Comparison of equine-induced pluripotent stem cell characteristics induced on different cell adhesion substrates

Abstract

This study evaluated the effects of cell adhesion substrates that lead to the generation of equine-induced pluripotent stem cells (eiPSC) from embryonic skin fibroblasts by lipofection of plasmid vectors expressing five reprogramming factors. The reprogramming efficiency of cells induced on the E8 fragment of laminin-511 (eiPSC-511) was higher than that on Geltrex containing laminin-111 as a major laminin (eiPSC-111), and supplementation with a cocktail of small molecular compounds increased the number of iPSC colonies on both substrates. In the cell proliferation assay, eiPSC-511 showed higher growth activity than eiPSC-111. Although no significant changes were observed in the expression of pluripotency markers between eiPSC-111 and eiPSC-511, the expression of DPPA3 was significantly upregulated in both iPSCs by reprogramming, suggesting that DPPA3 was a sensitive pluripotent marker for equine iPSC. While both iPSCs expressed high mRNA level of integrin alpha6 and beta1 subunits, mRNA level corresponding to ITGA3 and ITGA7 significantly increased in eiPSC-511 in comparison to those in eiPSC-111. These results suggested that the binding strength to the substrate in eiPSC-511 was stronger than that in eiPSC-111. On the contrary, although no significant differences were observed in the histology of teratomas, increased in vitro differentiation into three germ layers in eiPSC-111 was shown compared to those in eiPSC-511. Thus, these results contributed to the improved generation of iPSC in horses.