Vascularization by endothelial cells (ECs) is an important element in tissue engineering of organoids. Morphogenesis of these cells is regulated not only by biochemical properties of the extracellular matrix (ECM) but also by its mechanical properties. Here, we investigated the effect of adhesion substrate elasticity on the formation of capillary-like networks by ECs; in particular, we examined the three-dimensional (3D) configurations of the resulting networks. Bovine pulmonary microvascular ECs were cultured on a series of collagen gels of different elasticities but the same collagen concentration. The cells cultured in rigid and flexible gels formed 3D networks via different processes; cells formed dense, thin networks in the flexible gel, whereas thicker and deeper networks were formed in the rigid gel. Cross-sections of the networks revealed that those formed within the rigid gel had large lumens composed of multiple cells, whereas those formed within the flexible gel had small, intracellular vacuoles.
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