Fig. 2

The properties of senescence in lean hASCs and obese hASCs were determined. a Graph showing the cell growth curve of hASCs from lean donor and obese donor monitored and recorded by Realtime xCELLigence analysis. Histogram showing the significant difference of the cell index of hASCs from two groups. n = 3 different donors. Error bars represent SD. b Representative image of hASCs in the two groups immunostained with antibody against Ki-67. Histogram showing the difference of the positive cell percentage of Ki-67 from lean hASCs and obese hASCs. The number of fields was 22–24 in each group, respectively. n = 3 different donors. Data are presented as the means ± SEM. Scale bars, 50 μm. c Relative mRNA levels of cell cycle inhibitor genes (CDKN1A, CDKN2A, CDKN2B, and TP53) in lean hASCs and obese hASCs were determined by quantitative RT-PCR. The relative expression of each gene was normalized against 18S rRNA. Data are presented as the means ± SEM. n = 3 different donors. d Representative image of hASCs in the two groups for SA-β-Gal staining. Histogram showing the difference of the positive area percentage of SA-β-Gal from lean hASCs and obese hASCs. The number of fields was 15–19 in each group, respectively. n = 3 different donors. Data are presented as the means ± SEM. Scale bars, 50 μm. e Representative images of hASCs from lean and obese donors for immunostained with antibody against γ-H2AX. Histogram showing the difference of the relative intensity of γ-H2AX normalized to DAPI each nuclear of cells. Fold represents the relative intensity was normalized against lean hASCs from each independent experiment. The number of cells for the expression of γ-H2AX was 204–309 in each group, respectively. n = 3 different donors. Data are presented as the means ± SEM. Scale bars, 25 μm. f Relative mRNA levels of SASP-related genes (IL1R1, IGFBP3, TGFB1, and TGFBR2), and GLB1 in lean hASCs and obese hASCs were determined by quantitative RT-PCR. The relative expression of each gene was normalized against 18S rRNA. Data are presented as the means ± SEM. n = 3 different donors. Significant difference by unpaired two-tailed paired t test