| Abstract |
Transplanted endothelial progenitor cells (EPCs) repair blood vessels and exert regenerative effects on disorders such as lower limb ischemia. EPCs serve as a model for pathophysiological and pharmacokinetic studies, which is important for drug discovery. However, primary human EPCs are phenotypically unstable, which limits their clinical utility. Therefore, we employed human induced pluripotent stem (iPS) cells to circumvent this problem. Here we focused on human iPS cell-derived sac-like structures (iPS-sacs), which contain endothelial lineage cells and hematopoietic lineage cells. Previous studies isolated only hematopoietic lineage cells from iPS-sacs. Therefore, here we attempted to isolate EPCs. However, iPS-sacs generated by a published protocol did not contain sufficient EPCs. Therefore, to generate iPS-sacs highly enriched in EPCs, we added the glycogen synthase kinase 3 beta (GSK3β) inhibitor CHIR-99021 to the culture medium early during differentiation. The cells rapidly differentiated into mesoderm to yield abundant EPCs, and CHIR-99021 increased the proportion of EPCs contained in iPS-sacs. EPCs, which were purified using anti-platelet endothelial cell adhesion molecule (PECAM1) antibody-conjugated beads, expressed markers of immature endothelial cells. Purified EPCs formed tube-like structures and incorporated acetylated low density lipoprotein (Ac-LDL), reflecting endothelial phenotypes. The simple method described here will likely improve regenerative medicine and facilitate basic studies on the endothelial lineage.
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