Diabetic Myocardium Research Articles

Experimental diabetes mellitus exacerbates ischemia/reperfusion-induced myocardial injury by promoting mitochondrial fission: Role of down-regulation of myocardial Sirt1 and subsequent Akt/Drp1 interaction.

Diabetes mellitus (DM) has a negative impact on clinical outcomes for patients with myocardial infarction. The aim of the present study was to assess whether decreased myocardial levels of Sirtuin1 (Sirt1) contribute to the increased susceptibility of the diabetic myocardium to ischemia/reperfusion (I/R) injury. In vivo, myocardial levels of Sirt1 expression and activity were decreased in mice with STZ-induced DM. Increasing Sirt1 activity prevented the DM-induced exacerbation of myocardial mitochondrial fission, apoptosis and dysfunction elicited by I/R. In vitro, anoxia/reoxygenation (A/R) challenge of cardiomyocytes (CM) that were preconditioned with high glucose (HG-CM) resulted in an exacerbation of the A/R-induced mitochondrial fission, oxidant production and CM apoptosis; effects reversed by increasing Sirt1 protein/activity. Inhibition of Drp1 prevented the exacerbated CM mitochondrial fission and oxidant production after A/R challenge of HG-CM. Decreased Sirt1 in HG-CM was associated with decreased Akt phosphorylation. Inhibition of Akt had no effect on CM Sirt1 levels, but further increased Drp1 activation. Increasing Sirt1 levels prevented the decrease in Akt phosphorylation and Drp1 activation in A/R challenged HG-CM. In conclusion: our data indicate that the increased vulnerability of the diabetic myocardium to I/R-induced apoptosis/dysfunction is attributable, in part, to decreased myocardial Sirt1 activity which leads to a decrease in Akt activation, an increase in Drp1 activity, culminating in excessive mitochondrial fission and ROS production.

Peroxisome proliferator-activated receptor-gamma activation attenuates diabetic cardiomyopathy via regulation of the TGF-β/ERK pathway and epithelial-to-mesenchymal transition

Diabetic cardiomyopathy (DCM) is a kind of disease caused by metabolic disorders and microangiopathy. The main pathophysiological changes of DCM include fibrosis, myocardial cell apoptosis and autonomic neuropathy. Therefore, treatment aimed at these processes may benefit patients with DCM. We designed an experiment with the peroxisome proliferator-activated receptor-gamma (PPARγ) agonist GW 1929 to detect whether the activation of PPARγ could alleviate the degree of DCM. To further detect the mechanism of PPARγ in DCM, we used the PPARγ antagonist GW 9662 and ERK antagonist PD 098059 both in vitro and in vivo and found that PPARγ functioned by inhibiting ERK. We also performed Western blot, PCR, ELISA, immunohistochemistry, TUNEL assay, Sirius red staining and gelatin zymography to investigate inflammation, apoptosis, MMP activity and epithelial-to-mesenchymal transition (EMT). The results showed that the activation of PPARγ inhibited these reactions and inhibiting ERK also simulated this phenomenon. In conclusion, these results demonstrated that PPARγ activation in the diabetic myocardium of mice reduces myocardial fibrosis via regulation of the TGF-β/ERK pathway and EMT.

Abstract 536: Increased O-linked Glycosylation in Diabetic Myocardium of Mice and Human

Objectives: The modification of serine and threonine residues of nuclear and cytoplasmic proteins by O-linked β-N-acetylglucosamine (O-GlcNAc) has emerged as a highly dynamic post-translational modification that plays a critical role in regulating numerous biological processes. Our focus was to establish the difference of O-linked glycosylation in diabetic (DM) and Non-diabetic (ND) heart of mouse and human in the presence or absence of cardioplegic ischemia and cardiopulmonary bypass (CP/CPB). Methods and results: Right atrial tissue was harvested pre- and post-CP/CPB from the ND patients and patients with type-2 DM undergoing cardiac surgery (n = 4/group).The post-CP/CPB tissue samples were from a atrial tissue segment exposed to cold, hyperkalemic blood cardioplegia (ischemia) and a brief period (10 minutes) of reperfusion. Ventricular tissue samples were also obtained from mice with and without type-2 DM (n =3/group). Thin Layer Chromatography (TLC) was used to measure the contents of acetyl coenzyme A (acetyl-CoA) in the tissue lysates of mouse and human myocardium. Western blotting and immunohistochemistry analysis were employed to determine the expression/localization of O-Linked β-N-acetylglucosamine (O-GlcNAc) in the tissue lysates and paraffin-embedded tissue sections. Diabetes tends to increase the amounts of acetyl-CoA and O-GlcNAc (20% increase vs. ND, P<0.05) in the myocardium compared with ND. Post-CP/CPB further induced more acetyl-CoA and this effect was more pronounced in the diabetic atrial myocardium than that of ND. Immunofluorescence experiments with O-GlcNAc-specific antibody and paraffin-embedded tissue sections from human atria and mouse heart show a significant number of cells stain positively with the antibody in the diabetes sections (3 folds increases in optical density vs. ND, P <0.01). The protein oxidation and ratio of NADH/NAD in the DM myocardium were significantly increased compared with ND (P<0.05). Conclusion: Diabetes and cardioplegic ischemia are associated with an increase in O-linked glycosylation in the mouse and human myocardium, suggesting that diabetes and cardioplegic ischemia affect myocardial metabolism via hexosamine biosynthetic pathway.

Effect of Autophagy Inhibition on the Protection of Ischemia Preconditioning against Myocardial Ischemia/Reperfusion Injury in Diabetic Rats.

Background:Ischemia preconditioning (IPC) remains the most powerful intervention of protection against myocardial ischemia/reperfusion injury (IRI), but diabetes can weaken or eliminate its cardioprotective effect and detailed mechanisms remain unclear. In this study, we aimed to explore whether changes of autophagy in the diabetic condition are attributable to the decreased cardioprotective effect of IPC.Methods:Sixty diabetic male Sprague-Dawley rats were randomly divided into the control (C), IRI, rapamycin (R), wortmannin (W), rapamycin + IPC (R + IPC), and wortmannin + IPC (W + IPC) groups. The in vivo rat model of myocardial IRI was established by ligaturing and opening the left anterior descending coronary artery via the left thoracotomy. Durations of ischemia and reperfusion are 30 min and 120 min, respectively. Blood samples were taken at 120 min of reperfusion for measuring serum concentrations of troponin I (TnI) and creatine kinase isoenzyme MB (CK-MB) using the enzyme-linked immunosorbent assay. The infarct size was assessed by Evans blue and triphenyltetrazolium chloride staining. The expressions of LC3-II, beclin-1, phosphoinositide 3-kinase (PI3K), mammalian target of rapamycin (mTOR), and P-Akt/Akt ratio in the ischemic myocardium were assessed by Western blotting.Results:Compared to the IRI group, infarct size (56.1% ± 6.1% vs. 75.4 ± 7.1%, P < 0.05), serum cTnI (0.61 ± 0.21 vs. 0.95 ± 0.26 ng/ml, P < 0.05), and CK-MB levels (6.70 ± 1.25 vs. 11.51 ± 2.35 ng/ml, P < 0.05) obviously decreased in the W + IPC group. Compared with the C group, myocardial expressions of LC3-II (0.46 ± 0.04 and 0.56 ± 0.04 vs. 0.36 ± 0.04, P < 0.05) and beclin-1 (0.34 ± 0.08 and 0.38 ± 0.07 vs. 0.24 ± 0.03, P < 0.05) evidently increased, and myocardial expressions of mTOR (0.26 ± 0.08 and 0.25 ± 0.07 vs. 0.38 ± 0.06, P < 0.05), PI3K (0.29 ± 0.04 and 0.30 ± 0.03 vs. 0.38 ± 0.02, P < 0.05), and P-Akt/Akt ratio (0.49 ± 0.10 and 0.48 ± 0.06 vs. 0.72 ± 0.07, P < 0.05) markedly decreased in the IRI and R groups, indicating an increased autophagy. Compared with the IRI group, myocardial expression of beclin-1 (0.26 ± 0.03 vs. 0.34 ± 0.08, P < 0.05) significantly decreased, and myocardial expressions of mTOR (0.36 ± 0.04 vs. 0.26 ± 0.08, P < 0.05), PI3K (0.37 ± 0.03 vs. 0.29 ± 0.04, P < 0.05), and P-Akt/Akt ratio (0.68 ± 0.05 vs. 0.49 ± 0.10, P < 0.05) increased obviously in the W + IPC group, indicating a decreased autophagy.Conclusions:Increased autophagy in the diabetic myocardium is attributable to decreased cardioprotection of IPC, and autophagy inhibited by activating the PI3K-Akt-mTOR signaling pathway can result in an improved protection of IPC against diabetic myocardial IRI.

Protective effect of myocardial calcitonin gene-related peptide gene transduction on myocardial ischemia/reperfusion injury in diabetic rats

Objective To evaluate the feasibility of lentivirus (Lv) mediated calcitonin gene-related peptide(CGRP) gene transduction in diabetic rats by myocardial injection and whether this method has a protective effect on diabetic myocardium during ischemia/reperfusion(I/R) injury. Methods Sixty healthy male SD rats, weighing 200-220 g, were randomly divided into control group (group C), diabetes mellitus group (group DM), Sham transduction group (group Sham), myocardial injection control group (group MC) and myocardial injection experiment group(group ME), in accordance with a random number table. Rats in group Sham, group MC and group ME were given with normal saline, at a titer of 2×107 virus genomes(vg)/ml Lv-enhanced green fluorescent protein (eGFP) and Lv-CGRP-eGFP 50 μl respectively by myocardial injection. One week later, rats in group DM, group Sham, group MC and group ME received streptozocin(STZ) 50 mg/kg intraperitoneal injection to induce diabetes. Eight weeks after the injection of STZ, myocardial I/R was induced by occlusion and release of left anterior descending branch of coronary artery. The latency of tail flick reflection, the expression of eGFP, the transcription level of CGRP, the expression of CGRP in myocardium, spinal cord and dorsal root ganglion(DRG) and myocardial ischemia area, infarct size(IS) area were measured. Results Compared with group C, the latency of tail flick reflection in group DM, group Sham, group MC and group ME were significantly prolonged in the 4th week after diabetic model successfully manufactured(P<0.05). The transcription levels of CGRP in the ischemic myocardium, spinal cord T1-T5 and DRG T1-T5 in group DM, group Sham and group MC were lower than those in group C and group ME (P<0.05). The level of CGRP in ischemic myocardium and DRG T1-T5 of group ME was higher than that in group C(P<0.05). The content of CGRP in cord T1-T5, DRG T1-T5, ischemic myocardium and serum in group C and group ME were higher than those in group DM, group Sham and group MC (P<0.05). The content of CGRP in cord T1-T5, ischemic myocardium in group ME was higher than that in group C (P<0.05). The infarct size in group DM, group Sham, group MC were higher than that in group C and group ME(P<0.05), the infarct size of group ME was higher than that in group C(P<0.05). Conclusions Intramyocardial injection of CGRP gene can effectively enhance the expression of CGRP at I/R of diabetic rat′ heart. Increased CGRP may reduce cardiac I/R injury in diabetic rats. Key words: Calcitonin gene-related peptide; Diabetes mellitus; Myocardium; Ischemia-reperfusion

Impairment of the Akt pathway in transplanted Type 1 diabetic hearts is associated with post-transplant graft injury.

The use of 'marginal' hearts, such as from donors with diabetes mellitus (DM), could offer an opportunity to expand the donor pool in cardiac transplantation. Previous studies have shown that the phosphatidylinositol-3-kinase (PI3K)/Akt pathway is altered after ischaemia/reperfusion injury in the diabetic myocardium. We hypothesized that DM-induced cardiac dysfunction in donors is further impaired after heart transplantation and that PI3K/Akt-pathway alterations may be one of the underlying pathomechanisms. In the donor rats, DM was induced with a single dose of streptozotocin. Non-diabetic rats only received citrate buffer. After 8 weeks, the donor left ventricular (LV) cardiac function was measured. Then, the hearts were heterotopically transplanted into non-diabetic recipients. We evaluated LV graft function 1.5 h after transplantation via a Millar catheter system at different LV volumes. Histological analyses were performed, and the expression of 84 genes involved in PI3K/Akt signalling was profiled. DM was associated with significantly decreased LV contractility and impaired relaxation. After transplantation, in the DM group, the grafts' systolic function (LV systolic pressure 112 ± 31 vs 155 ± 60 mmHg; dP/dtmax 2676 ± 896 vs 3584 ± 1779 mmHg/s, P < 0.05) and diastolic function (dP/dtmin 924 ± 205 vs 1748 ± 512 mmHg/s, P < 0.05) were significantly reduced at an intraventricular volume of 170 µl. The expression of 10 genes involved in PI3K/Akt signalling, as well as the phosphorylated Akt/total Akt protein expression ratio, were significantly down-regulated in the diabetic heart after transplantation. DM-induced cardiac dysfunction is further impaired after transplantation. Targeting the PI3K/Akt pathway may result in a functional amelioration of the a priori-diseased myocardia, which could increase the number of potential cardiac donors.

A Crude 1-DNJ Extract from Home Made Bombyx Batryticatus Inhibits Diabetic Cardiomyopathy-Associated Fibrosis in db/db Mice and Reduces Protein N-Glycosylation Levels.

The traditional Chinese drug Bombyx Batryticatus (BB), which is also named the white stiff silkworm, has been widely used in Chinese clinics for thousands of years. It is famous for its antispasmodic and blood circulation-promoting effects. Cardiomyocyte hypertrophy, interstitial cell hyperplasia, and myocardial fibrosis are closely related to the N-glycosylation of key proteins. To examine the alterations of N-glycosylation that occur in diabetic myocardium during the early stage of the disease, and to clarify the therapeutic effect of 1-Deoxynojirimycin (1-DNJ) extracted from BB, we used the db/db (diabetic) mouse model and an approach based on hydrophilic chromatography solid-phase extraction integrated with an liquid Chromatograph Mass Spectrometer (LC-MS) identification strategy to perform a site-specific N-glycosylation analysis of left ventricular cardiomyocyte proteins. Advanced glycation end products (AGEs), hydroxyproline, connective tissue growth factor (CTGF), and other serum biochemical indicators were measured with enzyme-linked immunosorbent assays (ELISA). In addition, the α-1,6-fucosylation of N-glycans was profiled with lens culinaris agglutinin (LCA) lectin blots and fluorescein isothiocyanate (FITC)-labelled lectin affinity histochemistry. The results indicated that 1-DNJ administration obviously downregulated myocardium protein N-glycosylation in db/db mice. The expression levels of serum indicators and fibrosis-related cytokines were reduced significantly by 1-DNJ in a dose-dependent manner. The glycan α-1,6-fucosylation level of the db/db mouse myocardium was elevated, and the intervention effect of 1-DNJ administration on N-glycan α-1,6-fucosylation was significant. To verify this result, the well-known transforming growth factor-β (TGF-β)/Smad2/3 pathway was selected, and core α-1,6-fucosylated TGF-β receptor II (TGFR-βII) was analysed semi-quantitatively with western blotting. The result supported the conclusions obtained from LCA lectin affinity histochemistry and lectin blot analysis. The expression level of α-1,6-fucosyltransferase (FUT8) mRNA was also detected, and the results showed that 1-DNJ administration did not cause obvious inhibitory effects on FUT8 expression. Therefore, the mechanism of 1-DNJ for relieving diabetic cardiomyopathy (DCM)-associated fibrosis can be concluded as the inhibition of N-acetylglucosamine (N-GlcNAc) formation and the reduction of substrate concentration.

Simvastatin prevented myocardium of diabetes rats from apoptosis through inhibition of oxidative stress

To investigate the protective effects and the possible mechanisms of simvastatin on myocardial injury induced by diabetes. Twenty-four SD rats (180~220)g were randomly divided into control group (control, n=8) and modeled groups(n=16), the modeled groups were injected with streptozotocin intraperitoneally to induce diabetes. Then the modeled rats were randomly divided into diabetes mellitus group (DM group, n=8) and diabetes mellitus + simvastatin group (DM+S group, n=8). Rats in DM+S group were treated with simvastatin at the dose of 40 mg/(kg·d)by gavage for 4 weeks, and the other two groups were treated with the same amount of saline. At the end of experiments, the heart tissues were collected for further observation. The content of malondialdehyde (MDA) and the activity of superoxide dismutase (SOD) in heart tissues were measured by spectrophotometry; HE staining of rat heart slides was used to observe the pathological changes; TUNEL assay was used to determine the apoptosis index of myocardial cells in each groups; The distribution of p53 in the heart tissues was evaluated by immunohistochemistry; Western blot was used to detect the expressions of p53, p53-phospho-serine 15, Bax and Bcl-2 in the heart tissues. ①Compared with control group, the content of malondialdehyde (MDA) was increased while the activity of superoxide dismutase (SOD) was decreased significantly in DM group (P<0.01). After simvastatin administration, the activity of SOD was increased and the content of MDA was decreased significantly (P<0.01). ② HE staining results showed that the myocardial cells in the DM group were disorganized, with unclear morphological structure and a large number of inflammatory cells infiltration. Compared with DM group, the myocardial morphology in DM+S group was improved significantly. ③TUNEL staining results showed that the apoptosis index of myocardial cells in DM group was increased significantly compared with that of control group, and the apoptosis index was decreased significantly after the treatment of simvastatin (P<0.01).④ Immunohistochemistry showed that compared with control group,the expression of p53 in DM group was increased significantly, and was expressed in both cytoplasm and nucleus, while the expression of p53 in DM+S group was decreased and the expression of p53 in nucleus was decreased significantly (P<0.01). ⑤ The results of Western blot showed that the expression levels of p53, p53-phospho-serine15 and Bax were higher than those in control group, and the expression of Bcl-2 was lower than that in control group (P<0.01). After simvastatin administration, the expression levels of p53,p53-phospho-serine 15 (P<0.01) and Bax were decreased significantly (P<0.05) and the expression of Bcl-2 was increased (P<0.05). Simvastatin exerted protective effects on myocardial injury caused by diabetes through improving the abnormal morphological changes of diabetic myocardium, alleviating oxidative stress and inhibiting apoptosis of myocardial cells. The mechanism is related to the regulation of apoptosis pathway mediated by p53.

Anti-arrhythmogenic and anti-inflammatory effects of troxerutin in ischemia/reperfusion injury of diabetic myocardium

IntroductionMedicinal plants are increasingly used in the treatment of cardiovascular diseases due to their multifaceted properties. This study was designed to investigate anti-arrhythmic and anti-inflammatory potentials of the natural bioflavonoid, troxerutin (TXR) in myocardial ischemia/reperfusion (I/R) injury in diabetic rats. MethodsMale Wistar rats were randomly divided into 4 groups (control, control + TXR [150 mg/kg, daily], diabetic, and diabetic + TXR). Type-1 diabetes was induced by an intraperitoneal injection of streptozotocin (50 mg/kg) and lasted for 10 weeks. After mounting on the Langendorff apparatus, isolated hearts in all groups received a normal Krebs–Henseleit solution for 20 min of stabilization period, followed by 30 min of regional ischemia through ligation of the left anterior descending coronary artery, and 60 min of full reperfusion. During the experiment, the electrocardiograms were recorded and the arrhythmias [number, duration and incidence of premature ventricular complexes (PVC), ventricular tachycardia (VT), ventricular fibrillation (VF), and arrhythmia score] during I/R phases were assessed based on the Lambeth Convention. Ischemic left ventricular samples were used to determine the activities of lactate dehydrogenase (LDH), interleukin-1beta (IL-1β), and tumor necrosis factor (TNF-α). ResultsThe arrhythmias induced by I/R were not significantly changed in diabetic group as compared to the control group. However, pretreatment with TXR significantly reduced the number of PVC and duration and incidence of VF in ischemic phase in comparison to the untreated animals (P < 0.05). In addition, the duration, and incidence of most arrhythmias during reperfusion phase were significantly declined by TXR administration in both control and diabetic groups (P < 0.05). Pretreatment of rats with TXR significantly reduced myocardial inflammatory cytokines TNF-α and IL-1β levels after I/R insult in diabetic as well as control hearts (P < 0.05). ConclusionPreconditioning with TXR could provide cardioprotection by anti-arrhythmic and anti-inflammatory effects against I/R injury in rat hearts. This effect of TXR can introduce this material as a protective agent in cardiovascular diseases.

Signaling mediators modulated by cardioprotective interventions in healthy and diabetic myocardium with ischaemia-reperfusion injury.

Ischaemic heart diseases are one of the major causes of death in the world. In most patients, ischaemic heart disease is coincident with other risk factors such as diabetes. Patients with diabetes are more prone to cardiac ischaemic dysfunctions including ischaemia-reperfusion injury. Ischaemic preconditioning, postconditioning and remote conditionings are reliable interventions to protect the myocardium against ischaemia-reperfusion injuries through activating various signaling pathways and intracellular mediators. Diabetes can disrupt the intracellular signaling cascades involved in these myocardial protections, and studies have revealed that cardioprotective effects of the conditioning interventions are diminished in the diabetic condition. The complex pathophysiology and poor prognosis of ischaemic heart disease among people with diabetes necessitate the investigation of the interaction of diabetes with ischaemia-reperfusion injury and cardioprotective mechanisms. Reducing the outcomes of ischaemia-reperfusion injury using targeted strategies would be particularly helpful in this population. In this study, we review the protective interventional signaling pathways and mediators which are activated by ischaemic conditioning strategies in healthy and diabetic myocardium with ischaemia-reperfusion injury.

Protection by dimethyl fumarate against diabetic cardiomyopathy in type 1 diabetic mice likely via activation of nuclear factor erythroid-2 related factor 2

Oxidative stress and inflammation play key roles in the development of diabetic cardiomyopathy (DCM). Dimethyl fumarate (DMF), an FDA approved medicine for relapsing multiple sclerosis, has manifested its antioxidant and anti-inflammatory function mostly in the central nervous system. In this study, we investigated whether DMF could attenuate the development of DCM. Type 1 diabetes mouse model was established using multiple low-dose streptozotocin, and the diabetic mice were treated with DMF (10 mg/kg body weight) for 3 months. Cardiac functions were determined using echocardiography. Oxidative stress, pro-inflammatory cytokines and pro-fibrotic markers were determined with commercially available kits, real-time quantitative PCR or western blot techniques. DCM was characterized by diminished cardiac function, accompanied by oxidative stress and enhanced expression of pro-inflammatory cytokines. Diabetic cardiac tissue exhibited marked fibrosis, revealed by extracellular matrix deposition as determined by Sirius red staining of the myocardial tissues. Furthermore, Nrf2 and its downstream effectors were repressed in diabetic myocardium. On the contrary, diabetic animals treated with DMF exhibited blunted oxidative stress, inflammation, fibrosis and this correlated with Nrf2 activation. Our findings suggest that DMF could potentially thwart diabetes-induced myocardial tissue injury, likely via activation of Nrf2 function, providing firm impetus for future repurposing of DMF in the management of DCM.

Polydatin Protects Diabetic Heart against Ischemia-Reperfusion Injury via Notch1/Hes1-Mediated Activation of Pten/Akt Signaling.

Diabetes exacerbates oxidative/nitrative stress during myocardial ischemia-reperfusion (MI/R) injury. Recent studies highlighted the cardioprotective actions of polydatin. However, its effect on diabetic MI/R injury and the underlying mechanisms remain unknown. This work was undertaken to evaluate the effect of polydatin on diabetic MI/R injury with a focus on Notch1/Hes1 signaling and myocardial oxidative/nitrative stress. Streptozotocin- (STZ-) induced diabetic rats were administered with polydatin (20 mg/kg/d) in the absence or presence of DAPT (a γ-secretase inhibitor) or LY294002 (a PI3K/Akt inhibitor) and then subjected to MI/R injury. Polydatin administration preserved cardiac function and reduced myocardial infarct size. Moreover, polydatin ameliorated myocardial oxidative/nitrative stress damage as evidenced by decreased myocardial superoxide generation, malondialdehyde, gp91phox expression, iNOS expression, NO metabolite level, and nitrotyrosine content and increased eNOS phosphorylation. However, these effects were blocked by DAPT administration. DAPT also inhibited the stimulatory effect of polydatin on the Notch1/Hes1-Pten/Akt signaling pathway in a diabetic myocardium. Additionally, LY294002 not only abolished polydatin's antiapoptotic effect but also reversed its inhibitory effect on myocardial oxidative/nitrative stress. Polydatin effectively reduced MI/R injury and improved left ventricular functional recovery under diabetic condition by ameliorating oxidative/nitrative stress damage. Importantly, Notch1/Hes1-mediated activation of Pten/Akt signaling played a crucial role in this process.

Pathological prolongation of action potential duration as a cause of the reduced alpha-adrenoceptor-mediated negative inotropy in streptozotocin-induced diabetic mice myocardium

Effect of pathological prolongation of action potential duration on the α-adrenoceptor-mediated negative inotropy was studied in streptozotocin-induced diabetic mice myocardium. In streptozotocin-treated mouse ventricular myocardium, which had longer duration of action potential than that in control mice, the negative inotropic response induced by phenylephrine was smaller than that in control mice. 4-Aminopyridine prolonged the action potential duration and decreased the negative inotropy in control mice. Cromakalim shortened the action potential duration and increased the negative inotropy in streptozotocin-treated mice. These results suggest that the reduced α-adrenoceptor-mediated inotropy in the diabetic mouse myocardium is partly due to its prolonged action potential.

Combined postconditioning with ischemia and cyclosporine-A restore oxidative stress and histopathological changes in reperfusion injury of diabetic myocardium.

Objective(s):Chronic diabetes impedes cardioprotection in reperfusion injury and hence protecting the diabetic heart would have important outcomes. In this study, we evaluated whether combined postconditioning with ischemia and cyclosporine-A can restore oxidative stress and histopathological changes in reperfusion injury of the diabetic myocardium.Materials and Methods:Streptozocin-induced diabetic hearts and nondiabetic controls in eight subgroups (with or without receiving ischemic-postconditioning (IPostC), cyclosporine-A, an inhibitor of mitochondrial permeability transition, or both of them) suffered from 30 min regional ischemia followed by 45 min reperfusion on an isolated-heart Langendorff system. The levels of lactate dehydrogenase (LDH) in the coronary effluent, and the levels of oxidative stress markers including 8-isoprostane, superoxide dismutase (SOD), glutathione peroxidase (GPX), and total antioxidant capacity (TAC) in myocardial supernatant prepared from the ischemic zone were measured using specific kits, spectrophotometrically. Histopathological studies were performed -eosin staining method.Results:Administration of IPostC and cyclosporine-A (alone or together) in nondiabetic hearts potentially reduced the severity of histological changes and level of LDH release as compared with untreated-controls (P<0.05). of any procedures in diabetic hearts did not show significant cardioprotective effects (P>0.1). However, the combined postconditioning with ischemia and CsA exerted significant protective effects in diabetic hearts (P<0.05).Conclusion:By augmenting the protective effects of IPostC and CsA through their combined application, reperfusion injury and related oxidative stress are reduced in diabetic hearts similar to non-diabetics.

Effect of Ischemic Postconditioning on Myocardial Function and Infarct Size Following Reperfusion Injury in Diabetic Rats Pretreated With Vildagliptin.

Cardioprotective actions of ischemic postconditioning (IPostC) against ischemia/reperfusion (I/R) injury are abolished in diabetic hearts. This study has investigated the combined effects of IPostC and vildagliptin (Vilda) on myocardial function and infarct size (IS) against I/R injury in diabetic myocardium. Diabetes was induced by a high-fat diet/low dose of streptozotocin (35 mg/kg; intraperitoneally) in Wistar rats (200-250 g) and lasted for 12 weeks. Vilda (6 mg/kg/d) was orally administered for 5 weeks in diabetic groups after seventh week of diabetes. At the end of the 12-week period, the hearts of rats were removed and subjected to 35-minute regional ischemia (through left anterior descending ligation) followed by 60-minute reperfusion, on Langendorff apparatus. Ischemic postconditioning was induced by 6 repetitive cycles of 10-second ischemia and 10-second reperfusion, immediately at the onset of the reperfusion. Myocardial hemodynamic was measured throughout the experiment. The IS was assessed by triphenyltetrazolium chloride staining method. The myocardial contents of troponin-I (cTnI), interleukin-6 (IL-6), and 8-isoprostane were measured in the homogenate from ischemic zone of left ventricles by enzyme-linked immunosorbent assay kit. Pretreatment of the diabetic rats with Vilda significantly recovered the diabetes-induced reduction in left ventricular developed pressures and contractility at the baseline ( P < .05 to P < .01). After I/R injury, IPostC could not significantly improve the myocardial function, cTnI content, and IS of the diabetic hearts. However, in Vilda-treated hearts, concomitant application of IPostC significantly recovered the heart functions, returned cTnI content as well as myocardial IL-6 and 8-isoprostane levels back to the control values ( P < .01 to P < .001), and reduced IS more effectively (by 45%) in comparison to the diabetic group ( P < .001). Besides its glycemic and lipid profile controlling effects, Vilda has a protective effect on heart function and tends to restore cardioprotective effects of IPostC on diabetic hearts.

Resveratrol Ameliorates Cardiac Dysfunction by Inhibiting Apoptosis via the PI3K/Akt/FoxO3a Pathway in a Rat Model of Diabetic Cardiomyopathy

The aim of this study was to explore the effect and mechanism of action of resveratrol (RSV) on cardiac function in diabetic cardiomyopathy (DCM). Hyperglycemia-induced apoptosis contributes to the pathogenic changes in DCM. RSV treatment inhibited high glucose-induced apoptosis of neonatal rat ventricular myocytes. Additionally, high glucose decreased cell viability, prevented serine-threonine kinase (Akt) and FoxO3a phosphorylation, and suppressed cytoplasmic translocation of FoxO3a. However, these effects of apoptosis were reversed by 10 μM of RSV. The PI3K inhibitor LY294002 abolished the RSV protective effect in vitro. RSV (5 or 50 mg·kg·d orally for 8 weeks) prevented the deterioration of cardiac function and structural cardiomyopathy in a streptozotocin-induced rat model of diabetes and reduced apoptosis in diabetic myocardium. Furthermore, it restored streptozotocin-impaired phosphorylation of Akt and FoxO3a (p-Akt and p-FoxO3a) and suppressed nuclear translocation of FoxO3a in vivo. Together, these data indicate that RSV has therapeutic potential against DCM by inhibiting apoptosis via the PI3K/Akt/FoxO3a pathway.

P1589Metabolomic impact of glucagon-like peptide-1 on diabetic myocardium - glucagon-like peptide-1 accelerates myocardial glycolysis and utilizes more lactate as myocardial substrate

P1589Metabolomic impact of glucagon-like peptide-1 on diabetic myocardium - glucagon-like peptide-1 accelerates myocardial glycolysis and utilizes more lactate as myocardial substrate

Long Noncoding RNA H19 Acts as a Competing Endogenous RNA to Mediate CTGF Expression by Sponging miR-455 in Cardiac Fibrosis.

Cardiac fibrosis is closely related to multiple cardiovascular system diseases, and noncoding RNAs (ncRNAs), including long noncoding RNA (lncRNA) and microRNA (miRNA), have been reported to play a vital role in fibrogenesis. The present study aims to investigate the potential regulatory mechanism of lncRNA H19 and miR-455 on fibrosis-associated protein synthesis in cardiac fibroblasts (CFs). miRNA microarray assay revealed 34 significantly dysregulated miRNAs, including 13 upregulated miRNAs and 21 downregulated miRNAs. Among these aberrantly expressed miRNAs, we paid attention to miR-455, which was significantly downregulated in diabetic mouse myocardium and Ang II-induced CFs. Loss- and gain-of-function experiments showed that miR-455 expression levels were negatively correlated with collagen I and III expression in Ang II-induced CFs. Bioinformatic prediction programs (TargetScan, miRanda, starBase) predicted that miR-455 targeted connective tissue growth factor (CTGF) and H19 with complementary binding sites at the 3'-untranslated region, which was validated by luciferase reporter assay. Functional validation assay demonstrated that H19 knockdown could enhance the antifibrotic role of miR-455 and attenuate the CTGF expression and further decrease fibrosis-associated protein synthesis (collagen I, III, and α-SMA). The present study reveals a novel function of the H19/miR-455 axis targeting CTGF in cardiac fibrosis, suggesting its potential therapeutic role in cardiac diseases.

Inhibition of sarcolemmal FAT/CD36 by sulfo-N-succinimidyl oleate rapidly corrects metabolism and restores function in the diabetic heart following hypoxia/reoxygenation.

AimsThe type 2 diabetic heart oxidizes more fat and less glucose, which can impair metabolic flexibility and function. Increased sarcolemmal fatty acid translocase (FAT/CD36) imports more fatty acid into the diabetic myocardium, feeding increased fatty acid oxidation and elevated lipid deposition. Unlike other metabolic modulators that target mitochondrial fatty acid oxidation, we proposed that pharmacologically inhibiting fatty acid uptake, as the primary step in the pathway, would provide an alternative mechanism to rebalance metabolism and prevent lipid accumulation following hypoxic stress.Methods and resultsHearts from type 2 diabetic and control male Wistar rats were perfused in normoxia, hypoxia and reoxygenation, with the FAT/CD36 inhibitor sulfo-N-succinimidyl oleate (SSO) infused 4 min before hypoxia. SSO infusion into diabetic hearts decreased the fatty acid oxidation rate by 29% and myocardial triglyceride concentration by 48% compared with untreated diabetic hearts, restoring fatty acid metabolism to control levels following hypoxia-reoxygenation. SSO infusion increased the glycolytic rate by 46% in diabetic hearts during hypoxia, increased pyruvate dehydrogenase activity by 53% and decreased lactate efflux rate by 56% compared with untreated diabetic hearts during reoxygenation. In addition, SSO treatment of diabetic hearts increased intermediates within the second span of the Krebs cycle, namely fumarate, oxaloacetate, and the FAD total pool. The cardiac dysfunction in diabetic hearts following decreased oxygen availability was prevented by SSO-infusion prior to the hypoxic stress. Infusing SSO into diabetic hearts increased rate pressure product by 60% during hypoxia and by 32% following reoxygenation, restoring function to control levels.ConclusionsDiabetic hearts have limited metabolic flexibility and cardiac dysfunction when stressed, which can be rapidly rectified by reducing fatty acid uptake with the FAT/CD36 inhibitor, SSO. This novel therapeutic approach not only reduces fat oxidation but also lipotoxicity, by targeting the primary step in the fatty acid metabolism pathway.

A novel identified circular RNA, circRNA_010567, promotes myocardial fibrosis via suppressing miR-141 by targeting TGF-β1

Circular RNAs (circRNAs) are a novel type of endogenous noncoding RNA gaining research interest in recent years. Despite this increase in interest, the mechanism of circRNAs in the pathogenesis of multiple cardiovascular diseases, particularly myocardial fibrosis, is rarely reported. In the following study, the expression profiles and potential mechanisms of circRNAs in mice myocardial fibrosis models in vitro are investigated. Previous research examining circRNA expression profiles of diabetic db/db mice myocardium using circRNA microarray found 43 circRNAs were abnormally expressed, including 24 up-regulated circRNAs and 19 down-regulated circRNAs. Furthermore, circRNA_010567 was markedly up-regulated in diabetic mice myocardium and cardiac fibroblasts (CFs) treated with Ang II. Bioinformatics analysis predicted circRNA_010567, sponge miR-141 and miR-141 directly target TGF-β1, which was validated by dual-luciferase assay. Subsequently, functional experiments revealed circRNA_010567 silencing could up-regulate miR-141 and down-regulate TGF-β1 expression, and suppress fibrosis-associated protein resection in CFs, including Col I, Col III and α-SMA. Results demonstrate the circRNA_010567/miR-141/TGF-β1 axis plays an important regulatory role in the diabetic mice myocardial fibrosis model. The present study characterizes a new function of circRNA in the pathogenesis of myocardial fibrosis in a diabetic mouse model, providing novel insight for circRNA-miRNA-mRNA in cardiovascular disease.