Study On Alcohol-induced Cardiac Injury And The Preventive Effect Of Zinc Supplementation

Objective:This study was designed to determine the effect of long-term high-dose alcohol consumption on cardiac structure and function in rats,and to disclose its underlying mechanisms.We also investigated whether dietary zinc supplementation could provide protection from alcoholic heart damage.Methods:Eight-week-old male Wistar rats were randomly divided into four groups following 2×2 factorial design,alcohol×zinc,that is control group(group C,n=12), alcohol group(group A,n=12),alcohol and zinc supplementation group(group AZ, n=12)and zinc supplementation group(group Z,n=12).Group A received 10%(v/v) alcohol freely and zinc supplementation was done by adding zinc sulfate to the standard rat chow diet at 100mg zinc atom/Kg.At the end of the 5th month, transthoracic echocardiography was performed in all groups,and parameters HR,LVEDD,LVESD,LA,IVS,LVPW,LVM,EF,FS,SV,E,Ea,Aa,Ea/Aa and IVRT were obtained.Then,blood was drawn using a heparinized syringe from the left ventricle,and plasma was obtained by centrifugation.The heart was weighed, and the myocardial samples were processed for both pathological and biochemical analysis.Whole blood zinc concentrations were determined by atomic absorption spectrophotometry.The level of plasma MDA was assessed by thiobarbituric acid method modified by Yagi.Histopathological and ultrastructural changes of myocardium were examined by light and electron microscopy.DNA fragmentation was determined using a terminal deoxynucleotidyl transferase-mediated dUTP nick-end-labeling(TUNEL)assay.Total collagen content of the heart was quantified by examining hydroxyproline content.CollagenⅠ,CollagenⅢ,Caspase-3,Bax,and Bcl-2 gene expression was evaluated by semiquantitative reverse transcription-polymerase chain reaction.All data are expressed as mean±SE.The data were analyzed according to a 2×2 factorial design.The level of significance was considered at P＜0.05.Results:1.General condition:no mortality.The daily alcohol consumption of group A and group AZ was 5.28±0.25g/Kg and 5.35±0.33g/Kg,respectively.The daily zinc intake of group AZ and Z was 15.15±4.49mg/Kg and 15.01±3.57mg/Kg, respectively.No significant differences in these two parameters were measured between the two groups(P＞0.05).2.Echocardiographic data:LVEDD,LVESD,LA,Aa and IVRT values were greater in group A compared with the other three groups(P＜0.05).No differences were found in IVS,LVPW,LVM,EF,FS and SV values among four groups(P＞0.05). Ea,Ea/Aa and E values were decreased significantly in group A compared with the other three groups(P＜0.05).3.Whole blood and plasma test:Chronic alcohol feeding caused a significant decrease in the whole blood zinc concentrations in group A(P＜0.05),but dietary zinc supplementation prevented the alcohol-induced decrease in group AZ rats.Alcohol feeding significantly increased MDA concentrations in the plasma(P＜0.05)and zinc supplementation(group AZ)had no statistically differences compared with the other two groups(P＞0.05).4.Heart morphological alterations and tissue injury:Alcohol feeding increased the heart weight normalized to body weight significantly(P＜0.05),but zinc supplementation did not cause significant changes.Histological examination by HE staining identified hypertrophic cardiomyocytes,patchy inflammatory areas with interstitial neutrophil infiltration and myocardial fibrosis in the myocardium in group A.Zinc supplementation could significantly improve the above changes(only a few inflammatory infiltration and no fibrosis was found).Ultrastructural examination by electron microscopy revealed that alcohol could induce many ultrastructural changes involved nucleus,mitochondrion,lysosome,sarcolemma,myofibril,Z line and cell membrane.Zinc supplementation prevented the above alcohol-induced pathological changes shown in group AZ.TUNEL-positive nuclei increased significantly in group A compared with the other three groups(P＜0.05).Zinc supplementation significantly inhibited alcohol-induced cardiomyocyte apoptosis.5.Total collagen content:Compared with group C,AZ and Z,hydroxyproline content of group A significantly increased(P＜0.05),but there was no statistically difference among group C,AZ and Z(P＞0.05).6.RT-PCR data:The mRNA abundances of CollagenⅠ,CollagenⅢ,Caspase-3, Bax in alcohol group were increased significantly compared with the other three groups(P＜0.05),but Bcl-2 had no statistically differences(P＞0.05).No significant differences were found among group C,AZ and Z(P＞0.05).Conclusion:1.High-dose alcohol consumption of 5 months led to ventricular hypertrophy and diastolic dysfunction,whereas systolic function maintained normal in rats.2.Pathological study of both light and electron microscope found myocardial inflammatory infiltration and fibrosis,interstitial fibrosis after 5 months alcohol consumption.These changes were responsible for left ventricular diastolic dysfunction,and were prior to left ventricular systolic dysfunction.3.Alcohol-induced cardiac injury may be related to oxidative stress,zinc defect, cardiomyocyte apoptosis and myocardial fibrosis.4.Zinc supplementation prevented alcohol-induced cardiac injury by maintaining normal structure and function of heart,cardiomyocyte,and even its ultrastructure.5.Prevention of alcoholic cardiac damage by Zinc supplementation might benefit from attenuation of oxidative stress,keeping zinc concentration balance,and inhibition of cardiomyocyte apoptosis and myocardial fibrosis.