Sodium–glucose cotransporter 2 inhibitors (SGLT2i) have favorable cardiovascular outcomes in patients with diabetes. However, whether SGLT2i can improve obesity-related cardiac dysfunction is unknown. Sestrin2 is a novel stress-inducible protein that regulates AMPK–mammalian target of rapamycin (mTOR) and suppresses oxidative damage. The aim of this study was to determine whether empagliflozin (EMPA) improves obesity-related cardiac dysfunction via regulating Sestrin2-mediated pathways in diet-induced obesity. C57BL/6J mice and Sestrin2 knockout mice were fed a high-fat diet (HFD) for 12 weeks and then treated with or without EMPA (10 mg/kg) for 8 weeks. Treating HFD-fed C57BL/6J mice with EMPA reduced body weight and whole-body fat and improved metabolic disorders. Furthermore, EMPA improved myocardial hypertrophy/fibrosis and cardiac function and reduced cardiac fat accumulation and mitochondrial injury. Additionally, EMPA significantly augmented Sestrin2 levels and increased AMPK and endothelial nitric oxide synthase phosphorylation, but inhibited Akt and mTOR phosphorylation. These beneficial effects were partially attenuated in HFD-fed Sestrin2 knockout mice. Intriguingly, EMPA treatment enhanced the Nrf2/HO-1–mediated oxidative stress response, suggesting antioxidant and anti-inflammatory activity. Thus, EMPA improved obesity-related cardiac dysfunction via regulating Sestrin2-mediated AMPK-mTOR signaling and maintaining redox homeostasis. These findings provide a novel mechanism for the cardiovascular protection of SGLT2i in obesity.

Background— Elderly patients are more sensitive than younger patients to myocardial ischemia, which results in higher mortality. We investigated how aging affects the cardioprotective AMP-activated protein kinase (AMPK) signaling pathway. Methods and Results— Ischemic AMPK activation was impaired in aged compared with young murine hearts. The expression and secretion of the AMPK upstream regulator, macrophage migration inhibitory factor (MIF), were lower in aged compared with young adult hearts. Additionally, the levels of hypoxia-inducible factor 1α, a known transcriptional activator of MIF, were reduced in aged compared with young hearts. Ischemia-induced AMPK activation in MIF knockout mice was blunted, leading to greater contractile dysfunction in MIF-deficient than in wild-type hearts. Furthermore, intramyocardial injection of adenovirus encoding MIF in aged mice increased MIF expression and ischemic AMPK activation and reduced infarct size. Conclusions— An impaired MIF-AMPK activation response in senescence thus may be attributed to an aging-associated defect in hypoxia-inducible factor 1α, the transcription factor for MIF. In the clinical setting, impaired cardiac hypoxia-inducible factor 1α activation and consequent reduced MIF expression may play an important role in the increased susceptibility to myocardial ischemia observed in older cardiac patients.

The regulation of AMPK in the ischemic heart remains incompletely understood. Recent evidence implicates the role of Sestrin2 in the AMPK signaling pathway, and it is hypothesized that Sestrin2 plays an influential role during myocardial ischemia to promote AMPK activation. Sestrin2 protein was found to be expressed in adult cardiomyocytes and accumulated in the heart during ischemic conditions. Sestrin2 knockout (KO) mice were used to determine the importance of Sestrin2 during ischemia and reperfusion (I/R) injury. When wild‐type (WT) and Sestrin2 KO mice were subjected to in vivo I/R, myocardial infarct size was significantly greater in Sestrin2 KO compared with WT hearts. Similarly, Langendorff perfused hearts indicated exacerbated postischemic contractile function in Sestrin2 KO hearts compared with WT. Ischemic AMPK activation was found to be impaired in the Sestrin2 KO hearts. Immunoprecipitation of Sestrin2 demonstrated an association with AMPK. Moreover, liver kinase B1 (LKB1), a major AMPK upstream kinase, was associated with the Sestrin2‐AMPK complex in a time‐dependent manner during ischemia, whereas this interaction was nearly abolished in Sestrin2 KO hearts. Thus, Sestrin2 plays an important role in cardioprotection against I/R injury, serving as an LKB1‐AMPK scaffold to initiate AMPK activation during ischemic insults.—Morrison, A., Chen, L. Wang, J., Zhang, M., Yang, H., Ma, Y., Budanov, A., Lee, J. H., Karin, M., Li, J. Sestrin2 promotes LKB1‐mediated AMPK activation in the ischemic heart. FASEB J . 29, 408‐417 (2015). www.fasebj.org

Nucleotide-binding oligomerization domain-Like Receptor with a Pyrin domain 3 (NLRP3) inflammasome was emerged as a marker of metabolic dysregulation. We revealed that age-related Sirtuin-1 (SIRT1) modulates cardiac metabolism that medicated inflammatory response during ischemia and reperfusion (I/R) stress. We hypothesize that SIRT1 attenuates NLRP3 inflammasome-dependent inflammation and pyroptosis during myocardial I/R through metabolic modulation. C57BL/6J wild type (WT) mice, inducible cardiomyocyte specific SIRT1 knockout (icSIRT1 KO) and inducible cardiomyocyte specific PDH E1α knockout (icPDH E1α KO) mice were subjected to ligation and release of left anterior descending coronary artery for in vivo regional I/R models. The echocardiography measurement demonstrated that SIRT1 agonist SRT1720 (30 μg/g) improved cardiac systolic function during 45 min of ischemia and 6 h of reperfusion in C57BL/6J WT mice. The biochemical analysis showed that I/R triggered activation of cardiac pyruvate dehydrogenase (PDH), while SIRT1 agonist SRT1720 inhibited I/R-induced PDH activity and reduced production of reactive oxygen species (ROS) during myocardial I/R. Moreover, SRT1720 regulates PDH-related glucose oxidative metabolism to reduce NLRP3 inflammasome activation and pyroptosis in an Akt signaling dependent manner during I/R. Furthermore, an impaired Akt signaling was observed in icSIRT1 KO versus SIRT1fox/flox mice under I/R stress. Intriguingly, we observed lower levels of ROS generation, decreased NLRP3 levels and less pyroptosis occurred in the icPDH E1α KO versus PDH E1αflox/flox hearts during I/R. Taken together, the results indicate that SIRT1 agonism can inhibit activation of NLRP3 inflammasome via Akt-dependent metabolic regulation during ischemic insults by I/R.

A longevity gene, Sirtuin 1 (SIRT1) and energy sensor AMP-activated protein kinase (AMPK) have common activators such as caloric restriction, oxidative stress, and exercise. The objective of this study is to characterize the role of cardiomyocyte SIRT1 in age-related impaired ischemic AMPK activation and increased susceptibility to ischemic insults. Mice were subjected to ligation of left anterior descending coronary artery for in vivo ischemic models. The glucose and fatty acid oxidation were measured in a working heart perfusion system. The cardiac functions by echocardiography show no difference in young wild-type C57BL/6 J (WT, 4–6 months), aged WT C57BL/6 J (24–26 months), and young inducible cardiomyocyte-specific SIRT1 knockout (icSIRT1 KO) (4–6 months) mice under physiological conditions. However, after 45 mins ischaemia and 24-h reperfusion, the ejection fraction of aged WT and icSIRT1 KO mice was impaired. The aged WT and icSIRT1 KO hearts vs. young WT hearts also show an impaired post-ischemic contractile function in a Langendorff perfusion system. The infarct size of aged WT and icSIRT1 KO hearts was larger than that of young WT hearts. The immunoblotting data demonstrated that aged WT and icSIRT1 KO hearts vs. young WT hearts had impaired phosphorylation of AMPK and downstream acetyl-CoA carboxylase during ischaemia. Intriguingly, AMPK upstream LKB1 is hyper-acetylated in both aged WT and icSIRT1 KO hearts; this could blunt activation of LKB1, leading to an impaired AMPK activation. The working heart perfusion results demonstrated that SIRT1 deficiency significantly impaired substrate metabolism in the hearts; fatty acid oxidation is augmented and glucose oxidation is blunted during ischaemia and reperfusion. Adeno-associated virus (AAV9)-Sirt1 was delivered into the aged hearts via a coronary delivery approach, which significantly rescued the protein level of SIRT1 and the ischemic tolerance of aged hearts. Furthermore, AMPK agonist can rescue the tolerance of aged heart and icSIRT1 KO heart to ischemic insults. Cardiac SIRT1 mediates AMPK activation via LKB1 deacetylation, and AMPK modulates SIRT1 activity via regulation of NAD⁺ level during ischaemia. SIRT1 and AMPK agonists have therapeutic potential for treatment of aging-related ischemic heart disease.