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-1α (Peroxisome proliferator-activated receptor-γ coactivator).
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.
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 (TNFα) and natriuretic peptides (ANP, BNP) gene expression were also observed. Oxidative stress was manifested by low SOD and PGC-1α 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-1α (p < 0.04). Inhibition of SIRT1 activity in cardiomyocytes led to a marked reduction in both SIRT1 and PGC-1α. ROS levels were significantly (p < 0.03) increased by glucose and SIRT1 inhibition.
In the current study we present evidence of the cardioprotective effects of CR operating through SIRT1 and PGC-1 α, thereby decreasing oxidative stress, fibrosis and inflammation. Our results suggest that increasing SIRT1 and PGC-1α levels offer new therapeutic approaches for the protection of the diabetic heart.
Arachidonic acid (AA) is metabolized in mammals by enzymes of the CYP4A and 4F families to 20-hydroxyeicosatetraeonic acid (20-HETE) which plays an important role in the regulation of renal function, ...vascular tone and arterial pressure. In the vasculature, 20-HETE is a potent vasoconstrictor, the up-regulation of which contributes to inflammation, oxidative stress, endothelial dysfunction and an increase in peripheral vascular resistance in models of obesity, diabetes, ischemia/reperfusion, and vascular oxidative stress. Recent studies have established a role for 20-HETE in normal and pathological angiogenic conditions. We discuss in this review the synthesis of 20-HETE and how it and various autacoids, especially the renin-angiotensin system, interact to promote hypertension, vasoconstriction, and vascular dysfunction. In addition, we examine the molecular mechanisms through which 20-HETE induces these actions and the clinical implication of inhibiting 20-HETE production and activity.
Sirtuin 6 (SIRT6) is a protein associated with prolonged life expectancy. We investigated whether life extension is associated with cardioprotection against hypoxia. The proposed study is to develop ...approaches to reduce hypoxic damage through the use of the sirtuin pathway and to elucidate the mechanism involved. For that purpose we subjected cardiomyocytes from transgenic mice (TG) with over-expression of SIRT6, to hypoxic stress in cell cultures. We hypothesized that cardiomyocytes from transgenic mice subjected to prolonged hypoxia may release survival factors or fewer damage markers to protect them from hypoxic stress compared with wild type (WT) mice. Lactate dehydrogenase (LDH) and creatine kinase (CK) released to the medium and propidium iodide (PI) binding, were markedly decreased following hypoxia in TG cardiomyocytes. The protective mechanism of SIRT6 over-expression includes the activation of pAMPKα pathway, the increased protein level of B-cell lymphoma 2 (Bcl2), the inhibition of nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB), the decrease of reactive oxygen species (ROS) and the reduction in the protein level of phospho-protein kinase B (pAkt) during hypoxia. Together, all these processes impede the necrosis/apoptosis pathways leading to the improved survival of cardiomyocytes following hypoxia, which might explain life extension.
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•Sirtuin 6 is a protein associated with prolonged life expectancy.•Over-expression of sirtuin 6 protects cardiocytes from hypoxia and oxidative stress.•Over-expression of sirtuin 6 activates the pAMPKα pathway and the Bcl2 expression.•Over-expression of sirtuin 6 decreases ROS formation and pAkt level during hypoxia.•These pathways protect cardiocytes from hypoxia and might explain lifespan extension.
Abstract Toll-like receptors are expressed in immune cells and cardiac muscle. We examined whether the cardiac Toll-like receptor 4 (TLR4) is involved in the acute myocardial dysfunction caused by ...septic shock and myocardial ischemia (MI). We used wild type mice (WT), TLR4 deficient (TLR4-ko) mice and chimeras that underwent myeloablative bone marrow transplantation to dissociate between TLR4 expression in the heart (TLR4-ko/WT) and the immunohematopoietic system (WT/TLR4-ko). Mice were injected with lipopolysaccharide (LPS) (septic shock model) or subjected to coronary artery ligation (MI model) and tested in vivo and ex vivo , for function, histopathology proinflammatory cytokine and TLR4 expression. WT mice challenged with LPS or MI displayed reduced cardiac function, increased myocardial levels of IL-1β and TNF-α and upregulation of mRNA encoding TLR4 prior to myocardial leukocyte infiltration. TLR4 deficient mice sustained significantly smaller infarctions as compared to control mice at comparable areas at risk. The cardiac function of TLR4-ko mice was not affected by LPS and demonstrated reduced suppression by MI compared to WT. Chimeras deficient in myocardial TLR4 were resistant to suppression induced by LPS and the heart function was less depressed, compared to the TLR4-ko, following MI in the acute phase (4 h). In contrast, hearts of chimeras deficient in immunohematopoietic TLR4 expression were suppressed both by LPS and MI, exhibiting increased myocardial cytokine levels, similar to WT mice. We concluded that cardiac function of TLR4-ko mice and chimeric mice expressing TLR4 in the immunohematopoietic system, but not in the heart, revealed resistance to LPS and reduced cardiac depression following MI, suggesting that TLR4 expressed by the cardiomyocytes themselves plays a key role in this acute phenomenon.
Diabetic and obese patients are at higher risk of severe disease and cardiac injury in corona virus 2 (SARS-CoV-2) infections. Cellular entry of SARS-CoV-2 is mainly via the angiotensin-converting ...enzyme 2 (ACE2) receptor, which is highly expressed in normal hearts. There is a disagreement regarding the effect of factors such as obesity and diabetes on ACE2 expression in the human heart and whether treatment with renin-angiotensin system inhibitors or anti-diabetic medications increases ACE2 expression and subsequently the susceptibility to infection. We designed this study to elucidate factors that control ACE2 expression in human serum, human heart biopsies, and mice.
Right atrial appendage biopsies were collected from 79 patients that underwent coronary artery bypass graft (CABG) surgery. We investigated the alteration in ACE2 mRNA and protein expression in heart tissue and serum. ACE2 expression was compared with clinical risk factors: diabetes, obesity and different anti-hypertensive or anti-diabetic therapies. WT or db/db mice were infused with Angiotensin II (ATII), treated with different anti-diabetic drugs (Metformin, GLP1A and SGLT2i) were also tested.
ACE2 gene expression was increased in diabetic hearts compared to non-diabetic hearts and was positively correlated with glycosylated hemoglobin (HbA1c), body mass index (BMI), and activation of the renin angiotensin system (RAS), and negatively correlated with ejection fraction. ACE2 was not differentially expressed in patients who were on angiotensin converting enzyme inhibitors (ACEi) or angiotensin receptor blockers (ARBs) prior to the operation. We found no correlation between plasma free ACE2 and cardiac tissue ACE2 expression. Transmembrane serine protease 2 (TMPRSS2), metalloprotease ADAM10 and ADAM17 that facilitate viral-ACE2 complex entry and degradation were increased in diabetic hearts. ACE2 expression in mice was increased with ATII infusion and attenuated following anti-diabetic drugs treatment.
Patients with uncontrolled diabetes or obesity with RAS activation have higher ACE2 expressions therefore are at higher risk for severe infection. Since ACEi or ARBs show no effect on ACE2 expression in the heart further support their safety.
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.
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.
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<0.05). CoPP-treated diabetic animals improved cardiac function (43±2% p<0.01), reduced CK, Troponin T levels and infarct size compared to non-treated diabetic mice (P<0.01, P<0.001, P<0.01 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<0.05). 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<0.05). CoPP significantly increased the expression levels of pAKT and pGSK3β (p<0.05) in cardiomyocytes and in diabetic mice with MI. SnPP abolished CoPP's cardioprotective effects.
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.
Type 2 diabetes mellitus (DM2) leads to cardiomyopathy characterized by cardiomyocyte hypertrophy, followed by mitochondrial dysfunction and interstitial fibrosis, all of which are exacerbated by ...angiotensin II (AT). SIRT1 and its transcriptional coactivator target PGC-1α (peroxisome proliferator-activated receptor-γ coactivator), and heme oxygenase-1 (HO-1) modulates mitochondrial biogenesis and antioxidant protection. We have previously shown the beneficial effect of caloric restriction (CR) on diabetic cardiomyopathy through intracellular signaling pathways involving the SIRT1-PGC-1α axis. In the current study, we examined the role of HO-1 in diabetic cardiomyopathy in mice subjected to CR.
Cardiomyopathy was induced in obese diabetic (
) mice by AT infusion. Mice were either fed ad libitum or subjected to CR. In an in vitro study, the reactive oxygen species (ROS) level was determined in cardiomyocytes exposed to different glucose levels (7.5-33 mM). We examined the effects of Sn(tin)-mesoporphyrin (SnMP), which is an inhibitor of HO activity, the HO-1 inducer cobalt protoporphyrin (CoPP), and the SIRT1 inhibitor (EX-527) on diabetic cardiomyopathy.
Diabetic mice had low levels of HO-1 and elevated levels of the oxidative marker malondialdehyde (MDA). CR attenuated left ventricular hypertrophy (LVH), increased HO-1 levels, and decreased MDA levels. SnMP abolished the protective effects of CR and caused pronounced LVH and cardiac metabolic dysfunction represented by suppressed levels of adiponectin, SIRT1, PPARγ, PGC-1α, and increased MDA. High glucose (33 mM) increased ROS in cultured cardiomyocytes, while SnMP reduced SIRT1, PGC-1α levels, and HO activity. Similarly, SIRT1 inhibition led to a reduction in PGC-1α and HO-1 levels. CoPP increased HO-1 protein levels and activity, SIRT1, and PGC-1α levels, and decreased ROS production, suggesting a positive feedback between SIRT1 and HO-1.
These results establish a link between SIRT1, PGC-1α, and HO-1 signaling that leads to the attenuation of ROS production and diabetic cardiomyopathy. CoPP mimicked the beneficial effect of CR, while SnMP increased oxidative stress, aggravating cardiac hypertrophy. The data suggest that increasing HO-1 levels constitutes a novel therapeutic approach to protect the diabetic heart. Brief Summary: CR attenuates cardiomyopathy, and increases HO-1, SIRT activity, and PGC-1α protein levels in diabetic mice. High glucose reduces adiponectin, SIRT1, PGC1-1α, and HO-1 levels in cardiomyocytes, resulting in oxidative stress. The pharmacological activation of HO-1 activity mimics the effect of CR, while SnMP increased oxidative stress and cardiac hypertrophy. These data suggest the critical role of HO-1 in protecting the diabetic heart.
This study investigated modifications to the ubiquitin proteasome system (UPS) in a mouse model of type 2 diabetes mellitus (T2DM) and their relationship to heart complications. db/db mice heart ...tissues were compared with WT mice tissues using RNA sequencing, qRT-PCR, and protein analysis to identify cardiac UPS modifications associated with diabetes. The findings unveiled a distinctive gene profile in the hearts of db/db mice with decreased levels of nppb mRNA and increased levels of Myh7, indicating potential cardiac dysfunction. The mRNA levels of USP18 (deubiquitinating enzyme), PSMB8, and PSMB9 (proteasome β-subunits) were down-regulated in db/db mice, while the mRNA levels of RNF167 (E3 ligase) were increased. Corresponding LMP2 and LMP7 proteins were down-regulated in db/db mice, and RNF167 was elevated in Adult diabetic mice. The reduced expression of LMP2 and LMP7, along with increased RNF167 expression, may contribute to the future cardiac deterioration commonly observed in diabetes. This study enhances our understanding of UPS imbalances in the hearts of diabetic mice and raises questions about the interplay between the UPS and other cellular processes, such as autophagy. Further exploration in this area could provide valuable insights into the mechanisms underlying diabetic heart complications and potential therapeutic targets.
The limited regenerative capacity of the injured myocardium leads to remodeling and often heart failure. Novel therapeutic approaches are essential. Induced pluripotent stem cells (iPSC) ...differentiated into cardiomyocytes are a potential future therapeutics. We hypothesized that organ-specific reprogramed fibroblasts may serve an advantageous source for future cardiomyocytes. Moreover, exosomes secreted from those cells may have a beneficial effect on cardiac differentiation and/or function. We compared RNA from different sources of human iPSC using chip gene expression. Protein expression was evaluated as well as exosome micro-RNA levels and their impact on embryoid bodies (EBs) differentiation. Statistical analysis identified 51 genes that were altered (
≤ 0.05), and confirmed in the protein level, cardiac fibroblasts-iPSCs (CF-iPSCs) vs. dermal fibroblasts-iPSCs (DF-iPSCs). Several miRs were altered especially miR22, a key regulator of cardiac hypertrophy and remodeling. Lower expression of miR22 in CF-iPSCs vs. DF-iPSCs was observed. EBs treated with these exosomes exhibited more beating EBs
= 0.05. vs. control. We identify CF-iPSC and its exosomes as a potential source for cardiac recovery induction. The decrease in miR22 level points out that our CF-iPSC-exosomes are naïve of congestive heart cell memory, making them a potential biological source for future therapy for the injured heart.
Objective We compared the flow rates, reactivity, and morphology of the distal internal thoracic artery and its branches, the superior epigastric and musculophrenic arteries, to test their ...applicability as possible conduits in coronary artery bypass grafting surgeries. Methods Skeletonized internal thoracic artery and subdivisions of patients undergoing coronary artery bypass grafting were studied intraoperatively (n = 100) for flow and length measurements and in vitro in organ baths (n = 58) for active response to norepinephrine. Quantitative microscopic analysis of the muscle density and degree of intimal hyperplasia was performed. Results were analyzed according to age, gender, risk factors, and medications. Results Internal thoracic artery subdivisions contributed an average extra length of 2 cm. Free flow rates were 129 ± 45 mL/min, 114 ± 41 mL/min, and 93 ± 36 mL/min in the internal thoracic artery, superior epigastric artery, and musculophrenic artery, respectively. Sternum and internal thoracic artery length and free flow rates were significantly lower in women. The subdivisions were significantly more reactive to norepinephrine than the distal internal thoracic artery ( P ∼ .005), although sensitivity to norepinephrine was similar. Patients treated with beta-blockers had significantly decreased reactivity ( P = .009). Microscopic analysis suggests similar muscle content in the internal thoracic artery and subdivisions. Eccentric (28%) and concentric (62%) intimal hyperplasia were observed in 90% of specimens, with no evidence for atherosclerotic plaques. There was no significant difference in the degree of intimal hyperplasia between the distal internal thoracic artery and its subdivisions, and there was no correlation to risk factors. Conclusions Our results confirm the previous studies on the higher contractility in internal thoracic artery subdivisions, suggesting caution in the use of the bifurcation for revascularization. However, the extra length, sufficient flow, and favorable histologic properties suggest that the bifurcation may be appropriate for coronary revascularization in selected cases.
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