註釋 The human blood-brain barrier (BBB) represents the interface of microvasculature and the central nervous system, regulating the transport of nutrients and protecting the brain from external threats. To gain a deeper understanding of (patho)physiological processes affecting the BBB, sophisticated models mimicking the in vivo situation are required. Currently, most in vitro models are cultivated on stiff, semipermeable, and non-biodegradable Transwell® membrane inserts, not adequately mimicking the complexity of the extracellular environment of the native human BBB. To overcome these disadvantages, we developed three-dimensional electrospun scaffolds resembling the natural structure of the human extracellular matrix. The polymer fibers of the scaffold imitate collagen fibrils of the human basement membrane, exhibiting excellent wettability and biomechanical properties, thus facilitating cell adhesion, proliferation, and migration. Cultivation of human induced pluripotent stem cells (hiPSCs) on these scaffolds enabled the development of a physiological BBB phenotype monitored via the formation of tight junctions and validated by the paracellular permeability of sodium fluorescein, further accentuating the non-linearity of TEER and barrier permeability. The novel in vitro model of the BBB forms a tight endothelial barrier, offering a platform to study barrier functions in a (patho)physiologically relevant context. 