Autophagy induces G0/G1 arrest and apoptosis in menstrual blood-derived endometrial stem cells via GSK3-/-catenin pathway

Background/aimsMenstrual blood-derived endometrial stem cells (MenSCs) emerge as an ideal source for cell-based treatment in regenerative medicine and immunotherapy. However, the major obstacle is the low survival rate in tissues and the limited expansion number. Autophagy is an intracellular metabolic self-degradative process which plays important roles in normal cellular division and survival, and the present study aimed to explore the related mechanisms between autophagy and survival of MenSCs in vitro and in vivo.MethodsThe MenSCs were obtained from menstrual blood procured from healthy female donors. In vitro, MenSCs were exposed to rapamycin and Earle's balanced salts solution (EBSS). We evaluated the MenSCs immunophenotypic cell cycle distribution by propidium iodide (PI) staining and cell apoptosis by Annexin V/PI staining as well as their proliferative potential by the MTT assay. We also assessed the expression of genes associated with the cell cycle and Gsk3 signaling pathway by western blot analysis. We depressed Atg5 and Gsk3 expression by short hairpin RNA (shRNA) and undertook the experiments. Moreover, the labeled MenSCs were observed and counted with DiI after transplantation into the mice via the tail vein by microscopy in vivo.ResultsIn vitro, rapamycin and starvation induced autophagy of MenSCs. Hyperactive autophagy significantly induced G0/G1 arrest and slightly promoted apoptosis of MenSCs. Meanwhile, autophagy could stimulate p-GSK3 expression in MenSCs. Further, knockdown GSK3 can accelerate the proliferation of MenSCs by shRNA and CHIR99021. Moreover, the shGSK3 MenSCs showed strong proliferative activity in vitro and in vivo.ConclusionsOur results indicate that autophagy induced G0/G1 arrest and apoptosis of MenSCs via GSK3/-catenin pathway. Inhibiting autophagy or reduced GSK3 levels may improve survival rate in vivo, thus playing roles in MenSCs therapy.