Regulation of Diabetic Cardiomyopathy by Caloric Restriction is Mediated by Intracellular Signaling Pathways Involving \u27SIRT1 and PGC-1alpha\u27

BACKGROUND: Metabolic disorders such as obesity, insulin resistance and type 2 diabetes mellitus (DM2) are all linked to diabetic cardiomyopathy that lead to heart failure. Cardiomyopathy is initially characterized by cardiomyocyte hypertrophy, followed by mitochondrial dysfunction and fibrosis, both of which are aggravated by angiotensin. Caloric restriction (CR) is cardioprotective in animal models of heart disease through its catabolic activity and activation of the expression of adaptive genes. We hypothesized that in the diabetic heart; this effect involves antioxidant defenses and is mediated by SIRT1 and the transcriptional coactivator PGC-1alpha (Peroxisome proliferator-activated receptor-gamma coactivator). METHODS: Obese Leptin resistant (db/db) mice characterized by DM2 were treated with angiotensin II (AT) for 4 weeks to enhance the development of cardiomyopathy. Mice were concomitantly either on a CR diet or fed ad libitum. Cardiomyocytes were exposed to high levels of glucose and were treated with EX-527 (SIRT1 inhibitor). Cardiac structure and function, gene and protein expression and oxidative stress parameters were analyzed. RESULTS: AT treated db/db mice developed cardiomyopathy manifested by elevated levels of serum glucose, cholesterol and cardiac hypertrophy. Leukocyte infiltration, fibrosis and an increase in an inflammatory marker (TNFalpha) and natriuretic peptides (ANP, BNP) gene expression were also observed. Oxidative stress was manifested by low SOD and PGC-1alpha levels and an increase in ROS and MDA. DM2 resulted in ERK1/2 activation. CR attenuated all these deleterious perturbations and prevented the development of cardiomyopathy. ERK1/2 phosphorylation was reduced in CR mice (p = 0.008). Concomitantly CR prevented the reduction in SIRT activity and PGC-1alpha (p \u3c 0.04). Inhibition of SIRT1 activity in cardiomyocytes led to a marked reduction in both SIRT1 and PGC-1alpha. ROS levels were significantly (p \u3c 0.03) increased by glucose and SIRT1 inhibition. CONCLUSION: In the current study we present evidence of the cardioprotective effects of CR operating through SIRT1 and PGC-1 alpha, thereby decreasing oxidative stress, fibrosis and inflammation. Our results suggest that increasing SIRT1 and PGC-1alpha levels offer new therapeutic approaches for the protection of the diabetic heart

The Role of Neutrophils in Corneal Wound Healing in HO-2 Null Mice

Our studies demonstrated that Heme oxygenase (HO), in particular, the constitutive HO-2, is critical for a self-resolving inflammatory and repair response in the cornea. Epithelial injury in HO-2 null mice leads to impaired wound closure and chronic inflammation in the cornea. This study was undertaken to examine the possible relationship between HO-2 and the recruitment of neutrophils following a corneal surface injury in wild type (WT) and HO-2 knockout (HO-2−/−) mice treated with Gr-1 monoclonal antibody to deplete peripheral neutrophils. Epithelial injury was performed by removing the entire corneal epithelium. Infiltration of inflammatory cell into the cornea in response to injury was higher in HO-2−/− than in WT. However, the rate of corneal wound closure following neutrophil depletion was markedly inhibited in both WT and HO-2−/− mice by 60% and 85%, respectively. Neutropenia induced HO-1 expression in WT but not in HO-2−/− mice. Moreover, endothelial cells lacking HO-2 expressed higher levels of the Midkine and VE-cadherin and displayed strong adhesion to neutrophils suggesting that perturbation in endothelial cell function caused by HO-2 depletion underlies the increased infiltration of neutrophils into the HO-2−/− cornea. Moreover, the fact that neutropenia worsened epithelial healing of the injured cornea in both WT and HO-2−/− mice suggest that cells other than neutrophils contribute to the exaggerated inflammation and impaired wound healing seen in the HO-2 null cornea

Heme Oxygenase-1 Induction Improves Cardiac Function following Myocardial Ischemia by Reducing Oxidative Stress

Background Oxidative stress plays a key role in exacerbating diabetes and cardiovascular disease. Heme oxygenase-1 (HO-1), a stress response protein, is cytoprotective, but its role in post myocardial infarction (MI) and diabetes is not fully characterized. We aimed to investigate the protection and the mechanisms of HO-1 induction in cardiomyocytes subjected to hypoxia and in diabetic mice subjected to LAD ligation. Methods In vitro: cultured cardiomyocytes were treated with cobalt-protoporphyrin (CoPP) and tin protoporphyrin (SnPP) prior to hypoxic stress. In vivo: CoPP treated streptozotocin-induced diabetic mice were subjected to LAD ligation for 2/24 h. Cardiac function, histology, biochemical damage markers and signaling pathways were measured. Results HO-1 induction lowered release of lactate dehydrogenase (LDH) and creatine phospho kinase (CK), decreased propidium iodide staining, improved cell morphology and preserved mitochondrial membrane potential in cardiomyocytes. In diabetic mice, Fractional Shortening (FS) was lower than non-diabetic mice (35±1%vs.41±2, respectively p\u3c0.05). CoPP-treated diabetic animals improved cardiac function (43±2% p\u3c0.01), reduced CK, Troponin T levels and infarct size compared to non-treated diabetic mice (P\u3c0.01, P\u3c0.001, P\u3c0.01 respectively). CoPP-enhanced HO-1 protein levels and reduced oxidative stress in diabetic animals, as indicated by the decrease in superoxide levels in cardiac tissues and plasma TNFα levels (p\u3c0.05). The increased levels of HO-1 by CoPP treatment after LAD ligation led to a shift of the Bcl-2/bax ratio towards the antiapoptotic process (p\u3c0.05). CoPP significantly increased the expression levels of pAKT and pGSK3β (p\u3c0.05) in cardiomyocytes and in diabetic mice with MI. SnPP abolished CoPP\u27s cardioprotective effects. Conclusions HO-1 induction plays a role in cardioprotection against hypoxic damage in cardiomyocytes and in reducing post ischemic cardiac damage in the diabetic heart as proved by the increased levels of pAKT with a concomitant inhibition of pGSK3β leading to preserved mitochondrial membrane potential

The CYP/20-HETE/GPR75 Axis in Hypertension

20-Hydroxyeicosatetraenoic acid (20-HETE) is a bioactive lipid generated from the ω-hydroxylation of arachidonic acid (AA) by enzymes of the cytochrome P450 (CYP) family, primarily the CYP4A and CYP4F subfamilies. 20-HETE is most notably identified as a modulator of vascular tone, regulator of renal function, and a contributor to the onset and development of hypertension and cardiovascular disease. 20-HETE-mediated signaling promotes hypertension by sensitizing the vasculature to constrictor stimuli, inducing endothelial dysfunction, and potentiating vascular inflammation. These bioactions are driven by the activation of the G-protein coupled receptor 75 (GPR75), a 20-HETE receptor (20HR). Given the capacity of 20-HETE signaling to drive pro-hypertensive mechanisms, the CYP/20-HETE/GPR75 axis has the potential to be a significant therapeutic target for the treatment of hypertension and cardiovascular diseases associated with increases in blood pressure. In this chapter, we review 20-HETE-mediated cellular mechanisms that promote hypertension, highlight important data in humans such as genetic variants in the CYP genes that potentiate 20-HETE production and describe recent findings in humans with 20HR/GPR75 mutations. Special emphasis is given to the 20HR and respective receptor blockers that have the potential to pave a path to translational and clinical studies for the treatment of 20-HETE-driven hypertension, and obesity/metabolic syndrome

Sequencing of 640,000 Exomes Identifies GPR75 Variants Associated with Protection from Obesity

Large-scale human exome sequencing can identify rare protein-coding variants with a large impact on complex traits such as body adiposity. We sequenced the exomes of 645,626 individuals from the United Kingdom, the United States, and Mexico and estimated associations of rare coding variants with body mass index (BMI). We identified 16 genes with an exome-wide significant association with BMI, including those encoding five brain-expressed G protein-coupled receptors (CALCR, MC4R, GIPR, GPR151, and GPR75). Protein-truncating variants in GPR75 were observed in ~4/10,000 sequenced individuals and were associated with 1.8 kilograms per square meter lower BMI and 54% lower odds of obesity in the heterozygous state. Knock out of Gpr75 in mice resulted in resistance to weight gain and improved glycemic control in a high-fat diet model. Inhibition of GPR75 may provide a therapeutic strategy for obesity