Culture conditions of human embryonic stem cells for differentiation into retinal vascular structure.

ABSTRACT

OBJECTIVE: This study sought to identify the suitable cell culture conditions for the in vitro-induced differentiation of human embryonic stem cells (hESCs) into retinal vascular tissue cell types. MATERIALS AND METHODS: To do this, we established four treatment groups. Group A was designed to culture hESCs in a three-dimensional system. The feeder cells and leukemia inhibitory factor (LIF) were removed in Group B. In group C, hESCs were cultured with a variety of pro-angiogenic growth factors. In group D, hESCs were cultured with intact retinal support cells and extracellular matrix. On days 15 and 30, the expression of platelet endothelial cell adhesion molecule 1 (PECAM1), α-smooth muscle actin (αSMA), and macrophage marker F4/80 were detected by immunofluorescence staining. ELISA was used to detect the expression of stromal cell-derived factor-1 (SDF-1). RESULTS: At both 15 and 30 day timepoints, the highest PECAM1, αSMA, and F4/80 positive rates and SDF-1 expression levels were observed in group D, followed by group C, group B, with group A presenting the lowest expression of these proteins (p<0.05). Also, group D showed obvious angiogenesis structures. CONCLUSIONS: Our study indicates that hESCs can differentiate into retinal vascular-like structures. The presence of intact retinal support cells, a variety of cytokines, and extracellular matrix components were essential to facilitate this differentiation.