Electrical Stimulation Directs Formation of Perfused Vasculature in Engineered Tissues

Effective, rapid, and functionally perfusable vascularization remains a major challenge in tissue engineering. Current approaches to generating vasculature in vitro require multipart fabrication methods or complex and costly media supplements, limiting their scalability. Here, we demonstrate that exogenous electrical stimulation (ESTIM) offers a promising alternative by enhancing 3D vasculogenesis in engineered human tissues. Exposing 3D endothelial-fibroblast cocultures to pulsed ESTIM promoted the formation of dense and branched vascular networks. In a microfluidic device model, we show that ESTIM induces the formation of a perfusable, interconnected vascular network, whereas unstimulated networks remain less mature. Importantly, we demonstrate that upon implantation, ESTIM-pretreated vascular grafts exhibit elevated anastomosis with the host and perfusion with blood relative to the untreated grafts. In addition, we use ESTIM to promote engraftment of a vascularized 3D liver construct. Mechanistically, we find that ESTIM induces membrane hyperpolarization in endothelial cells via voltage-gated potassium channels (K_V). Inhibiting K_Vs abrogated ESTIM's pro-vasculogenic effects in endothelial cells. Conversely, pharmacologically activating hyperpolarization induced endothelial responses even without ESTIM, directly linking K_V-mediated hyperpolarization as a key mechanism by which ESTIM drives vascular assembly and function. Ultimately, our work establishes ESTIM as a new orthogonal approach to promote the formation of perfusable vasculature in engineered tissues.