Corneal endothelial expansion using human umbilical cord mesenchymal stem cell-derived conditioned medium

Rabbit corneal endothelial cells are frequently used in pharmacological experiments and are useful for corneal transplant experiments. We performed the present study to analyze the effect of conditioned medium (CM) derived from human umbilical cord mesenchymal stem cells (HUMSCs) on the growth of rabbit corneal endothelial cells (RCECs) and to establish a program for expansion of RCECs in vitro. RCECs were cultured using a CM derived from HUMSCs (HUMSCs-CM) in vitro. The proliferation ability of RCECs cultured in the presence of HUMSCs-CM was evaluated by conducting 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide, colony formation, and scratch migration assays. The proliferation ability of RCECs maintained in HUMSCs-CM was significantly enhanced as compared to RCECs cultivated in the control group. Immunofluorescence indicated that zonula occludens-1 (ZO-1) and N-cadherin were located at intercellular junctions. Real-time PCR and western blot analyses demonstrated that the CEC-relative functional markers were expressed in RCECs maintained in HUMSCs-CM. Flow cytometry analyses demonstrated that HUMSCs-CM promoted the G0/G1 entrance to the S phase in RCECs. Our results demonstrated that HUMSCs-CM induced the proliferation of RCECs in vitro and maintained the necessary characteristic phenotypes. The expanded RCECs may provide a promising cell source for experimental research and clinical therapy.