Hydrogen sulfide attenuates high fat diet-induced cardiac dysfunction via the suppression of endoplasmic reticulum stress

Abstract

Diabetic cardiomyopathy is a significant contributor to the morbidity and mortality associated with diabetes and metabolic syndrome. However, the underlying molecular mechanisms that lead to its development have not been fully elucidated. Hydrogen sulfide (H2S) is an endogenously produced signaling molecule that is critical for the regulation of cardiovascular homeostasis. Recently, therapeutic strategies aimed at increasing its levels have proven cardioprotective in models of acute myocardial ischemia-reperfusion injury and heart failure. The precise role of H2S in the pathogenesis of diabetic cardiomyopathy has not yet been established. Therefore, the goal of the present study was to evaluate circulating and cardiac H2S levels in a murine model of high fat diet (HFD)-induced cardiomyopathy. Diabetic cardiomyopathy was produced by feeding mice HFD (60% fat) chow for 24 weeks. HFD feeding reduced both circulating and cardiac H2S and induced hallmark features of type-2 diabetes. We also observed marked cardiac dysfunction, evidence of cardiac enlargement, cardiac hypertrophy, and fibrosis. H2S therapy (SG-1002, an orally active H2S donor) restored sulfide levels, improved some of the metabolic perturbations stemming from HFD feeding, and attenuated HFD-induced cardiac dysfunction. Additional analysis revealed that H2S therapy restored adiponectin levels and suppressed cardiac ER stress stemming from HFD feeding. These results suggest that diminished circulating and cardiac H2S levels play a role in the pathophysiology of HFD-induced cardiomyopathy. Additionally, these results suggest that H2S therapy may be of clinical importance in the treatment of cardiovascular complications stemming from diabetes.

Keywords: Diabetes; Diabetic cardiomyopathy; ER stress; Hydrogen sulfide.

Fig. 1

Representative immunoblots and densitometric analysis of cystathionine beta synthase (CBS), cystathionine gamma lyase (CSE), and 3-mercaptopyruvate sulfutransferase (3-MST) (A-B). Circulating (C-D) and cardiac (E-F) levels of free hydrogen sulfide (H₂S) and sulfane sulfure. All samples (blood and hearts) were collected from control, high-fat diet (HFD), and HFD-fed mice supplemented with SG-1002 (20 mg/kg/day). One group of mice received the SG-1002 supplemented HFD chow for the entire 24 weeks (HFD-S). We delayed the treatment in another group mice (HFD-D). These mice received the HFD chow for 12 weeks before being switched to the SG-1002 supplemented HFD chow for the final 12 weeks of the study. Results are expressed as mean ± SEM. Numbers in bars represent the sample size. *p<0.05, **p<0.01 and ***p<0.001 vs. Control.

Fig 2

Left Ventricular End Systolic Diameter (LVESD) (A), LV Ejection Fraction (B), LV Fractional Shortening (C) relaxation constant Tau (D), Max dP/dT and Min dP/dT, (E) and Circumferential Stress (F) were evaluated in control, HFD, HFD-S, and HFD-D mice following 24 weeks of HFD feeding. Results are expressed as mean ± SEM.In panels A-C, ***p<0.001 vs. Baseline. In panels D-F, *p<0.05, **p<0.01 and ***p<0.001 vs. Control.

Fig 3

Heart weight to body weight ratios (A), Representative photomicrographs of wheat germ agglutinin stained hearts (B), Summary of myocyte cell surface area (C) and cross sectional area measurements (D) of wheat germ agglutinin stained hearts, Representative photomicrographs of Picrosirius Red stained hearts (E), and summary of fibrosis area as % of the LV calculated from the Picrosirius Red sections (F), in control, HFD, HFD-S, and HFD-D mice following 24 weeks of HFD feeding. Scale bar equals 100 μm in panels B and D, respectively. Results are expressed as mean ± SEM. *p<0.05, **p<0.01 and ***p<0.001 vs. Control.

Fig. 4

Circulating adiponectin levels (A), representative immunoblots (B) and densitometric analysis of total AMP-activated protein kinase (t-AMPK) and the ratio of phosphorylated AMPK (p-AMPK) to t-AMPK (C), glucose transporter 4 (GLUT4) (D), total acetyl-coenzyme A carboxylase (t-ACC) and the ratio of phosphorylated ACC (p-ACC) to t-ACC (E), and cardiac triglyceride levels (F) in control, HFD, HFD-S, and HFD-D mice following 24 weeks of HFD feeding. Results are expressed as mean ± SEM. *p<0.05, **p<0.01 and ***p<0.001 vs. Control.

Fig. 5

Representative immunoblots (A) and densitometric analysis of inositol-requiring enzyme 1 (IRE1) (B), activating transcription factor 6 (ATF6) (C), and ATF4 (D). All samples were collected from the hearts of control, HFD, HFD-S, and HFD-D mice following 24 weeks of HFD feeding. Results are expressed as mean ± SEM. *p<0.05, **p<0.01 and ***p<0.001 vs. Control.

Fig. 6

Representative immunoblots (A) and densitometric analysis of X-box– binding protein-1 (XBP1) (B), total JNK (t-JNK) (C), ratio of phosphorylated JNK (p-JNK) to t-JNK (D), and ratio of p-JNK to total protein load (E). All samples were collected from the hearts of control, HFD, HFD-S, and HFD-D mice following 24 weeks of HFD feeding. Results are expressed as mean ± SEM. *p<0.05, **p<0.01 and ***p<0.001 vs. Control.

Fig. 7

Representative immunoblots (A) and densitometric analysis of glucose-regulated protein 94 (GRP94) (B), GRP78 (C), and cleaved caspase-12 (D). All samples were collected from the hearts of control, HFD, HFD-S, and HFD-D mice following 24 weeks of HFD feeding. Results are expressed as mean ± SEM. *p<0.05 and ***p<0.001 vs. Control.