Single-cell transcriptomics of vascularized human brain organoids decipher lineage-specific stress adaptation in fetal hypoxia-reoxygenation injury.

Fetal hypoxia, a major contributor to neonatal mortality, induces complex neurovascular disruptions in developing brains, yet human-specific cellular mechanisms remain poorly understood due to limitations in existing models. This study establishes an advanced vascularized human cortical organoid (vhCO) model to decode cell type-specific injury mechanisms and therapeutic targets during hypoxia-reoxygenation. We developed vhCOs by integrating cortical and vascular organoids, recapitulating mid-to-late gestational neurodevelopment with diverse lineages-neural progenitors, neurons, microglia, and functional vasculature with blood-brain barrier properties. Hypoxia-reoxygenation experiments were conducted on vhCOs, followed by single-cell transcriptomic profiling to dissect cellular responses. Key findings include: (1) Lineage-specific vulnerabilities: astrocyte precursors exhibited developmental arrest, while immature GABAergic neurons (Subtype I) underwent neurogenic collapse. Microglia displayed a biphasic inflammatory response-initially suppressed, then hyperactivated post-reoxygenation, diverging from animal models; (2) Hypoxia memory persisted in non-neural cells (pericytes, fibroblasts), driving compartment-specific vascular remodeling via Notch signaling and collagen deposition; (3) Rewired neural-non-neural crosstalk networks (e.g., IGF2-IGF2R, LGALS3-MERTK, Wnts-SFRP2) revealed novel repair targets inaccessible to conventional models. By prioritizing single-cell resolution, this study delineates human-specific neurovascular pathophysiology and stress adaptation networks in hypoxic brain injury. The vhCO platform bridges translational gaps, offering a paradigm for precision therapeutics and advancing research on developmental brain disorders.