Fig. 1

Culture of vASCs at 35 °C attenuates NOX1-dependent oxidative stress and apoptosis, enabling long-term expansion. vASCs were propagated at 35 °C or 37 °C. A ROS accumulation was detected by DCFDA staining and analyzed by flow cytometry at the indicated passages (P). (AI) Representative FACS analyses. (AII) Summary results of three independent repeats at P2–4. Error bars represent SD. (AIII) qRT-PCR analysis of NOX1 RNA levels in cells grown at 35 °C (black line) versus 37 °C (gray line) at the indicated passages (n = two independent repeats). B The percentage of apoptotic cells at each passage of cells grown at 37 °C and 35 °C was determined by PI/Annexin staining, followed by FACS analysis. (BI) Representative FACS analyses. (BII) Images of representative 37 °C and 35 °C vASCs at P1 and P5. (BIII) Mean of FACs analyses by-passage rate of apoptosis in 37 °C and 35 °C vASC cultures (n = two independent repeats). C A growth curve comparing the ability of visceral ASCs to undergo long-term expansion at 37 °C versus 35 °C as evident by the number of passages reached against the time in culture in days (n = four independent repeats). Cells were passaged only when reaching confluence. Error bars represent standard deviation (SD). D The doubling time of 37OC and 35OC vASC was compared (n=two independent repeats). The doubling time of 37 °C and 35 °C vASC was compared (n = two independent repeats). Student’s two-tailed t test for equal variance: **p < 0.01; *p < 0.05