Blood-Brain Barrier Aging and Neurodegeneration — From Leakage to Neuronal Loss

SUMMARY

# Blood-Brain Barrier Aging and Neurodegeneration — From Leakage to Neuronal Loss ## Background and Rationale Blood-brain barrier (BBB) integrity deteriorates with aging and is increasingly recognized as a contributing factor to neurodegeneration across multiple diseases including Alzheimer's, Parkinson's, and ALS. This validation study employs in vitro BBB models using human brain microvascular endothelial cell lines to investigate age-related changes in barrier function and their mechanistic l

METHODOLOGY NOTES

**Phase 1: Cell Culture Establishment (Days 1-7)** • Establish primary human brain microvascular endothelial cells (hBMECs) and co-culture with pericytes and astrocytes in transwell inserts • Seed hBMECs at 2×10^5 cells/cm² on collagen IV-coated transwell membranes (0.4 μm pore size) • Add pericytes (1×10^4 cells/cm²) and astrocytes (5×10^4 cells/cm²) to basal compartment • Maintain in endothelial growth medium with 10% FBS for 5-7 days until confluent monolayer formation • Confirm barrier integrity by transendothelial electrical resistance (TEER) >150 Ω·cm² **Phase 2: Aging Induction Protocol (Days 8-21)** • Apply aging stress conditions: chronic oxidative stress (100 μM H₂O₂, 2h daily), inflammatory cytokines (TNF-α 10 ng/ml, IL-1β 5 ng/ml), and advanced glycation end products (AGEs, 100 μg/ml) • Establish control groups (n=24 per condition): young BBB model (no treatment), aged BBB model (aging cocktail), and positive controls • Monitor TEER daily and replace media every 48 hours •