Rapamycin Treatment Is Beneficial for the Generation of Rabbit-Induced Pluripotent Stem-Like Cells

Autophagy could promote the generation of induced pluripotency stem cells (iPSCs) in humans and mice. However, little was known whether it had similar effects in other species, the detailed mechanism and the features of formed iPSC colonies were also not clear. In this study, we first established the doxycycline (DOX)-inducible tetO lentiviral vector system suitable for the generation of rabbit iPSCs. Rapamycin, a mechanistic target of rapamycin (mTOR) inhibitor, was added during rabbit embryonic fibroblasts induction to improve the autophagy level. The colony formation efficiency and the expression of autophagy- and pluripotent-related genes were detected. The results showed that the established DOX-inducible tetO lentiviral system was successfully used to induce rabbit iPS-like cells. Compared with the untreated group, the number of alkaline phosphatase (AP)-positive colonies was increased 5.5-fold, when 0.5 nM rapamycin was added on days 1-3 after transduction, the colony morphology was improved and the iPS-like cells could be passaged >10 generations. The expression of autophagy-related genes (ATG), ATG5, ATG7, LC3, and ULK1 was increased with different patterns during the induction process, expression of OCT4, SOX2, and KLF4 significantly increased (p < 0.05). The mentioned results indicate that rapamycin treatment is beneficial for the generation of rabbit iPSCs by regulating autophagy and pluripotency-related gene expression.