Progression Of Diabetic Cardiomyopathy Research Articles

Cellular interplay between cardiomyocytes and non-myocytes in diabetic cardiomyopathy.

Patients with Type 2 diabetes mellitus (T2DM) frequently exhibit a distinctive cardiac phenotype known as diabetic cardiomyopathy. Cardiac complications associated with T2DM include cardiac inflammation, hypertrophy, fibrosis, and diastolic dysfunction in the early stages of the disease, which can progress to systolic dysfunction and heart failure. Effective therapeutic options for diabetic cardiomyopathy are limited and often have conflicting results. The lack of effective treatments for diabetic cardiomyopathy is due in part, to our poor understanding of the disease development and progression, as well as a lack of robust and valid preclinical human models that can accurately recapitulate the pathophysiology of the human heart. In addition to cardiomyocytes, the heart contains a heterogeneous population of non-myocytes including fibroblasts, vascular cells, autonomic neurons, and immune cells. These cardiac non-myocytes play important roles in cardiac homeostasis and disease, yet the effect of hyperglycaemia and hyperlipidaemia on these cell types is often overlooked in preclinical models of diabetic cardiomyopathy. The advent of human-induced pluripotent stem cells provides a new paradigm in which to model diabetic cardiomyopathy as they can be differentiated into all cell types in the human heart. This review will discuss the roles of cardiac non-myocytes and their dynamic intercellular interactions in the pathogenesis of diabetic cardiomyopathy. We will also discuss the use of sodium-glucose cotransporter 2 inhibitors as a therapy for diabetic cardiomyopathy and their known impacts on non-myocytes. These developments will no doubt facilitate the discovery of novel treatment targets for preventing the onset and progression of diabetic cardiomyopathy.

Plin5 Bidirectionally Regulates Lipid Metabolism in Oxidative Tissues.

Cytoplasmic lipid droplets (LDs) can store neutral lipids as an energy source when needed and also regulate the key metabolic processes of intracellular lipid accumulation, which is associated with several metabolic diseases. The perilipins (Plins) are a family of proteins that associate with the surface of LDs. As a member of Plins superfamily, perilipin 5 (Plin5) coats LDs in cardiomyocytes, which is significantly related to reactive oxygen species (ROS) production originated from mitochondria in the heart, consequently determining the progression of diabetic cardiomyopathy. Plin5 may play a bidirectional function in lipid metabolism which is in a state of dynamic balance. In the basic state, Plin5 inhibited the binding of comparative gene identification-58 (CGI-58) to adipose triglyceride lipase (ATGL) by binding CGI-58, thus inhibiting lipolysis. However, when the body is under stress (such as cold, fasting, exercise, and other stimuli), protein kinase A (PKA) phosphorylates and activates Plin5, which then causes Plin5 to release the binding site of CGI-58 and ATGL, prompting CGI-58 to bind to ATGL and activate ATGL activity, thus accelerating the lipolysis process, revealing the indispensable role of Plin5 in lipid turnover. Here, the purpose of this review is to summarize the present understanding of the bidirectional regulation role of Plin5 in oxidative tissues and to reveal its potential role in diabetic cardiomyopathy protection.

Effect of Metformin on T2D-Induced MAM Ca2+ Uncoupling and Contractile Dysfunction in an Early Mouse Model of Diabetic HFpEF.

Diabetic cardiomyopathy (DCM) is a leading complication in type 2 diabetes patients. Recently, we have shown that the reticulum-mitochondria Ca2+ uncoupling is an early and reversible trigger of the cardiac dysfunction in a diet-induced mouse model of DCM. Metformin is a first-line antidiabetic drug with recognized cardioprotective effect in myocardial infarction. Whether metformin could prevent the progression of DCM remains not well understood. We therefore investigated the effect of a chronic 6-week metformin treatment on the reticulum-mitochondria Ca2+ coupling and the cardiac function in our high-fat high-sucrose diet (HFHSD) mouse model of DCM. Although metformin rescued the glycemic regulation in the HFHSD mice, it did not preserve the reticulum-mitochondria Ca2+ coupling either structurally or functionally. Metformin also did not prevent the progression towards cardiac dysfunction, i.e., cardiac hypertrophy and strain dysfunction. In summary, despite its cardioprotective role, metformin is not sufficient to delay the progression to early DCM.

Sleeve Gastrectomy-Induced AMPK Activation Attenuates Diabetic Cardiomyopathy by Maintaining Mitochondrial Homeostasis via NR4A1 Suppression in Rats.

Diabetic cardiomyopathy (DCM) is characterized by impaired diastolic and systolic myocardial performance and is a major cause of morbidity and mortality in patients with diabetes. Surgical bariatric procedures, such as sleeve gastrectomy (SG), result in remission of type 2 diabetes (T2DM) and have benefits with myocardial function. Maintaining cardiac mitochondrial homeostasis is a promising therapeutic strategy for DCM. However, whether SG surgery affects mitochondrial function and its underlying mechanism remains unclear. This study aimed to investigate the effect of SG surgery on mitochondrial homeostasis and intracellular oxidative stress in rats with DCM. We also examined cellular phenotypes and molecular mechanisms in high glucose and high fat-stimulated myocytes. The rat model of DCM was established by high-fat diet feeding and low-dose streptozotocin injection. We observed a remarkably metabolic benefit of SG, including a reduced body weight, food intake, blood glucose levels, and restored glucose tolerance and insulin sensitivity post-operatively. Also, SG ameliorated the pathological cardiac hypertrophy, myocardial fibrosis and the dysfunction of myocardial contraction and diastole, consequently delayed the progression of DCM. Also, SG restored the mitochondrial dysfunction and fragmentation through the AMPK signaling activation mediated nuclear receptor subfamily 4 group A member 1 (NR4A1)/DRP1 suppression in vivo. H9c2 cardiomyocytes showed that activation of AMPK could reverse the mitochondrial dysfunction somehow. Collectively, our study provided evidence that SG surgery could alleviate mitochondrial dysfunction in DCM. Moreover, AMPK-activated NR4A1/DRP1 repression might act as a significant reason for maintaining mitochondrial homeostasis in the myocardium, thus contributing to morphological and functional alleviation of DCM.

Lipotoxicity-induced mtDNA release promotes diabetic cardiomyopathy by activating the cGAS-STING pathway in obesity-related diabetes

Diabetic cardiomyopathy (DCM) is characterized by lipid accumulation, mitochondrial dysfunction, and aseptic inflammatory activation. Mitochondria-derived cytosolic DNA has been reported to induce inflammation by activating cyclic GMP-AMP synthase (cGAS)/the stimulator of interferon genes (STING) pathway in the adipose, liver, and kidney tissues. However, the role of cytosolic mtDNA in the progression of DCM is unclear. In this study, with an obesity-related DCM mouse model established by feeding db/db mice with a high-fat diet (HFD), we observed increased mtDNA in the cytosol and activated cGAS-STING signaling pathway during DCM, as well as the downstream targets, IRF3, NF-κB, IL-18, and IL-1β. In a further study with a palmitic acid (PA)-induced lipotoxic cell model established in H9C2 cells, we revealed that the cytosolic mtDNA was the result of PA-induced overproduction of mitochondrial ROS, which also led to the activation of the cGAS/STING system and its downstream targets. Notably, treatment of extracted mtDNA alone was sufficient to activate the cGAS-STING signaling pathway in cultured H9C2 cells. Besides, both knockdown of STING in PA-induced H9C2 cells and inhibition of STING by C-176 injection in the DCM mouse model could remarkably block the inflammation and apoptosis of cardiomyocytes. In conclusion, our study elucidated the critical role of cytosolic mtDNA-induced cGAS-STING activation in the pathogenesis of obesity-related DCM and provided preclinical validation for using a STING inhibitor as a new potential therapeutic strategy for the treatment of DCM.Graphical abstract

Biological Functions and Clinical Prospects of Extracellular Non-Coding RNAs in Diabetic Cardiomyopathy: an Updated Review.

Diabetic cardiomyopathy (DCM) is one of the major causes of heart failure in diabetic patients. However, the pathogenesis of diabetic cardiomyopathy has not been fully elucidated. Diagnosis and therapeutic strategy of DCM is still challenging. Various non-coding RNAs (ncRNA) are implicated in the onset and progression of DCM. Interestingly, ncRNAs not only are regulators intracellularly, but also can exist and function in extracellular space. Recent evidences have demonstrated that extracellular ncRNAs play emerging roles in both intracardiac and inter-organ communication during the pathogenesis of DCM; thus, extracellular ncRNAs are attractive diagnostic biomarkers and potential therapeutic targets for DCM. This article will review the current knowledge of the roles of extracellular ncRNAs in DCM, especially focusing on their physio-pathological properties and perspectives of potential clinical translation for biomarkers and therapies. Recent evidences have demonstrated that extracellular ncRNA play emerging roles in both intracardiac and inter-organ communication involved in the pathogenesis of diabetic cardiomyopathy (DCM), thus shown as attractive diagnostic biomarkers and potential therapeutics for DCM. In the current review, we first summarize the progress regarding the paracrine role of extracellular ncRNA in DCM. miRNAs and circRNAs have been shown to mediate the communication among cardiomyocytes, endothelial cells, and vascular smooth muscle cells in the diabetic heart. Subsequently, we systematically describe that extracellular ncRNAs contribute to the crosstalk between the heart and other organs in the context of diabetes. Researches have indicated that miRNAs acted as hepatokines and adipokines to mediates the injure effect of distal organs on hearts. As for clinical application, extracellular ncRNAs are promising biomarker and have therapeutic potential. (Created with BioRender.com).

Myeloid differential protein-2 inhibition improves diabetic cardiomyopathy via p38MAPK inhibition and AMPK pathway activation

Myeloid differential protein-2 (MD2) has been shown to play a critical role in the progression of diabetic cardiomyopathy (DCM). This study aims to explore the non-inflammatory mechanisms mediated by MD2 in DCM and to test the therapeutic effects of MD2 inhibitor C30 on DCM. Streptozotocin (STZ) was used to construct DCM model in wild-type and MD2 knockout mice. The collected heart samples were subjected to RNA-sequencing assay. Gene set enrichment analysis of the RNA-seq data indicated that MD2 knockout was associated with energy metabolism pathways in diabetic mouse heart. Further data showed that AMPK pathway was impaired under high glucose condition, which was mediated by p38MAPK activation. MD2 knockout or pharmacological inhibitor C30 completely rescued AMPK signaling through p38MAPK inhibition. Importantly, C30 treatment significantly prevented myocardial damage and dysfunction in T1DM mice evidenced by improved cardiac function and reduced cardiomyocyte apoptosis and cardiac fibrosis. Furthermore, the therapeutic effect of C30 on DCM was correlated to p38MAPK inhibition and AMPK pathway activation in vivo and in vitro. In conclusion, MD2 inhibition exhibits therapeutic effects on DCM through p38MAPK inhibition and AMPK activation, which enables MD2 a promising target for DCM treatment by suppressing metaflammation and improving cardiac metabolism.

Inhibition of Sestrin2 overexpression in diabetic cardiomyopathy ameliorates cardiac injury via restoration of mitochondrial function.

Mitochondrial dysfunction-induced apoptosis plays a crucial role in the progression of diabetic cardiomyopathy (DCM). Sestrin2 is an important oxidative stress response protein and is involved in the maintenance of mitochondrial function, especially under stress. The aim of the present study was to investigate the role of Sestrin2 in DCM and to explore the underlying mechanisms. H9c2 cardiomyocytes were induced with high glucose (HG) medium (33 mmol/l glucose) for an in vitro DCM model. C57BL/6 mice were induced for the in vivo DCM model by intraperitoneal streptozotocin injection. H9c2 cardiomyocytes were exposed to HG and infected with lentiviruses to express Sestrin2 short hairpin RNA (shRNA). The study found that cell viability and mitochondrial function were impaired while cell apoptosis and oxidative stress were increased in DCM. Sestrin2 was significantly upregulated in myocardial tissues of DCM mice and H9c2 cardiomyocytes in HG conditions. Downregulation of Sestrin2 increased cell viability, decreased cell apoptosis, and attenuated oxidative stress in H9c2 cells exposed to HG. Moreover, HG-induced mitochondrial injury was alleviated by Sestrin2 silencing. In conclusion, our finding indicated that the inhibition of enhanced Sestrin2 expression ameliorates cardiac injury in DCM, which might be largely attributed to the restoration of mitochondrial function.

CircularRNA circ_0071269 knockdown protects against from diabetic cardiomyopathy injury by microRNA-145/gasdermin A axis

ABSTRACT Circular RNAs (circRNAs) are involved in the development and progression of diabetic cardiomyopathy (DCM). However, the specific function and underlying mechanism of circ_0071269 in DCM remains unclear. In our study, mRNA and miRNA expression was detected by real-time quantitative PCR (qRT-PCR). RNase R and actinomycin D treatment were applied to test the characteristics of circ_0071269. Cell Counting Kit-8 (CCK-8) assay, lactate dehydrogenase (LDH) and enzyme-linked immunosorbent assay (ELISA) kits were performed to determine the cell viability, cell LDH content and interleukin (IL)-1β and IL-18 levels, respectively. Cell death rate was determined by Flow cytometry, and Western blotting was for the protein expression levels. In addition, luciferase reporter and RNA pull-down assays were performed to confirm the binding relationship between miR-145 and circ_0071269 or gasdermin A (GSDMA). Echocardiography, Hematoxylin and Eosin (HE) Staining, and Immunohistochemical (IHC) Staining were performed to evaluate myocardial damage in vivo. We found that circ_0071269 was significantly overexpressed in H9c2 cells upon treatment with high glucose. Knockdown of circ_0071269 promoted cell viability and inhibited the inflammatory response, cytotoxicity, and pyroptosis of H9c2 cells in vitro. Moreover, circ_0071269 sponges miR-145 to upregulate GSDMA. A miR-145 inhibitor antagonized the effects of circ_0071269 knockdown on the cellular functions of H9c2 cells, while the effects of miR-145 were abrogated by the overexpression of GSDMA. Meanwhile, knockdown of circ_0071269 attenuated cardiac dysfunction of DM mice. Hence, circ_0071269 may promote the development of DCM through the miR-145/GSDMA axis and thus provide a novel marker for the treatment of DCM.

METTL14 suppresses pyroptosis and diabetic cardiomyopathy by downregulating TINCR lncRNA

N6-methyladenosine (m6A) is one of the most important epigenetic regulation of RNAs, such as lncRNAs. However, the underlying regulatory mechanism of m6A in diabetic cardiomyopathy (DCM) is very limited. In this study, we sought to define the role of METTL14-mediated m6A modification in pyroptosis and DCM progression. DCM rat model was established and qRT-PCR, western blot, and immunohistochemistry (IHC) were used to detect the expression of METTL14 and TINCR. Gain-and-loss functional experiments were performed to define the role of METTL14-TINCR-NLRP3 axis in pyroptosis and DCM. RNA pulldown and RNA immunoprecipitation (RIP) assays were carried out to verify the underlying interaction. Our results showed that pyroptosis was tightly involved in DCM progression. METTL14 was downregulated in cardiomyocytes and hear tissues of DCM rat tissues. Functionally, METTL14 suppressed pyroptosis and DCM via downregulating lncRNA TINCR, which further decreased the expression of key pyroptosis-related protein, NLRP3. Mechanistically, METTL14 increased m6A methylation level of TINCR gene, resulting in its downregulation. Moreover, the m6A reader protein YTHDF2 was essential for m6A methylation and mediated the degradation of TINCR. Finally, TINCR positively regulated NLRP3 by increasing its mRNA stability. To conclude, our work revealed the novel role of METTL14-mediated m6A methylation and lncRNA regulation in pyroptosis and DCM, which could help extend our understanding the epigenetic regulation of pyroptosis in DCM progression.

Mitochondrial quality control in diabetic cardiomyopathy: from molecular mechanisms to therapeutic strategies.

In diabetic cardiomyopathy (DCM), a major diabetic complication, the myocardium is structurally and functionally altered without evidence of coronary artery disease, hypertension or valvular disease. Although numerous anti-diabetic drugs have been applied clinically, specific medicines to prevent DCM progression are unavailable, so the prognosis of DCM remains poor. Mitochondrial ATP production maintains the energetic requirements of cardiomyocytes, whereas mitochondrial dysfunction can induce or aggravate DCM by promoting oxidative stress, dysregulated calcium homeostasis, metabolic reprogramming, abnormal intracellular signaling and mitochondrial apoptosis in cardiomyocytes. In response to mitochondrial dysfunction, the mitochondrial quality control (MQC) system (including mitochondrial fission, fusion, and mitophagy) is activated to repair damaged mitochondria. Physiological mitochondrial fission fragments the network to isolate damaged mitochondria. Mitophagy then allows dysfunctional mitochondria to be engulfed by autophagosomes and degraded in lysosomes. However, abnormal MQC results in excessive mitochondrial fission, impaired mitochondrial fusion and delayed mitophagy, causing fragmented mitochondria to accumulate in cardiomyocytes. In this review, we summarize the molecular mechanisms of MQC and discuss how pathological MQC contributes to DCM development. We then present promising therapeutic approaches to improve MQC and prevent DCM progression.

Association of Fibroblast Growth Factor 21 with Lipid Profile, MDA and AOPP in Patients with Type 2 Diabetes Mellitus

Background: Diabetes mellitus is rising all over the world due to population growth, aging, urbanisation, and the increase of obesity due to physical inactivity, characterized by persistent high blood glucose levels associated with aberrations in lipid, carbohydrate, and protein metabolisms leading to water and electrolyte imbalance. Cardiovascular diseases are the leading causes of mortality in diabetic patients. Mechanisms such as oxidative stress, lipid metabolism imbalance, as well as myocardial cell apoptosis are key factors to facilitate the progression of Diabetic cardiomyopathy.
 Aim: The aim of this study was to assess FGF-21 levels and their association with lipid profile parameters and oxidative stress in patients with type 2 diabetes mellitus.
 Methods: A patient based cross-sectional study was conducted among the subjects with history of type 2 DM for the past 10 years.
 Results: Variations in FBS, T.C, TG, LDL, HDL, VLDL, FGF-21, MDA and AOPP levels among cases and controls were depicted in Table 2. There was an increase in all these parameters in cases compared to controls whereas HDL showed a decrease among cases.
 Conclusion: Our study concluded that there is a significant correlation between fibroblast growth factor 21 (FGF-21), oxidative stress, and abnormal lipid profile in type 2 diabetic patients. We would recommend further studies to explore the role of FGF21 as an important marker in predicting cardiovascular risk in diabetic patients.

In vitro and in vivo studies reveal the beneficial effects of chlorogenic acid against ER stress mediated ER-phagy and associated apoptosis in the heart of diabetic rat

Endoplasmic reticulum (ER) and associated signaling pathways are involved in diabetic cardiomyopathy (DCM) however, detailed studies are not available. The present study investigated the role of ER stress and related pathways such as ER-phagy, apoptosis and their underlying mechanisms using appropriate models. Beneficial effect of chlorogenic acid was also evaluated against ER stress mediated DCM. H9c2 cells with high glucose (33 mM, in vitro model of hyperglycemia) showed significant activation of ER stress response (GRP78, PERK, IRE1α, ATF6α) and altered its regulatory proteins (PDI, ERO1α). Also, it enhanced ER-phagy through upregulation of Sec62, RTN3 and downregulation of FAM134B. High glucose caused apoptosis via increased levels of CHOP, caspase 12 and calnexin. All these proteins (PERK, IRE1α, ATF6α, RTN3, Sec62 and FAM134B) have been found to have a significant role in the functioning of heart such as excitation contraction coupling and we expect these alterations to induce cardiomyopathy during diabetes. This was confirmed in in vivo study too. High fat, high fructose diet with mild streptozotocin induced diabetic rats showed an increased expression of BNP confirming cardiac injury. We also noticed severe ER stress in the heart of diabetic animals. All these have contributed significantly into alterations in histopathology and increase of weight of the hearts. These findings clearly show that ER stress plays a vital protagonist in the progression of DCM. We also found chlorogenic acid is effective against hyperglycemia induced pathological alteration both in vitro as well as in vivo.

Abstract 11457: Overexpression of Thioredoxin-1 Ameliorates the Progression of Diabetic Cardiomyopathy by Preservation of Cardiac Function, Increased Survivability and Decreased Fibrosis in Aged Transgenic Mice

Introduction: Increased production of reactive oxygen species contributes to the etiology of diabetic complications. It acts as an intracellular antioxidant by scavenging reactive oxygen species (ROS) and protect the cells against oxidative stress and cell death. Hypothesis: To determine the long-term effect of streptozotocin (STZ) induced type 1 diabetes on aging Trx-1 transgenic mice related to survivability, heart function, fibrosis, and apoptosis up to 180 days (d) compared to its corresponding wild-type (WT C57Bl/6J). Method: WT and Trx-1 overexpressed mice (Trx-1 Tg/+ ) (8-12 weeks old) were injected with STZ (i.p. 50mg/kg.body weight(bw)-male; 75mg/kg.bw-female for 5 consecutive days) for induction of diabetes (Dia). Fasting blood sugar level (FBS) was measured 30d after the final injection, and FBS >220mg/dL is used as the cut-off for diabetes. Echocardiography, Survival, FBS measurements were performed on 30, 60, 90, 120, 150, and 180d. Immunohistochemistry was performed on heart tissues (30, 90, and 180d). Results: Dia-Trx-1 Tg/+ mice maintained EF at 60d (61.14±2.3%), 120d (56.1±1.94%) and 180d (60.8±4.3%), when compared to 30d (64.4±1.94%) representing preserved cardiac functions. On the other hand, Dia-WT mice show significant progressive depression of EF at 60d (53.8±1.9%), 120d (48.8±2.1%), and 180d (48.9±1.5%), (p<0.05) when compared to 30d (68.4±6.1%) Dia-WT. Moreover, Dia-Trx-1 Tg/+ mice show significantly higher EF than their corresponding Dia-WT counterparts at each time point measured after diabetic induction (n=8-12, p<0.05). Dia-Trx-1 Tg/+ mice (n=71) also exhibited increased survival rate over Dia-WT (n=47) counterparts with a median survival of 150 days compared to 60 days (Log Rank test: p=0.004). Dia-Trx-1 Tg/+ mice also showed a significant reduction in cardiac fibrosis (4.18±1.08% vs. 17.5±5.1%; p=0.03, n=4) and TUNEL positive cells (3.81±1.49 vs. 7.66±1.46; p=0.03, n=4) when compared to their counterpart (Dia-WT) after 180d of diabetes. Conclusion: These results show that Trx-1 Tg/+ mice preserved cardiac functions, increased survival, decreased apoptosis and fibrosis up to 180d under diabetic conditions, and hence suggesting Trx-1 based treatment strategy could prevent heart failure in aging.

Abstract 9022: Chronic Beta3-Adrenergic Receptor Antagonist Therapy Rescues Diabetic Cardiomyopathy Through Ca 2+ /Calmodulin-Dependent Protein Kinase II Inhibition in a Type II Diabetes Murine Model

Background: Recent evidence showed that CaMKII inhibition with KN93 protects against diabetic cardiomyopathy (DCM). However, the off-target effects have limited its use. There are no CaMKII inhibitors available clinically. It may be best to use some complementary approaches for CaMKII inhibition. Previously we showed that diabetes produced upregulation of cardiac β 3 -AR-mediated inhibitory pathway promoting DCM progression. There is a significant crosstalk between β-AR signaling and CaMKII activation presenting CaMKII as a possible downstream mediator of detrimental β 3 -AR signaling in DCM. It is possible that the benefit of β 3 -AR antagonists (β 3 ANT) might involve modulation of CaMKII. We tested the hypothesis that chronic L-748-337, a selective β 3 ANT may inhibit CaMKII activity and limit DCM. Methods: LV myocyte CaMKIIδ expression and activity and myocyte function were determined in 3 groups (7/group) wild-type female mice over 14 weeks (W): Type 2 DM ( T2 ), 14 W fed high-fat diet (HFD), but after HFD for 4 W receiving streptozotocin (STZ, 40 mg/kg/day, i.p. 5 days); T2β 3 ANT , T2 mice at 10 W received L-748,337 (10 -7 M/kg/day, mini-pump) for 4 W; and Vehicle controls (C). Results: Versus C, T2 diabetes was induced in mice received HFD and low dose STZ with significantly elevated blood glucose. But only T2 myocytes had significantly increased protein levels of CaMKIIδ (T2: 29%, 0.53 vs C: 0.41) and CaMKIIδ phosphorylation (at Thr287) (PCaMKIIδ) (79%, 0.52 vs 0.29) followed by significantly reduced cell contraction (dL/dt max , 75 vs 134 μm/s), relaxation (dR/dt max , 60 vs 114 μ m/s) and [Ca 2+ ] iT (0.16 vs 0.21). The isoproterenol (ISO, 10 -8 M)-stimulated increases in dL/dt max (38% vs 58%), dR/dt max and [Ca 2+ ] iT (19% vs 30%) were also significantly reduced. By contrary, versus C, T2β 3 ANT myocytes had similar levels of CaMKIIδ (0.37) and PCaMKIIδ (0.31) with preserved cell contractility (131 μm/s), relaxation (109 μm/s) and [Ca 2+ ] iT (0.21). ISO caused increases in dL/dt max (57%), dR/dt max (52%) and [Ca 2+ ] iT (30%) were also closed to control values. Conclusions: Chronic β 3 ANT prevents Type II diabetes-caused upregulation of cardiac CaMKIIδ, thereby leading to the preservation of normal myocyte function, [Ca 2+ ] iT , and β-adrenergic reserve in a DCM murine model.

Therapeutic Approach of Flavonoid in Ameliorating Diabetic Cardiomyopathy by Targeting Mitochondrial-Induced Oxidative Stress.

Diabetes cardiomyopathy is one of the key factors of mortality among diabetic patients around the globe. One of the prior contributors to the progression of diabetic cardiomyopathy is cardiac mitochondrial dysfunction. The cardiac mitochondrial dysfunction can induce oxidative stress in cardiomyocytes and was found to be the cause of majority of the heart morphological and dynamical changes in diabetic cardiomyopathy. To slow down the occurrence of diabetic cardiomyopathy, it is crucial to discover therapeutic agents that target mitochondrial-induced oxidative stress. Flavonoid is a plentiful phytochemical in plants that shows a wide range of biological actions against human diseases. Flavonoids have been extensively documented for their ability to protect the heart from diabetic cardiomyopathy. Flavonoids’ ability to alleviate diabetic cardiomyopathy is primarily attributed to their antioxidant properties. In this review, we present the mechanisms involved in flavonoid therapies in ameliorating mitochondrial-induced oxidative stress in diabetic cardiomyopathy.

Soluble Klotho-integrin β1/ERK1/2 pathway ameliorates myocardial fibrosis in diabetic cardiomyopathy.

Soluble Klotho (sKL) is closely related to insulin resistance, which is a major factor in the progression of diabetic cardiomyopathy (DCM). The purpose of this study was to investigate the role of sKL in the regulation of DCM and the mechanism involved. A mouse model of type 2 diabetes was induced by high-fat diet and streptozotocin injection. An insulin-resistant cardiac fibroblast model was established by high glucose and high insulin. KL gene overexpression was achieved in vivo and vitro through transfection with an adenovirus-harboring KL-cDNA. Gene overexpression was used to evaluate the role of sKL in the pathophysiologic characteristics of DCM. Insulin-resistant cardiac fibroblasts reduced sKL expression and collagen deposition. Diabetic mice constructed by streptozotocin exhibited severe insulin resistance, inflammation, fibrosis, left ventricular dysfunction, and sKL downregulation. The overexpression of sKL mitigated insulin resistance and metabolic disturbance; inflammation, fibrosis, and upregulated collagen I/III content ratio in diabetic state were significantly reduced. Our findings were accompanied by notable moderation of cardiac function. Further, blunted phosphorylation of Akt was restored with sKL gene overexpression, and activated phosphorylation of extracellular signal-regulated kinase 1/2 in DCM was reduced. Our results suggest that sKL protein overexpression exerts a defensive measure by ameliorating selective insulin resistance in mouse DCM, thus revealing its underlying mechanism for potential human DCM treatment.

Abstract P319: SARM1 NAD Hydrolase Deficiency Protects Hearts Against Diabetic Cardiomyopathy

NAD depletion is associated with the pathogenesis of diseases such as heart failure. Strategies to replenish cellular NAD levels by activating NAD synthesis pathways have shown promises to treat heart disease. However, how NAD consumption mechanisms lead to NAD depletion are less understood. SARM1 is a novel intracellular NAD hydrolase that degrades NAD and promotes axonal degeneration in neurons. We recently showed that NAD redox imbalance and depletion promote the progression of diabetic cardiomyopathy. Therefore, we hypothesized that SARM1 deficiency might protect hearts against diabetic cardiomyopathy. 16-week diabetic stress initiated by streptozotocin (STZ) injections was applied to wild-type C57BL/6 (WT) and whole-body SARM1-KO mice. Cardiac function was measured after 8-week or 16-week of diabetic stress, and cardiac tissue and plasma samples from these mice were harvested after 16 weeks of diabetes. SARM1 mRNA or protein levels were suppressed in SARM1-KO hearts. STZ induced similar hyperglycemia (~600 mg/dL) in both WT and SARM1-KO male mice after 16 weeks. Chronic diabetic stress led to progressive decline in systolic function (Baseline fractional shortening: 50%; 16-week diabetes: 35%; P<0.05; n=5) in WT male mice, which was ameliorated in SARM1-KO male mice (16-week diabetes: 48%; P<0.05; n=5). Progressive decline in diastolic function induced by chronic diabetes (Baseline WT E’/A’: 1.53; WT 16-week diabetes: 1.12; n=5) was also improved in diabetic SARM1-KO mice (KO E’/A’ ratio at 16-week diabetes: 1.5; P<0.05; n=5). Heart weights were similar in diabetic WT or diabetic SARM1-KO hearts. A similar study is on-going in a female cohort. Despite loss of SARM1 expression, no compensatory changes in expressions of other NAD hydrolases (i.e. Cd38 or Bst1) were observed in diabetic SARM1-KO hearts, while expression levels of genes related to NAD consumption and synthesis pathways were mostly unchanged except Qprt and Haao. Cardiac fibrosis was induced in diabetic WT hearts and were suppressed in diabetic SARM1-KO hearts after 16-week diabetic stress, but these changes were not observed in tissues harvested after 8-week diabetic stress. The results suggest fibrosis is a later event in the progression of diabetic cardiomyopathy. WT and SARM1-KO mice have also been challenged with high fat diet feeding (HFD) for 16 weeks, and increased fasting glucose levels and body weights were similarly observed in HFD-WT and HFD-SARM1-KO mice. Longitudinal cardiac function analyses are on-going. Our data thus far support the protective role of SARM1 deficiency in metabolic stress-induced cardiomyopathy, while pathogenic mechanisms of SARM1 in diabetic hearts remain to be determined.

High glucose‑induced upregulation of CD36 promotes inflammation stress via NF‑κB in H9c2 cells.

Cardiac inflammation serves an important role in the progression of diabetic cardiomyopathy. CD36 (cluster of differentiation 36) mediates inflammation stress in a variety of disease states. The present study investigated CD36 expression in high glucose (HG)-induced H9c2 cells, whether CD36 upregulation promotes inflammatory stress, and its potential mechanism. HG induced CD36 expression in a time-dependent manner in cells, which was blocked following CD36 knockout or treatment with N-acetylcysteine or MitoTEMPO. CD36 translocation to the cell membrane was increased at 72 h by HG stimulation of H9c2 cells. Moreover, CD36 knockout inhibited HG-induced reactive oxygen species (ROS) generation, tumor necrosis factor-α, interleukin (IL)-6 and IL-1β expression, and nuclear factor (NF)-κB pathway activation. Further, CD36 knockout reversed metabolic reprogramming, lipid accumulation and AMP-activated protein kinase activation caused by HG. The aforementioned data suggest that HG-induced upregulation of CD36 promotes inflammatory stress via NF-κB in H9c2 cells, mediated by metabolism reprogramming, lipid accumulation and enhanced ROS generation.

Abstract MP211: NAD Redox Imbalance Exacerbates Diabetic Cardiomyopathy

Diabetes and heart failure are linked to NAD redox imbalance, whose role in diabetic cardiomyopathy has not been directly tested. Streptozotocin-induced diabetes in WT mice for 16 weeks promoted declines in systolic and diastolic function, which associated with lowered cardiac NAD/NADH ratio (NAD redox imbalance). To test the hypothesis that , we employed mouse models with cardiac-specific manipulations of NAD redox states. Cardiac-specific Ndufs4-KO mice (cKO) exhibit lowered cardiac NAD/NADH ratio with normal baseline function, geometry and energetics. Control and cKO mice were challenged with 8-week diabetic stress. Metabolomic analyses of plasma collected after the diabetic stress showed similar hyperglycemia and dyslipidemia stresses in diabetic control and diabetic cKO mice. Chronic diabetic stress promoted systolic and diastolic dysfunctions in control mice, which were further exacerbated in diabetic cKO mice in both male and female cohorts. Collagen levels and transcript analyses of fibrosis and extracellular matrix-dependent pathways showed no change in diabetic cKO hearts, suggesting that cardiomyocyte dysfunction is a likely culprit for the exacerbated dysfunction. Increased protein acetylation, including SOD2-K68Ac, was observed in diabetic cKO hearts. Inhibited antioxidant function by SOD2-K68Ac promoted protein oxidation in diabetic cKO hearts, suggesting oxidative stress as a pathogenic mechanism. We next examined phosphorylation status of myofilament proteins in these diabetic hearts. MyBPC-S282Pi levels are suppressed in failing hearts and remained unchanged in diabetic cKO hearts. TnI-S150Pi increases myofilament calcium sensitivity and prolongs calcium dissociation, while TnI-S23/24Pi imposes the opposite effects. TnI-S150Pi levels were elevated in diabetic cKO hearts, while TnI-S23/24Pi levels unchanged. Therefore, exacerbated diastolic dysfunction in diabetic cKO hearts is due to the selective phosphorylation at TnI-S150. AMPK is activated by energetic stress and phosphorylates TnI-S150. ATP levels decreased, and AMP/ATP ratio increased in diabetic cKO hearts, implicating impaired energetics to promote TnI-S150Pi and dysfunction. Elevation of NAD levels normalized cardiac NAD redox balance in diabetic cKO hearts. Elevated levels of SOD2-K68Ac and TnI-S150Pi, exacerbated systolic and diastolic dysfunction in diabetic cKO hearts were all reversed by elevation of NAD levels. Dysfunction in diabetic control hearts was also ameliorated by elevation of NAD levels. These data collectively conclude that NAD redox imbalance is a positive mediator of the progression of diabetic cardiomyopathy.