Rat Induced Pluripotent Stem Cells Protect H9C2 Cells from Cellular Senescence via a Paracrine Mechanism

Abstract

Objectives: Cellular senescence may play an important role in the pathology of heart aging. We aimed to explore whether induced pluripotent stem cells (iPSCs) could inhibit cardiac cellular senescence via a paracrine mechanism. Methods: We collected iPSC culture supernatant, with or without oxidative stress, as conditioned medium (CM) for the rat cardiomyocyte-derived cell line H9C2. Then we treated H9C2 cells, cultured with or without CM, with hypoxia/reoxygenation to induce cellular senescence and measured senescence-associated β-galactosidase (SA-β-gal) activity, G1 cell proportion and expression of the cell cycle regulators p16^INK4a, p21^Waf1/Cip1 and p53 at mRNA and protein levels in H9C2 cells. In addition, we used Luminex-based analysis to measure concentrations of trophic factors in iPSC-derived CM. Results: We found that iPSC-derived CM reduced SA-β-gal activity, attenuated G1 cell cycle arrest and reduced the expression of p16^INK4a, p21^Waf1/Cip1 and p53 in H9C2 cells. Furthermore, the CM contained more trophic factors, e.g. tissue inhibitor of metalloproteinase-1 and vascular endothelial growth factor, than H9C2-derived CM. Conclusions: Paracrine factors released from iPSCs prevent stress-induced senescence of H9C2 cells by inhibiting p53-p21 and p16-pRb pathways. This is the first report demonstrating that antisenescence effects of stem cell therapy may be a novel therapeutic strategy for age-related cardiovascular disease.