Gap junction communication mediates transforming growth factor-β activation and endothelial-induced mural cell differentiation

During blood vessel assembly, endothelial cells recruit mesenchymal progenitors and induce their differentiation into mural cells via contact-dependent transforming growth factor-beta (TGF-beta) activation. We investigated whether gap junction channels are formed between endothelial cells and recruited mesenchymal progenitors and whether intercellular communication is necessary for endothelial-induced mural cell differentiation. Mesenchymal progenitors from Cx43(-/-) murine embryos and Cx43(+/+) littermates were cocultured with prelabeled endothelial cells. Intracellular dye injection and dual whole-cell voltage clamp revealed that endothelial cells formed gap junction channels with Cx43(+/+) but not Cx43(-/-) progenitors. In coculture with endothelial cells, Cx43(-/-) progenitors did not undergo mural cell differentiation as did Cx43(+/+) cells. Stable reexpression of Cx43 in Cx43(-/-) cells (reCx43) restored their ability to form gap junctions with endothelial cells and undergo endothelial-induced mural cell differentiation. Cocultures of endothelial cells and either Cx43(+/+) or reCx43 mesenchymal cells produced activated TGF-beta; endothelial-Cx43(-/-) cocultures did not. However, Cx43(-/-) cells did produce latent TGF-beta and undergo mural cell differentiation in response to exogenous TGF-beta1. These studies indicate that gap junction communication between endothelial and mesenchymal cells mediates TGF-beta activation and subsequent mural cell differentiation.