Abstract
Autologous human cardiac stem/progenitor cell (hCPC) therapy is a promising treatment that has come into use in recent years for patients with cardiomyopathy. Though innovative in theory, a major hindrance to the practical application of this treatment is that the hCPCs of elderly patients, who are most susceptible to myocardial disease, are senescent and prone to cell death. Rejuvenating hCPCs from elderly patients may help overcome this obstacle, and can be accomplished by reversing entry into the cellular stage of senescence. p16(INK4A), a cyclin dependent kinase inhibitor, is an important player in the regulation of cell senescence. In this study, we investigated whether knockdown of p16(INK4A) will rejuvenate aging hCPCs to a youthful phenotype. Our data indicated that upregulation of p16(INK4A) is associated with hCPC senescence. Both cell proliferation and survival capacity were significantly increased in hCPCs infected with lentivirus expressing p16(INK4A) shRNA when compared to control hCPCs. The knockdown of p16(INK4A) also induced antioxidant properties as indicated by a 50% decrease in ROS generation at basal cell metabolism, and a 25% decrease in ROS generation after exposure to oxidative stress. Genes associated with cell senescence (p21(CIP1)), anti-apoptosis (BCL2 and MCL1), anti-oxidant (CYGB, PRDX1 and SRXN1), and NFκB signal pathway (p65, IKBKB, HMOX1, etc.), were significantly upregulated after the p16(INK4A) knockdown. Knocking down the NFĸB-p65 expression also significantly diminished the cytoprotective effect caused by the p16(INK4A) knockdown. Our results suggest that genetic knockdown of p16(INK4A) may play a significant role in inducing antioxidant effects and extending lifespan of aging hCPCs. This genetic modification may enhance the effectiveness of autologous hCPC therapy for repair of infarcted myocardium.