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This investigation builds upon compelling evidence that membrane lipid composition, particularly the presence of anionic phospholipids such as phosphatidylserine and phosphatidylinosit","entities":{"genes":["CERS2/CLDN1/DNMT1"],"diseases":["Parkinson's Disease"]},"model_system":"human","_schema_version":1,"experiment_type":"validation","primary_outcome":"Validate Validation: Membrane-Nucleation in iPSC Neurons","methodology_notes":"**Phase 1: iPSC-Derived Neuron Preparation (Weeks 1-6)**\n• Differentiate human iPSCs (n=6 cell lines, 3 healthy controls + 3 PD patients with SNCA mutations) into dopaminergic neurons using dual SMAD inhibition protocol\n• Validate neuronal identity at day 35-42 using immunofluorescence (TH+, MAP2+, β-tubulin III+)\n• Confirm dopaminergic phenotype with HPLC analysis of dopamine production\n• Seed neurons at 50,000 cells/well in 96-well imaging plates for membrane composition analysis\n\n**Phase 2: Membrane Lipid Composition Manipulation (Week 7)**\n• Treat neurons with lipid supplementation medium containing: PS (25-100 μM), PIP2 (10-50 μM), or vehicle control\n• Include positive control with rotenone (100 nM) and negative control with cholesterol supplementation\n• Incubate for 48-72 hours with daily medium changes\n• Validate membrane incorporation using mass spectrometry lipidomics (LC-MS/MS)\n\n**Phase 3: Alpha-Synuclein Analysis (Week 8)**\n• Fix cells at 24h, 48h, and 72h timepoints for imm","replication_status":"single_study","extraction_metadata":{"backfill_at":"2026-04-16T01:00:16.906323","needs_review":true,"extraction_notes":"Backfilled from wiki source (no PMID 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