N-acetylcysteine attenuates myocardial dysfunction and postischemic injury by restoring caveolin-3/eNOS signaling in diabetic rats

Cardiovasc Diabetol. 2016 Oct 12;15(1):146. doi: 10.1186/s12933-016-0460-z.

Abstract

Background: Patients with diabetes are prone to develop cardiac hypertrophy and more susceptible to myocardial ischemia-reperfusion (I/R) injury, which are concomitant with hyperglycemia-induced oxidative stress and impaired endothelial nitric oxide (NO) synthase (eNOS)/NO signaling. Caveolae are critical in the transduction of eNOS/NO signaling in cardiovascular system. Caveolin (Cav)-3, the cardiomyocytes-specific caveolae structural protein, is decreased in the diabetic heart in which production of reactive oxygen species are increased. We hypothesized that treatment with antioxidant N-acetylcysteine (NAC) could enhance cardiac Cav-3 expression and attenuate caveolae dysfunction and the accompanying eNOS/NO signaling abnormalities in diabetes.

Methods: Control or streptozotocin-induced diabetic rats were either untreated or treated with NAC (1.5 g/kg/day, NAC) by oral gavage for 4 weeks. Rats in subgroup were randomly assigned to receive 30 min of left anterior descending artery ligation followed by 2 h of reperfusion. Isolated rat cardiomyocytes or H9C2 cells were exposed to low glucose (LG, 5.5 mmol/L) or high glucose (HG, 25 mmol/L) for 36 h before being subjected to 4 h of hypoxia followed by 4 h of reoxygenation (H/R).

Results: NAC treatment ameliorated myocardial dysfunction and cardiac hypertrophy, and attenuated myocardial I/R injury and post-ischemic cardiac dysfunction in diabetic rats. NAC attenuated the reductions of NO, Cav-3 and phosphorylated eNOS and mitigated the augmentation of O2-, nitrotyrosine and 15-F2t-isoprostane in diabetic myocardium. Immunofluorescence analysis demonstrated the colocalization of Cav-3 and eNOS in isolated cardiomyocytes. Immunoprecipitation analysis revealed that diabetic conditions decreased the association of Cav-3 and eNOS in isolated cardiomyocytes, which was enhanced by treatment with NAC. Disruption of caveolae by methyl-β-cyclodextrin or Cav-3 siRNA transfection reduced eNOS phosphorylation. NAC treatment attenuated the reductions of Cav-3 expression and eNOS phosphorylation in HG-treated cardiomyocytes or H9C2 cells. NAC treatment attenuated HG and H/R induced cell injury, which was abolished during concomitant treatment with Cav-3 siRNA or eNOS siRNA.

Conclusions: Hyperglycemia-induced inhibition of eNOS activity might be consequences of caveolae dysfunction and reduced Cav-3 expression. Antioxidant NAC attenuated myocardial dysfunction and myocardial I/R injury by improving Cav-3/eNOS signaling.

Keywords: Caveolin-3; Diabetes; Diabetic cardiomyopathy; Myocardial ischemia–reperfusion injury; N-acetylcysteine.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acetylcysteine / pharmacology*
  • Animals
  • Antioxidants / pharmacology*
  • Cardiomegaly / chemically induced
  • Cardiomegaly / enzymology
  • Cardiomegaly / physiopathology
  • Cardiomegaly / prevention & control*
  • Caveolae / drug effects
  • Caveolae / enzymology
  • Caveolae / pathology
  • Caveolin 3 / genetics
  • Caveolin 3 / metabolism*
  • Cell Hypoxia
  • Cell Line
  • Cytoprotection
  • Diabetes Mellitus, Experimental / chemically induced
  • Diabetes Mellitus, Experimental / drug therapy*
  • Diabetic Cardiomyopathies / chemically induced
  • Diabetic Cardiomyopathies / enzymology
  • Diabetic Cardiomyopathies / physiopathology
  • Diabetic Cardiomyopathies / prevention & control*
  • Heart Rate / drug effects
  • Male
  • Myocardial Reperfusion Injury / chemically induced
  • Myocardial Reperfusion Injury / enzymology
  • Myocardial Reperfusion Injury / physiopathology
  • Myocardial Reperfusion Injury / prevention & control*
  • Myocytes, Cardiac / drug effects*
  • Myocytes, Cardiac / enzymology
  • Myocytes, Cardiac / metabolism
  • Myocytes, Cardiac / pathology
  • Nitric Oxide / metabolism
  • Nitric Oxide Synthase Type III / genetics
  • Nitric Oxide Synthase Type III / metabolism*
  • Oxidative Stress / drug effects
  • Phosphorylation
  • RNA Interference
  • Rats, Sprague-Dawley
  • Signal Transduction / drug effects*
  • Streptozocin
  • Transfection
  • Ventricular Function, Left / drug effects

Substances

  • Antioxidants
  • Cav3 protein, rat
  • Caveolin 3
  • Nitric Oxide
  • Streptozocin
  • Nitric Oxide Synthase Type III
  • Nos3 protein, rat
  • Acetylcysteine