity may obviate the necessity to create small-molecule mimics of NGF and may be applicable to neurotrophins, engineered multifunctional chimeric neurotrophins, antioxidant enzymes, and other therapeutic proteins with specific clinical application to neurological diseases.\n\nID: 9365024\nTitle: Putrescine-modified catalase with preserved enzymatic activity exhibits increased permeability at the blood-nerve and blood-brain barriers.\nAbstract: Much evidence exists in support of the hypothesis that free radicals contribute to the pathogenesis of several neurodegenerative disorders and that mechanisms of free radical generation occur both intracellularly and extracellularly. Previous studies in this laboratory have shown that covalent modification of growth factors and antioxidant enzymes with the naturally occurring polyamine, putrescine, increases their permeability at the blood-nerve and blood-brain barriers (BNB and BBB), but does not significantly inhibit bioactivity. Furthermore, putrescine-modified superoxide dismutase (SOD) was shown to reduce neurodegeneration in a rat model of global cerebral ischemia. The purpose of the present study was to modify the antioxidant enzyme, catalase (CAT), with putrescine (PUT) at carboxylic acid groups whose ionization, and hence reactivity, was controlled with pH and investigate the effects on permeability and enzymatic activity. Modification of CAT with PUT increased its permeability 2-3-fold and preserved 67% of its enzymatic activity compared to native CAT and 137% compared to lyophilized CAT. The results of this study indicate that modification of CAT with putrescine increases its permeability while preserving enzymatic activity. PUT-SOD administered in combination with PUT-CAT may eliminate both the superoxide radical and the H2O2 produced from the dismutation of superoxide, respectively, and thus prevent the formation of hydroxyl radicals. This combination may exhibit increased neuroprotective effects, compared to native enzymes, following systemic administration for the treatment of free radical associated neurodegenerative disorders.\n\nID: 9191211\nTitle: N-methyl-D-aspartate receptor-mediated events contribute to neurovascular breakdown during experimental allergic encephalomyelitis.\nAbstract: \n\nID: 42553012\nTitle: Microbiota-Neuroinflammation Crosstalk in Primary Brain Tumors: Focus on Glioblastoma.\nAbstract: Glioblastoma (GBM) is the most aggressive primary brain tumor in adults and remains difficult to treat because of diffuse invasion, immunosuppression, metabolic adaptability, and therapy resistance. This review evaluates how gut microbiota and microbiota-associated neuroinflammatory signaling may contribute to GBM biology and therapeutic response. We synthesized mechanistic, preclinical, translational, and emerging clinical evidence on microbiota-neuroinflammation interactions in GBM. The review focused on gut-brain axis pathways, microbial metabolites, blood-brain barrier (BBB) regulation, glial and myeloid immune activity, tumor-associated microbial signatures, microbial peptide-HLA presentation, and microbiome-informed biomarker or therapeutic strategies. Current evidence suggests that microbiota-related signals may influence GBM through systemic immune modulation, short-chain fatty acids, tryptophan-derived metabolites, polyamines, BBB effects, and altered microglial and tumor-associated myeloid cell function. Polyamine metabolism may sustain myeloid-cell-mediated immunosuppression in the acidic GBM tumor microenvironment, whereas microglial GLUT5-dependent fructose metabolism may limit inflammatory antigen presentation and adaptive antitumor immunity. Sequencing-based studies have reported bacterial and fungal nucleic acid signatures in brain tumor specimens, but these findings require careful interpretation because of low biomass, contamination risk, and methodological variability. Preclinical models further indicate that microbiome modulation can alter inflammatory tone, tumor growth, immune-cell infiltration, and response to immune checkpoint blockade. Microbiota-regulated neuroinflammation is a biologically plausible contributor to GBM progression, immune suppression, and treatment resistance. However, most evidence remains preclinical or early translational. Well-controlled, spatially resolved, multi-omic studies are required before microbiome-based biomarkers or interventions can be clinically implemented for patient stratification and future precision clinical neuro-oncology applications.\n\nID: 42528048\nTitle: Exosome-Mediated Delivery of PROTACs for Targeted Protein Degradation in Cancer, Neurodegenerative, Infectious, and Inflammatory Diseases.\nAbstract: Proteolysis-targeting chimeras (PROTACs) are heterobifunctional molecules that hijack the ubiquitin-proteasome system to drive catalytic, sub-stoichiometric degradation of disease-associated proteins, offering a mechanistic advantage over occupancy-driven inhibitors and access to 'undruggable' targets. However, their clinical translation is constrained by high molecular weight, poor solubility, low oral bioavailability, inefficient membrane permeability, nonspecific biodistribution, off-target degradation, and the concentration-dependent 'hook effect.' Exosomes, nanoscale extracellular vesicles with innate biocompatibility, low immunogenicity, prolonged circulation, and the ability to cross barriers such as the blood-brain barrier, offer a biologically integrated platform to overcome these limitations. This review traces the evolution of PROTAC technology, delineates the challenges of conventional delivery, and evaluates the rationale for exosomal encapsulation, including cargo protection, intracellular trafficking, endosomal escape, and release kinetics. We examine natural and engineered exosomes spanning source selection, active loading strategies, and surface functionalization for tissue-specific homing and synthesize therapeutic applications across viral infections, cancer, neurodegenerative disorders, and inflammatory diseases. Proof-of-concept studies, such as camel milk-derived exosomes delivering the BRD4-targeting PROTAC ARV-825, demonstrate enhanced permeability, lower IC50 values, and improved oral bioavailability. Finally, we discuss key hurdles to clinical translation: scalable production, purification, and standardization, and outline future directions for exosome-mediated targeted protein degradation.\n\nID: 42508392\nTitle: Targeting the Microbiota-Butyrate-BHB Axis As a Potential Metabolic Therapeutic Strategy for Alzheimer's Disease.\nAbstract: Alzheimer's disease (AD) is increasingly linked to metabolic and microbial dysregulations, with butyrate and \u03b2-hydroxybutyrate (BHB) identified as key modulators. BHB is a ketone body primarily produced by the liver during periods of low glucose availability, and BHB levels are reduced in the AD brain and peripheral blood. Butyrate, a microbiota-derived short-chain fatty acid, is also reduced in the peripheral blood in association with gut microbiota dysbiosis. The reduction of butyrate and BHB, together with gut microbiota dysbiosis, is commonly observed in AD patients and correlates with cognitive decline and AD-related pathologies. Reduced levels of butyrate and BHB may impair energy metabolism, exacerbating amyloid-beta (A\u03b2) and tau pathologies. In contrast, preliminary evidence from interventional and ketogenic studies suggests that increasing butyrate and BHB levels may attenuate AD-related pathologies, such as A\u03b2 and tau abnormalities, while improving cognitive function. Emerging evidence suggests that both butyrate and BHB can cross the blood-brain barrier (BBB), suppressing oxidative stress and pro-inflammatory cytokines and potentially mitigating A\u03b2 aggregation and tau hyperphosphorylation. Accordingly, therapeutic strategies targeting the microbiota-butyrate-BHB axis represent a promising and biologically plausible approach that warrants further rigorous clinical investigation for the prevention and treatment of AD. This review comprehensively examines the roles of butyrate and BHB in ameliorating key AD-related pathologies, including A\u03b2 aggregation, tau hyperphosphorylation, neuroinflammation, and neurodegeneration. Additionally, the review explores the bidirectional association between butyrate/BHB and gut dysbiosis in AD, including how dysbiosis reduces butyrate/BHB levels while elevating them may reverse dysbiosis and improve gut-brain axis function. Collectively, these findings suggest that butyrate and BHB emerge as promising candidates for potential novel adjunctive therapies for AD, although supporting human evidence remains preliminary and heterogeneous.\n\nID: 42494471\nTitle: Glucose Transporter 1 in Health and Disease.\nAbstract: Glucose Transporter 1 (GLUT1) is the quintessential facilitator of basal glucose uptake, indispensable for maintaining cellular energy homeostasis, particularly across the blood-brain barrier. Beyond physiological necessity, GLUT1 dysregulation drives a broad pathological spectrum. While genetic haploinsufficiency precipitates severe neurological energy crises like Glut1 deficiency syndrome (Glut1DS), oncogenic networks hyperactivate GLUT1 as the central executor of the Warburg effect to fuel malignant proliferation. Despite its immense therapeutic potential, severe on-target toxicity in normal tissues and adaptive metabolic plasticity remain critical roadblocks to systemic GLUT1 inhibition. This review comprehensively synthesizes GLUT1's multidimensional regulatory networks in health and disease, dissecting how its overexpression fundamentally remodels the tumor microenvironment (TME). We elucidate how GLUT1-driven \"metabolic competition\" fosters metabolic immune exclusion and drives therapeutic resistance. Furthermore, we map the paradigm shift from traditional systemic blockades to emerging precision interventions. Specifically, we highlight \"Trojan horse\" glycan-functionalized nanocarriers, targeted protein degradation technologies like PROTACs, and metabolically engineered CAR-T cells. By conceptualizing GLUT1 as the linchpin of the immunosuppressive ecosystem, this work provides a strategic roadmap for precision metabolic immuno-oncology, guiding the development of novel therapies that maximize durable efficacy while minimizing collateral physiological damage.\n\nID: 42478899\nTitle: The liver-brain axis: A multidimensional regulatory network implicated in Alzheimer's disease pathogenesis and clinical implications.\nAbstract: Alzheimer's disease (AD) is a progressive neurodegenerative disorder characterized by amyloid-\u03b2 (A\u03b2) deposition. The liver-brain axis underscores the central role of the liver in modulating cognitive function through multidimensional regulatory mechanisms. As a core metabolic and detoxification organ, the liver also helps maintain cerebral homeostasis via pathways including the urea cycle, antioxidant systems, ketone body metabolism, and bile acid regulation. Dysfunction of these processes may lead to ammonia accumulation, exacerbated oxidative stress, and A\u03b2 clearance, thereby accelerating the pathological progression of AD. Liver-derived factors such as apolipoprotein E (APOE), C-reactive protein (CRP), fibroblast growth factor 21 (FGF21), and insulin-like growth factor 1 (IGF-1) significantly increase the risk of AD through dual mechanisms-inhibiting A\u03b2 clearance and activating neuroinflammation, thereby directly affecting cognitive function via modulation of inflammation, metabolism, and blood-brain barrier (BBB) integrity. Neural interfaces formed by the hypothalamic-pituitary-target gland axis and the vagus nerve enable communication from the liver to the brain, with emerging evidence also supporting a reverse influence from the brain to the liver. Emerging technologies such as molecular tracing and nanocarriers provide new tools for deciphering dynamic interactions within the liver-brain axis. Liver-targeted metabolic interventions show potential for reversing cognitive impairment. Unlike previous reviews that mainly focused on single pathways, this review conceptualizes the liver-brain axis as a multidimensional regulatory network in AD. By clearly linking network nodes to potential therapeutic interventions, it provides us with a novel framework that not only describes the various mechanisms but also focuses on identifying actionable targets for disease prevention and treatment.\n\nID: 42465365\nTitle: Challenges and Solutions in Quantifying Brain \u03b2-Hydroxybutyrate (BHB) with 1 H-MRS Following Oral Keto-Ester Consumption.\nAbstract: \u03b2-hydroxybutyrate (BHB), a ketone body and alternative cerebral energy substrate, can be measured in vivo using J-difference edited proton magnetic resonance spectroscopy ( 1 H-MRS). Oral ketone supplementation with substrates such as the ketone monoester (R)-3-hydroxybutyl-(R)-3-hydroxybutyrate (KME) and 1,3-butanediol (BD) have gained attention as a mechanism to elevate circulating BHB and induce ketosis without dietary restrictions. Elevated brain ketone availability is of growing therapeutic interest as a strategy to support neuronal energetics in conditions such as epilepsy, neurodegenerative disease, and alcohol use disorder (AUD). However, both pathways introduce BD into the bloodstream, which crosses the blood-brain barrier. Critically, BD exhibits a spectral signature that closely resembles the prominent BHB peak in JDE-MR spectroscopic imaging (MRSI), identified in a pilot AUD study. Two separate JDE-MRSI acquisitions tailored for BHB and BD editing were implemented, exploiting frequency separation between the BHB (4.14ppm) and BD (3.95ppm) coupling partners of the observed 1.2ppm resonance to independently quantify each metabolite. Brain BD concentrations (0.25-0.58mM) were comparable to or exceeded corresponding BHB concentrations (0.20-0.27mM) in all volunteers after consumption of a single dose of the KME, indicating that BD constitutes a major fraction of the signal conventionally attributed to BHB. Combined BHB+BD concentrations (\u223c0.45-0.85mM) were consistent with brain BHB values reported in prior studies employing similar doses of the KME, indicating that those measurements likely reflect a combined BHB+BD signal. Separate quantification of the two metabolites is important for interpreting brain ketone studies and for understanding the full pharmacology of KME supplementation.\n\nID: 42427667\nTitle: Shared Genomic Architecture Between Schizophrenia and Multiple Sclerosis Identifies an Un-Drugged HCAR1 Neuroimmune Checkpoint.\nAbstract: Multiple Sclerosis (MS) pathogenesis is contingent upon the hyper-proliferative infiltration of peripheral macrophages across the blood-brain barrier. While front-line therapeutics, such as Dimethyl Fumarate, achieve clinical efficacy by agonizing the HCAR2 immune cooling switch, the tandemly duplicated HCAR1 lactate sensor has remained entirely unexplored. Here, by cross-referencing MS and Schizophrenia (SCZ) genomic architectures, we identify a massive shared structural fracture strictly localized to the HCAR tandem regulatory domain. We demonstrate that this locus acts as a highly specific neuroimmune ignition switch: it drives disease susceptibility but is unequivocally unassociated with MS severity or classical systemic autoimmune phenotypes (Crohn's Disease, Lupus, Rheumatoid Arthritis, and Psoriasis). Crucially, utilizing high-resolution eQTL mapping in purified human immune lineages, we reveal that the shared MS/SCZ risk allele drives a profound, state-independent transcriptomic collapse of HCAR1 exclusively in peripheral macrophages. This enhancer failure renders activated macrophages physically \"lactate blind\"-unable to sense their own glycolytic exhaust to engage the cAMP-suppressing negative feedback loop required to halt immune proliferation. By bridging psychiatric genetics and neuroimmunology, this study reframes the HCAR tandem array as a master neuroimmune bifurcation point and introduces the un-drugged HCAR1 lactate brake as a critical therapeutic checkpoint for arresting demyelinating disease.\n\nID: 42423995\nTitle: Unveiling the role of CB2 receptor in beta-hydroxybutyrate mediated modulation of.\nAbstract: The cannabinoid receptor type 2 (CB2R), primarily expressed in microglia, the brain's resident immune cells, acts as a central regulator of neuroinflammatory responses. When CB2R is activated, it triggers anti-inflammatory signaling, making it a promising target for modulating microglial function in neuroinflammatory diseases. The ketone body, \u03b2-hydroxybutyrate (BHB), is gaining attention as a therapeutic agent for neurodegenerative disorders due to its ability to modulate neuroinflammation and preserve blood-brain barrier integrity. One mechanism by which BHB exerts anti-inflammatory effects is through regulation of microglial function; however, the precise mechanisms remain unclear. Since the role of BHB in this context is unexplored, we used two neuroinflammation models to test the hypothesis that CB2R-associated signaling contributes to the effects of BHB. In a mouse model of diet-induced obesity (DIO), characterized by chronic low-grade neuroinflammation, BHB treatment promoted ramified microglial morphology and enhanced debris clearance while sparing synaptic elements. These changes were accompanied by alterations in CB2R-related signaling markers and a slight increase in hydroxycarboxylic acid receptor 2 (HCA2), a known BHB target. When primary microglial cultures were challenged with lipopolysaccharide (LPS), BHB helped restore their function. However, that benefit disappeared when CB2R was pharmacologically blocked. Importantly, BHB increased the expression of arginase 1 (Arg1), a hallmark of anti-inflammatory responses, a change reversed by CB2R blockade. Moreover, BHB reduced NF-\u03baB signaling, and CB2R inhibition attenuated this effect, suggesting that CB2R-associated signaling contributes to BHB's anti-inflammatory actions. Collectively, our findings demonstrate that BHB's anti-inflammatory effects are mediated, at least in part, through CB2R signaling, providing new insight into its therapeutic potential for neuroinflammation.\n\nID: 42402305\nTitle: The brain renin-angiotensin system in Parkinson's disease: Friend or foe? mechanistic insights and therapeutic implications.\nAbstract: The renin-angiotensin system (RAS), classically known for its role in cardiovascular and fluid homeostasis, also regulates neuronal homeostasis in the central nervous system (CNS), where its dysregulation contributes to PD pathogenesis. The emerging evidence links excessive activation of the brain RAS in PD, where sustained activation of the angiotensin II (Ang II)/angiotensin type-1 receptor (AT1R) axis promotes oxidative stress, neuroinflammation, mitochondrial dysfunction, and blood-brain barrier (BBB) disruption that leads to progressive dopaminergic neurodegeneration. This AngII-AT1R signaling increases the production of reactive oxygen species (ROS) mediated by NADPH oxidase, primes microglia to a chronic pro-inflammatory state, disrupts the proteostatic regulation of nigrostriatal neuronal \u03b1-synuclein clearance, and intensifies the selective vulnerability of nigrostriatal neurons. The counter-regulatory ACE2/angiotensin (1-7)/Mas and AT2R pathway seems to have neuroprotective effects; however, it reverses the negative effects of Ang II. In preclinical, epidemiological, and emerging clinical evidence, pharmacological modulation of the RAS, particularly BBB-penetrant angiotensin receptor blockers (ARBs) and angiotensin-converting enzyme inhibitors (ACEIs), has shown promise as neuroprotective agents. In the current area of research, RAS-targeted interventions represent a promising and mechanistically grounded strategy for disease modification rather than symptomatic management alone. This review explores molecular, cellular, and system-level insights into RAS dysregulation in PD, integrates translational evidence supporting RAS-modulating therapies, and highlights emerging biomarkers and precision medicine approaches that may guide therapeutic optimization. This review also highlights the brain RAS as a key mediator linking redox imbalance, neuroinflammation, and multisystem dysfunction in PD and makes it a promising therapeutic axis for slowing the disease progression.\n\nID: 42400441\nTitle: Probing Cinnamamides and Benzamides as Anthelmintics: Discovery of Potent Drug-Like Agents Against Angiostrongylus cantonensis.\nAbstract: Emergent helminthiases are increasingly impacting global health in both humans and animals, especially given the limited efficacy of existing drugs against these infections. Neuroangiostrongyliasis, an eosinophilic meningitis caused by Angiostrongylus cantonensis, currently lacks effective treatment, highlighting the need for novel anthelmintics. We previously identified cinnamoyl-benzylpiperazine, a simplified analogue of cinnarizine, as an effective anthelmintic agent against first (L1) and third-stage (L3) larvae of A. cantonensis in\u00a0vitro. In the present work, structural modifications on the active prototype cinnamoyl-benzylpiperazine were performed, prioritizing the improvement of solubility and the provision of balanced physicochemical properties compatible with blood-brain barrier permeability, alongside anthelmintic activity. A set of 31 compounds divided into two series (I-cinnamoyl and II-benzoyl) was synthesized and tested against L1 and L3 larvae, yielding EC50 values ranging from 4.1 to 27.6\u2009\u03bcM. SAR analyses revealed that the activity of set I is strongly associated with balanced electronic density in both the cinnamamide and amine regions (described by ionization potential descriptors), whereas modifications in the charge distribution of the molecules (indicated by topological charge descriptors) appear to determine the anthelmintic activity of set II. None of the compounds displayed significant toxicity to HaCaT mammalian cells (up to 200\u2009\u03bcM) or Caenorhabditis elegans worms (up to 1000\u2009\u03bcM), denoting specific activity against A. cantonensis. The balanced polarity of compound 4a-I (EC50 L1 4.7\u2009\u03bcM; L3 10.2\u2009\u03bcM) and the localized charge density provided by the methoxy group in compounds 1c-II (EC50 L1 5.5\u2009\u03bcM; L3 10.9\u2009\u03bcM) and 5c-II (EC50 L1 4.9\u2009\u03bcM; L3 11.9\u2009\u03bcM) seem important for interacting with the putative target in the helminth. Collectively, the substituents in these molecules provided improved drug-likeness over the previous set of compounds, and represent noteworthy derivatives for further investigation against A. cantonensis in\u00a0vitro.\n\nID: 42387117\nTitle: Leveraging Carnitine-functionalized Lipid Nanocarrier based Targeted Delivery of A1874 PROTAC for Glioblastoma.\nAbstract: Glioblastoma(GBM) is highly aggressive and therapeutically refractory brain malignancy. Despite maximal intervention, patient prognosis remains dismal. A major obstacle in its management is the limited permeation of therapeutic agents across the blood-brain barrier(BBB), further intensified by resistance to standard chemotherapy such as temozolomide(TMZ). Proteolysis-targeting chimeras(PROTACs), offer new opportunities by enabling selective degradation of oncogenic proteins. A1874, a heterobifunctional molecule, has demonstrated potent, selective degradation of oncogenic driver-BRD4 in pancreatic, breast, and colon cancer. BRD4 drives GBM cell proliferation, survival, and resistance to therapy; therefore, we investigated the therapeutic potential of A1874 in brain cancer. Herein, we developed brain-targeted self-nanoemulsifying drug delivery system, termed PRONano, designed to enhance the targeted delivery of A1874. The system is functionalized with Palmitoyl-DL-carnitine chloride(PC) to facilitate transport across the BBB. Physicochemical characterization was performed to assess particle size. In-vitro cytotoxicity, qualitative and quantitative cellular uptake was analyzed. Mechanistic validation was conducted using western blot and qPCR, while 3D spheroid assay was employed to assess efficacy in tumor-mimicking microenvironment. PRONano exhibited nanoscale particle size and significantly enhanced intracellular uptake of A1874. Formulation exhibited enhanced cytotoxicity in temozolomide-sensitive and resistant GBM cells. Effective BRD4 protein degradation was identified in the Western blot. PRONano significantly inhibited 3-D spheroid tumor growth suggesting better penetration and efficacy in tumor-like microenvironment. PRONano is brain-targeted, rationally designed nanoformulation which can overcome major A1874 delivery constraints. This strategy augmented PROTAC delivery and therapeutic potential in GBM, supporting future preclinical development.\n\nID: 42369646\nTitle: Integrating Bone-Brain Axis Modulation and Tea Consumption for Enhancing Neurovascular Resilience and Patient Rehabilitation Education.\nAbstract: Intracerebral hemorrhage (ICH) is one of the most devastating subtypes of stroke, with limited preventive options and a challenging recovery process. This study presents a translational framework that integrates tea polyphenols (TPPs) and exercise-induced metabolites as dual modulators of neurovascular stability, with a focus on patient education for enhancing post-stroke recovery. By synthesizing preclinical and clinical evidence, we demonstrate how TPPs, particularly epigallocatechin gallate (EGCG), and key exercise metabolites such as lactate, \u03b2-hydroxybutyrate, and short-chain fatty acids (SCFAs) interact with shared redox-sensitive and inflammatory signaling pathways (Nrf2/NF-\u03baB/AMPK axis) to reinforce endothelial integrity, preserve blood-brain barrier function, and maintain cerebral perfusion. These interventions also reshape the gut microbiota, promoting an SCFA-enriched, anti-inflammatory profile that fosters bidirectional gut-brain communication, further stabilizing vascular homeostasis. Multi-omics evidence suggests that TPPs and exercise metabolites may jointly regulate metabolic and immune pathways, enhancing resilience against oxidative and inflammatory injuries in the vasculature. We propose a mechanistic model in which TPPs and exercise-derived metabolites synergistically support neurovascular function and reduce neurovascular vulnerability associated with ICH, while promoting cognitive recovery and metabolic health. Incorporating these findings into patient rehabilitation education may help individuals make informed decisions about lifestyle changes that enhance vascular health. Future research should explore the dose-response relationship, the optimal timing between tea and exercise, and individual variations, using metabolomic, microbiomic, and imaging biomarkers to personalize cerebrovascular prevention strategies.\n\nID: 42314466\nTitle: Rewiring cell death and evading repair: Evolution of temozolomide and emerging strategies against resistant glioblastoma.\nAbstract: Glioblastoma (GBM) is the most common and aggressive primary malignant brain tumor, characterized by a universally dismal prognosis. For over two decades, the alkylating agent temozolomide (TMZ) has remained the cornerstone of the standard-of-care \"Stupp protocol\" largely owing to its unique pH-dependent activation and favorable blood-brain barrier penetration. However, the clinical efficacy of TMZ is profoundly compromised by complex intrinsic and acquired resistance mechanisms. These are primarily driven by O6-methylguanine-DNA-methyltransferase (MGMT) overexpression, mismatch repair (MMR) deficiency, and hyperactive base excision repair (BER) pathways. In this review, the chemical properties and classical cytotoxic mechanisms of TMZ are comprehensively summarized, followed by a systematic exploration of emerging medicinal chemistry strategies designed to overcome this multifaceted resistance. Specifically, recent advances in circumventing MGMT-mediated repair through epigenetic modulation and targeted protein degradation are highlighted. Furthermore, we discuss the rational structural modifications of the imidazotetrazine scaffold to induce steric hindrance, alongside a paradigm shift toward rewiring cell death mechanisms-such as transitioning from DNA monomethylation to interstrand cross-linking (ICLs) and the activation of non-apoptotic pathways, including ferroptosis and autophagy. Finally, systemic interventions exploiting \"BRCAness\"-induced synthetic lethality via PARP inhibitors are examined, culminating in future perspectives on the integration of artificial intelligence (AI) and spatiotemporally responsive nanocarriers for the design of next-generation anti-GBM therapeutics.\n\nID: 42312138\nTitle: A patent review of cyclin-dependent kinase 5 (CDK5) inhibitors (1999-2025).\nAbstract: Cyclin-dependent kinase 5 (CDK5) is a critical regulator of neuronal development and function, whose hyperactivation exacerbates neurodegenerative disorders and certain cancers. While CDK5 is a compelling therapeutic target, achieving selective inhibition is challenging due to high structural homology with cell-cycle CDKs and poor blood-brain barrier penetrance. The article provides the first comprehensive analysis of patented CDK5 inhibitors disclosed between 1999 and 2025. We examine the chemical diversity, selectivity profiles, and therapeutic claims of major chemotypes, including purine analogs (e.g., roscovitine), pyrazole derivatives (e.g., dinaciclib, milciclib), indolobenzazepinones, indirubin derivatives, and emerging modalities such as peptides. Furthermore, we discuss major obstacles such as overcoming off-target toxicity against other CDKs, ensuring sufficient CNS exposure, and identifying reliable biomarkers. Lastly, we speculate on how future success will depend on new strategies such as p25-specific modulation, targeted protein degradation, and advanced delivery systems to translate CDK5' therapeutic potential into the clinic.\n\nID: 42234836\nTitle: Peptide-Based Delivery Systems: Selected Insights into Cell-Penetrating Peptides.\nAbstract: The blood-brain barrier (BBB) is a highly selective biological interface that regulates molecular transport between the bloodstream and the central nervous system (CNS). Its primary function is to protect the brain from harmful substances while maintaining neural homeostasis. This regulatory capacity is attributed to its complex, multilayered structure, comprising nonfenestrated endothelial cells, astrocytic end-feet, pericytes, specialized transporters, and efflux pumps, along with dynamic mechanisms that adapt to physiological changes. Despite its essential protective role, the BBB presents a significant obstacle to the delivery of therapeutics targeting CNS disorders, limiting the ability of many drugs to reach their intended sites of action. Numerous invasive and noninvasive strategies have been explored to enhance CNS drug delivery; however, many are hindered by limitations such as low efficacy, poor specificity, and immunogenicity. Peptides, as endogenous short chains of amino acids, offer a promising alternative due to their high biocompatibility, ease of synthesis, and structural versatility. Their cationic charge facilitates interactions with negatively charged proteoglycans on cell surfaces, while their amphiphilic nature enhances membrane permeability, enabling efficient cellular uptake and translocation. These properties make peptide-based vectors particularly suitable for transporting diverse therapeutic cargoes across biological barriers. This review discusses the structural characteristics, origins, and classifications of peptide vectors, as well as the mechanisms underlying their cellular internalization. It further evaluates their advantages and current limitations in clinical applications, and highlights emerging strategies aimed at optimizing peptide-mediated delivery across the BBB.\n\nID: 42215997\nTitle: Convergence of neuroinflammation across major neurotropic viral exposomes in AD and ADRD.\nAbstract: Alzheimer's disease (AD) and Alzheimer's disease-related dementias (ADRD) are multifactorial neurodegenerative disorders driven by complex interactions among genetic susceptibility, aging, and environmental exposures. Growing epidemiological and mechanistic evidence implicates neurotropic viral exposomes, defined as cumulative lifetime viral infections, as significant contributors to AD risk. Viral encephalitis and common viral infections, including herpes simplex virus type 1 (HSV-1), human immunodeficiency virus (HIV), cytomegalovirus (CMV), SARS-CoV-2, and influenza, have been associated with an increased incidence of AD/ADRD; however, the molecular mechanisms underlying these associations remain incompletely understood. A systematic literature review was conducted using PubMed, Web of Science, Scopus, and Google Scholar (1990-2025) to identify epidemiological, experimental, and mechanistic studies linking viral infections to AD-related pathology. Systems biology approaches were applied using Cytoscape, STRING, KEGG, WikiPathways, and Ingenuity Pathway Analysis to construct protein-protein interaction networks and identify convergent biological processes shared between AD and viral host-response pathways. Functional enrichment analyses focused on neuroinflammation, amyloid-\u03b2 (A\u03b2) metabolism, tau pathology, autophagy, and blood-brain barrier (BBB) integrity. Across diverse viral infections, strong convergence was observed in innate immune activation pathways, including microglial priming and NLRP3 inflammasome signaling, accompanied by chronic production of proinflammatory cytokines (IL-1\u03b2, TNF-\u03b1, IFN-\u03b3). Multiple viruses modulated amyloidogenic APP processing, impaired A\u03b2 clearance, promoted tau hyperphosphorylation, disrupted autophagy-lysosomal systems, and compromised BBB integrity. Systems-level analyses revealed overlapping signaling hubs, including NF-\u03baB, MAPK, PI3K-Akt, and cGAS-STING that amplify neurodegenerative cascades, with effects most pronounced in genetically susceptible populations such as APOE4 carriers. Collectively, current evidence supports a mechanistic link between viral exposomes and AD/ADRD mediated through convergent neuroinflammatory, and proteostatic pathways. Although viral infections alone are unlikely to be sufficient to cause AD, recurrent or persistent viral exposures may act as potent disease modifiers that accelerate neurodegenerative processes. Integrating viral biomarkers, genetic risk stratification, and systems biology approaches offers promising opportunities for early diagnosis, prevention, and development of mechanism-guided therapeutic strategies.\n\nID: 42150270\nTitle: PROTACs for central nervous system disorders: challenges and opportunities.\nAbstract: Central nervous system (CNS) disorders, particularly neurodegenerative diseases and brain tumors, pose substantial and long-standing challenges to the global public health system. Current therapeutic approaches for CNS disorders are primarily confined to symptomatic relief and generally fail to halt or reverse disease progression. However, proteolysis-targeting chimeras (PROTACs), as an emerging therapeutic strategy, have introduced new prospects for effective intervention in this domain. Although several studies have demonstrated the use of PROTACs in degrading pathogenic proteins associated with CNS disorders, their clinical translation remains hindered by multiple challenges, including insufficient degradation efficiency, potential off-target toxicity, and limited blood-brain barrier (BBB) permeability. In response to the above challenges, researchers are actively pursuing various strategies-ranging from structural optimization of PROTACs to enhance their degradative activity, to the implementation of novel nano-delivery systems and targeted delivery approaches to improve BBB permeability and tissue selectivity. These strategic advancements have demonstrated significant potential in preclinical research for the treatment of CNS disorders. This review provides a comprehensive overview of the research and development of various PROTACs targeting CNS disorders, and highlights potential application strategies for advancing protein degrader therapeutics in the CNS field. By synthesizing current advances and challenges, it offers researchers in related disciplines a well-defined theoretical framework and forward-looking strategic guidance, thereby facilitating in-depth investigation and transformative applications in this rapidly evolving domain.\n\nID: 42130715\nTitle: Blood-Brain Barrier (BBB)-Penetrable Androgen Receptor (AR) Degrader as a Potential Therapeutic Agent for Glioblastoma.\nAbstract: Androgen receptor (AR) contributes to the progression of glioblastoma (GBM), which is consistent with the sex difference in GBM, which has a higher incidence in males than in females. Therefore, targeting AR is a potential therapeutic approach for GBM treatment. However, AR mutation commonly occurs in GBM, which makes conventional AR antagonists less effective. AR degraders abolish AR at the protein level regardless of the mutation status of AR, which makes it a better strategy in GBM. Compound A is an analog of the cyclooxygenase-2 (COX-2) inhibitor Nimesulide. Mechanistically, compound A targets HSP27, disrupts the HSP27-AR complex, and thereby promotes AR degradation in GBM cells at 1 \u03bcM, leading to inhibition of AR-overexpressing GBM cell growth with IC50s around 0.2 \u03bcM. In a GBM patient-derived cell line, DI318, compound A (1 \u03bc\u039c) also significantly decreases AR protein levels. The compound significantly inhibits GBM xenograft growth at 20 mg/kg and does not cause toxicity in mice up to 200 mg/kg. Pharmacokinetic studies reveal that compound A has a half-life (t 1/2) of 3.11 h and a BBB penetration of 52%, which is even higher than the standard chemotherapy Temozolomide. These results suggest that the AR degrader has great potential as a novel GBM treatment.\n\nID: 42065711\nTitle: Targeted Protein Degradation and Delivery Strategies in the Context of Neurological Disorders.\nAbstract: Neurological disorders represent a leading cause of global mortality and disability, yet treatment options remain limited due to the challenges of targeting pathogenic proteins, particularly those considered \"undruggable\" by conventional small molecules. Targeted protein degradation (TPD) has expanded the druggable proteome by harnessing proteasomal and lysosomal pathway to eliminate these targets, offering the advantages of lower toxicity and reduced resistance compared to traditional modulation. This review systematically delineates TPD mechanisms according to their degradation pathways, including proteasomal, endosomal-lysosomal, and autophagy-lysosomal systems, and highlights their unique applications in brain diseases. However, the translation of TPD to neurological disease is limited by physicochemical liabilities, cell-type dependence, risks associated with whole-protein ablation, the blood-brain barrier (BBB) and poor brain bioavailability. To address these translational barriers, we emphasize the integration of TPD with drug delivery systems (DDS) as a pivotal strategy. By optimizing pharmacokinetics, stability, and BBB penetration, nano-DDS significantly enhances brain targeting and therapeutic precision. Finally, we evaluate recent progress in nano-TPD systems and offer critical insights into their future trajectory in treating complex brain disorders.\n\nID: 42059004\nTitle: A new hope: Clinical advances in targeted therapies for pediatric diffuse midline glioma.\nAbstract: Diffuse midline glioma (DMG) is an aggressive pediatric brain tumor with a 1%-2% overall survival, largely due to the ineffectiveness of conventional treatments such as chemotherapy and radiotherapy, as well as the inoperability of the tumors because of their critical location and infiltrative diffuse growth. Recent advances in targeted therapies offer new hope, particularly those addressing key molecular characteristics and newly identified cancer dependencies. Among these are histone deacetylase inhibitors (HDACis), receptor tyrosine kinase inhibitors, and novel agents such as ONC201 and unesbulin that target metabolic and epigenetic pathways respectively. In addition, emerging therapies like FACT inhibitors and polyamine pathway inhibitors are showing promise by disrupting critical cancer cell processes. Immunotherapies, including CAR-T cells targeting surface antigens such as GD2 and B7-H3, cancer vaccines, monoclonal antibodies, and oncolytic viruses, are also gaining traction, offering a new approach by harnessing the immune system to attack tumor cells. Despite these advances, challenges such as drug delivery across the blood-brain barrier and therapeutic resistance persist, necessitating the development of combination therapies and innovative delivery methods. Ongoing research is focused on refining these strategies and exploring additional molecular and immunological targets to improve outcomes for children with DMG.\n\nID: 42031360\nTitle: Engineering brain-penetrant PROTACs: Bridging molecular design and CNS delivery.\nAbstract: The drug development for central nervous system (CNS) disorders, particularly neurodegenerative diseases, such as Alzheimer's disease, Parkinson's disease, and Huntington's disease, faces formidable challenges. While proteolysis-targeting chimeras (PROTACs) represent a paradigm-shifting modality by redefining target engagement mechanisms, their clinical translation remains hindered by limited blood-brain barrier (BBB) permeability and suboptimal pharmacokinetic profiles. In recent years, a range of CNS-targeted delivery strategies have emerged, advancing PROTAC research toward more translatable therapeutic applications. This review highlights recent advances and persistent challenges in noninvasive BBB-penetrant delivery systems, including viral vectors, engineered exosomes, functionalized nanocarriers, and cell membrane-derived biomimetic vehicles, with a particular emphasis on intranasal administration as a direct route to the brain. Parallel progress in rational molecular engineering, encompassing E3 ligase selection, linker polarity and rigidity modulation, and optimization of target-binding ligands, has further enhanced PROTAC drug-likeness and BBB transport efficiency. Current CNS-directed PROTAC designs increasingly incorporate cell-penetrating peptides, nanoparticles, and prodrug formulations to balance stability, selectivity, and brain exposure. Future advanced PROTAC delivery platforms require integrating multifunctional nanocarriers with rational structural optimization to enhance BBB permeability. Further artificial intelligence-accelerated molecular design and targeted protein degradation technologies offer novel avenues for addressing undruggable CNS targets.\n\nID: 42003242\nTitle: Enhanced autophagy drives endothelial tight junction loss, BBB disruption, and behavioral deficits during inflammation.\nAbstract: The blood-brain barrier (BBB) protects the brain but becomes compromised during systemic inflammatory conditions such as sepsis. The mechanisms driving BBB disruption remain incompletely understood. Here, we identified a significant enrichment of the macroautophagy/autophagy-lysosome-related pathway in the upregulated proteome using quantitative proteomics on brain microvessels from mice after cecal ligation and puncture (CLP) that induces polymicrobial sepsis. CLP progressively induced autophagic flux in brain endothelial cells, peaking at 24\u2009h post-procedure before subsiding. Similarly, an mRFP-GFP-LC3 reporter assay and immunoblotting showed that lipopolysaccharide (LPS) treatment increased autophagic flux in bEnd.3 cells in a time- and dose-dependent manner. Mice intraperitoneally (IP) injected with the autophagy inhibitors chloroquine (CQ) or 3-methyladenine (3-MA) were resistant to BBB disruption caused by CLP or IP injection of LPS, whereas those injected with the autophagy inducer rapamycin (Rapa) were more susceptible. CQ and 3-MA reduced, while Rapa increased, CLP-induced lethality in mice. These effects were confirmed in vitro using a dextran infiltration assay on bEnd.3 cell transwell cultures. CQ alleviated both the acute disruption of the tight junction proteins TJP1/ZO-1 and CLDN5 in brain microvessels and the long-term memory and anxiety deficits in LPS-challenged mice. siRNA-mediated knockdown of the SNARE protein STX17, which inhibits autophagosome-lysosome fusion, attenuated LPS-induced tight junction protein degradation in bEnd.3 cells. Importantly, inhibition of TLR4 or its downstream kinase TBK1 reduced LPS-induced autophagy and preserved tight junction proteins, implicating TLR4-TBK1 signaling in endothelial autophagy activation. These results suggest that excessive autophagy in endothelial cells drives BBB damage and cognitive dysfunction in sepsis.Abbreviations: 3-MA: 3-methyladenine; ATG5: autophagy related 5; BBB: blood-brain barrier; bEnd.3 cells: brain-derived endothelial cells.3; CLP: cecal ligation and puncture; CQ: chloroquine; EPM, elevated plus maze; GO: Gene Ontology; IP: intraperitoneal; LPS: lipopolysaccharide; MAP1LC3/LC3-II: microtubule associated protein 1 light chain 3-II; MWM: Morris Water Maze; NOR: novel object recognition test; OFT: open field test; Rapa: rapamycin; SAE: sepsis-associated encephalopathy; SNARE: soluble N-ethylmaleimide-sensitive factor attachment protein receptor; TBK1: TANK binding kinase 1; TICAM1/TRIF: TIR domain containing adaptor molecule 1; TLR4: toll like receptor 4; TMT: tandem mass tag.\n\nID: 41980980\nTitle: Dysregulation of macrophage lipid metabolism underlies intracellular bacterial neuroinvasion.\nAbstract: Acute central nervous system infection is highly lethal, yet the mechanisms by which intracellular bacteria infiltrate the brain remain unclear. Phagocytes are central to host defense, but how infected cells facilitate bacterial access to the brain is poorly defined. In this study, we characterize a CD36+ Fabp4+ Pparg+ macrophage subset that mediates bacterial penetration of the brain without disrupting the blood-brain barrier. Biomechanical analysis reveals that CD36+ macrophages exhibit abundant protrusions and adhesion molecules, enabling resistance to blood flow shear stress and promoting endothelial adhesion. Metabolomic profiling reveals dysregulated lipid metabolism during neuroinvasion, with \u03b2-hydroxybutyrate promoting the differentiation and survival of CD36+ macrophages. Importantly, ketogenesis exacerbates symptoms during bacterial neuroinvasion, which could be halted by physiological glucose supplementation. Here, we show that intracellular bacteria exploit metabolically reprogrammed macrophages to access the brain, highlighting glycolipid metabolic homeostasis as a potential therapeutic target in bacterial neuroinvasion.\n\nID: 41941974\nTitle: Intelligent delivery of autophagy-targeting chimeric peptides by engineered exosomes for the degradation of \u03b1-synuclein.\nAbstract: Targeted degradation of the aggregated \u03b1-synuclein holds tremendous potential for treating Parkinson's disease (PD). However, most of the developed aggregated \u03b1-synuclein-specific degraders, e.g., autophagy-targeting chimeric peptides, are limited by the blood-brain barrier (BBB), substantia nigra (SN) neuron targetability, and intracytoplasmic release. To overcome these obstacles, we constructed an engineered exosome (EXO) equipped with surficial glucose-regulated protein 94 (GRP94)-targeting peptide N, luminal \u03b1-synuclein-degrading peptide P1, and cathepsin-B-cleavable GFLG as the linker between the exosome skeleton protein and P1, termed NEXOGFLG-P1. We verified that the NEXOGFLG-P1 exosomes could cross the BBB and target diseased SN neurons in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine hydrochloride (MPTP)-induced PD model mice. Following fusion with endosomes, the exposed P1 was released into the cytoplasm by cytoplasmic cathepsin B-mediated GFLG cleavage to degrade \u03b1-synuclein. Collectively, the NEXOGFLG-P1 exosomes exhibit a significant degradation effect on \u03b1-synuclein aggregates, providing a proof-of-concept platform for treating PD. STATEMENT OF SIGNIFICANCE: Targeted degradation of \u03b1-synuclein aggregates holds tremendous potential for the etiological treatment of Parkinson's disease (PD). However, most of current \u03b1-synuclein-specific degraders are stuck with low blood-brain barrier permeability, poor targetability for diseased cells, and uncontrolled release. Notably, \u03b1-synuclein predominantly affects neurons in the substantia nigra (SN) region rather than the whole brain. To overcome these obstacles, we constructed an engineered exosome, termed NEXOGFLG-P1, to specially deliver and release autophagy-targeting chimeric peptide to degrade \u03b1-synuclein in the diseased SN neurons through the autophagy-lysosomal pathway. The engineered exosomes exhibit the great potential in targeting diseased SN neurons and degrading \u03b1-synuclein aggregates, providing a proof-of-concept therapeutic platform for treating PD.\n\nID: 41921838\nTitle: L-type amino acid transporter 1 targeting self-assembly polyelectrolyte nanocomplex with enhanced nose to brain delivery of oxytocin.\nAbstract: L-type amino acid transporter 1 (LAT1) is essential for the transport of large neutral amino acids (AA) across the blood-brain barrier (BBB), which plays a crucial role and is widely employed in brain drug delivery. Thus, in this study, tryptophan (Trp), leucine (Leu) and tyrosine (Tyr) were selected and various amino acid grafted chitosan (CTS) was first synthesized and characterized by Fourier transform infrared spectroscopy (FTIR) and 1H nuclear magnetic resonance (1H NMR) spectrum. Furthermore, oxytocin (OT) loaded polyelectrolyte complex (PEC) based on various amino acid grafted chitosan and chondroitin sulfate (CS) as well as OT loaded chitosan-chondroitin sulfate PEC was fabricated and optimized by particle size, zeta potential and encapsulation efficiency (EE%). The morphologies and OT conformation stability of different PEC formulations were investigated by transmission electron microscopy (TEM) and circular dichroism (CD) spectroscopy respectively. Moreover, to further reveal the superiority of amino acid grafted chitosan PECs on mucosal permeation and brain targeting via LAT1 transporter, the ex vivo permeation study, in vivo pharmacokinetics behavior as well as brain distribution of OT in different PECs were also carried out by intranasal administration. It was found that both chitosan-chondroitin sulfate and amino acid grafted chitosan-chondroitin sulfate PECs exhibited comparable particle size ranging from 180 to 210\u00a0nm with a positive charge. The PEC formulation presented a sustained release behavior compared to the OT solution and better nasal mucosal permeation was observed for amino acid grafted chitosan-chondroitin sulfate PECs. Both pharmacokinetics behavior and brain distribution results demonstrated that amino acid grafted chitsaon could significantly improve the OT absorption and brain accumulation in the order of Trp\u00a0>\u00a0Tyr\u00a0>\u00a0Leu, compared to naked chitosan-chondroitin sulfate PECs and OT solution. The behavioral and immunofluorescence results also revealed that amino acid grafted chitosan-chondroitin sulfate PECs exhibited a superior inhibitory effect on oxycodone (Oxy) addiction. In conclusion, amino acid grafted chitosan-chondroitin sulfate PECs could significantly promote brain drug transportation and could be a potential vehicle for protein or peptide delivery via nasal to brain pathway.\n\nID: 41882776\nTitle: Streptococcus suis disrupts the blood-brain barrier through inducing ubiquitin-proteasome-mediated degradation of KAT2A.\nAbstract: Streptococcus suis (S. suis) induces host cell death and has the ability to invade the blood-brain barrier (BBB). However, S. suis-induced BBB disruption has not been completely elucidated. In this study, we focused on the regulatory role of lysine acetyltransferase 2A (KAT2A) on BBB, which is a key regulator of cell death. We found that S. suis strain SC19 induces cell death in human cerebral microvascular endothelial cell line D3 (hCMEC/D3) by membrane disruption and LDH release. During SC19 infection, KAT2A protein expression was markedly reduced but its mRNA expression was not affected. Further study demonstrated that SC19-induced KAT2A reduction was though ubiquitin-proteasome-mediated protein degradation pathway. KAT2A inhibition using significantly exacerbated SC19-induced cell permeability disruption by transwell infection model. Consistently, KAT2A knockdown and pharmacological inhibition significantly aggravated SC19-induced downregulation of tight junction proteins including ZO-1 and occludin, whereas KAT2A overexpression partially restored their levels. Similarly, KAT2A inhibition aggravated SC19-induced downregulation of ZO-1 in mice brain and increased mice death rate, indicating the protective role of KAT2A on BBB. Mechanistically, KAT2A was found to regulate necroptosis in hCMEC/D3. KAT2A knockdown aggravated necroptosis and upregulated phosphorylation of RIPK1, whereas RIPK1 inhibitor Nec-1 rescued SC19-induced necroptosis, indicating that KAT2A regulates RIPK1-dependent necroptosis during SC19 infection. Furthermore, quantitative proteomic analysis identified a network of putative KAT2A-mediated downstream targets and pathways that contribute to BBB integrity. Collectively, these findings reveal the underlying mechanism by which SC19 disrupts BBB through inducing KAT2A-mediated necroptosis and provide a potential therapeutic target for the treatment of bacterial meningitis.\n\nID: 41828397\nTitle: Impact of the Combination of Epigallocatechin Gallate and Ellagic Acid Supplemented with Ketone Bodies on Energetic Restoration of Mitochondrial Dysfunction and Metabolic Inefficiencies in Patients with Multiple Sclerosis: A Review.\nAbstract: Multiple sclerosis (MS) is characterized by progressive mitochondrial dysfunction affecting complexes I, III, and IV of the electron transport chain, contributing to axonal energy failure and neurodegeneration. This review examines the potential of combining \u03b2-hydroxybutyrate (\u03b2HB), epigallocatechin-3-gallate (EGCG), and ellagic acid (EA) as a multi-target therapeutic strategy to restore mitochondrial function in patients with MS. Experimental and clinical studies demonstrate that each compound exerts complementary mechanisms. Ketone bodies provide an alternative energy substrate and restore complex I activity via sirtuin-dependent pathways. EGCG acts predominantly at the peripheral level by reducing systemic inflammation and oxidative stress. EA-derived urolithins effectively cross the blood-brain barrier to directly enhance mitochondrial biogenesis and respiratory chain function in the central nervous system. Clinical trials have reported improvements in fatigue, cognition, mood, and muscle function following supplementation with these compounds. The convergence of their actions on energy restoration, reactive oxygen species reduction, and epigenetic modulation of protective pathways suggests their synergistic potential. Optimized delivery strategies, including exogenous ketone salts, liposomal EGCG, and microencapsulated EA, may overcome bioavailability limitations and interindividual variability in the gut microbiota metabolism.\n\nID: 41802693\nTitle: IGFBP7 protects against endothelial apoptosis and blood-brain barrier disruption following oxygen-glucose deprivation/reoxygenation.\nAbstract: The blood-brain barrier (BBB) is a critical interface maintained by cerebral microvascular endothelial cells, which rely on tight junction (TJ) proteins like ZO-1 and occludin to regulate permeability. Following ischemic stroke, BBB disruption and subsequent cerebral edema are major contributors to poor clinical outcomes. Insulin-like growth factor-binding protein 7 (IGFBP7) is highly expressed in these endothelial cells and has been implicated in both BBB formation and vascular repair. However, its specific role in stroke-induced endothelial injury has remained inconclusive. In this study, we investigated the function of IGFBP7 in an in vitro model of oxygen-glucose deprivation and reoxygenation (OGD/R) using mouse microvascular endothelial (bEnd.3) cells. We manipulated IGFBP7 levels through overexpression and siRNA-mediated silencing. Our results show that OGD/R treatment significantly decreased IGFBP7 expression, which led to increased BBB permeability, degradation of tight junction proteins, and cellular apoptosis. Importantly, overexpressing IGFBP7 mitigated these effects, restoring the expression of ZO-1 and reducing apoptosis. Conversely, silencing IGFBP7 exacerbated both tight junction protein degradation and cell apoptosis. Endothelial IGFBP7 exerts a protective role in ischemic stroke by blunting OGD/R-mediated BBB damage and thus, it may represent an interesting novel therapeutic target to be explored in future clinical investigation.\n\nID: 41785850\nTitle: Efficient amyloid-\u03b2 degradation in Alzheimer's disease using SPYTACs.\nAbstract: Clearance of aberrant cerebral amyloid-\u03b2 (A\u03b2) deposits represents a promising therapeutic strategy for Alzheimer's disease (AD), yet current anti-A\u03b2 immunotherapy raises safety concerns due to frequent adverse effects. Extracellular targeted protein degradation (eTPD) offers an approach for safe and efficient clearance of disease-causing proteins. Here, we develop a next-generation eTPD platform, synthetic peptide-programmed lysosome-targeting chimeras (SPYTACs), using entirely synthesized bispecific peptides. Leveraging low-density lipoprotein receptor-related protein 1 (LRP1), SPYTACs effectively facilitate targeted degradation of extracellular proteins and enable transcytosis across the blood-brain barrier. In vivo administration of SPYTACs effectively reduces peripheral and cerebral A\u03b2 burden, attenuates synapse loss, and improves cognitive function in 5\u00d7FAD mice at both prodromal and symptomatic stages. Notably, SPYTAC treatment shows fewer side effects, including intracerebral hemorrhage and inflammation, compared with conventional immunotherapies. The high modularity and genetic encodability enable SPYTACs to target customized disease-causing proteins, underscoring their therapeutic versatility and translational promise across diverse diseases driven by pathogenic proteins.\n\nID: 42249126\nTitle: The significance of the ketolytic gene OXCT1 in metabolism, intracellular signaling and disease development.\nAbstract: OXCT1 (3-oxoacid CoA-transferase 1), encoding the mitochondrial enzyme SCOT (succinyl-CoA:3-ketoacid CoA transferase), is a key enzyme in ketone body utilization. It catalyzes the reversible transfer of CoA from succinyl-CoA to acetoacetate, generating acetoacetyl-CoA, which is subsequently converted into acetyl-CoA for entry into the tricarboxylic acid (TCA) cycle and mitochondrial energy production. Although classically regarded as a metabolic enzyme, emerging evidence indicates that OXCT1 participates in broader regulatory networks. In addition to its protein-coding transcript, the OXCT1 locus produces regulatory non-coding RNAs, including circ-OXCT1 and lncRNA OXCT1-AS1, which provides additional levels of regulation, as for now reported in various types of cancer. Moreover, the review summarizes current knowledge on OXCT1 biochemical function, regulation, and tissue distribution, with emphasis on transcriptional control, post-translational modifications such as lysine succinylation and redox-dependent regulation, and integration with nutrient-sensing and stress-response pathways. By combining recent literature with bioinformatics analysis, we demonstrate that OXCT1 displays highly dynamic expression across diverse cancer types and metabolic states, consistent with a role in metabolic plasticity and disease progression. Furthermore, we discuss here the OXCT1 role in other pathological conditions including metabolic disorders, neurological disease, and cardiomyopathy, implicating both metabolic dysfunction and aberrant protein modification in disease mechanisms. Collectively, these findings establish OXCT1 as a central regulator of metabolic adaptation and a potential therapeutic target.\n\nID: 41213123\nTitle: \u03b2-Hydroxybutyrylation Links Ketone Metabolism to Mitochondrial Remodeling in Diabetic Cardiomyopathy.\nAbstract: Diabetic cardiomyopathy (DbCM) is characterized by metabolic remodeling and energetic stress independent of coronary artery disease. Increased reliance on fatty acid and ketone body metabolism has been observed in DbCM, but the regulatory mechanisms linking altered substrate use to myocardial dysfunction remain poorly understood. In particular, lysine \u03b2-hydroxybutyrate (Kbhb), a ketone body-derived, posttranslational modification, has emerged as a potentially critical regulator but has not been fully investigated. We conducted a comprehensive multiomics study integrating metabolomics, transcriptomics, proteomics, and Kbhb-specific proteomics on myocardial tissues in a well-established mouse model of DbCM. Kbhb-modified proteins were systematically mapped and quantified, followed by motif, subcellular localization, and protein-protein interaction analyses. DbCM cardiac tissue exhibited coordinated upregulations of fatty acid \u03b2-oxidation, ketone metabolism, and tricarboxylic acid cycle activity at the transcriptomic, proteomic, and metabolomic levels. Kbhb profiling revealed extensive mitochondrial protein modification, with Atp5f1a-K239 identified as a key modification site strongly correlated with \u03b2-hydroxybutyrate and isocitric acid concentrations. This study identifies Kbhb as a potential metabolic-epigenetic modifier linking ketone body availability to the regulation of mitochondrial proteins in DbCM. Our findings provide novel insights into metabolic-epigenetic cross talk and identify potential therapeutic targets for interventions to restore mitochondrial function in alleviating diabetic heart disease. We performed a multiomics study to better understand dysfunctions in diabetic cardiomyopathy (DbCM) and specifically identify links between lysine \u03b2-hydroxybutyrylation (Kbhb), a ketone body-derived, posttranslational modification, and cardiac dysfunction. DbCM cardiac tissue exhibited coordinated upregulations of fatty acid \u03b2-oxidation, ketone metabolism, and tricarboxylic acid cycle activity at the transcriptomic, proteomic, and metabolomic levels. Mitochondrial proteins showed that high Kbhb modification and modification of the Atp5f1a-K239 site were strongly correlated with high \u03b2-hydroxybutyrate and isocitric acid concentrations. This study identifies Kbhb modification of mitochondrial proteins as a potential mechanism linking ketone body availability to mitochondrial function in DbCM.\n\nID: 39626664\nTitle: \u03b2-hydroxybutyrate is a metabolic regulator of proteostasis in the aged and Alzheimer disease brain.\nAbstract: Loss of proteostasis is a hallmark of aging and Alzheimer disease (AD). We identify \u03b2-hydroxybutyrate (\u03b2HB), a ketone body, as a regulator of protein solubility. \u03b2HB primarily provides ATP substrate during periods of reduced glucose availability, and regulates other cellular processes through protein interactions. We demonstrate \u03b2HB-induced protein insolubility is not dependent on covalent protein modification, pH, or solute load, and is observable in mouse brain in\u00a0vivo after delivery of a ketone ester. This mechanism is selective for pathological proteins such as amyloid-\u03b2, and exogenous \u03b2HB ameliorates pathology in nematode models of amyloid-\u03b2 aggregation toxicity. We generate libraries of the \u03b2HB-induced protein insolublome using mass spectrometry proteomics, and identify common protein domains and upstream regulators. We show enrichment of neurodegeneration-related proteins among \u03b2HB targets and the clearance of these targets from mouse brain. These data indicate a metabolically regulated mechanism of proteostasis relevant to aging and AD.\n\nID: 39587264\nTitle: The tRNA methyltransferase Mettl1 governs ketogenesis through translational regulation and drives metabolic reprogramming in cardiomyocyte maturation.\nAbstract: After birth, the heart undergoes a shift in energy metabolism and cytoarchitecture to enhance efficient energy production and cardiac contraction, which is essential for postnatal development and growth. However, the precise mechanisms regulating this process remain elusive. Here we show that the RNA modification enzyme Mettl1 is a critical regulator of postnatal metabolic reprogramming and cardiomyocyte maturation in mice, primarily through its influence on the translation of the rate-limiting ketogenesis enzyme Hmgcs2. Our findings reveal that ketogenesis is vital for the postnatal transition of fuel from glucose to fatty acids in cardiomyocytes, achieved by modulating tricarboxylic acid cycle-related enzymatic activity via lysine \u03b2-hydroxybutyrylation protein modification. Loss of Mettl1 results in aberrant metabolic reprogramming and cardiomyocyte immaturity, leading to heart failure, although some clinical features can be rescued by \u03b2-hydroxybutyrate supplementation. Our study provides mechanistic insights into how Mettl1 regulates metabolic reprogramming in neonatal cardiomyocytes and highlights the importance of ketogenesis in cardiomyocyte maturation.\n\nID: 38865959\nTitle: Dual-mode upconversion sensors for detecting differently charged biotargets based on the oxidase-mimicking activity of Ce4+ and electrostatic control.\nAbstract: Heparin is a highly negatively charged sulfated linear polymer glycosaminoglycan that has been widely used as an anticoagulant in medicine. Protamine is a cationic protein rich in arginine that is used to treat the blood-brain barrier during excess heparin surgery. Trypsin is the most important digestive enzyme-encoding generated by the pancreas and can specifically cleave the carboxyl ends of arginine and lysine residues. Heparin, protamine, and trypsin interact and constrain each other, and their fluctuations reflect the body's dysfunction. Therefore, it is necessary to develop a fast, sensitive, and highly selective assay for regularly monitoring the levels of heparin, protamine, and trypsin in serum. Herein, a fluorescent and colorimetric dual-mode upconversion nanoparticle (UCNP) biosensor was used for the determination of heparin, protamine, and trypsin based on the oxidase-mimicking activity of Ce4+ and electrostatic control. The biosensor exhibited sensitive detection of heparin, protamine, and trypsin with low limits of detection (LODs) of 16\u00a0ng/mL, 87\u00a0ng/mL and 31\u00a0ng/mL, respectively. Furthermore, the designed biosensor could eliminate autofluorescence, which not only effectively increased the accuracy of the sensor but also provided a new sensing pathway for the detection of differently charged biotargets.\n\nID: 37984250\nTitle: Knockdown of Smox protects the integrity of the blood-brain barrier through antioxidant effect and Nrf2 pathway activation in stroke.\nAbstract: Once an ischemic stroke occurs, reactive oxygen species (ROS) and oxidative stress degrade the tight connections between cerebral endothelial cells resulting in their damage. The expression of antioxidant genes may be enhanced, and ROS formation may be reduced following Nrf2 activation, which is associated with protection against ischemic stroke. Overexpression of spermine oxidase (Smox) in the neocortex led to increased H2O2 production. However, how Smox impacts the regulation of the blood-brain barrier (BBB) through antioxidants has not been examined yet. We conducted experiments both in the cell level and in the transient middle cerebral artery occlusion (tMCAO) model to evaluate the effect of Smox siRNA lentivirus (si-Smox) knockdown on BBB protection against ischemic stroke. Mice treated with si-Smox showed remarkably decreased BBB breakdown and reduced endothelial inflammation following stroke. The treatment with si-Smox significantly elevated the Bcl-2 to Bax ratio and decreased the production of cleaved caspase-3 in the tMCAO model. Further investigation revealed that the neuroprotective effect was the result of the antioxidant properties of si-Smox, which reduced oxidative stress and enhanced CD31+ cells in the peri-infarct cortical areas. Of significance, si-Smox activated Nrf2 in both bEnd.3 cells and tMCAO animals, and blocking Nrf2 with brusatol diminished the protective effects of si-Smox. The study findings suggest that si-Smox exerts neuroprotective effects and promotes angiogenesis by activating the Nrf2 pathway, thus decreasing oxidative stress and apoptosis caused by tMCAO. As a result, si-Smox may hold potential as a therapeutic candidate for preserving BBB integrity while treating ischemic stroke.\n\nID: 37461525\nTitle: \u03b2-hydroxybutyrate is a metabolic regulator of proteostasis in the aged and Alzheimer disease brain.\nAbstract: Loss of proteostasis is a hallmark of aging and Alzheimer disease (AD). Here, we identify \u03b2-hydroxybutyrate (\u03b2HB), a ketone body, as a regulator of protein solubility in the aging brain. \u03b2HB is a small molecule metabolite which primarily provides an oxidative substrate for ATP during hypoglycemic conditions, and also regulates other cellular processes through covalent and noncovalent protein interactions. We demonstrate \u03b2HB-induced protein insolubility across in vitro, ex vivo, and in vivo mouse systems. This activity is shared by select structurally similar metabolites, is not dependent on covalent protein modification, pH, or solute load, and is observable in mouse brain in vivo after delivery of a ketone ester. Furthermore, this phenotype is selective for pathological proteins such as amyloid-\u03b2, and exogenous \u03b2HB ameliorates pathology in nematode models of amyloid-\u03b2 aggregation toxicity. We have generated a comprehensive atlas of the \u03b2HB-induced protein insolublome ex vivo and in vivo using mass spectrometry proteomics, and have identified common protein domains within \u03b2HB target sequences. Finally, we show enrichment of neurodegeneration-related proteins among \u03b2HB targets and the clearance of these targets from mouse brain, likely via \u03b2HB-induced autophagy. Overall, these data indicate a new metabolically regulated mechanism of proteostasis relevant to aging and AD.\n\nID: 36587768\nTitle: Carnitine octanoyltransferase is important for the assimilation of exogenous acetyl-L-carnitine into acetyl-CoA in mammalian cells.\nAbstract: In eukaryotes, carnitine is best known for its ability to shuttle esterified fatty acids across mitochondrial membranes for \u03b2-oxidation. It also returns to the cytoplasm, in the form of acetyl-L-carnitine (LAC), some of the resulting acetyl groups for posttranslational protein modification and lipid biosynthesis. While dietary LAC supplementation has been clinically investigated, its effects on cellular metabolism are not well understood. To explain how exogenous LAC influences mammalian cell metabolism, we synthesized isotope-labeled forms of LAC and its analogs. In cultures of glucose-limited U87MG glioma cells, exogenous LAC contributed more robustly to intracellular acetyl-CoA pools than did \u03b2-hydroxybutyrate, the predominant circulating ketone body in mammals. The fact that most LAC-derived acetyl-CoA is cytosolic is evident from strong labeling of fatty acids in U87MG\u00a0cells by exogenous 13C2-acetyl-L-carnitine. We found that the addition of d3-acetyl-L-carnitine increases the supply of acetyl-CoA for cytosolic posttranslational modifications due to its strong kinetic isotope effect on acetyl-CoA carboxylase, the first committed step in fatty acid biosynthesis. Surprisingly, whereas cytosolic carnitine acetyltransferase is believed to catalyze acetyl group transfer from LAC to coenzyme A, CRAT-/- U87MG\u00a0cells were unimpaired in their ability to assimilate exogenous LAC into acetyl-CoA. We identified carnitine octanoyltransferase as the key enzyme in this process, implicating a role for peroxisomes in efficient LAC utilization. Our work has opened the door to further biochemical investigations of a new pathway for supplying acetyl-CoA to certain glucose-starved cells.\n\nID: 36358597\nTitle: Spermidine/Spermine N1-Acetyltransferase 1 (SAT1)-A Potential Gene Target for Selective Sensitization of Glioblastoma Cells Using an Ionizable Lipid Nanoparticle to Deliver siRNA.\nAbstract: Spermidine/spermine N1-acetyltransferase 1 (SAT1) responsible for cell polyamine catabolism is overexpressed in glioblastoma multiforme (GB). Its role in tumor survival and promoting resistance towards radiation therapy has made it an interesting target for therapy. In this study, we prepared a lipid nanoparticle-based siRNA delivery system (LNP-siSAT1) to selectively knockdown (KD) SAT1 enzyme in a human glioblastoma cell line. The LNP-siSAT1 containing ionizable DODAP lipid was prepared following a microfluidics mixing method and the resulting nanoparticles had a hydrodynamic size of around 80 nm and a neutral surface charge. The LNP-siSAT1 effectively knocked down the SAT1 expression in U251, LN229, and 42MGBA GB cells, and other brain-relevant endothelial (hCMEC/D3), astrocyte (HA) and macrophage (ANA-1) cells at the mRNA and protein levels. SAT1 KD in U251 cells resulted in a 40% loss in cell viability. Furthermore, SAT1 KD in U251, LN229 and 42MGBA cells sensitized them towards radiation and chemotherapy treatments. In contrast, despite similar SAT1 KD in other brain-relevant cells no significant effect on cytotoxic response, either alone or in combination, was observed. A major roadblock for brain therapeutics is their ability to cross the highly restrictive blood-brain barrier (BBB) presented by the brain microcapillary endothelial cells. Here, we used the BBB circumventing approach to enhance the delivery of LNP-siSAT1 across a BBB cell culture model. A cadherin binding peptide (ADTC5) was used to transiently open the BBB tight junctions to promote paracellular diffusion of LNP-siSAT1. These results suggest LNP-siSAT1 may provide a safe and effective method for reducing SAT1 and sensitizing GB cells to radiation and chemotherapeutic agents.\n\nID: 33391334\nTitle: Metabolomic and Proteomic Profiles Associated With Ketosis in Dairy Cows.\nAbstract: Ketosis is a common metabolic disease in dairy cows during early lactation. However, information about the metabolomic and proteomic profiles associated with the incidence and progression of ketosis is still limited. In this study, an integrated metabolomics and proteomics approach was performed on blood serum sampled from cows diagnosed with clinical ketosis (case, \u2265 2.60 mmol/L plasma \u03b2-hydroxybutyrate; BHBA) and healthy controls (control, < 1.0 mmol/L BHBA). Samples were taken 2 weeks before parturition and 2 weeks after parturition from 19 animals (nine cases, 10 controls). All serum samples (n = 38) were subjected to Liquid Chromatography-Mass Spectrometry (LC-MS) based metabolomic analysis, and 20 samples underwent Data-Independent Acquisition (DIA) LC-MS based proteomic analysis. A total of 97 metabolites and 540 proteins were successfully identified, and multivariate analysis revealed significant differences in both metabolomic and proteomic profiles between cases and controls. We investigated clinical ketosis-associated metabolomic and proteomic changes using statistical analyses. Correlation analysis of statistically significant metabolites and proteins showed 78 strong correlations (correlation coefficient, R \u2265 0.7) between 38 metabolites and 25 proteins, which were then mapped to pathways using IMPaLA. Results showed that ketosis altered a wide range of metabolic pathways, such as metabolism, metabolism of proteins, gene expression and post-translational protein modification, vitamin metabolism, signaling, and disease related pathways. Findings presented here are relevant for identifying molecular targets for ketosis and biomarkers for ketosis detection during the transition period.\n\nID: 27105115\nTitle: Metabolic Regulation of Gene Expression by Histone Lysine \u03b2-Hydroxybutyrylation.\nAbstract: Here we report the identification and verification of a \u03b2-hydroxybutyrate-derived protein modification, lysine \u03b2-hydroxybutyrylation (Kbhb), as a new type of histone mark. Histone Kbhb marks are dramatically induced in response to elevated \u03b2-hydroxybutyrate levels in cultured cells and in livers from mice subjected to prolonged fasting or streptozotocin-induced diabetic ketoacidosis. In total, we identified 44 histone Kbhb sites, a figure comparable to the\u00a0known number of histone acetylation sites. By ChIP-seq and RNA-seq analysis, we demonstrate that histone Kbhb is a mark enriched in active gene promoters and that the increased H3K9bhb levels that occur during starvation are associated with genes upregulated in starvation-responsive metabolic pathways. Histone \u03b2-hydroxybutyrylation thus represents a new epigenetic regulatory mark that couples metabolism to gene expression, offering a\u00a0new avenue to study chromatin regulation and diverse functions of \u03b2-hydroxybutyrate in the context of important human pathophysiological states, including diabetes, epilepsy, and neoplasia.\n\nID: 25029034\nTitle: Synthesis and in vitro evaluation of BBB permeability, tumor cell uptake, and cytotoxicity of a series of carboranylporphyrin conjugates.\nAbstract: A series of tri[(p-carboranylmethylthio)tetrafluorophenyl]porphyrin conjugates of linear and branched polyamines, glucose, arginine, tri(ethylene glycol), and Tyr-D-Arg-Phe-\u03b2-Ala (YRFA) peptide were synthesized. These conjugates were investigated for their BBB permeability in human hCMEC/D3 brain endothelial cells, and their cytotoxicity and uptake were assessed using human glioma T98G cells. For comparison purposes, a symmetric tetra[(p-carboranylmethylthio)tetrafluorophenyl]porphyrin was also synthesized, and its crystal structure was obtained. All porphyrin conjugates show low dark cytotoxicity (IC50>400 \u03bcM) and low phototoxicity (IC50>100 \u03bcM at 1.5 J/cm2) toward T98G cells. All conjugates were efficiently taken up by T98G cells, particularly the cationic polyamine and arginine conjugates, and were localized in multiple cellular organelles, including mitochondria and lysosomes. All compounds showed relatively low in vitro BBB permeability compared with that of lucifer yellow because of their higher molecular weight, hydrophobicity, and tendency for aggregation in solution. Within this series, the branched polyamine and YRFA conjugates showed the highest permeability coefficient, whereas the glucose conjugate showed the lowest permeability coefficient.\n\nID: 24105845\nTitle: RBE4 cells are highly resistant to paraquat-induced cytotoxicity: studies on uptake and efflux mechanisms.\nAbstract: Paraquat (PQ) is a widely used, highly toxic and non-selective contact herbicide, which has been associated with central neurotoxic effects, namely the development of Parkinson's disease, but whose effects to the blood-brain barrier (BBB) itself have rarely been studied. This work studied the mechanisms of PQ uptake and efflux in a rat's BBB cell model, the RBE4 cells. PQ is believed to enter cells using the basic or neutral amino acid or polyamine transport systems or through the choline-uptake system. In contrast, PQ efflux from cells is reported to be mediated by P-glycoprotein. Therefore, we evaluated PQ-induced cytotoxicity and the effect of some substrates/blockers of these transporters (such as arginine, L-valine, putrescine, hemicholinium-3 and GF120918) on such cytotoxicity. RBE4 cells were shown to be extremely resistant to PQ after 24\u2009h of exposure; even at concentrations as high as 50\u2009mM approximately 45% of the cells remained viable. Prolonging exposure until 48\u2009h elicited significant cytotoxicity only for PQ concentrations above 5\u2009mM. Although hemicholinium-3, a choline-uptake system inhibitor, significantly protected cells against PQ-induced toxicity, none of the effects were observed for arginine, L-valine or putrescine. Meanwhile, inhibiting the efflux pump P-glycoprotein using GF120918 significantly enhanced PQ-induced cytotoxicity. In conclusion, PQ used the choline-uptake system, instead of the transporters for the basic or neutral amino acids or for the polyamines, to enter RBE4 cells. P-glycoprotein extrudes PQ back to the extracellular medium. However, this efflux mechanism only partially explains the observed RBE4 resistance to PQ.\n\nID: 18726697\nTitle: CNS delivery via adsorptive transcytosis.\nAbstract: Adsorptive-mediated transcytosis (AMT) provides a means for brain delivery of medicines across the blood-brain barrier (BBB). The BBB is readily equipped for the AMT process: it provides both the potential for binding and uptake of cationic molecules to the luminal surface of endothelial cells, and then for exocytosis at the abluminal surface. The transcytotic pathways present at the BBB and its morphological and enzymatic properties provide the means for movement of the molecules through the endothelial cytoplasm. AMT-based drug delivery to the brain was performed using cationic proteins and cell-penetrating peptides (CPPs). Protein cationization using either synthetic or natural polyamines is discussed and some examples of diamine/polyamine modified proteins that cross BBB are described. Two main families of CPPs belonging to the Tat-derived peptides and Syn-B vectors have been extensively used in CPP vector-mediated strategies allowing delivery of a large variety of small molecules as well as proteins across cell membranes in vitro and the BBB in vivo. CPP strategy suffers from several limitations such as toxicity and immunogenicity--like the cationization strategy--as well as the instability of peptide vectors in biological media. The review concludes by stressing the need to improve the understanding of AMT mechanisms at BBB and the effectiveness of cationized proteins and CPP-vectorized proteins as neurotherapeutics.\n\nID: 18712585\nTitle: Development of a smart nano-vehicle to target cerebrovascular amyloid deposits and brain parenchymal plaques observed in Alzheimer's disease and cerebral amyloid angiopathy.\nAbstract: To design a smart nano-vehicle (SNV) capable of permeating the blood-brain barrier (BBB) to target cerebrovascular amyloid formed in both Alzheimer's disease (AD) and cerebrovascular amyloid angiopathy (CAA). SNV consists of a chitosan polymeric core prepared through ionic gelation with tripolyphosphate. A polyamine modified F(ab') portion of IgG4.1, an anti-amyloid antibody, was coated as a biosensor on the SNV surface. A similar polymeric core coated with bovine serum albumin (BSA) served as a control nano-vehicle (CNV). The BBB uptake of (125)I-SNVs and (125)I-CNVs was evaluated in mice. The uptake and transcytosis of SNVs and CNVs across bovine brain microvascular endothelial cells (BBMECs) was evaluated using flow cytometry and confocal microscopy. Plasma clearance of (125)I-SNVs was nine times higher than that of the (125)I-CNVs. However, the uptake of (125)I-SNVs in various brain regions was about 8 to 11 times higher than that of (125)I-CNVs. The uptake of FITC-BSA loaded SNVs in BBMECs was twice the uptake of FITC-BSA loaded CNVs. Confocal micrographs demonstrated the uptake and transcytosis of Alexa Fluor 647 labeled SNVs, but not CNVs, across the BBMEC monolayer. SNVs are capable of carrying a payload of model protein across the BBB to target cerebral amyloid.\n\nID: 9824696\nTitle: Lack of evidence for direct involvement of NMDA receptors or polyamines in blood-brain barrier injury after cerebral ischemia in rats.\nAbstract: It is hypothesized that after various types of brain injury, blood-brain barrier (BBB) opening and vasogenic edema result from excessive neuronal release of glutamate and stimulation of capillary N-methyl-d-aspartate (NMDA) receptors linked to polyamine (putrescine) synthesis in endothelial cells. We produced cerebral ischemia in rats and measured BBB opening 6 h later as the increase in regional transfer constants (Ki) for blood to brain diffusion of [3H]sucrose. Such BBB opening was not mitigated by drugs which block NMDA receptors (MK801 or AR-R 15896AR) or polyamine synthesis (difluoromethylornithine). These results question generality of the capillary NMDA receptor/polyamine hypothesis.\n\nID: 8149897\nTitle: Pharmacokinetics and saturable blood-brain barrier transport of biotin bound to a conjugate of avidin and a monoclonal antibody to the transferrin receptor.\nAbstract: The delivery of biotinylated therapeutics through the blood-brain barrier (BBB) may be facilitated by the use of avidin-based chimeric peptide conjugates. The latter are formed by conjugating avidin to a BBB drug delivery vector, which is a protein that undergoes receptor-mediated transcytosis through the BBB. The murine OX26 monoclonal antibody to the rat transferrin receptor undergoes receptor-mediated transport through the BBB, and previous studies have shown that a [3H]biotin/avidin-OX26 conjugate is effectively transported through the BBB. However, avidin is a cationic protein, which causes a marked increase in the systemic clearance of avidin-based conjugates from the plasma compartment. The present studies describe attempts to elevate the reduced plasma area under the curve (AUC) of [3H]biotin/avidin-OX26 by preloading or coloading with unconjugated OX26 antibody or unconjugated avidin. Both systemic clearance and BBB transport of avidin-OX26 were equally affected by OX26 preloading or coloading; this had inverse effects on the plasma AUC and the BBB permeability surface area product with no resulting change in the fractional delivery of [3H]biotin to brain. Conversely, avidin coloading preferentially reduced brain clearance of the [3H]biotin/avidin-OX26 conjugate, without substantial alteration in the plasma AUC and greatly reduced the fractional delivery of [3H]biotin to brain. In summary, these studies show that the use of avidin-based vectors results in rapid systemic clearance, which causes a reduction in the delivery of [3H]biotin to brain, despite a comparable BBB permeability coefficient for either the unconjugated OX26 antibody or the avidin-OX26 conjugate.\n\nID: 1602382\nTitle: Transport of recombinant CD4 through the rat blood-brain barrier in vivo.\nAbstract: One class of potential acquired immunodeficiency syndrome therapeutics are derivatives of recombinant CD4 (rCD4). Therefore, the present investigations use in vivo techniques to measure the rate at which [3H]rCD4 is transported through the blood-brain barrier (BBB). In addition, the binding of labeled rCD4 to isolated human and bovine brain capillaries is measured. These studies show that [3H]CD4 is removed rapidly from the bloodstream with a half-time of 12.6 +/- 0.9 min. The volume of distribution (Vd) of the protein in brain increases with time and reaches a Vd that is 11.1 +/- 1.1-fold greater than the brain Vd of plasma marker, native rat serum albumin. In addition, [3H]rCD4 is extracted rapidly by the kidney and the ratio of rCD4 Vd to native rat serum albumin Vd in the rat kidney reaches 99 +/- 5 at 60 min after i.v. injection. rCD4 is shown to undergo transcytosis through the BBB using an internal carotid artery perfusion/capillary depletion method coupled with gel filtration fast protein liquid chromatography. In conclusion, these studies report the unexpected finding that rCD4 is transportable through the BBB. rCD4 is a cationic protein and the mechanism of rCD4 transport through the BBB may be analogous to the absorptive-mediated transcytosis of other polycationic proteins.\n\nID: 2600816\nTitle: Transport of histone through the blood-brain barrier.\nAbstract: The present studies were designed to determine if the endogenous cationic protein, e.g., histone, is capable of penetrating the blood-brain barrier (BBB) in vivo. Calf thymus histone was iodinated with [125I]iodine and was found to be taken up rapidly by isolated bovine brain capillaries used as an in vitro model system of the BBB via a time- and temperature-dependent mechanism. The binding was saturable and a Scatchard plot of the binding data was linear, yielding a KD = 15.2 +/- 2.8 microM and a maximal binding = 7.7 +/- 1.0 nmol/mg of protein. Other polycations such as protamine or polylysine markedly inhibited uptake of [125I] histone, but cationized albumin demonstrated minimal inhibition and cationized immunoglobulin caused no inhibition of bovine brain capillary uptake of [125I]histone. The in vivo brain VD of [125I] histone reached 159 +/- 70 microliters/g by 10 min of carotid arterial perfusion as compared to the 10-min VD for [3H]albumin, 17 +/- 7 microliter/g. Most of this uptake represented sequestration by the vasculature, but approximately 8% of the total histone taken up by brain was found to be transported unmetabolized (based on trichloroacetic acid precipitability of brain supernatant [( 125I]) into brain interstitium. These studies demonstrate that histone is transported through the BBB in vivo via absorptive-mediated transport. Thus, histone is an endogenous protein that is capable of transport through the BBB and may be a potential vector for pharmaceutical delivery through the BBB.\n\nID: 3097421\nTitle: Blood-brain barrier breakdown by cold injury. Polyamine signals mediate acute stimulation of endocytosis, vesicular transport, and microvillus formation in rat cerebral capillaries.\nAbstract: Polyamines have been previously implicated in the mediation of blood-brain barrier breakdown induced by cryogenic injury (H Koenig, AD Goldstone, CY Lu, Biochem Biophys Res Commun 116:1039, 1983). We studied acute (less than 5 minute) changes in capillary ultrastructure, microvascular permeability, and the levels of polyamines and their rate regulating synthetic enzyme ornithine decarboxylase (ODC) in rat cerebral cortex after focal cold injury. Microvascular permeability was measured by relative transport of intravenously administered fluorescein. Capillary ultrastructure was studied by quantitative stereology and morphometry after intravenous administration of horseradish peroxidase. Focal cold injury induced a 2.5-, 3.8-, 1.7-, and 1.4-fold increase in the levels of ODC, putrescine, spermidine and spermine, and a 46-fold increase in fluorescein uptake in perilesional cortex. Few capillaries in control cortex contained endocytic pits or horseradish peroxidase-positive vesicles, whereas most capillaries near lesions showed these structures. Cryoinjury induced a 5-fold increase in the relative volume of microvilli and horseradish peroxidase vesicles, a 2.3-fold increase in area of luminal endocytic pits, and a 6.3-fold increase in area of abluminal exocytic pits. The ODC inhibitor alpha-difluoromethylornithine blocked the cryoinjury-induced changes in ODC, polyamines, fluorescein uptake, and capillary ultrastructure. Putrescine negated the effect of alpha-difluoromethylornithine or capillary ultrastructure, and was previously shown to nullify the alpha-difluoromethylornithine effects on polyamines and fluorescein permeability (cited above). These data link rapid changes in ODC and polyamines to blood-brain barrier breakdown, and suggest that the abnormal permeability is associated with an acute, polyamine-mediated stimulation of microvillus formation, endocytosis, and vesicular transport in capillary endothelium.\n\nID: 6842259\nTitle: Eosinophil cationic protein (ECP) in the cerebrospinal fluid.\nAbstract: ECP (eosinophil cationic protein) has been measured by means of a specific radioimmunoassay in the cerebrospinal fluid (CSF) from 210 individuals with various diseases affecting the central nervous system. In the same specimens lactoferrin and albumin were measured as well, as indicators of neutrophil-involved inflammation and damage to the blood-brain barrier. From a patient reference group (n = 39) the upper \"normal\" limit for ECP was estimated to 1.7 microgram/l. In patients with acute cerebrovascular disease (n = 108) ECP levels were elevated in 38% of the cases which was a significantly (P less than 0.001) greater proportion than seen for lactoferrin (7%). In patients with acute infections of the CNS (n = 30) 67% had raised ECP levels with significantly higher levels (P less than 0.001) in those having bacterial infections. The ECP levels were significantly correlated (P less than 0.001) to the lactoferrin-levels in the whole infectious group. In patients with tumours (n = 25) raised levels of ECP were found in 67% of those with malignant and in 6% of those having benign tumours. This difference was statistically significant (P = 0.001). The ECP levels were closely related to those of lactoferrin (P less than 0.001) and albumin (P less than 0.005). Of the patients with multiple sclerosis (n = 19) 25% had raised ECP levels. This proportion was not significantly different from those having raised lactoferrin levels. In three patients extremely high ECP levels (70-455 micrograms/l) were found and a causal relationship between ECP and the brain tissue damage in these patients is suggested. In comparison with the neutrophil-related data the findings suggest a preferential involvement of eosinophils in some diseases affecting the central nervous system.\n\nID: 42498022\nTitle: Genome-resolved characterization of candidate thermotolerance traits and predicted protein conformational behavior in Calditerricola during hyperthermophilic composting of organic wastes.\nAbstract: Hyperthermophilic composting (HC) can generate temperatures above 80\u00a0\u00b0C without external heating, thereby accelerating organic-waste stabilization; however, how dominant heat-adapted microorganisms maintain cellular function under such extreme conditions remains unclear. Here, we integrated metagenomics, metagenome-assembled genome reconstruction, Calditerricola-resolved functional profiling, partial least squares path modeling, and molecular dynamics simulations to investigate candidate thermotolerance-related traits associated with Calditerricola enrichment during HC. The pile temperature reached 82.6\u00a0\u00b0C on day 2 and peaked at 86.6\u00a0\u00b0C on day 4, accompanied by progressive humification, with humic substances increasing from 40.45 to 51.28\u00a0mg/g and HA/FA reaching 3.45. Microbial communities differed significantly among composting phases (R2\u00a0=\u00a00.975, P\u00a0=\u00a00.004), and Calditerricola increased from 0.02% in the initial phase to 6.1% in the thermophilic phase before declining to 0.7% in maturation. Community-level profiles showed comparatively modest variation in selected thermotolerance-related pathways, whereas the independently normalized Calditerricola profile displayed clearer phase-associated increases in functions linked to polyamine metabolism, membrane/envelope homeostasis, proteostasis, and DNA maintenance. Path modeling revealed consistent positive associations between Calditerricola enrichment and polyamine synthesis, membrane stability, and proteostasis. qPCR further revealed phase-associated increases in the community-level copy numbers of representative target genes, particularly polA and speE. During 100-ns simulations at 360\u00a0K, the predicted apo structures of PolA, AtpD, SpeE, and FabH retained their overall folds and comparatively stable catalytic-residue geometries. Together, these results define an association-based multi-module framework of candidate traits linked to Calditerricola persistence during HC, providing a genome-resolved basis for prioritizing testable thermotolerance targets in engineered high-temperature waste-treatment systems.\n\nID: 42467639\nTitle: N-acetyl-L-leucine normalizes Transcription Factor EB activity by stereospecific bidirectional modulation in a HeLa cell model of Niemann-Pick disease type C.\nAbstract: Levacetylleucine (Aqneursa\u2122), an acetylated derivative and pro-drug of L-leucine, is the only FDA-approved monotherapy for Niemann-Pick disease type C (NPC). Its acetyl group enables transport via monocarboxylate transporters, supporting blood-brain barrier penetration and efficient cellular uptake. Inside cells, levacetylleucine is metabolised by acylases, generating elevated levels of L-leucine that enhance mitochondrial bioenergetics and is thought to ameliorate lysosomal dysfunction indirectly. Here, we describe a direct effect of levacetylleucine on lysosomal regulation through modulation of TFEB, the master transcription factor for lysosomal and autophagy genes. Levacetylleucine rapidly alters TFEB translocation between the cytoplasm and the nucleus in a biphasic, homeostasis-restoring manner. In wild-type HeLa cells, levacetylleucine promotes TFEB activation and nuclear localisation. However, in NPC1 disease models, where we show that TFEB is over-activated and enriched in the nucleus due to lysosomal stress, levacetylleucine reduces nuclear TFEB and restores a more normal cytoplasmic-to-nuclear balance. These effects occur at clinically relevant concentrations associated with lysosomal storage reduction. The effects of the drug are stereospecific: while the L-enantiomer is active, the D-enantiomer and racemate show no effect, revealing the antagonistic properties of the D-enantiomer. This bidirectional normalisation of TFEB activity highlights a direct mechanism through which levacetylleucine modulates lysosomal and autophagic pathways in the HeLa cell model, giving mechanistic insight into its therapeutic potential in NPC, and also across diverse neurological and neurodevelopmental disorders.\n\nID: 42429378\nTitle: Integrated multi-omics analysis reveals a pH-driven metabolic and translational switch in Ureaplasma parvum.\nAbstract: Human ureaplasmas are minimal-genome bacteria and pathobionts of the urogenital tract. They must adapt to fluctuating pH conditions despite the absence of canonical transcriptional regulatory systems. However, the mechanisms underlying these responses remain unclear. This study aimed to construct a system-level model of pH adaptation in this minimal pathogen. We used an integrated multi-omics platform combining proteomics, metabolomics, and RNA modification profiling to construct a system-level model of pH adaptation. The results revealed a bifurcated strategy governed by the differential activation of preexisting, co-regulated functional modules. Under neutral pH conditions (pH 7), Ureaplasma parvum activated energy metabolism and upregulated ATP synthesis while forming a stress-counteracting proteostasis pathway. This may suggest a biological energy state under high stress conditions. Conversely, under acidic stress (pH 5), it activated biosynthesis/translation, showing significant upregulation of ribosomal proteins and accumulation of translation precursors and the polyamine spermidine. This may represent a state of expanded translational capacity. This adaptive switch is accompanied by dynamic reorganization of the epitranscriptome, highlighting the importance of post-transcriptional regulation. This study suggests mechanisms by which minimal organisms achieve adaptive plasticity through sophisticated post-transcriptional and metabolic control, providing a new framework for understanding Ureaplasma physiology and the biology of genome-reduced organisms.IMPORTANCEMinimal bacteria challenge canonical views of cellular regulation. In organisms with radically reduced genomes and sparse transcription factors, how adaptive plasticity is achieved remains a core question. Our study proposes a model in which a simple physicochemical cue-extracellular pH-selects among prewired cellular programs, while post-transcriptional and epitranscriptomic layers fine-tune execution. The findings of this study suggest a multi-omics scheme for how organisms adapt to environmental changes and ensure survival without inducing new circuits or complex transcriptional regulation. Conceptually, it proposes regulation via RNA modifications in processes, such as metabolism, proteostasis, and translation. This framework may be generalizable to other genome-reduced microorganisms. Beyond microbiology, it provides design principles for synthetic biology and offers a mechanistic interpretation of phenotypic tolerance to stress factors. It may encourage the use of pH-linked epitranscriptome signals as measurable indicators of cellular state.\n\nID: 42012729\nTitle: Spermidine in the aging brain: mechanisms, preclinical evidence, and clinical perspectives.\nAbstract: Spermidine, a naturally occurring polyamine, has emerged as a candidate modulator of brain aging because it regulates autophagy, mitochondrial function, oxidative balance, and neuroinflammatory signaling. This review summarizes current knowledge on spermidine biology, including its synthesis, catabolism, transport, and cell-specific handling in neurons and glia, with emphasis on ATP13A2/ATP13A4-dependent trafficking and EP300-linked autophagy. Preclinical studies consistently show that spermidine enhances proteostasis, reduces soluble amyloid-beta, improves microglial phagocytic activity, preserves mitochondrial fitness, and attenuates inflammatory responses in models of aging and neurodegeneration. Human evidence is encouraging but still limited: observational studies associate higher dietary spermidine intake with better cognitive outcomes, whereas interventional trials suggest possible benefits in subjective cognitive decline and mild dementia, with variable dose-dependent effects. Overall, spermidine is a biologically plausible nutraceutical for healthy brain aging, but larger, well-designed clinical trials are needed to define efficacy, bioavailability, safety, and optimal therapeutic use across prodromal, early-stage, and disease-specific patient populations.\n\nID: 41890048\nTitle: Distinctive Lacritin Cleavage-Potentiated Bactericidal Alteration of the P. aeruginosa Transcriptome.\nAbstract: Lacritin is a tear, saliva, plasma and cerebrospinal fluid glycoprotein with broad polypharmacology. Selective deficiency of its bioactive monomeric form appears to be deleterious for ocular surface health for which replacement therapy is beneficial. Its cleavage-potentiated C-terminus represented by the N-104 proteoform in tears is bactericidal and synergizes with the tear thrombin peptide GKY20. In the pathogenic and multidrug resistant PA14 strain of P. aeruginosa, we recently discovered that N-104 binds to the outer-membrane lipoprotein YaiW to gain access to the periplasm where it targets and inhibits the inner-membrane ferrous iron transporter FeoB (of FeoABC) as well as PotH, a subunit of the polyamine transporter PotFGHI. Further, PA14 gene expression shifts toward anaerobic respiratory pathways. Here we explore N-104-associated transcriptional changes over a broader range of functional categories pointing to a reduction in: (i) virulence by suppressed gene expression of virulence factors AprA and LasA; and Hcp1 and PsrA necessary for the respective assembly of type VI and III secretion systems, (ii) fitness (less AtsC, MgtA), (iii) metabolism (less AdhA, AtsC, GcvH2, GcvP2, FadE1, SsuD, SsuF, TauB, TauD, UspK, UspN), (iv) stress response (less UnG, PfpI, RmF), (v) proteostasis (less ClpB, GrpE, HtpG), (vii) quorum sensing (less CifR, GcvH2, GcvP2, PsrA, QuiP), and (viii) survival under anaerobic conditions (less AdhA, MhR, ModA, UspKLNO). Upregulated genes are directed towards enhancing PA14: (i) multidrug (more OprJ of MexCD-OprJ) and (ii) tellurite (more TerC) efflux, coupled with a seemingly PA14 survival attempt at (iii) anaerobic respiration (more NosR), (iv) translational fidelity (more QueE, RimP, TrmD) and (v) metabolism (CysT, MoaA1, Sbp, SsuA, SsuE). The overlap with aminoglycosides (4.3%), \u03b2-lactams (0%), cyclic peptides (2.5%), fluoroquinilones (0%) and macrolide (1.9%) classes of antibiotics in P. aeruginosa was minimal. Thus, N-104 appears to widely perturb PA14 fundamental processes in a distinctive manner.\n\nID: 41875963\nTitle: Mechanisms of blood-brain barrier penetration: A molecular dynamics study on R9 and MPG peptide translocation.\nAbstract: One of the major obstacles in treating diseases that affect the central nervous system is delivering drugs across the blood-brain-barrier (BBB). Cell-penetrating peptides (CPPs) can be used as delivery vectors, but their translocation mechanism is still poorly understood, in part due to the simplistic membrane models applied to their interpretation. Here we investigate the translocation mechanism of two CPPs, R9 and MPG, using molecular dynamics and enhanced sampling techniques on a realistic membrane model of human brain microvascular endothelial cells. The results suggest that R9 induces greater membrane disruption compared to MPG, yet that both face a significant free energy barrier to translocation. In both peptides the first interactions were initiated by the N-terminus and prominently involved arginine residues even for MPG. The crucial role of the plasticity of both partners (BBB bending, partial CPP unfolding) on the translocation energetics was also explored by sampling ad hoc collective variables, revealing the important role of long polyunsaturated acyl chain lipids. Together, these findings provide mechanistic insight into CPP-mediated transport and offer guidelines for rational design.\n\nID: 41756429\nTitle: Polyamine metabolic enzyme SAT1 remodels the neuronal transcriptome and rescues \u03b1-synuclein toxicity in Drosophila.\nAbstract: Polyamine homeostasis is tightly regulated by interconversion and catabolic pathways and has been increasingly implicated in neurodegenerative disorders, including Parkinson's disease (PD), where accumulation of \u03b1-synuclein (\u03b1-Syn) perturbs neuronal homeostasis. Spermidine/spermine N1-acetyltransferase 1 (SAT1) occupies a central position in polyamine interconversion, and alterations in SAT1 activity have been linked to \u03b1-Syn toxicity and PD-related neuropathology. To investigate how SAT1 activity influences \u03b1-Syn-associated neurodegeneration, we employed a Drosophila model of neuronal \u03b1-Syn expression. SAT1 overexpression reduced \u03b1-Syn protein levels, altered its subcellular distribution within the brain, and mitigated \u03b1-Syn-induced lifespan shortening. Transcriptomic analyses showed that SAT1 modulates stress-associated gene expression in the \u03b1-Syn background, including attenuation of chaperone and ubiquitin-related responses and coordinated changes in pathways linked to mitochondrial function and amino acid metabolism. SAT1 co-expression attenuated \u03b1-Syn-associated alterations in genes involved in mitochondrial quality control, including USP30, Uch-L5R, RNF185, and the mitochondrial ornithine carrier SLC25A15. At the protein level, SAT1 increased mitochondrial-associated signal, enhanced LC3 association with mitochondrial compartments, restored LC3-II/LC3-I ratios in mitochondrial fractions and reduced mitochondrial accumulation of \u03b1-Syn. Our findings indicate that SAT1 activity is associated with reduced \u03b1-Syn toxicity and altered mitochondrial-associated proteostasis during \u03b1-Syn expression.\n\nID: 41496478\nTitle: Cell swelling and upright mounting-based imaging for high-resolution visualization of intracellular trafficking across the BBB using conventional confocal microscopy.\nAbstract: Receptor-mediated transcytosis (RMT) represents a promising strategy for delivering macromolecular and colloidal therapeutics across the blood-brain barrier (BBB). However, mechanistic elucidation of RMT remains limited by the difficulty of visualizing subcellular trafficking pathways. Conventional imaging approaches either lack sufficient spatial resolution or require costly, technically complex instrumentation. Here, we report a cell swelling and upright mounting-based (CSUM-based) imaging approach that reorients the Z-axis into the high-resolution XY-plane using standard confocal microscopy, enabling direct RMT visualization without computational reconstruction or specialized hardware. We tracked intracellular trafficking of transferrin (Tf) and anti-transferrin receptor antibody (anti-TfR Ab) as model cargos using our CSUM-based imaging approach via compartment-specific markers and time-resolved co-localization analysis. This approach resolved cargo-containing vesicles traversing from the apical to basolateral membranes. Tf completed transcytosis within 15\u2009min, whereas anti-TfR Ab initially entered the endolysosomal pathway before rerouting to transcytosis under receptor saturation conditions. The CSUM approach provides a simple yet effective platform for high-resolution visualization of membrane transport and vesicle dynamics, offering broad applicability to drug delivery research and the design of brain-targeted therapeutics.\n\nID: 41330616\nTitle: Age-related immune states and PD-1 blockade: mechanisms and strategies for the elderly.\nAbstract: Aging impairs antitumor immunity and may reduce the efficacy of immune checkpoint inhibitors (ICIs). However, the underlying mechanisms remain unclear. Building on our recent findings, we review three key mechanisms of CD8+ T-cell aging: elevated T-cell receptor (TCR) activation thresholds, mitochondrial dysfunction, and disruption of proteostasis. Studies in aged mice have revealed that aged na\u00efve T cells exhibit defective priming due to increased CD45 expression, which raises the TCR activation threshold and restricts effector differentiation. Aging also impairs mitochondrial metabolism, particularly fatty acid oxidation. Furthermore, we highlight the role of proteostasis collapse, including defective autophagy and increased endoplasmic reticulum stress, as a contributor to T-cell dysfunction. Spermidine, a polyamine that declines with age, has the potential to modulate both mitochondrial function and proteostasis. Its supplementation has been shown to improve programmed cell death-1 blockade responsiveness in aged mice. Although clinical studies in humans have yielded inconsistent results regarding the effect of chronological age on ICI efficacy, identifying patients with \"age-related\" immune microenvironments may enable stratified therapeutic approaches based on insights from preclinical aging models.\n\nID: 41309058\nTitle: Autophagy and GLUT1 trafficking: an overview of molecular mechanisms.\nAbstract: Autophagy is a catabolic process that enables cellular metabolic adaptation in response to nutrient deprivation. It facilitates the degradation of proteins and cellular components within lysosomes to generate essential metabolites. The glucose transporter 1 (GLUT1) is among the proteins that can undergo autophagy-mediated degradation in response to metabolic stimuli. GLUT1 is essential for cellular glucose supply in several tissues. Notably, GLUT1 facilitates glucose transport across the blood-brain barrier, creating a concentration gradient from the bloodstream into the brain's interstitial fluid. The presence of GLUT1, at the plasma membrane, is the first step in initiating glucose uptake and driving glycolysis inside the cell. Glycolysis can be initiated in response to several stimuli, including glucose availability, autophagy inhibition, and growth factor accessibility. In this review, we highlight recently described mechanisms that govern the subcellular distribution of GLUT1 with a focus on autophagy-mediated trafficking. Understanding how autophagy coordinates GLUT1 sorting in response to metabolic demands may uncover novel therapeutic targets for metabolic disorders characterized by dysregulated GLUT1 trafficking.\n\nID: 41286441\nTitle: Polyamines sustain epithelial regeneration in aged intestines by modulating protein homeostasis.\nAbstract: Ageing dampens the regenerative potential of intestinal epithelium across species including humans, yet the underlying causes remain elusive. Here we characterized the temporal dynamics of regeneration following injury induced by 5-fluorouracil, a commonly used chemotherapeutic agent, using proteomic and metabolomic profiling of intestinal tissues together with functional assays. The comparison of regeneration dynamics in mice of different ages revealed the emergence of proteostasis stress and increased levels of polyamines following injury exclusively in old epithelia. We show that delayed regeneration is an intrinsic feature of aged epithelial cells that display reduced protein synthesis and the accumulation of ubiquitylated proteins. The inhibition of the polyamine pathway in vivo further delays regeneration in old mice, whereas its activation by dietary intervention or supplementation of polyamines is sufficient to enhance the regenerative capacity of aged intestines. Our findings highlight the promising epithelial targets for interventions aimed at tackling the decline in tissue repair mechanisms associated with ageing.\n\nID: 41013206\nTitle: Anti-P antibodies that impair memory perturb hippocampal glutamatergic receptor trafficking, synapse structure and microglia.\nAbstract: Anti-ribosomal P protein autoantibodies (anti-P) are associated with psychosis and cognitive dysfunction in patients with systemic lupus erythematosus (SLE), yet the underlying mechanisms remain undefined, hindering targeted therapies. Anti-P cross-react with a neuronal surface protein (NSPA), alter glutamatergic synaptic transmission and plasticity in hippocampal slices, and impair spatial memory in a short-term passive transfer mouse model. NSPA knockout mice display spatial memory deficit linked to reduced NMDAR activity and postsynaptic density (PSD) levels, along with an increased membrane-associated tyrosine phosphatase PTPMEG, suggesting disrupted glutamatergic receptor trafficking. Here, we investigated the acute effects of anti-P on receptor cell surface expression and trafficking in cultured hippocampal neurons and their long-term impact on hippocampal components and spatial memory in anti-P(\u2009+) immunized mice. NMDAR and AMPAR surface expression and NMDAR recycling were assessed in 21-24 DIV primary hippocampal neurons by immunofluorescence and FRAP using SEP-tagged receptors under the effects of rabbit anti-P IgG fractions. In vivo, female C57BL/6 mice were immunized with recombinant P0 ribosomal protein to induce anti-P, followed by lipopolysaccharide (LPS) intraperitoneal administration to breach the blood-brain-barrier (BBB). Spatial memory was evaluated with a water maze memory flexibility test. Hippocampal synaptosomal membranes and PSD-enriched fractions were analyzed by immunoblotting. Neuronal density, microglia and dendritic architecture were evaluated using Cresyl Violet, Iba1 and Golgi staining, respectively. Anti-P treatment of cultured neurons reduced GluN2A and GluA1 surface levels and impaired SEP-GluN2A and SEP-GluN2B recycling. Anti-P(\u2009+) mice showed spatial memory deficits persisting up to 24\u00a0days post-LPS, along with hippocampal alterations that include reduced levels of NMDAR, AMPAR, and PSD-95 in PSD fractions; increased membrane-associated PTPMEG;\u2009~\u20097% neuronal loss; higher number of microglia with reduced ramifications, and diminished dendritic width and spine density. Notably, increased PTPMEG levels were already detectable by day 10 post-LPS. Anti-P antibodies acutely impair glutamatergic receptor recycling and surface expression, while their long-term effects lead to sustained memory impairment associated with altered neuronal and microglial architecture, and PTPMEG increased levels preceding PSD protein loss. These findings provide mechanistic insight into anti-P-mediated cognitive dysfunction and may inform therapeutic strategies for neuropsychiatric SLE.\n\nID: 40883288\nTitle: Retrograde transport of neurotrophin receptor TrkB-FL induced by excitotoxicity regulates Golgi stability and is a target for stroke neuroprotection.\nAbstract: Excitotoxicity, aberrant function of survival pathways dependent on brain-derived neurotrophic factor (BDNF), and disruption of the Golgi complex are shared pathological hallmarks in relevant neurological diseases, including stroke. However, the precise interdependence among these mechanisms is not completely defined, knowledge essential for developing neuroprotective strategies. For ischemic stroke, a leading cause of death, disability, and dementia, interfering with excitotoxicity-the major mechanism of neuronal death in the penumbra area-has shown promising results. We are exploring neuroprotection by promoting survival cascades dependent on the BDNF receptor, full-length tropomyosin-related kinase B (TrkB-FL), as these pathways become aberrant after excitotoxicity. We previously developed MTFL457, a blood-brain barrier (BBB) permeable neuroprotective peptide containing a TrkB-FL sequence, which efficiently prevents excitotoxicity-induced receptor processing and preserves BDNF-dependent pathways in an ischemia model, where it decreases infarct size and improves neurological outcome. In this work, using cellular and animal models, we demonstrate that excitotoxicity-induced TrkB-FL downregulation is secondary to receptor endocytosis, interaction with the endosomal protein hepatocyte growth factor-regulated tyrosine kinase substrate (Hrs), retrograde transport to the Golgi, and subsequent disruption of this organelle. Interestingly, peptide MTFL457 interferes with the TrkB-FL/Hrs interaction and receptor trafficking-processes required for excitotoxic Golgi fragmentation and TrkB-FL cleavage-demonstrating a central role for TrkB-FL in controlling Golgi stability. These results suggest the potential for peptide MTFL457 to preserve the function of this organelle and critical neuronal survival pathways in stroke and possibly other neurodegenerative diseases associated with excitotoxicity.\n\nID: 40241296\nTitle: Starvation Induces Upregulation of Monocarboxylate Transport in Glial Cells at the Drosophila Blood-Brain Barrier.\nAbstract: Living organisms can sense and adapt to constant changes in food availability. Maintaining a homeostatic supply of energy molecules is crucial for animal survival and normal organ functioning, particularly the brain, due to its high-energy demands. However, the mechanisms underlying brain adaptive responses to food availability have not been completely established. The nervous system is separated from the rest of the body by a physical barrier called the blood-brain barrier (BBB). In addition to its structural role, the BBB regulates the transport of metabolites and nutrients into the nervous system. This regulation is achieved through adaptive mechanisms that control the transport of nutrients, including glucose and monocarboxylates such as lactate, pyruvate, and ketone bodies. In Drosophila melanogaster , carbohydrate transporters increase their expression in glial cells of the BBB in response to starvation. However, changes in the expression or activity of Drosophila monocarboxylate transporters (dMCTs) at the BBB have not yet been reported. Here, we show that neuronal ATP levels remain unaffected despite reduced energy-related metabolites in the hemolymph of Drosophila larvae during starvation. Simultaneously, the transport of lactate and beta-hydroxybutyrate increases in the glial cells of the BBB. Using genetically encoded sensors, we identified Yarqay as a proton-coupled monocarboxylate transporter whose expression is upregulated in the subperineurial glia of the BBB during starvation. Our findings reveal a novel component of the adaptive response of the brain to starvation: the increase in the transport of monocarboxylates across the BBB, mediated by Yarqay, a novel dMCT enriched in the BBB.\n\nID: 40149801\nTitle: Molecular Motors in Blood-Brain Barrier Maintenance by Astrocytes.\nAbstract: The blood-brain barrier (BBB) comprises distinct cell types, including endothelial cells, pericytes, and astrocytes, and is essential for central nervous system (CNS) homeostasis by selectively regulating molecular transport and maintaining integrity. In particular, astrocytes are essential for BBB function, as they maintain BBB integrity through their end-feet, which form a physical and biochemical interface that enhances endothelial cell function and barrier selectivity. Moreover, they secrete growth factors like vascular endothelial growth factor (VEGF) and transforming growth factor-beta (TGF-\u03b2), which regulate tight junction (TJ) proteins (e.g., claudins and occludins) crucial for limiting paracellular permeability. Molecular motors like kinesins, dynein, and myosins are essential for these astrocyte functions. By facilitating vesicular trafficking and protein transport, they are essential for various functions, including trafficking of junctional proteins to support BBB integrity, the proper mitochondria localization within astrocyte processes for efficient energy supply, the polarized distribution of aquaporin (AQP)-4 at astrocyte end-feet for regulating water homeostasis across the BBB, and the modulation of neuroinflammatory responses. Moreover, myosin motors modulate actomyosin dynamics to regulate astrocyte process outgrowth, adhesion, migration, and morphology, facilitating their functional roles. Thus, motor protein dysregulation in astrocytes can compromise BBB function and integrity, increasing the risk of neurodegeneration. This review explores the complex interplay between astrocytes and molecular motors in regulating BBB homeostasis, which represents an attractive but poorly explored area of research.\n\nID: 39954801\nTitle: Microglia-derived sEV: Friend or foe in the pathogenesis of cognitive impairment.\nAbstract: As immune cells, microglia serve a dual role in cognition. Microglia-derived sEV actively contribute to the development of cognitive impairment by selectively targeting specific cells through various substances such as proteins, RNA, DNA, lipids, and metabolic waste. In recent years, there has been an increasing focus on understanding the pathogenesis and therapeutic potential of sEV. This comprehensive review summarizes the detrimental effects of M1 microglial sEV on pathogenic protein transport, neuroinflammation, disruption of the blood-brain barrier (BBB), neuronal death and synaptic dysfunction in relation to cognitive damage. Additionally, it highlights the beneficial effects of M2 microglia on alleviating cognitive impairment based on evidence from cellular experiments and animal studies. Furthermore, since microglial-secreted sEV can be found in cerebrospinal fluid or cross the BBB into plasma circulation, they play a crucial role in diagnosing cognitive impairment. However, using sEV as biomarkers is still at an experimental stage and requires further clinical validation. Future research should aim to explore the mechanisms underlying microglial involvement in various nervous system disorders to identify novel targets for clinical interventions.\n\nID: 39289695\nTitle: Truncated mini LRP1 transports cargo from luminal to basolateral side across the blood brain barrier.\nAbstract: The most crucial area to focus on when thinking of novel pathways for drug delivery into the CNS is the blood brain barrier (BBB). A number of nanoparticulate formulations have been shown in earlier research to target receptors at the BBB and transport therapeutics into the CNS. However, no mechanism for CNS entrance and movement throughout the CNS parenchyma has been proposed yet. Here, the truncated mini low-density lipoprotein receptor-related protein 1 mLRP1_DIV* was presented as blood to brain transport carrier, exemplified by antibodies and immunoliposomes using a systematic approach to screen the receptor and its ligands' route across endothelial cells in vitro. The use of mLRP1_DIV* as liposomal carrier into the CNS was validated based on internalization and transport assays across an in vitro model of the BBB using hcMEC/D3 and bEnd.3 cells. Trafficking routes of mLRP1_DIV* and corresponding cargo across endothelial cells were analyzed using immunofluorescence. Modulation of \u03b3-secretase activity by immunoliposomes loaded with the \u03b3-secretase modulator BB25 was investigated in co-cultures of bEnd.3 mLRP1_DIV* cells and CHO cells overexpressing human amyloid precursor protein (APP) and presenilin 1 (PSEN1). We showed that while expressed in vitro, mLRP1_DIV* transports both, antibodies and functionalized immunoliposomes from luminal to basolateral side across an in vitro model of the BBB, followed by their mLRP1_DIV* dependent release of the cargo. Importantly, functionalized liposomes loaded with the \u03b3-secretase modulator BB25 were demonstrated to effectively reduce toxic A\u00df42 peptide levels after mLRP1_DIV* mediated transport across a co-cultured endothelial monolayer. Together, the data strongly suggest mLRP1_DIV* as a promising tool for drug delivery into the CNS, as it allows a straight transport of cargo from luminal to abluminal side across an endothelial monolayer and it's release into brain parenchyma in vitro, where it exhibits its intended therapeutic effect.\n\nID: 38666466\nTitle: Adaptive Metabolic Responses Facilitate Blood-Brain Barrier Repair in Ischemic Stroke via BHB-Mediated Epigenetic Modification of ZO-1 Expression.\nAbstract: Adaptive metabolic responses and innate metabolites hold promising therapeutic potential for stroke, while targeted interventions require a thorough understanding of underlying mechanisms. Adiposity is a noted modifiable metabolic risk factor for stroke, and recent research suggests that it benefits neurological rehabilitation. During the early phase of experimental stroke, the lipidomic results showed that fat depots underwent pronounced lipolysis and released fatty acids (FAs) that feed into consequent hepatic FA oxidation and ketogenesis. Systemic supplementation with the predominant ketone beta-hydroxybutyrate (BHB) is found to exert discernible effects on preserving blood-brain barrier (BBB) integrity and facilitating neuroinflammation resolution. Meanwhile, blocking FAO-ketogenesis processes by administration of CPT1\u03b1 antagonist or shRNA targeting HMGCS2 exacerbated endothelial damage and aggravated stroke severity, whereas BHB supplementation blunted these injuries. Mechanistically, it is unveiled that BHB infusion is taken up by monocarboxylic acid transporter 1 (MCT1) specifically expressed in cerebral endothelium and upregulated the expression of tight junction protein ZO-1 by enhancing local \u03b2-hydroxybutyrylation of H3K9 at the promoter of TJP1 gene. Conclusively, an adaptive metabolic mechanism is elucidated by which acute lipolysis stimulates FAO-ketogenesis processes to restore BBB integrity after stroke. Ketogenesis functions as an early metabolic responder to restrain stroke progression, providing novel prospectives for clinical translation.\n\nID: 38164612\nTitle: Hyphenation of lipophilic ruthenium(II)-diphosphine core with 5-fluorouracil: an effective metallodrug against glioblastoma brain cancer cells.\nAbstract: Glioblastoma multiforme (GBM) is the most common highly aggressive malignant brain tumor, with a very limited chance for survival post-diagnosis and post-treatment. Despite significant advancement in GBM genomics implicated in molecularly targeted chemotherapies, the prognosis remains poor and requires new drug discovery approaches. We used fluoropyrimidine 5-fluorouracil (5-FU), an antimetabolite anticancer drug conjugated or 'caged' within a lipophilic Ru(II)-diphosphine (dppe) core formulated as [RuII(dppe)2(5-FU)]PF6 (Ru-DPPE-5FU), where dppe = 1,2-bis(diphenylphosphino)ethane, and evaluated its in vitro cytotoxicity in depth with aggressive GBM cells (LN229). The hydrophilic nature of 5-FU limits its passage through the blood-brain barrier (BBB), which prevents its effective accumulation and efficacy for GBM tumors. Herein, we attempted to modulate the lipophilicity of 5-FU by inserting it within a well-designed lipophilic {Ru(dppe)2}-core with anticipated higher efficiency towards GBM. The physicochemical properties of [RuII(dppe)2(5-FU)]PF6 (Ru-DPPE-5FU) were studied using various spectroscopic and analytical techniques. The molecular structure was determined using X-ray crystallography, showing a distorted {RuP4NO} octahedral geometry with bidentate (N, O) binding of 5-FU and its aromatization in the Ru(II)-bound form. The 31P-NMR spectra of Ru-DPPE-5FU showed four closely spaced distinct 31P-signals, indicating four unique chemical environments around P, and the strong coupling constants between them make it a second-order spectrum. The RuII/RuIII redox potential in Ru-DPPE-5FU shifted by \u223c0.91 V towards the anodic region as compared to its precursor complex cis-[Ru(dppe)2Cl2] (Ru-DPPE-Cl). DFT-based theoretical calculations have been performed to correlate the experimental electronic absorption spectra and redox behaviours of the complexes. The electrostatic potential (ESP) plots indicate the delocalization of the charge density on the O-/F-atom from the 5-FU ligand towards Ru(II) upon its complexation. The antioxidant properties of all the compounds were quantified by a 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging assay. The hyphenation of the 5-fluorouracil (5-FU) ligand to the lipophilic {Ru(dppe)2}-core endowed lipophilicity to Ru-DPPE-5FU with higher in vitro cytotoxicity (IC50 = 2.37 \u03bcM) against the LN229 GBM cells as compared to the hydrophilic 5-FU, suggesting efficient cellular uptake. Further biological assays indicated that the complex is highly potent in inhibiting significant proliferation and spheroid formation and restricting the migratory potentials of the GBM cells. Increased caspase 3/7 activity and the presence of apoptotic bodies at the center of 3-D GBM spheroids as revealed by AO/EB dual staining indicated a deeper penetration of the lipophilic complex. The Ru-DPPE-5FU complex displayed lower cytotoxicity in HaCaT normal cells (IC50 = 7.27 \u03bcM) in comparison to LN229 cancer cells with a selectivity index (S.I.) of \u22653. Overall, the synergism and caging of 5-FU within the hydrophobic {Ru(dppe)2}-core improves the pharmacokinetic profile of Ru-DPPE-5FU as a potent anticancer agent for glioblastoma.\n\nID: 38058025\nTitle: How S100B crosses brain barriers and why it is considered a peripheral marker of brain injury.\nAbstract: S100B is a 21-kDa protein that is produced and secreted by astrocytes and widely used as a marker of brain injury in clinical and experimental studies. The majority of these studies are based on measurements in blood serum, assuming an associated increase in cerebrospinal fluid and a rupture of the blood-brain barrier (BBB). Moreover, extracerebral sources of S100B are often underestimated. Herein, we will review these interpretations and discuss the routes by which S100B, produced by astrocytes, reaches the circulatory system. We discuss the concept of S100B as an alarmin and its dual activity as an inflammatory and neurotrophic molecule. Furthermore, we emphasize the lack of data supporting the idea that S100B acts as a marker of BBB rupture, and the need to include the glymphatic system in the interpretations of serum changes of S100B. The review is also dedicated to valorizing extracerebral sources of S100B, particularly adipocytes. Furthermore, S100B per se may have direct and indirect modulating roles in brain barriers: on the tight junctions that regulate paracellular transport; on the expression of its receptor, RAGE, which is involved in transcellular protein transport; and on aquaporin-4, a key protein in the glymphatic system that is responsible for the clearance of extracellular proteins from the central nervous system. We hope that the data on S100B, discussed here, will be useful and that it will translate into further health benefits in medical practice.\n\nID: 37603183\nTitle: In Vitro and Ex Vivo Methodologies for T-Cell Trafficking Through Blood-Brain Barrier After TLR Activation.\nAbstract: This chapter describes ex vivo isolation of human T cells and of na\u00efve splenocytes respectively collected from multiple sclerosis patients and healthy controls and experimental autoimmune encephalomyelitis-affected mice. After the magnetic sorting of na\u00efve and activated T helper lymphocytes, we provide details about the cell cultures to measure the interaction with extracellular matrix proteins using standard cell invasion or hand-made in vitro assays, upon different stimuli, through Toll-like receptor(s) ligands, T-cell activators, and cell adhesion molecules modulators. Finally, we describe the methods to harvest and recover T cells to evaluate the properties associated with their trafficking ability.\n\nID: 36919582\nTitle: Blood-brain barrier integrity impacts the use of plasma amyloid-\u03b2 as a proxy of brain amyloid-\u03b2 pathology.\nAbstract: Amyloid-\u03b2 (A\u03b2) and tau can be quantified in blood. However, biological factors can influence the levels of brain-derived proteins in the blood. The blood-brain barrier (BBB) regulates protein transport between cerebrospinal fluid (CSF) and blood. BBB altered permeability might affect the relationship between brain and blood biomarkers. We assessed 224 participants in research (TRIAD, n\u00a0=\u00a096) and clinical (BIODEGMAR, n\u00a0=\u00a0128) cohorts with plasma and CSF/positron emission tomography A\u03b2, p-tau, and albumin measures. Plasma A\u03b242/40 better identified CSF A\u03b242/40 and A\u03b2-PET positivity in individuals with high BBB permeability. An interaction between plasma A\u03b242/40 and BBB permeability on CSF A\u03b242/40 was observed. Voxel-wise models estimated that the association of positron emission tomography (PET), with plasma A\u03b2 was most affected by BBB permeability in AD-related brain regions. BBB permeability did not significantly impact the relationship between brain and plasma p-tau levels. These findings suggest that BBB integrity may influence the performance of plasma A\u03b2, but not p-tau, biomarkers in research and clinical settings. BBB permeability affects the association between brain and plasma A\u03b2 levels. BBB integrity does not affect the association between brain and plasma p-tau levels. Plasma A\u03b2 was most affected by BBB permeability in AD-related brain regions. BBB permeability increases with age but not according to cognitive status.\n\nID: 35889137\nTitle: Plasmodium falciparum S-Adenosylmethionine Synthetase Is Essential for Parasite Survival through a Complex Interaction Network with Cytoplasmic and Nuclear Proteins.\nAbstract: S-adenosylmethionine synthetase (SAMS) is a key enzyme for the synthesis of the lone methyl donor S-adenosyl methionine (SAM), which is involved in transmethylation reactions and hence required for cellular processes such as DNA, RNA, and histone methylation, but also polyamine biosynthesis and proteostasis. In the human malaria parasite Plasmodium falciparum, PfSAMS is encoded by a single gene and has been suggested to be crucial for malaria pathogenesis and transmission; however, to date, PfSAMS has not been fully characterized. To gain deeper insight into the function of PfSAMS, we generated a conditional gene knockdown (KD) using the glmS ribozyme system. We show that PfSAMS localizes to the cytoplasm and the nucleus of blood-stage parasites. PfSAMS-KD results in reduced histone methylation and leads to impaired intraerythrocytic growth and gametocyte development. To further determine the interaction network of PfSAMS, we performed a proximity-dependent biotin identification analysis. We identified a complex network of 1114 proteins involved in biological processes such as cell cycle control and DNA replication, or transcription, but also in phosphatidylcholine and polyamine biosynthesis and proteasome regulation. Our findings highlight the diverse roles of PfSAMS during intraerythrocytic growth and sexual stage development and emphasize that PfSAMS is a potential drug target.\n\nID: 35820539\nTitle: Microenvironment-tailored micelles restrain carcinoma-astrocyte crosstalk for brain metastasis.\nAbstract: Breast-to-brain metastatic cells can interact with the surrounding cells, including astrocytes and microglia, to generate a pro-tumorigenic niche. Breast-to-brain metastasis can be treated using a dual strategy of eliminating metastatic tumor cells and normalizing their localized microenvironment. The effective accumulation of drugs at the action site of metastasis is crucial to realizing the above strategy, especially when dealing with the blood-brain barrier (BBB)-penetrating and tumor-targeting tactics. Here, we establish an in-situ microenvironment-tailored micelle (T-M/siRNA) to co-deliver therapeutic siRNA and paclitaxel (PTX) into the breast-to-brain metastasis. Anchored with a D-type cyclic peptide, T-M/siRNA can penetrate the BBB and subsequently target the brain metastases. Upon internalization by metastatic tumor cells, T-M/siRNA can release PTX in the high-level glutathione (GSH), resulting in killing cancer cells. Meanwhile, the micellar structure is dissociated, resulting in lowering the charge density to release the loaded siRNA that can targeted downregulate the expression of protocadherin 7 (PCDH7). Treatment of model mice revealed that T-M/siRNA can inhibit the abnormal activation of astrocytes and immunosuppressive activation of microglia, resulting in significantly enhanced synergistic anti-tumor efficacy. This study indicates that the micelle system can serve as a hopeful strategy to treat breast-to-brain metastasis.\n\nID: 35472275\nTitle: Benzene and Borazine, so Different, yet so Similar: Insight from Experimental Charge Density Analysis.\nAbstract: Although benzene and borazine are isoelectronic and isostructural, they have very different electronic structures, mainly due to the polar nature of the B-N bond. Herein, we present an experimental study of the charge density distribution obtained from the multipole model formalism and Hirshfeld atom refinement (HAR) based on high-resolution X-ray diffraction data of borazine B3N3H6 (1) and B,B',B\u2033-trichloroborazine (2) crystals. These data are compared to those obtained from HAR for benzene (4) and 1,3,5-trichlorobenzene (5) and further compared with values obtained from density functional theory calculations in the gas phase, where N,N',N\u2033-trichloroborazine (3) was also included. The results confirm that, unlike benzene, borazines are only weakly aromatic with an island-like electronic delocalization within the B3N3 ring involving only the nitrogen atoms. Furthermore, delocalization indices and interacting quantum atom energy for bonded and non-bonded atoms were found to be highly suitable indicators capable of describing the origin of the discrepancies observed when the degree of aromaticity in 2 and 3 is evaluated using common aromaticity indices. Additionally, analysis of intermolecular interactions in the crystals brings further evidence of a weakly aromatic character of the borazines as it reveals surprising similarities between the crystal packing of borazine and benzene and also between B,B',B\u2033-trichloroborazine and 1,3,5-trichlorobenzene.\n\nID: 35165441\nTitle: A human brain vascular atlas reveals diverse mediators of Alzheimer's risk.\nAbstract: The human brain vasculature is of great medical importance: its dysfunction causes disability and death1, and the specialized structure it forms-the blood-brain barrier-impedes the treatment of nearly all brain disorders2,3. Yet so far, we have no molecular map of the human brain vasculature. Here we develop vessel isolation and nuclei extraction for sequencing (VINE-seq) to profile the major vascular and perivascular cell types of the human brain through 143,793 single-nucleus transcriptomes from 25 hippocampus and cortex samples of 9 individuals with Alzheimer's disease and 8 individuals with no cognitive impairment. We identify brain-region- and species-enriched genes and pathways. We reveal molecular principles of human arteriovenous organization, recapitulating a gradual endothelial and punctuated mural cell continuum. We discover two subtypes of human pericytes, marked by solute transport and extracellular matrix (ECM) organization; and define perivascular versus meningeal fibroblast specialization. In Alzheimer's disease, we observe selective vulnerability of ECM-maintaining pericytes and gene expression patterns that implicate dysregulated blood flow. With an expanded survey of brain cell types, we find that 30 of the top 45 genes that have been linked to Alzheimer's disease\u00a0risk by genome-wide association studies (GWASs) are expressed in the human brain vasculature, and we confirm this by immunostaining. Vascular GWAS genes map to endothelial protein transport, adaptive immune and ECM pathways. Many are microglia-specific in mice, suggesting a partial evolutionary transfer of Alzheimer's disease risk. Our work uncovers the molecular basis of the human brain vasculature, which will inform our understanding of overall brain health, disease and therapy.\n\nID: 35002279\nTitle: Dysregulation of the Retromer Complex in Brain Endothelial Cells Results in Accumulation of Phosphorylated Tau.\nAbstract: Transport through endothelial cells of the blood-brain barrier (BBB) involves a complex group of structures of the endo-lysosome system such as early and late endosomes, and the retromer complex system. Studies show that neuronal dysregulation of the vacuolar protein sorting 35 (VPS35), the main component of the retromer complex recognition core, results in altered protein trafficking and degradation and is involved in neurodegeneration. Since the functional role of VPS35 in endothelial cells has not been fully investigated, in the present study we aimed at characterizing the effect of its downregulation on these pathways. Genetic silencing of VPS35 in human brain endothelial cells; measurement of retromer complex system proteins, autophagy and ubiquitin-proteasome systems. VPS35-downregulated endothelial cells had increased expression of LC3B2/1 and more ubiquitinated products, markers of autophagy flux and impaired proteasome activity, respectively. Additionally, compared with controls VPS35 downregulation resulted in significant accumulation of tau protein and its phosphorylated isoforms. Our findings demonstrate that in brain endothelial cells retromer complex dysfunction by influencing endosome-lysosome degradation pathways results in altered proteostasis. Restoration of the retromer complex system function should be considered a novel therapeutic approach to rescue endothelial protein transport.\n\nID: 34668776\nTitle: The HIV-1 Matrix Protein p17 Does Cross the Blood-Brain Barrier.\nAbstract: Human immunodeficiency virus type 1 (HIV-1)-associated neurocognitive disorder (HAND) remains an important neurological manifestation in HIV-1-infected (HIV+) patients. Furthermore, detection of the HIV-1 matrix protein p17 (p17) in the central nervous system (CNS) and its ability to form toxic assemblies in the brain have been recently confirmed. Here, we show for the first time, using both an in vitro blood-brain barrier (BBB) model and in vivo biodistribution studies in healthy mice, that p17 can cross the BBB. There is rapid brain uptake with 0.35%\u2009\u00b1\u20090.19% of injected activity per gram of tissue (IA/g) 2 min after administration, followed by brain accumulation with 0.28%\u2009\u00b1\u20090.09% IA/g after 1 h. The interaction of p17 with chemokine receptor 2 (CXCR2) at the surface of brain endothelial cells triggers transcytosis. The present study supports the hypothesis of a direct role of free p17 in neuronal dysfunction in HAND by demonstrating its intrinsic ability to reach the CNS. IMPORTANCE The percentage of patients affected by HIV-1-associated neurocognitive disorder (HAND) ranges from 30% to 50% of HIV-infected (HIV+) patients. The mechanisms leading to HAND development need to be elucidated, but the roles of secreted viral proteins, chemokines, and proinflammatory molecules appear to be clear. In particular, the blood-brain barrier (BBB) represents a route for entry into the central nervous system (CNS) and thus plays an important role in HAND. Several findings suggest a key role for the HIV-1 matrix protein p17 (p17) as a microenvironmental factor capable of inducing neurocognitive disorders. Here, we show the ability of the p17 to cross the BBB and to reach the CNS, thus playing a crucial role in neuronal dysfunction in HAND.\n\nID: 34579098\nTitle: A Proton-Coupled Transport System for \u03b2-Hydroxy-\u03b2-Methylbutyrate (HMB) in Blood-Brain Barrier Endothelial Cell Line hCMEC/D3.\nAbstract: \u03b2-Hydroxy-\u03b2-methylbutyrate (HMB), a leucine metabolite, is used as a nutritional ingredient to improve skeletal muscle health. Preclinical studies indicate that this supplement also elicits significant benefits in the brain; it promotes neurite outgrowth and prevents age-related reductions in neuronal dendrites and cognitive performance. As orally administered HMB elicits these effects in the brain, we infer that HMB crosses the blood-brain barrier (BBB). However, there have been no reports detailing the transport mechanism for HMB in BBB. Here we show that HMB is taken up in the human BBB endothelial cell line hCMEC/D3 via H+-coupled monocarboxylate transporters that also transport lactate and \u03b2-hydroxybutyrate. MCT1 (monocarboxylate transporter 1) and MCT4 (monocarboxylate transporter 4) belonging to the solute carrier gene family SLC16 (solute carrier, gene family 16) are involved, but additional transporters also contribute to the process. HMB uptake in BBB endothelial cells results in intracellular acidification, demonstrating cotransport with H+. Since HMB is known to activate mTOR with potential to elicit transcriptomic changes, we examined the influence of HMB on the expression of selective transporters. We found no change in MCT1 and MCT4 expression. Interestingly, the expression of LAT1 (system L amino acid transporter 1), a high-affinity transporter for branched-chain amino acids relevant to neurological disorders such as autism, is induced. This effect is dependent on mTOR (mechanistic target of rapamycine) activation by HMB with no involvement of histone deacetylases. These studies show that HMB in systemic circulation can cross the BBB via carrier-mediated processes, and that it also has a positive influence on the expression of LAT1, an important amino acid transporter in the BBB.\n\nID: 34544422\nTitle: Investigating receptor-mediated antibody transcytosis using blood-brain barrier organoid arrays.\nAbstract: The pathways that control protein transport across the blood-brain barrier (BBB) remain poorly characterized. Despite great advances in recapitulating the human BBB in vitro, current models are not suitable for systematic analysis of the molecular mechanisms of antibody transport. The gaps in our mechanistic understanding of antibody transcytosis hinder new therapeutic delivery strategy development. We applied a novel bioengineering approach to generate human BBB organoids by the self-assembly of astrocytes, pericytes and brain endothelial cells with unprecedented throughput and reproducibility using micro patterned hydrogels. We designed a semi-automated and scalable imaging assay to measure receptor-mediated transcytosis of antibodies. Finally, we developed a workflow to use CRISPR/Cas9 gene editing in BBB organoid arrays to knock out regulators of endocytosis specifically in brain endothelial cells in order to dissect the molecular mechanisms of receptor-mediated transcytosis. BBB organoid arrays allowed the simultaneous growth of more than 3000 homogenous organoids per individual experiment in a highly reproducible manner. BBB organoid arrays showed low permeability to macromolecules and prevented transport of human non-targeting antibodies. In contrast, a monovalent antibody targeting the human transferrin receptor underwent dose- and time-dependent transcytosis in organoids. Using CRISPR/Cas9 gene editing in BBB organoid arrays, we showed that clathrin, but not caveolin, is required for transferrin receptor-dependent transcytosis. Human BBB organoid arrays are a robust high-throughput platform that can be used to discover new mechanisms of receptor-mediated antibody transcytosis. The implementation of this platform during early stages of drug discovery can accelerate the development of new brain delivery technologies.\n\nID: 34471008\nTitle: [Development of Biomembrane-mimetic Nanoparticles for the Treatment of Ischemic Stroke].\nAbstract: Increase in vascular permeability of the blood-brain barrier (BBB) is a distinct pathology following ischemic stroke. In previous studies, we demonstrated that liposomal drug delivery system (DDS)-based delivery of neuroprotectants is useful for treating cerebral ischemia/reperfusion injury. Additionally, our previous studies reported that combination therapy with liposomal fasudil plus tissue plasminogen activator (t-PA), a thrombolytic agent, brings about decrease in the risk of t-PA-derived cerebral hemorrhage and prolong the therapeutic time window of t-PA for treating acute ischemic stroke. However, accumulation of systemically administered liposomes into the brain parenchyma is still limited, and new technologies are needed that can overcome the BBB. The unique properties of leukocytes and exosomes, one of the extracellular vesicles, make them promising candidates for overcoming biological barriers (including the BBB) for drug delivery, as leukocytes and certain exosomes were reported to be able to permeate through the inflamed BBB in the ischemic stroke area. We prepared leukocyte-mimetic liposomes (LM-Lipo) via transfer of leukocyte membrane proteins by employing a phenomenon called intermembrane protein transfer, and demonstrated that LM-Lipo could pass through the layer of inflamed endothelial cells via regulation of intercellular junctions, like leukocytes. Additionally, we showed that LM-Lipo efficiently penetrated into spheroids of cancer cells, and inhibited their growth by entrapped doxorubicin. Taken together, it was suggested that imparting leukocyte-like characteristics to liposomes may be an effective approach to overcoming biological barriers. Herein, we summarize our findings regarding liposomal DDS for ischemic stroke therapy and recent approaches to develop biomembrane-mimetic DDS using leukocytes and exosomes.\n\nID: 34376696\nTitle: NEK1-mediated retromer trafficking promotes blood-brain barrier integrity by regulating glucose metabolism and RIPK1 activation.\nAbstract: Loss-of-function mutations in NEK1 gene, which encodes a serine/threonine kinase, are involved in human developmental disorders and ALS. Here we show that NEK1 regulates retromer-mediated endosomal trafficking by phosphorylating VPS26B. NEK1 deficiency disrupts endosomal trafficking of plasma membrane proteins and cerebral proteome homeostasis to promote mitochondrial and lysosomal dysfunction and aggregation of \u03b1-synuclein. The metabolic and proteomic defects of NEK1 deficiency disrupts the integrity of blood-brain barrier (BBB) by promoting lysosomal degradation of A20, a key modulator of RIPK1, thus sensitizing cerebrovascular endothelial cells to RIPK1-dependent apoptosis and necroptosis. Genetic inactivation of RIPK1 or metabolic rescue with ketogenic diet can prevent postnatal lethality and BBB damage in NEK1 deficient mice. Inhibition of RIPK1 reduces neuroinflammation and aggregation of \u03b1-synuclein in the brains of NEK1 deficient mice. Our study identifies a molecular mechanism by which retromer trafficking and metabolism regulates cerebrovascular integrity, cerebral proteome homeostasis and RIPK1-mediated neuroinflammation.\n=======================================================\n\n### [CUSTOM DATAPOINTS]\nCRITICAL EXTRACTION DIRECTIVE: You MUST extract the following custom datapoints as root-level key/value pairs inside your final JSON block:\n- \"suggested_experiments\": generate 1-3 suggested experiments\n- \"suggested_studies\": generate 1-3 suggested studies\n- \"swansons_literature_based_discovery_candidates\": You are an advanced Literature-Based Discovery (LBD) system executing Swanson\u2019s complementary-but-disjoint (A-B-C) model. Your goal is to find hidden, unpublished connections across the provided dataset. Strict Discovery Protocol: 1. Identify distinct, isolated sub-literatures (Domain A and Domain C) within the dataset that share NO direct citations, co-mentions, or common contextual paragraphs. 2. Find an intermediate biological mechanism, protein, path, or entity (Bridge B) that appears independently in both isolated domains (A-to-B and B-to-C). 3. Synthesize a novel, unstated hypothesis (A-to-C). Negative Constraint (Crucial): DO NOT output any connection if the relationship between Concept A and Concept C is explicitly mentioned, paired, or summarized anywhere in the source text. If a connection (like \"OMN resilience to SMN stabilization\") is already explicitly stated or grouped as a concept in the data, it is considered \"already known\" and must be disqualified. Format your output exactly as follows: - Discovered Hypothesis (A to C): [Clear, novel statement] - Literature A (Origin): [Entity/Concept and source context] - Literature C (Target): [Entity/Concept and source context] - The Intersecting Bridge B: [The shared mechanism/protein linking them] - Biological Rationale: [1-2 sentences explaining why this hidden connection is mechanistically plausible]\n- \"contradictions_between_evidences\": Identify conflicting evidence within the evidence set (if any) and flag the dispute here\n- \"repurposed_solutions\": identify and explain repurposed Solution potentials\n- \"charge_density_threshold\": Identify if a specific range of zeta potential or positive charge density per aliphatic carbon chain length correlates with maximum permeability coefficient-surface area (PS) values.\n- \"metabolic_synergy\": Investigate if \u03b2HB-induced autophagic clearance rates are influenced by the specific polyamine-modified charge profile of pathological proteins.\n- \"transporter_interaction\": Determine if the decrease in PS values for higher-charge polyamines implies engagement of distinct, potentially inhibitory, transport pathways compared to the putrescine-modified uptake mechanism.\n\n\nFormat Requirement:\nRAG AMNESIA IS ACTIVE: You must ONLY use the provided context literature. Do not use outside prior knowledge. If the evidence is missing, insufficient, or requires gap-filling to fully evaluate the claim, you MUST explicitly state the gaps and missing evidence in your justification. Under no circumstances should you invent or hallucinate citations or quotes.\n\nFirst provide disclaimer such as \"Even though this fact check looked at unique up-to-date abstracts, new evidence may refute this answer in the future. Although 'Zero Hallucinated Moneyshot Quotes' is programmatically enforced, AI is not always immune to inadvertently/erroneously misinterpreting data. This is not medical or professional advice, but instead, is an opinion calculated by AI based on the literature evaluated.\"\n---\nWrite in a highly academic, formal thesis tone.\nFormat your readable response using these exact academic headers:\n###[CLAIM EVALUATED AND ANSWER TO USER]\n(Exact wording of the claim evaluated)\n### [ABSTRACT & REWRITTEN CLAIM]\n(Scientific synthesis)\n### [INTRODUCTION & JUSTIFICATION]\n(Mechanistic explanation utilizing the 'moneyshot quotes' you will use in the EVIDENCE, METHODOLOGY & CITATIONS section later as well)\n### [DISCUSSION: NOVEL & OVERLOOKED]\n(5-10 bullet points of surprising facts)\n### [EVIDENCE, METHODOLOGY & CITATIONS]\n(Numbered list matching inline citations) For example \"1. ID: 12345 - Application: The text discusses ... and since no other evidence provided proves nor disproves the claim, the lowest rating allowed across all evidences is required. ID:12345 indicates the claim is overall plausible (Alignment with this ID: 3) - [copied/verbatim Quote text]\"\n\n**CRITICAL: You must include the exact quote you used in the [copied/verbatim Quote text] section.\n\nIf the prompt says \"at least 20 quotes\" then there must be at least 20 matching citations. You must actually use the quotes you select within the conext of the preprint publication you write.\n\nEvaluation Schema:\nRAG AMNESIA IS ACTIVE: You must ONLY use the provided context literature. Do not use outside prior knowledge. If the evidence is missing, insufficient, or requires gap-filling to fully evaluate the claim, you MUST explicitly state the gaps and missing evidence in your justification. Under no circumstances should you invent or hallucinate citations or quotes.\n\n###critical: WRAP YOUR THOUGHTS WITH The 'Charge Density Threshold' remains an undefined parameter within the current literature corpus [ID: Run2_Eval1_synthesis]. Clinical observation indicates a parabolic relationship between cationic charge and systemic efficacy: while increased cationic charge facilitates blood-brain barrier (BBB) penetration, excessive systemic charge\u2014exemplified by molecules such as avidin\u2014triggers accelerated systemic clearance [ID: Run2_Eval1_synthesis].
Identified Gaps: The absence of a quantified threshold suggests a critical knowledge gap in pharmacokinetics and barrier-crossing efficiency, requiring further titration studies to isolate the optimal charge-to-clearance ratio.
"
},
{
"type": "bottlenecks",
"title": "Critical Literature Gaps"
},
{
"type": "comparison_matrix",
"title": "Charge Impact Matrix",
"headers": [
"Parameter",
"Low Charge",
"Excessive Charge"
],
"rows": [
[
"BBB Binding",
"Insufficient",
"Optimal to High"
],
[
"Systemic Clearance",
"Baseline",
"Rapid/Accelerated"
]
]
},
{
"type": "gap_distribution",
"title": "Evidence Sufficiency Map"
}
]
}
},
{
"id": "mvc_dp_metabolic_synergy_1786460723347",
"title": "Metabolic Synergy Report",
"plan": {
"title": "METABOLIC SYNERGY : CUSTOM ANALYSIS",
"evidence_tier": "EVALUATED",
"panels": [
{
"type": "metrics",
"title": "Data Integrity Scorecard"
},
{
"type": "synthesis",
"title": "Executive Analysis: Metabolic Synergy",
"content": "The current dataset establishes a clear link between BHB (\u03b2-hydroxybutyrate) and the induction of protein insolubility and autophagic clearance mechanisms [ID: 39626664]. Furthermore, it identifies that polyamine-modified proteins undergo efficient transcytosis. However, a critical knowledge gap exists regarding the specific regulatory modulation of protein insolubility on the rate or efficiency of polyamine-modified protein transcytosis. This interaction remains unmapped in current literature."
},
{
"type": "gap_distribution",
"title": "Knowledge Gap Assessment"
},
{
"type": "bottlenecks",
"title": "Identified Mapping Bottlenecks"
},
{
"type": "logic_network",
"title": "Metabolic Pathway Mapping"
},
{
"type": "node_centrality",
"title": "Primary Entity Relevance"
}
]
}
},
{
"id": "mvc_dp_transporter_interaction_1786460736866",
"title": "Transporter Interaction Report",
"plan": {
"title": "TRANSPORTER INTERACTION : CUSTOM ANALYSIS",
"evidence_tier": "EVALUATED",
"panels": [
{
"type": "metrics",
"title": "Data Integrity Scorecard"
},
{
"type": "synthesis",
"title": "Executive Summary: Transporter Interaction",
"content": "The analysis of 'Transporter Interaction' reveals a critical knowledge gap. Current literature distinguishes between specific transport systems, namely choline-uptake, basic amino acid, and polyamine transport mechanisms [Run2_Eval1_synthesis]. However, there is insufficient evidence to determine the inhibitory interplay between these high-charge carriers. This lack of mechanistic resolution prevents the establishment of a comprehensive interaction model."
},
{
"type": "gap_distribution",
"title": "Evidence Gap Density"
},
{
"type": "bottlenecks",
"title": "Identified Literature Gaps"
},
{
"type": "comparison_matrix",
"title": "Transport System Categorization",
"headers": [
"System Type",
"Status",
"Interaction Evidence"
],
"rows": [
[
"Choline-Uptake",
"Categorized",
"None"
],
[
"Basic Amino Acid",
"Categorized",
"None"
],
[
"Polyamine",
"Categorized",
"None"
]
]
}
]
}
}
],
"aggregatedDatapoints": {
"suggested_experiments": [
{
"pentamatrix": "Run1_Eval1_synthesis",
"data": [
"Quantify BBB permeability of polyamine-modified therapeutic proteins using specific transporter knockouts to confirm the non-electrostatic flux hypothesis.",
"Perform mass spectrometry-based profiling of the protein insolublome in the presence of various synthetic polyamine derivatives to map charge-density vs. solubility outcomes."
]
},
{
"pentamatrix": "Run2_Eval1_synthesis",
"data": [
"Test the effect of varying arginine vs. lysine-based polyamine conjugation on BBB flux in hCMEC/D3 models.",
"Evaluate the impact of \u03b2HB on the endolysosomal clearance kinetics of cationic vs. native proteins in human brain endothelial cell cultures.",
"Assess if specific polyamine lengths affect the stability of the endosomal-lysosomal fusion process (SNARE protein-mediated)."
]
}
],
"suggested_studies": [
{
"pentamatrix": "Run1_Eval1_synthesis",
"data": [
"Comparative analysis of ketone ester delivery vs. polyamine-modified peptide therapy in transgenic AD mouse models to evaluate synergistic effects on proteostasis.",
"Evaluation of pH-dependent protein stability shifts using circular dichroism and fluorescence spectroscopy to decouple charge-based aggregation from \u03b2HB-mediated effects."
]
},
{
"pentamatrix": "Run2_Eval1_synthesis",
"data": [
"Quantify the optimal charge density threshold for transcytosis versus systemic clearance for therapeutic proteins.",
"Investigate the interplay between H3K9 \u03b2-hydroxybutyrylation and the expression of endothelial transport proteins under glucose-deprivation conditions."
]
}
],
"swansons_literature_based_discovery_candidates": [
{
"pentamatrix": "Run1_Eval1_synthesis",
"data": {
"Discovered Hypothesis (A to C)": "Metabolic regulation of autophagy by \u03b2HB may act in tandem with polyamine-modified chaperone delivery to accelerate the clearance of pre-aggregated amyloid-\u03b2 plaques.",
"Literature A (Origin)": "\u03b2HB-induced autophagic clearance of pathological proteins (Source ID 37461525)",
"Literature C (Target)": "Polyamine-mediated BBB transport of therapeutic peptides (Source ID 8627316)",
"The Intersecting Bridge B": "Intracellular protein transport and degradation pathways (autophagy)",
"Biological Rationale": "Since \u03b2HB increases the clearance of neurodegeneration-related proteins via autophagy and polyamines increase the flux of therapeutic molecules into the brain, combining these approaches might optimize both the targeting of plaques and the metabolic capacity for their removal."
}
},
{
"pentamatrix": "Run2_Eval1_synthesis",
"data": "- Discovered Hypothesis (A to C): \u03b2-Hydroxybutyrate treatment restores blood-brain barrier integrity in metabolic disorder models by promoting retromer-mediated protein trafficking (VPS35). - Literature A (Origin): BHB's role in endothelial ZO-1 expression and integrity (ID: 38666466). - Literature C (Target): VPS35 and endosomal retromer complex maintenance of BBB integrity (ID: 35002279). - The Intersecting Bridge B: Endothelial cell proteostasis (autophagy/lysosomal degradation pathways). - Biological Rationale: BHB has been shown to induce protein solubility and autophagic clearance in the brain (ID: 37461525), and retromer dysfunction causes tau-associated accumulation (ID: 35002279). It is mechanistically plausible that BHB-induced autophagic flux mitigates the downstream proteostasis collapse caused by retromer deficiency in cerebral endothelial cells."
}
],
"contradictions_between_evidences": [
{
"pentamatrix": "Run1_Eval1_synthesis",
"data": "There is a seeming contradiction regarding electrostatic interactions: cationic protein modification increases permeability (ID 8627316), yet increased positive charge density along the polyamine chain paradoxically decreases permeability, suggesting a complex, saturable, or non-electrostatic transport mechanism."
},
{
"pentamatrix": "Run2_Eval1_synthesis",
"data": "There is a slight conflict regarding whether polyamines always enhance uptake or if they can induce excitotoxicity or barrier disruption if ODC is hyperactivated, suggesting a delicate homeostasis for polyamine-mediated transport (ID: 3097421, ID: 28867747)."
}
],
"repurposed_solutions": [
{
"pentamatrix": "Run1_Eval1_synthesis",
"data": "The use of polyamine-modified peptides, originally studied for drug delivery (Source ID 8627316, 10363910), could be repurposed to target and destabilize existing amyloid deposits rather than just inhibiting their initial formation."
},
{
"pentamatrix": "Run2_Eval1_synthesis",
"data": "Polyamine-modified catalase and growth factors are already repurposed vectors for CNS delivery, and BHB-mediated metabolic reprogramming is now suggested as a strategy to enhance barrier integrity post-ischemia."
}
],
"charge_density_threshold": [
{
"pentamatrix": "Run2_Eval1_synthesis",
"data": "Not explicitly defined in the provided literature; however, cationic charge is universally shown to increase BBB binding, while excessive systemic charge (e.g., avidin) leads to rapid clearance."
}
],
"metabolic_synergy": [
{
"pentamatrix": "Run2_Eval1_synthesis",
"data": "Evidence confirms BHB induces protein insolubility/autophagic clearance (ID: 39626664) and that polyamine-modified proteins are efficiently transcytosed; the exact modulation of the former on the latter remains unmapped."
}
],
"transporter_interaction": [
{
"pentamatrix": "Run2_Eval1_synthesis",
"data": "Insufficient data; literature differentiates between choline-uptake, basic amino acid, and polyamine transport systems, but the specific inhibitory interplay between these high-charge carriers is currently unknown."
}
]
},
"stats": {
"promptTokens": 209912,
"completionTokens": 24721,
"totalTokens": 234633
},
"zenodo_doi": "10.5281/zenodo.21891046"
}