Myocardial Injury Caused by Chronic Alcohol Exposure-A Pilot Study Based on Proteomics.

Chronic alcohol exposure can cause myocardial degenerative diseases, manifested as cardiac insufficiency, arrhythmia, etc. These are defined as alcoholic cardiomyopathy (ACM). Alcohol-mediated myocardial injury has previously been studied through metabolomics, and it has been proved to be involved in the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway concerning unsaturated fatty acids biosynthesis and oxidative phosphorylation, which tentatively explored the mechanism of ACM induced by chronic drinking. To further study alcohol-induced myocardial injury, myocardial specimens from a previously successfully established mouse model of ACM were subjected to histological, echocardiographic, and proteomic analyses, and validated by real-time quantitative polymerase chain reaction (qPCR). Results of histopathology and echocardiography showed the hypertrophy of cardiomyocytes, the dilation of ventricles, and decreased cardiac function. Proteomic results, available via ProteomeXchange with identifier PXD032949, revealed 56 differentially expressed proteins (DEPs) were identified, which have the potential to be involved in the KEGG pathway related to fatty acid biosynthesis disorders, lipid metabolism disorders, oxidative stress, and, ultimately, in the development of dilated cardiomyopathy (DCM). The present study further elucidates the underlying effects of myocardial injury due to chronic alcohol intake, laying a foundation for further studies to clarify the potential mechanisms of ACM.

Polydatin attenuates chronic alcohol consumption-induced cardiomyopathy through a SIRT6-dependent mechanism.

Polydatin has attracted much attention as a potential cardioprotective agent against ischemic heart disease and diabetic cardiomyopathy. However, the effect and mechanism of polydatin supplementation on alcoholic cardiomyopathy (ACM) are still unknown. This study aimed to determine the therapeutic effect of polydatin against ACM and to explore the molecular mechanisms with a focus on SIRT6-AMP-activated protein kinase (AMPK) signaling and mitochondrial function. The ACM model was established by feeding C57/BL6 mice with an ethanol Lieber-DeCarli diet for 12 weeks. The mice received polydatin (20 mg kg-1) or vehicle treatment. We showed that polydatin treatment not only improved cardiac function but also reduced myocardial fibrosis and dynamin-related protein 1 (Drp-1)-mediated mitochondrial fission, and enhanced PTEN-induced putative kinase 1 (PINK1)-Parkin-dependent mitophagy in alcohol-treated myocardium. Importantly, these beneficial effects were mimicked by SIRT6 overexpression but abolished by the infection of recombinant serotype 9 adeno-associated virus (AAV9) carrying SIRT6-specific small hairpin RNA. Mechanistically, alcohol consumption induced a gradual decrease in the myocardial SIRT6 level, while polydatin effectively activated SIRT6-AMPK signaling and modulated mitochondrial dynamics and mitophagy, thus reducing oxidative stress damage and preserving mitochondrial function. In summary, these data present new information regarding the therapeutic actions of polydatin, suggesting that the activation of SIRT6 signaling may represent a new approach for tackling ACM-related cardiac dysfunction.

Ectopic accumulation of ceramide in cardiomyocytes modulates alcoholic cardiomyopathy via the TLR4-dependent pathway.

BACKGROUND AND AIMS: Excessive alcohol consumption predisposes drinkers to develop alcoholic cardiomyopathy. Although cardiomyocyte loss is the hallmark of cardiomyopathy, the underlying mechanism remains elusive. This study examined the potential mechanism of alcohol-induced cardiomyocyte death in a mouse model of alcoholic cardiomyopathy. METHODS: We established the alcoholic cardiomyopathy mouse model using C57BL/6J mice and confirmed it via echocardiography and histological examination. The cardiac ceramide content and profile were analyzed with a triple-quadrupole mass spectrometer. The molecular mechanism underlying the accumulation of ceramide due to chronic alcohol consumption and ceramide-induced cardiomyocyte death were investigated by in vivo and in vitro models. Finally, we established a TLR4 mutation model to explore the function of TLR4 in CH3/HeJ mice. RESULTS: Cardiac lipotoxicity that followed alcohol exposure resulted mainly in C16:0-, C18:0-, and C24:1-ceramide aggregation. Genes encoding the sphingosine hydrolysis enzymes (SMPD1 and SMPD2) rather than de novo synthetic biomarkers were markedly upregulated. Exogenous ceramide mimics (C6-ceramide) werenderlying the accumulation of ceramide observed to cause H9C2 cardiomyocyte-like cell death, which was consistent with results under palmate acid (PA) treatment. As a ceramide precursor, PA induces intracellular ceramide generation through TLR4 signaling, which can be abolished by an inhibitor of ceramide synthesis. Furthermore, mechanistic investigations demonstrated that pharmacological or genetic inhibition of TLR4 attenuated PA-induced cell death and corresponding ceramide production. Moreover, global mutation of TLR4 in CH3/HeJ mice significantly reduced the accumulation of C24:0, C24:1, OH_C24:1, and total ceramide following alcohol challenge. CONCLUSIONS: Our findings demonstrate that ceramide accumulation plays a crucial role in alcoholic cardiomyopathy, effects that are partially mediated through the TLR4-dependent pathway.

Characterization of myocardial oxidative metabolism and myocardial external efficiency in high-risk alcohol cardiotoxicity and alcoholic cardiomyopathy via dynamic 11C-Acetate positron emission tomography.

INTRODUCTION: The purpose of this study was to evaluate subjects with high-risk alcohol cardiotoxicity and patients with alcoholic cardiomyopathy (ACM) via dynamic 11C-Acetate positron emission tomography (PET) imaging as a myocardial oxidative metabolic probe. METHODS AND RESULTS: We recruited 37 subjects with chronic alcohol consumption [18 with moderate consumption (MC), 19 with heavy consumption (HC)], 5 ACM patients, and 12 healthy controls to receive dynamic 11C-Acetate PET scans. PET imaging data were analyzed to calculate kinetic parameters (e.g., Kmono, K1 and k2) based on the mono-exponential and one-tissue compartmental models. Myocardial oxygen consumption (MVO2) and myocardial external efficiency (MEE) were then derived from these kinetic parameters. MVO2 was significantly lowered in the HC group and in ACM patients (0.121± 0.018 and 0.111 ± 0.017 mL·g-1·min-1, respectively) compared with those in healthy controls and MC subjects (0.144 ± 0.023 and 0.146 ± 0.027 mL·g-1·min-1, respectively; P < .01). MEE was significantly reduced in ACM patients (13.0% ± 4.3%) compared with those of healthy controls (22.4% ± 4.6%, P < .01), MC subjects (20.1% ± 4.5%, P < .05), and HC subjects (22.3% ± 4.5%, P < .001). CONCLUSION: Functional assessment via dynamic 11C-Acetate PET imaging may represent a clinically feasible probe for identifying cohorts with high-risk cardiotoxicity due to addictive alcohol consumption and ACM.

Epigenetic modification in alcohol use disorder and alcoholic cardiomyopathy: From pathophysiology to therapeutic opportunities.

Alcohol consumption prompts detrimental psychological, pathophysiological and health issues, representing one of the major causes of death worldwide. Alcohol use disorder (AUD), which is characterized by compulsive alcohol intake and loss of control over alcohol usage, arises from a complex interplay between genetic and environmental factors. More importantly, long-term abuse of alcohol is often tied with unfavorable cardiac remodeling and contractile alterations, a cadre of cardiac responses collectively known as alcoholic cardiomyopathy (ACM). Recent evidence has denoted a pivotal role for ethanol-triggered epigenetic modifications, the interface between genome and environmental cues, in the organismal and cellular responses to ethanol exposure. To-date, three major epigenetic mechanisms (DNA methylation, histone modifications, and RNA-based mechanisms) have been identified for the onset and development of AUD and ACM. Importantly, these epigenetic changes induced by alcohol may be detectable in the blood, thus offering diagnostic, therapeutic, and prognostic promises of epigenetic markers for AUD and alcoholic complications. In addition, several epigenetic drugs have shown efficacies in the management of alcohol abuse, loss of control for alcohol usage, relapse, drinking-related anxiety and behavior in withdrawal. In this context, medications targeting epigenetic modifications may hold promises for pharmaceutical management of AUD and ACM.

On the Mechanism of Cardioprotective Effect of Fabomotizole in Alcoholic Cardiomyopathy.

The molecular mechanisms underlying the cardioprotective effect of fabomotizole were studied using the translational rat model of alcoholic cardiomyopathy developed by us. It was shown that intraperitoneal administration of fabomotizole (15 mg/kg) for 28 days to animals with alcoholic cardiomyopathy contributes to normalization of the expression of mRNA of genes of regulatory proteins СаМ (p=0.00001), Ерас1 (p=0.021), and Ерас2 (p=0.018) and receptors RyR2 (p=0.0031) and IP3R2 (p=0.006) in the myocardium of the myocardium of the left ventricle that is enhanced in control animals (p<0.05). These changes were accompanied by echocardiographically documented decrease in the degree of left ventricle remodeling and improvement of its inotropic function.

A Pilot Metabolomic Study on Myocardial Injury Caused by Chronic Alcohol Consumption-Alcoholic Cardiomyopathy.

Chronic alcohol consumption leads to myocardial injury, ventricle dilation, and cardiac dysfunction, which is defined as alcoholic cardiomyopathy (ACM). To explore the induced myocardial injury and underlying mechanism of ACM, the Liber-DeCarli liquid diet was used to establish an animal model of ACM and histopathology, echocardiography, molecular biology, and metabolomics were employed. Hematoxylin-eosin and Masson's trichrome staining revealed disordered myocardial structure and local fibrosis in the ACM group. Echocardiography revealed thinning wall and dilation of the left ventricle and decreased cardiac function in the ACM group, with increased serum levels of brain natriuretic peptide (BNP) and expression of myocardial BNP mRNA measured through enzyme-linked immunosorbent assay and real-time quantitative polymerase chain reaction (PCR), respectively. Through metabolomic analysis of myocardium specimens, 297 differentially expressed metabolites were identified which were involved in KEGG pathways related to the biosynthesis of unsaturated fatty acids, vitamin digestion and absorption, oxidative phosphorylation, pentose phosphate, and purine and pyrimidine metabolism. The present study demonstrated chronic alcohol consumption caused disordered cardiomyocyte structure, thinning and dilation of the left ventricle, and decreased cardiac function. Metabolomic analysis of myocardium specimens and KEGG enrichment analysis further demonstrated that several differentially expressed metabolites and pathways were involved in the ACM group, which suggests potential causes of myocardial injury due to chronic alcohol exposure and provides insight for further research elucidating the underlying mechanisms of ACM.

Astaxanthin attenuates alcoholic cardiomyopathy via inhibition of endoplasmic reticulum stress-mediated cardiac apoptosis.

Chronic excessive ethanol consumption is associated with a high incidence of mortality due to ethanol-induced dilated cardiomyopathy, known as alcoholic cardiomyopathy (ACM). Mechanistic studies have demonstrated that apoptosis is key to the pathogenesis of ACM, and endoplasmic reticulum (ER) stress-associated apoptosis contributes to various ethanol-related diseases. Astaxanthin (AST) is a natural carotenoid that exerts an anti-ER stress effect. Importantly, strong evidence has shown that AST induces beneficial effects in various cardiovascular diseases. The present study aimed to investigate whether AST induces beneficial effects on ACM by suppressing cardiac apoptosis mediated by ER stress. We showed that after 2 months of chronic excessive ethanol consumption, mice displayed obvious cardiac dysfunction and morphological changes associated with increased fibrosis, oxidative stress, ER stress and apoptosis. However, cardiac damage above was attenuated in response to AST treatment. The cardioprotective effect of AST against ethanol toxicity was also confirmed in both H9c2 cells and primary cardiomyocytes, indicating that AST-induced protection directly targets cardiomyocytes. Both in vivo and in vitro studies showed that AST inhibited all three ER stress signaling pathways activated by ethanol. Furthermore, administration of the ER stress inhibitor sodium 4-phenylbutyrate (4-PBA) strongly suppressed ethanol-induced cardiomyocyte damage. Interestingly, AST induced further anti-apoptotic effects once co-treated with 4-PBA, indicating that AST protects the heart from ACM partially by attenuating ER stress, but other mechanisms still exist. This study highlights that administration of AST ablated chronic excessive ethanol consumption-induced cardiomyopathy by suppressing cardiac ER stress and subsequent apoptosis.

Cardiac Mitochondrial PTEN-L determines cell fate between apoptosis and survival during chronic alcohol consumption.

Chronic alcohol consumption induces myocardial damage and a type of non-ischemic cardiomyopathy termed alcoholic cardiomyopathy, where mitochondrial ultrastructural damages and suppressed fusion activity promote cardiomyocyte apoptosis. The aim of the present study is to determine the role of mitochondrial fission proteins and/or other proteins that localise on cardiac mitochondria for apoptosis upon ethanol consumption. In vivo and in vitro chronic alcohol exposure increased mitochondrial Drp1 levels but knockdown of the same did not confer cardioprotection in H9c2 cells. These cells displayed downregulated expression of MFN2 and OPA1 for Bak-mediated cytochrome c release and apoptosis. Dysregulated PTEN/AKT cell survival signal in both ethanol treated and Drp1 knockdown cells augmented oxidative stress by promoting  mitochondrial PTEN-L and MFN1 interaction. Inhibiting this interaction with VO-OHpic, a reversible PTEN inhibitor, prevented Bak insertion into the mitochondria and release of cytochrome c to cytoplasm. Thus, our study provides evidence that Drp1-mediated mitochondrial fission is dispensable for ethanol-induced cardiotoxicity and that stress signals induce mitochondrial PTEN-L accumulation for structural and functional dyshomeostasis. Our in vivo results also demonstrates the therapeutic potential of VO-OHpic for habitual alcoholics developing myocardial dysfunction.

Myocardial tissue and metabolism characterization in men with alcohol consumption by cardiovascular magnetic resonance and 11C-acetate PET/CT.

BACKGROUND: Chronic alcohol consumption initially leads to asymptomatic left ventricular dysfunction, but can result in myocardial impairment and heart failure if ongoing. This study sought to characterize myocardial tissues and oxidative metabolism in asymptomatic subjects with chronic alcohol consumption by quantitative cardiovascular magnetic resonance (CMR) and 11C-acetate positron emission tomography (PET)/computed tomography (CT). METHODS: Thirty-four male subjects (48.8 ± 9.1 years) with alcohol consumption > 28 g/day for > 10 years and 35 age-matched healthy male subjects (49.5 ± 9.7 years) underwent CMR and 11C-acetate PET/CT. Native and post T1 values and extracellular volume (ECV) from CMR and Kmono and K1 from PET imaging were measured. Quantitative measurements by CMR and PET imaging were compared between subjects with moderate to heavy alcohol consumption and healthy controls, and their correlations were also analyzed. RESULTS: Compared to healthy controls, subjects with alcohol consumption showed significantly shorter native T1 (1133 ± 65 ms vs. 1186 ± 31 ms, p < 0.001) and post T1 (477 ± 42 ms vs. 501 ± 38 ms, p = 0.008) values, greater ECV (28.2 ± 2.2% vs. 26.9 ± 1.3%, p = 0.003), marginally lower Kmono (57.6 ± 12.1 min- 1 × 10- 3 vs. 63.0 ± 11.7 min- 1 × 10- 3, p = 0.055), and similar K1 (0.82 ± 0.13 min- 1 vs. 0.83 ± 0.15 min- 1, p = 0.548) after adjusting for confounding factors. There were no significant differences in CMR measurements and K1 between subjects with heavy and moderate alcohol consumption (all p > 0.05). In contrast, subjects with heavy alcohol consumption showed significantly lower Kmono values compared to those with moderate alcohol consumption (52.9 ± 12.1 min- 1 × 10- 3 vs. 63.7 ± 9.2 min- 1 × 10- 3, p = 0.012). Strong and moderate correlations were found between K1 and ECV in healthy controls (r = 0.689, p = 0.013) and subjects with moderate alcohol consumption (r = 0.518, p = 0.048), respectively. CONCLUSION: Asymptomatic men with heavy alcohol consumption have detectable structural and metabolic changes in myocardium on CMR and 11C-acetate PET/CT. Compared with quantitative CMR, 11C-acetate PET/CT imaging may be more sensitive for detecting differences in myocardial damage among subjects with moderate to heavy alcohol consumption.

(Pro)renin Receptor is Involved in Myocardial Damage in Alcoholic Cardiomyopathy.

BACKGROUND: (Pro)renin receptor (PRR), a novel member of the renin-angiotensin system, participates in various cardiovascular diseases. However, the role of PRR in alcoholic cardiomyopathy (ACM), which is caused by alcohol intake and manifests as myocardial damage and cardiac dysfunction, remains unclear. METHODS: PRR gene silencing was achieved by transfecting recombinant adenovirus expressing anti-PRR short hairpin RNA (PRR-shRNA). In vitro, primary rat cardiac fibroblasts (CFs) were cultured with the stimulation of alcohol (200 mM), with or without PRR-shRNA and PD98059. Immunofluorescence, RT-PCR, and Western blot were used to measure the protein and messenger (mRNA) expression of PRR, fibrotic factors, and members of related signaling pathways. In vivo, Wistar rats were fed a diet containing 9% (v/v) alcohol or a normal diet for 3 months, with or without PRR-shRNA. Sirius Red staining, immunohistochemical staining, and toluidine blue staining were used to evaluate myocardial fibrosis, oxidative stress, and inflammation response. RESULTS: Alcohol markedly increased PRR mRNA and protein expression in a time- and concentration-dependent manner in CFs. The increased expression of fibrotic factors induced by alcohol was prevented by PRR-shRNA and PD98059. Moreover, PRR-shRNA decreased the phosphorylation of extracellular regulated protein kinases (ERK) 1/2 in CFs. Furthermore, PRR-shRNA decreased cardiac fibrosis, reduced oxidative stress, and alleviated inflammation response in the myocardial tissue. CONCLUSIONS: Our results show that PRR-ERK1/2 signaling was involved in the development of ACM and that PRR could be a new target for the treatment of ACM.

Effects of the (Pro)renin Receptor on Cardiac Remodeling and Function in a Rat Alcoholic Cardiomyopathy Model via the PRR-ERK1/2-NOX4 Pathway.

Alcoholic cardiomyopathy (ACM) caused by alcohol consumption manifests mainly as by maladaptive myocardial function, which eventually leads to heart failure and causes serious public health problems. The (pro)renin receptor (PRR) is an important member of the local tissue renin-angiotensin system and plays a vital role in many cardiovascular diseases. However, the mechanism responsible for the effects of PRR on ACM remains unclear. The purpose of this study was to determine the role of PRR in myocardial fibrosis and the deterioration of cardiac function in alcoholic cardiomyopathy. Wistar rats were fed a liquid diet containing 9% v/v alcohol to establish an alcoholic cardiomyopathy model. Eight weeks later, rats were injected with 1 × 109v.g./100 µl of recombinant adenovirus containing EGFP (scramble-shRNA), PRR, and PRR-shRNA via the tail vein. Cardiac function was assessed by echocardiography. Cardiac histopathology was measured by Masson's trichrome staining, immunohistochemical staining, and dihydroethidium staining. In addition, cardiac fibroblasts (CFs) were cultured to evaluate the effects of alcohol stimulation on the production of the extracellular matrix and their underlying mechanisms. Our results indicated that overexpression of PRR in rats with alcoholic cardiomyopathy exacerbates myocardial oxidative stress and myocardial fibrosis. Silencing of PRR expression with short hairpin RNA (shRNA) technology reversed the myocardial damage mediated by PRR. Additionally, PRR activated phosphorylation of ERK1/2 and increased NOX4-derived reactive oxygen species and collagen expression in CFs with alcohol stimulation. Administration of the ERK kinase inhibitor (PD98059) significantly reduced NOX4 protein expression and collagen production, which indicated that PRR increases collagen production primarily through the PRR-ERK1/2-NOX4 pathway in CFs. In conclusion, our study demonstrated that PRR induces myocardial fibrosis and deteriorates cardiac function through ROS from the PRR-ERK1/2-NOX4 pathway during ACM development.

AIN-93 Diet as an Alternative Model to Lieber-DeCarli Diet for Alcoholic Cardiomyopathy.

BACKGROUND: The Lieber-DeCarli alcoholic liquid diet is a classical method for establishing animal models of alcoholic cardiomyopathy (ACM). No study has reported whether the AIN-93 diet, which is widely used as a standard diet for both long-term and short-term studies with laboratory animals, could be used to construct the ACM animal model. The present study intended to investigate whether the AIN-93 diet could be used to establish a mouse ACM model. METHODS: Twenty-four C57BL/6 male mice were randomly divided into 4 equally sized groups. In ethanol (EtOH)-fed groups, mice were fed a 4%-EtOH (w/v, 28% of total calories) alcoholic liquid diet of Lieber-DeCarli or the AIN-93 diet for chronic alcohol exposure for 180 days. In control-fed groups, mice were fed with non-EtOH liquid diets with the same calories as EtOH-fed groups. Morphological observations of the hearts and molecular investigation of the brain natriuretic peptide (BNP) were carried out by echocardiography, hematoxylin and eosin (H&E) and immunohistochemistry (IHC) staining, real-time quantitative polymerase chain reaction (qPCR), and enzyme-linked immunosorbent assay. RESULTS: Echocardiography showed that mice fed with either the 4%-EtOH Lieber-DeCarli diet or the 4%-EtOH AIN-93 diet had dilated ventricles and poor cardiac function. IHC staining of BNP, qPCR of BNP mRNA, and plasma concentration of BNP showed an up-regulated expression in mice fed with both the 4%-EtOH Lieber-DeCarli and 4%-EtOH AIN-93 diets. Less fatty liver was also observed in mice fed the AIN-93 alcoholic diet than those fed the Lieber-DeCarli alcoholic diet. CONCLUSIONS: The AIN-93 alcoholic liquid diet can be used to establish ACM animal models, as with the conventional Lieber-DeCarli alcoholic liquid diet.

Atorvastatin reduces alcohol-induced endoplasmic reticulum stress in AC16 cardiomyocytes.

OBJECTIVES: To investigate the effects of atorvastatin on the ultrastructure and lipid metabolism of AC16 cardiomyocytes in response to alcohol-induced endoplasmic reticulum stress (ERS). DESIGN: The expression of the ERS-related factor GRP78 in the established ERS model was determined by western blotting. Alcohol-exposed cardiomyocytes were treated with various concentrations of atorvastatin, and GRP78 expression was measured. Cardiomyocyte ultrastructure was observed and SREBP-1c and triglyceride (TG) levels were evaluated. RESULTS: Exposure to ethanol for 0, 12, 24, and 48 h significantly affected GRP78 expression (0.19 ± 0.02, 0.27 ± 0.03, 0.39 ± 0.01, and 0.64 ± 0.02, respectively). GRP78 expression in the 1, 10, and 100 µmol L-1 atorvastatin-treated groups was 0.50 ± 0.04, 0.38 ± 0.03, and 0.24 ± 0.01, respectively, and significantly different from control group expression (0.19 ± 0.02); the expression in the alcohol group was 0.64 ± 0.02. Alcohol-treated AC16 cells had significantly larger and fewer mitochondria and disorganized cristae, often replaced by vacuoles. These aberrations decreased with increasing atorvastatin concentrations. SREBP-1c expression also differed significantly among all atorvastatin-treated and control groups (0.47 ± 0.04, 0.39 ± 0.03, and 0.31 ± 0.02; normal 0.25 ± 0.02; alcohol 0.56 ± 0.03). TG expression differed significantly between the 10 and 100 µmol L-1 groups (26.84 ± 1.63, 23.11 ± 2.05) and the alcohol group (36.35 ± 2.41). CONCLUSIONS: Atorvastatin inhibited the expression of the ERS-related factor GRP78 in response to alcohol exposure, improved cell morphology, and enhanced lipid metabolism in a cellular model of alcoholic cardiomyopathy.

Epac Proteins and Calmodulin as Possible Arrhythmogenesis Trigger in Alcoholic Cardiomyopathy.

The expression of Epac proteins (exchange protein directly activated by cAMP) and calmodulin (CaM) was assessed by the content of the corresponding mRNA in biopsy specimens of cardiac atrium, left ventricle, and thoracic aorta of rats with alcoholic cardiomyopathy. In the myocardium, overexpression of Еpac1, Ерас2, and СаМ mRNA was found. The content of Epac2 mRNA in the left ventricle was elevated by 2.9 times (p=0.000001), in the left atrium by 3.2 times (p=0.00001), in the right atrium by 3 times (p=0.00001). In contrast to the myocardial tissue, the content of CaM mRNA in the thoracic aorta was not increased, but showed a tendency to decrease, when compared to the control values, while the level of Epac1 and Epac2 mRNA was increased. The assumption is made that regulatory proteins Epac and CaM can play a key role in arrhythmogenesis development under conditions of alcoholic cardiomyopathy.

Comparison among Reconstruction Algorithms for Quantitative Analysis of 11C-Acetate Cardiac PET Imaging.

Objective: Kinetic modeling of dynamic 11C-acetate PET imaging provides quantitative information for myocardium assessment. The quality and quantitation of PET images are known to be dependent on PET reconstruction methods. This study aims to investigate the impacts of reconstruction algorithms on the quantitative analysis of dynamic 11C-acetate cardiac PET imaging. Methods: Suspected alcoholic cardiomyopathy patients (N = 24) underwent 11C-acetate dynamic PET imaging after low dose CT scan. PET images were reconstructed using four algorithms: filtered backprojection (FBP), ordered subsets expectation maximization (OSEM), OSEM with time-of-flight (TOF), and OSEM with both time-of-flight and point-spread-function (TPSF). Standardized uptake values (SUVs) at different time points were compared among images reconstructed using the four algorithms. Time-activity curves (TACs) in myocardium and blood pools of ventricles were generated from the dynamic image series. Kinetic parameters K1 and k2 were derived using a 1-tissue-compartment model for kinetic modeling of cardiac flow from 11C-acetate PET images. Results: Significant image quality improvement was found in the images reconstructed using iterative OSEM-type algorithms (OSME, TOF, and TPSF) compared with FBP. However, no statistical differences in SUVs were observed among the four reconstruction methods at the selected time points. Kinetic parameters K1 and k2 also exhibited no statistical difference among the four reconstruction algorithms in terms of mean value and standard deviation. However, for the correlation analysis, OSEM reconstruction presented relatively higher residual in correlation with FBP reconstruction compared with TOF and TPSF reconstruction, and TOF and TPSF reconstruction were highly correlated with each other. Conclusion: All the tested reconstruction algorithms performed similarly for quantitative analysis of 11C-acetate cardiac PET imaging. TOF and TPSF yielded highly consistent kinetic parameter results with superior image quality compared with FBP. OSEM was relatively less reliable. Both TOF and TPSF were recommended for cardiac 11C-acetate kinetic analysis.

ALDH2 protects against alcoholic cardiomyopathy through a mechanism involving the p38 MAPK/CREB pathway and local renin-angiotensin system inhibition in cardiomyocytes.

BACKGROUND: Angiotensin II (Ang II) in the local cardiac renin-angiotensin system (RAS) is closely associated with alcoholic cardiomyopathy (ACM). Inhibition of local cardiac RAS has great significance in the treatment of ACM. Although aldehyde dehydrogenase 2 (ALDH2) has been demonstrated to protect against ACM through detoxification of aldehydes, the precise mechanisms are largely unknown. In the present study, we determined whether ALDH2 improved cardiac damage by inhibiting the local RAS in ACM and investigated the related regulatory mechanisms. METHODS AND RESULTS: Adult male mice were fed with 5% ethanol or a control diet for 2months, with or without the ALDH2 activator Alda-1. Heavy ethanol consumption induced cardiac damage, increased angiotensinogen (AGT) and Ang II and decreased myocardial ALDH2 activity in hearts. ALDH2 activation improved ethanol-induced cardiac damage and decreased AGT and Ang II in hearts. In vitro, ALDH2 activation or overexpression decreased AGT and Ang II in cultured cardiomyocytes treated with 400mmol/L ethanol for 24h. Furthermore, p38 MAP kinase (p38 MAPK)/cyclic adenosine monophosphate response element-binding protein (CREB) pathway activation by ethanol increased AGT and Ang II in cardiomyocytes. In addition, ALDH2 activation or overexpression inhibited the p38 MAPK/CREB pathway leading to decreased AGT and Ang II in cardiomyocytes. We also found that p38 MAPK activation effectively mitigated Alda-1-decreased AGT and Ang II, the effect of which was reversed by inhibition of CREB. CONCLUSIONS: ALDH2 decreased AGT and Ang II in the local cardiac RAS via inhibiting the p38 MAPK/CREB pathway in ACM, thus improving ethanol-induced cardiac damage.

Hydrogen sulfide alleviates myocardial fibrosis in mice with alcoholic cardiomyopathy by downregulating autophagy.

Myocardial fibrosis is one of the most important pathological features of alcoholic cardiomyopathy (ACM). Hydrogen sulfide (H2S) exerts protective effects in various types of cardiovascular disease, which has been demonstrated by many previous studies. However, there is a lack of adequate research on the effect of H2S on myocardial fibrosis in ACM. The present study aimed to investigate the etiopathogenic role of H2S in myocardial fibrosis induced by chronic alcohol intake. An ACM mouse model was induced by consumption of 4% ethanol solution in drinking water for 12 weeks. Sodium hydrosulfide (NaHS) was used as a donor to provide exogenous H2S. Twelve weeks later, mice were sacrificed to calculate the heart to body weight ratio. The degree of myocardial collagen deposition was evaluated by Masson's and Van Gieson's staining, the expression level of collagen â…  was measured by immunohistochemistry and autophagosomes were observed by transmission electron microscopy. In addition, the expression levels of autophagy­associated proteins and fibrosis-associated proteins were detected by western blotting, and the expression levels of miR-21 and miR-211 were detected by reverse transcription-quantitative polymerase chain reaction. The outcomes of the study revealed that chronic alcohol intake results in myocardial fibrosis, enhanced myocardial collagen deposition and increased expression levels of collagen I, autophagy, autophagy-associated proteins (Beclin 1, Atg3 and Atg7) and fibrosis-associated proteins (MMP8, MMP13, MMP14, MMP17 and TGF-ß1), as well as miR-21 and miR-221. These results were markedly reversed following treatment with H2S. The present study confirmed that H2S relieves myocardial fibrosis in mice with ACM, and the underlying mechanism may involve the downregulation of autophagy and miR-21 and miR-211 expression levels.

From the Cover: Alcohol Inhibition of the Enzymatic Activity of Glyceraldehyde 3-Phosphate Dehydrogenase Impairs Cardiac Glucose Utilization, Contributing to Alcoholic Cardiomyopathy.

Heavy consumption of alcohol induces cardiomyopathy and is associated with metabolic changes in the heart. The role of altered metabolism in the development of alcoholic cardiomyopathy remains largely unknown but is examined in the present study. The effect of chronic alcohol consumption on cardiac damage was examined in mice fed an alcohol or isocaloric control diet for 2 months. Signaling pathways of alcohol-induced metabolic alteration and pathologic changes were examined in both animal hearts and H9c2 cell cultures. Compared with controls, the hearts from the alcohol-fed mice exhibited cardiac oxidative stress, cell death, a fibrotic response, hypertrophic remodeling, and the eventual development of cardiac dysfunction. All these detrimental effects could be ameliorated by superoxide dismutase mimic Mn (111) tetrakis 1-methyl 4-pyridylporphyrin pentachloride (MnTMPyP) therapy. A mechanistic study showed that chronic alcohol exposure enhanced the expression of proteins regulating fatty acid uptake but impaired the expression of proteins involved in mitochondrial fatty acid oxidation, which compensatively geared the heart to the suboptimal energy source, glucose. However, chronic alcohol exposure also impaired the glycolytic energy production step regulated by glyceraldehyde-3-phosphate dehydrogenase, which further feeds back to enhance glucose uptake signaling and the accumulation of glycolytic intermediate product fructose, resulting in aggravation of alcohol-induced cardiac oxidative stress, cell death, and remodeling. All these dysmetabolic alterations could be normalized by MnTMPyP treatment, along with significant improvement in cardiac cell death and remodeling. These results demonstrate that alcohol-induced oxidative stress and altered glucose metabolism are causal factors for the development of alcoholic cardiomyopathy.

Etiology of alcoholic cardiomyopathy: Mitochondria, oxidative stress and apoptosis.

Putative mechanisms leading to the development of alcoholic cardiomyopathy (ACM) include the interrelated cellular processes of mitochondria metabolism, oxidative stress and apoptosis. As mitochondria fuel the constant energy demands of this continually contracting tissue, it is not surprising that alcohol-induced molecular changes in this organelle contribute to cardiac dysfunction and ACM. As the causal relationship of these processes with ACM has already been established, the primary objective of this review is to provide an update of the experimental findings to more completely understand the aforementioned mechanisms. Accordingly, recent data indicate that alcohol impairs mitochondria function assessed by membrane potential and respiratory chain activity. Indictors of oxidative stress including superoxide dismutase, glutathione metabolites and malondialdehyde are also adversely affected by alcohol oftentimes in a sex-dependent manner. Additionally, myocardial apoptosis is increased based on assessment of TUNEL staining and caspase activity. Recent work has also emerged linking alcohol-induced oxidative stress with apoptosis providing new insight on the codependence of these interrelated mechanisms in ACM. Attention is also given to methodological differences including the dose of alcohol, experimental model system and the use of males versus females to highlight inconsistencies and areas that would benefit from establishment of a consistent model.