The Role Of P62 On Chronic Oxidative Stress‐Induced Myocytes Ca handling

Abstract

IntroductionRecent studies have shown that the activity of autophagy impairs, and the accumulation of p62 protein, autophagy substrate, in the heart observes in heart failure patients and mouse models of cardiomyopathy. Accumulations of p62 protein are known to activate the intrinsic cellular stress. Abnormal Ca release is partly responsible for oxidative stress‐induced contractile dysfunction of myocytes in ischemic heart disease. Myocytes Ca release is controlled by a function of sarcoplasmic reticulum (SR) Ca store. However, the role of p62 protein on myocytes Ca regulation in the ischemic heart is not clear.PurposeTo investigate the role of p62 on SR Ca regulation in myocytes isolated from mice with the Isoproterenol‐induced myocardial injury.MethodsTo test our hypothesis, both wild‐type (WT) and p62 knockout (KO) mice were used at 10–12 weeks of age. Myocytes isolated from WT and p62 KO mice (n=3 per group) injected with 200 mg/kg Isoproterenol (ISO; beta agonist, twice per day) for 0 (Base), 5 days, and 10 days. SR Ca handling and contractility measured in field‐stimulated single myocytes loaded with fluorescent Ca indicators (Fura‐2AM; cytosolic Ca loading). Amplitudes of caffeine (10 mM, 5 sec)‐induced Ca transients used as estimates of SR Ca store. Sets of heart from WT and p62 mice were processed for western blot (n=3 per group).ResultsThe p62‐deficient myocytes had no difference of myocytes contractility, and SR Ca handling at the baseline compared to WT myocytes (n=35–45 cells per group). After acute ISO exposure (1μM for 1 min), WT myocytes increased systolic Ca release whereas p62‐deficient myocytes did not. An exposure of a submaximal oxidative stress (1μM; H2O2) in p62‐deficient myocytes, however, increased myocytes contractility and the fraction of Ca released from the SR during each beat (fractional Ca release; systolic Ca release over SR Ca store) after acute ISO stimulation (p<0.05). In WT mice injected with ISO for 5 days and 10 days, myocytes had a biphasic effect on myocytes contractility and fractional Ca release (n=15–45 cells per group, p<0.05). Interestingly, in p62‐deficient myocytes further increased myocytes contractility and fractional Ca release (Figure) on chronic ISO injections from 5 days to 10 days. WT mice hearts with chronic exposure to ISO for 10 days increased p62 protein expressions and suppressed the conversion of LC3 I to LC3 II.ConclusionTaken together, our results suggest that accumulations of p62 protein may directly cause SR Ca defects via beta‐adrenergic signaling in myocytes under chronic myocardial injury.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.