Figure 4. Autophagy blockage abolishes the protective effects of HMGA2 overexpression against H2O2-induced loss of potential regenerative power and pluripotency. hTSCs transfected with lenti-HMGA2 (HMGA2 OE) or the empty virus (nc) were incubated with 3-MA and rapamycin for 24 h, alone or in combination as indicated, and subsequently exposed to 0.5 mM H2O2 for 24 h. Untransfected cells (mock) were included for comparison. (A) Clonogenicity was assessed by the colony formation assay. (B) The quantitative expression of stemness markers Nanog, Nucleostemin, Oct-4, and SSEA-4 were measured by qRT-PCR. (C–E) The multilineage differentiation capability and the expression of markers for lineage-specific differentiation (PPARγ for adipogenesis, Runx-2 for osteogenesis and Sox-9 for chondrogenesis) was evaluated by qRT-PCR. Scale bar = 50 μm. The data shown are from three replicates and are indicated as mean ± SD. *p < 0.05, **p < 0.01.