Abstract
Objective: The focus of this study was to investigate the role of connexin (Cx) 45 in endothelial-induced mural cell differentiation. Methods and results: We created mural cell precursors that stably express only Cx45 in Cx43-deficient mesenchymal cells (ReCx45), and used our in vitro model of blood vessel assembly to assess the capacity of this Cx to support endothelial-induced mural cell differentiation. Lucifer Yellow dye injection and dual whole-cell patch clamping revealed that functional gap junctions exhibiting properties of Cx45-containing channels formed among ReCx45 transfectants, and between ReCx45 and endothelial cells. Heterocellular Cx45-containing gap junction channels enabled transforming growth factor-β activation and promoted the upregulation of mural cell-specific proteins in the mesenchymal precursors. Conclusions: These studies reveal a critical role for Cx45 in the regulation of endothelial-induced mural cell differentiation, which is consistent with the phenotype of Cx45-deficient embryos that exhibit dysregulated transforming growth factor-β and lack mural cell development.