Effect Of Curcumin On Insulin Resistance Accompanied By Endoplasmic Reticulum Stress In Cardiomyocytes Induced By Alcohol

Introduction: Heavy alcohol consumption for a long time can result in cardiomegaly,arrhythmia and congestive heart failure.This phenomenon characterized by cardiomyopathy change is called alcoholic cardiomyopathy(ACM).The prevalence of alcoholic cardiomyopathy rises year by year and it has become a kind of increasing disease in cardiovascular field especially in developed countries and many developing countries,which introduces a serious threat to human health and consume huge medical resources.The pathogenesis of alcoholic cardiomyopathy,endoplasmic reticulum stress and energy metabolism have become hotspots in the last few years.Alcohol induces endoplasmic reticulum stress,and then,endoplasmic reticulum stress induces insulin signaling disorders,these induced pathological changes initiate energy metabolism disorders in myocardial cells,leading to cardiac dysfunction and degressive cardiac function,and finally resulted in alcoholic cardiomyopathy.Therefore,to explore effective drug prevention and treatment to alcohol-induced endoplasmic reticulum stress injury,inhibition of myocardial cell insulin signaling pathway disorders,and to improve cardiac function has an important clinical significance.Curcumin(C21H20O6)is a polyphenolic compound extracted from the roots of Chinese traditional medicine Turmeric,and has pharmacological properties such as anti-oxidation,anti-cancer,anti-angiogenesis,antiproliferation and anti-inflammatory.In recent years,it has been reported that in many in vivo and in vitro experiments,curcumin can activate the insulin signaling pathway,but also as an endoplasmic reticulum stress antagonist,play a role in alleviating endoplasmic reticulum stress.It has been reported in the literature that curcumin has the effect of inhibiting adipocyte degeneration in alcoholic liver disease.We take this as a starting point to explore alcoholic cardiomyopathy.Curcumin can inhibit the endoplasmic reticulum stress and then relieve insulin resistance,and thus relieve myocardial glucose metabolism disorder.This experiment intends to use AC16 cardiomyocytes to establish an alcoholic cardiomyocytes model,and to observe whether in vitro curcumin administration can reduce the occurrence of alcohol-induced endoplasmic reticulum stress and glucose metabolism disorder.Materials and Methods:1.AC16 cells as the research object,five groups are established: a normal control group(without any intervention)?The control group(insulin)?Alcohol(400 mmol / l),curcumin(50 ?mol / l)+ alcohol group and 4-PBA + alcohol group.Concentration is determined according to the literature and pre-experiment.2.Using MTS assay to examine different concentrations of alcohol on the survival rate of myocardial cells and the choice of the optimal alcohol and curcumin concentrations for subsequent experiments.3.RT-PCR method was used to detect the changes of endoplasmic reticulum stress relating m RNA expression in each group.4.Western blot was used to detect the expression of GRP78,IRE1 and IRS-1,AKT,p-IRS and p-AKT in each group.5.Transmission electron microscopy was used to observe the ultrastructural structures of endoplasmic reticulum stress.6.Glucose oxidase method was used to detect glucose consumption in each group.7.The data of experimental group were expressed as mean ± standard deviation(`x±s),using SPSS 21.0 analysis,the differences between groups were compared using one-way ANOVA.P <0.05 is statistically significance.Results:1.Cell survival rate: Alcohol leds to a decreased cell viability via a concentration-dependent manner,cell survival rate decreased more obviously at the concentration of 400mmol/L which is used as the damage conditions in the follow-up experiments.Different concentrations of curcumin 0,5,10,20,50,100 umol/l interfering cells for 48 hours resulting in unobvious effect on cell survival rate.Curcumin inhibites the declining in cell viability caused by alcohol via concentration-dependent manner,survival rate was significantly enhanced at the concentration of 50 umol /l,so 50umol/l was used as the cndition to study the protective effect of curcumin.2.Compared with the control group,the expression of GRP78 m RNA and IRE1 m RNA in alcohol group increased significantly.The expression of GRP78 m RNA and IRE1 m RNA decreased after curcumin and 4-PBA pretreatment,the difference was significant compared with alcohol group.3.The structure of endoplasmic reticulum in the control group was obviously abnormal detected by transmission electron microscope.The structure of endoplasmic reticulum was abnormal in alcohol group,charactered as swollen,full of unfolded protein inside,and increased number of endoplasmic reticulum.The abnormalities of the structure of the cytoplasm were relieved compared with the alcohol group.4.Western blot detection of endoplasmic reticulum stress-related protein GRP78,IRE1 expression increased significantly in alcohol group compared with the control group.GRP78 and IRE1 expression decreased in curcumin and 4-PBA pretreatment group,compared with alcohol group.The protein expressions of P-IRS-1(Thr308)and p-AKT(Thr308)were significantly decreased in alcohol group.The expressions of IRS-1(Thr308)and p-AKT(Thr308)in curcumin and 4-PBA group were higher than those in alcohol group.5.Glucose consumption was significantly lower in alcohol group than that in control(insulin)group.Compared with alcohol group,curcumin and 4-PBA pretreatment group showed higher glucose consumption.Conclusion: Alcohol can stimulate endoplasmic reticulum stress,increase insulin resistance related factor expression and induce abnormal glucose metabolism.In addition to these,curcumin and endoplasmic reticulum stress antagonist can play a similar role,by reducing endoplasmic reticulum stress and thus promoting glucose metabolism.The recovery function for alcoholic cardiomyopathy presented by Curcumin may provide us with new ideas for drug treatment in the future.