Study Of The Mechanism Of Alcohol-aggravated Stress Stress-induced Heart Failure

Purpose:Heart failure(HF)is a global epidemic caused by myocardial damage caused by various causes,resulting in changes in the structure and function of the heart muscle,and eventually leading to impaired ventricular pumping or filling function.Long-term excessive pressure overload(PO)on the heart can promote myocardial damage.Long-term alcohol consumption can exacerbate this stress-load-induced heart failure,and the underlying mechanism by which alcohol causes this remains poorly understood.Histone acetylation refers to the modification that occurs when Acetyl-Co A is deposited on its residue by the action of histone acetyltransferase on histone lysine,which requires two basic conditions,one is the activation of histone acetyltransferase(HAT),and the other is the need for sufficient acetyl-Co A(Acetyl-Co A)as a substrate.A large amount of clinical and experimental evidence suggests that alcohol can be metabolized into acetaldehyde and then converted into acetic acid in the liver,and the acetyl group in acetyl-acetic acid synthesizes a large amount of Acetyl-Co A under the action of acetyl-Co A synthetase 2(ACSS2)and is deposited directly on histones,rapidly promoting the acetylation of histones.The aim of this article was to explore how chronic alcohol consumption can aggravate stress-induced heart failure by promoting histone H3 acetylation.Method:1.Alcohol(Ethanol,Et OH)aggravates stress load-induced heart failure(1)The pressure load model was constructed by performing abdominal aortic coarctation(AC)on C57BL/6J male mice,and then feeding AC mice to alcoholic liquid feed to construct an alcoholic mouse animal model.(2)The 18 mice were randomly divided into three groups: Sham group,AC-8W group,and AC+Et OH group.Mice are euthanized 8 weeks after AC surgery,their heart weight and body weight are recorded,and serum and heart tissue from mice are collected.Western Blot,Realtime-q PCR detection of mice,Masson staining and other experimental techniques were used to detect the protein and m RNA expression levels of heart failure and fibrosis related indexes in mouse left ventricular tissue,respectively.Alcohol consumption has been shown to worsen stress-induced heart failure.2.Acetic acid generated by alcohol metabolism promotes acetylation of histone H3(1)The acetic acid detection kit was used to determine the concentration of acetic acid in the serum of mice fed with alcoholic feed,and it was clear that the metabolism of alcohol in mice increased the level of acetic acid concentration in serum,and Western Blot was used to detect the level of histone H3 acetylation in the left ventricular tissue of mice,which proved that alcohol promoted histone acetylation of myocardial tissue.(2)A mouse model of high concentration of acetic acid in vivo was constructed by feeding mice drinking water containing small molecules of acetate.The 12 male mice were randomly divided into three groups: AC-14 group,AC+fasting group,AC+fasting+Acetate group.Mice are euthanized 14 days after AC surgery,mice tail vein blood glucose are measured and mice are collected serum and left ventricular tissue.Western Blot,Realtime-q PCR and other technologies were used to investigate the effect of acetic acid on histone acetylation of mouse myocardium,and the effect of acetic acid on the expression and transcription level of myocardial heart failure-related index proteins in mice.Acetic acid has been shown to aggravate stress-induced heart failure by promoting histone acetylation of the myocardium.(3)Neonatal Rat Ventricular Cardiomyocytes(NRVMs)were cultured,phenylephrine(PE)was used to stimulate NRVMs to construct a hypertrophic model of primary cardiomyocytes,and then NRVMs were treated with acetic acid small molecules,and the histone H3 acetylation level of NRVMs after treatment with different concentrations of acetic acid was observed by Western Blot technology.3.Acetic acid-dependent ACSS2 promotes histone acetylation of the myocardium(1)An animal model of ACSS2 inactivation was constructed by intraperitoneally injecting mice with a small molecule ACSS2 inhibitor(Ac-Co A Synthase Inhibitor1(ACSS2i).The experimental groupings were as follows: AC+Fasting,AC+Fasting+Acetate,AC+Fasting+Acetate+ACSSi,left ventricular tissue was collected after AC surgery 4 days later,and the effect of acetic acid on myocardial histone H3 acetylation and heart failure-related gene expression in mice after ACSS2 inactivation was detected by Western Blot technology.(2)Using PE and ACSS2 i to construct cell models of cell hypertrophy and ACSS2 inactivation.The experimental groups were as follows: control group(NT),cell hypertrophy induction group(PE),PE+ sugar-free group,PE+ sugar-free + acetic acid group,PE + sugar-free + acetic acid + ACSS2 i group,and the effect of acetic acid on the expression of NRVMs histone H3 acetylation and heart failure-related genes after in vitro inhibition of ACSS2 was observed by Western Blot technology.4.Inhibition of histone H3 acetylation can inhibit heart failure(1)An animal model with down-regulated histone H3 acetylation levels was constructed using histone acetyltransferase p300 and the small molecule inhibitor A485 of CBP.Thirteen male mice were randomly divided into three groups: Sham group,AC-28 group,AC+A485 group.After 28 days of AC surgery,mice were euthanized,the weight of the mice,the weight of the heart and their left ventricular tissue were collected,and the expression of heart failure-related gene proteins and transcription and the level of myocardial fibrosis after inhibition of histone acetylation in vivo were observed by Western Blot and Masson.(2)A cell model with down-regulated histone H3 acetylation levels was constructed using A485,a small molecule inhibitor of HAT.The effects of inhibition of histone acetylation on the expression levels of heart failure-related gene proteins and m RNA were observed by Western Blot and Realtime-q PCR.Detection of intranuclear histone H3 acetylation levels by cellular immunofluorescence.results:1.Alcohol aggravates stress load-induced heart failure(1)Compared with the Sham group,the ratio of heart to body weight of AC8-week mice was significantly increased;Compared with mice in the AC 8-week group,the ratio of heart to body weight in the AC+Et OH group was further increased.(2)The results of Western Blot test showed that compared with the AC 8-week group,the expression level of heart failure and fibrosis related genes(BNP,FN1)protein in the left ventricular tissue of mice in the AC+Et OH group was significantly upregulated.(3)Realtime-q PCR test results showed that compared with the AC 8-week group,the m RNA expression levels of Nppa and Nppb in the left ventricular tissue of mice in the AC+Et OH group were significantly increased.(4)The results of Masson staining showed that compared with the AC 8-week group,the fibrotic area of left ventricular tissue in the AC+Et OH group increased significantly.2.Acetic acid generated by alcohol metabolism promotes acetylation of histone H3(1)The results of the detection of acetic acid concentration in mouse serum showed that the metabolism of alcohol in mice would increase the concentration of acetic acid in mouse serum.(2)Compared with AC 8-week mice,mice in the AC+Et OH group further promoted AC-induced histone H3 acetylation;(3)Compared with the AC+Fasting group,the acetylation level and BNP protein expression level of left ventricular tissue histone H3 in mice in the AC+Fasting+Acetate group increased significantly.In addition,acetate-induced histone acetylation of NRVMs was regulated upward with the increase of acetic acid concentration.(4)Realtime-q PCR test results showed that acetic acid could promote the m RNA expression levels of Nppa,Nppb and FN1 in the left ventricular tissue of mice.3.Acetic acid relies on ACSS2 to promote histone acetylation of myocardiumWestern Blot results demonstrated that acetic acid induces histone hyperacetylation of cardiomyocytes at both in vivo and in vitro levels,and ACSS2i inhibits acetic acid-induced histone H3 acetylation.It was proved that acetic acid is used to induce histone acetylation modification of the myocardium through the action of ACSS2.4.Inhibition of histone H3 acetylation can inhibit heart failure(1)In vivo level: Compared with the AC group,the ratio of heart to body weight of mice in the A485 group was significantly reduced,the histone H3 acetylation level of left ventricular tissue was also significantly inhibited,and the expression levels of Nppa,Nppb and FN1 were also significantly down-regulated.(2)In vitro level: Compared with the PE group,A485 inhibited the histone H3 acetylation of NRVMs,and inhibited the expression of Nppa and Nppb.Cellular immunofluorescence assays also suggest that A485 can inhibit intranuclear levels of H3 acetylation of the histone of NRVMs.Conclusion:Alcohol is metabolized to produce acetic acid,which induces histone H3 acetylation of the myocardium under the action of ACSS2,thereby aggravating stress-induced heart failure.Histone acetylation participates in the occurrence and development of heart failure by regulating the transcription and protein expression levels of heart failure-related genes,and inhibits histone acetylation to inhibit the occurrence and development of heart failure in mice.