Redox-Resistant SERCA [Sarco(endo)plasmic Reticulum Calcium ATPase] Attenuates Oxidant-Stimulated Mitochondrial Calcium and Apoptosis in Cardiac Myocytes and Pressure Overload-Induced Myocardial Failure in Mice.

BACKGROUND: SERCA [sarco(endo)plasmic reticulum calcium ATPase] is regulated by oxidative posttranslational modifications at cysteine 674 (C674). Because sarcoplasmic reticulum (SR) calcium has been shown to play a critical role in mediating mitochondrial dysfunction in response to reactive oxygen species, we hypothesized that SERCA oxidation at C674 would modulate the effects of reactive oxygen species on mitochondrial calcium and mitochondria-dependent apoptosis in cardiac myocytes. METHODS: Adult rat ventricular myocytes expressing wild-type SERCA2b or a redox-insensitive mutant in which C674 is replaced by serine (C674S) were exposed to H2O2 (100 µmol/Lµ). Free mitochondrial calcium concentration was measured in adult rat ventricular myocytes with a genetically targeted fluorescent probe, and SR calcium content was assessed by measuring caffeine-stimulated release. Mice with heterozygous knock-in of the SERCA C674S mutation were subjected to chronic ascending aortic constriction. RESULTS: In adult rat ventricular myocytes expressing wild-type SERCA, H2O2 caused a 25% increase in mitochondrial calcium concentration that was associated with a 50% decrease in SR calcium content, both of which were prevented by the ryanodine receptor inhibitor tetracaine. In cells expressing the C674S mutant, basal SR calcium content was decreased by 31% and the H2O2-stimulated rise in mitochondrial calcium concentration was attenuated by 40%. In wild-type cells, H2O2 caused cytochrome c release and apoptosis, both of which were prevented in C674S-expressing cells. In myocytes from SERCA knock-in mice, basal SERCA activity and SR calcium content were decreased. To test the effect of C674 oxidation on apoptosis in vivo, SERCA knock-in mice were subjected to chronic ascending aortic constriction. In wild-type mice, ascending aortic constriction caused myocyte apoptosis, LV dilation, and systolic failure, all of which were inhibited in SERCA knock-in mice. CONCLUSIONS: Redox activation of SERCA C674 regulates basal SR calcium content, thereby mediating the pathologic reactive oxygen species-stimulated rise in mitochondrial calcium required for myocyte apoptosis and myocardial failure.

SGLT2 inhibitor ertugliflozin decreases elevated intracellular sodium, and improves energetics and contractile function in diabetic cardiomyopathy.

BACKGROUND: Elevated myocardial intracellular sodium ([Na+]i) was shown to decrease mitochondrial calcium ([Ca2+]MITO) via mitochondrial sodium/calcium exchanger (NCXMITO), resulting in decreased mitochondrial ATP synthesis. The sodium-glucose co-transporter 2 inhibitor (SGLT2i) ertugliflozin (ERTU) improved energetic deficit and contractile dysfunction in a mouse model of high fat, high sucrose (HFHS) diet-induced diabetic cardiomyopathy (DCMP). As SGLT2is were shown to lower [Na+]i in isolated cardiomyocytes, we hypothesized that energetic improvement in DCMP is at least partially mediated by a decrease in abnormally elevated myocardial [Na+]i. METHODS: Forty-two eight-week-old male C57BL/6J mice were fed a control or HFHS diet for six months. In the last month, a subgroup of HFHS-fed mice was treated with ERTU. At the end of the study, left ventricular contractile function and energetics were measured simultaneously in isolated beating hearts by 31P NMR (Nuclear Magnetic Resonance) spectroscopy. A subset of untreated HFHS hearts was perfused with vehicle vs. CGP 37157, an NCXMITO inhibitor. Myocardial [Na+]i was measured by 23Na NMR spectroscopy. RESULTS: HFHS hearts showed diastolic dysfunction, decreased contractile reserve, and impaired energetics as reflected by decreased phosphocreatine (PCr) and PCr/ATP ratio. Myocardial [Na+]i was elevated > 2-fold in HFHS (vs. control diet). ERTU reversed the impairments in HFHS hearts to levels similar to or better than control diet and decreased myocardial [Na+]i to control levels. CGP 37157 normalized the PCr/ATP ratio in HFHS hearts. CONCLUSIONS: Elevated myocardial [Na+]i contributes to mitochondrial and contractile dysfunction in DCMP. Targeting myocardial [Na+]i and/or NCXMITO may be an effective strategy in DCMP and other forms of heart disease associated with elevated myocardial [Na+]i.

Effects of Sodium-Glucose Linked Transporter 2 Inhibition With Ertugliflozin on Mitochondrial Function, Energetics, and Metabolic Gene Expression in the Presence and Absence of Diabetes Mellitus in Mice.

Background Inhibitors of the sodium-glucose linked transporter 2 improve cardiovascular outcomes in patients with or without type 2 diabetes mellitus, but the effects on cardiac energetics and mitochondrial function are unknown. We assessed the effects of sodium-glucose linked transporter 2 inhibition on mitochondrial function, high-energy phosphates, and genes encoding mitochondrial proteins in hearts of mice with and without diet-induced diabetic cardiomyopathy. Methods and Results Mice fed a control diet or a high-fat, high-sucrose diet received ertugliflozin mixed with the diet (0.5 mg/g of diet) for 4 months. Isolated mitochondria were assessed for functional capacity. High-energy phosphates were assessed by 31P nuclear magnetic resonance spectroscopy concurrently with contractile performance in isolated beating hearts. The high-fat, high-sucrose diet caused myocardial hypertrophy, diastolic dysfunction, mitochondrial dysfunction, and impaired energetic response, all of which were prevented by ertugliflozin. With both diets, ertugliflozin caused supernormalization of contractile reserve, as measured by rate×pressure product at high work demand. Likewise, the myocardial gene sets most enriched by ertugliflozin were for oxidative phosphorylation and fatty acid metabolism, both of which were enriched independent of diet. Conclusions Ertugliflozin not only prevented high-fat, high-sucrose-induced pathological cardiac remodeling, but improved contractile reserve and induced the expression of oxidative phosphorylation and fatty acid metabolism gene sets independent of diabetic status. These effects of sodium-glucose linked transporter 2 inhibition on cardiac energetics and metabolism may contribute to improved structure and function in cardiac diseases associated with mitochondrial dysfunction, such as heart failure.

Differential Effects of Sacubitril/Valsartan on Diastolic Function in Mice With Obesity-Related Metabolic Heart Disease.

Mice with obesity and metabolic heart disease (MHD) due to a high-fat, high-sucrose diet were treated with placebo, a clinically relevant dose of sacubitril (SAC)/valsartan (VAL), or an equivalent dose of VAL for 4 months. There were striking differences between SAC/VAL and VAL with regard to: 1) diastolic dysfunction; 2) interstitial fibrosis; and to a lesser degree; 3) oxidative stress-all of which were more favorably affected by SAC/VAL. SAC/VAL and VAL similarly attenuated myocardial hypertrophy and improved myocardial energetics. In mice with obesity-related MHD, neprilysin inhibition exerts favorable effects on diastolic function.