Figure 3.

Characterization of hCMVECs/hMSCs co-culture derived in situ de novo vascular structures by immunofluorescence microscopy. Day 14 vasculogenic hCMVECs/hMSCs CCC co-cultures generated not only extensive plexuses of capillary structures lined simply by endothelial cells but also vascular structures resembling that of small to medium sized muscular arteries (A–O). These thin and delicately intertwined microvascular structures expressed both endothelial and smooth muscle cell markers, VWF and α-SMA, respectively, and revealed apparent areas of co-localizations (A–C). The PECAM1 positive endothelial cells formed a linear solid cord-like structure, and were uniformly surrounded by the α-SMA positive smooth muscle cells, and these α- SMA positive cells were oriented in a direction that was perpendicular to the direction of linearly assembled endothelial cells (D–F). Further morphogenesis of these endothelium lined tubular structures illustrated the dynamic process of evolving tunica media, the hMSC-derived α-SMA positive cells were wrapping around the entire circumference of these endothelial outgrowth (G–I). Subsequent stages of morphogenesis revealed emergence of greater caliber vessels, having sleeve-like outer smooth muscle cells encasing the endothelium lined tubular structure (J–O). Merged images (A–O). (A–O, scale bar 100 μm).