Objective:

Ischemic stroke remains a leading cause of death and disability worldwide. However, conventional in vitro models often lack the cellular complexity and human relevance required for effective translational research. This study aims to establish a human-based in vitro platform for ischemic stroke modelling by differentiating induced pluripotent stem cells (iPSCs) into key neurovascular cell types and assessing their functional properties under ischemic-like conditions.

Methods:

Human iPSCs were differentiated into cortical neurons, brain microvascular endothelial cells and astrocytes, using lineage-specific protocols. Cell identity was confirmed by immunocytochemistry and gene expression profiling. Functional characterization included calcium imaging for neurons, glutamate uptake assays for astrocytes, and barrier integrity measurements for endothelial cells. To model ischemic injury, neurons and endothelial cells were exposed to oxygen-glucose deprivation (OGD), and their responses were assessed by morphological analysis and cell viability assays.

Results:

Differentiation protocols yielded cortical neurons expressing MAP2 and βIII-tubulin, astrocytes expressing GFAP and S100β, and endothelial cells positive for CD31 and VE-cadherin. Functional assays confirmed the maturity and physiological relevance of each cell type. Under OGD conditions, both neurons and endothelial cells exhibited measurable stress responses, including morphological alterations and reduced viability, validating the model’s potential to mimic key aspects of ischemic injury. Development of neurovascular co-culture systems is ongoing.

Conclusion:

We successfully established and functionally validated iPSC-derived human neurons, astrocytes, and endothelial cells as components of an in vitro stroke model. Preliminary OGD experiments highlight the model’s potential for studying cell-type-specific responses to ischemic stress. This platform lays the groundwork for future mechanistic studies and therapeutic screening in a human-relevant context.