Soft nanofiber modified micropatterned substrates enhance native-like endothelium maturation via CXCR4/calcium-mediated actin cytoskeleton assembly

Regeneration and maturation of native-like endothelium is crucial for material-guided small-diameter vascular regeneration. Although parallel-microgroove-patterned (micropatterned) substrates are capable of promoting endothelial regeneration with native-like endothelial cell (EC) alignment, their unbefitting high-stiffness acutely inhibits cell—matrix interaction and endothelial maturation. Given that the sufficient softness of nanofibers allows cells to deform the local matrix architecture to satisfy cell survival and functional requirements, in this study, an effective strategy of decorating micropatterned substrate with soft nanofibers was exploited to enhance cell—matrix interaction for engineering healthy endothelium. Results demonstrated that the micropatterned nanofibrous membranes were successfully obtained with high-resolution parallel microgrooves (groove width: ∼ 15 µm; groove depth: ∼ 5 µm) and adequate softness (bulk modulus: 2.27 ± 0.18 MPa). This particular substrate markedly accelerated the formation and maturation of confluent native-like endothelium by synchronously increasing cell—cell and cell—matrix interactions. Transcriptome analysis revealed that compared with smooth features, the microgrooved pattern was likely to promote endothelial remodeling via integrin α5-mediated microtubule disassembly and type I interleukin 1 receptor-mediated signaling pathways, whereas the nanofibrous pattern was likely to guide endothelial regeneration via integrin α5β8-guided actin cytoskeleton remodeling. Nevertheless, endowing micropatterned substrate with soft nanofibers was demonstrated to accelerate endothelial maturation via chemokine (C-X-C motif) receptor 4/calcium-mediated actin cytoskeleton assembly. Furthermore, numerical simulation results of hemodynamics indicated the positive impact of the micropatterned nanofibers on maintaining stable hemodynamics. Summarily, our current work supports an affirmation that the micropatterned nanofibrous substrates can significantly promote regeneration and maturation of native-like endothelium, which provides an innovative method for constructing vascular grafts with functional endothelium.