Regulatory Mechanism Of Histone Acetylation On Anacardic Acid Attenuates Pressure-Overload Cardiac Hypertrophy Of Mice

Part?Effectsof pressure overload on histone acetylation modification in myocardial hypertrophy related genes of miceObjective:To investigate the role of histone acetylase modification imbalance mediated by histone acetylases(HATs)p300(E1A-binding protein 300kD)and P300/CBP-associated factor(PCAF)on overexpession of related genes in myocardial hypertrophy mice induced by pressure overload.Methods: Kunming mice old 8 weeks were selected as the research object,and were randomly divided into normal group,sham group and thoracic aortic constriction(TAC)group according to random number table method for 6 mice in each group.Thoracic aortic was partially ligated in the TAC group,only the thoracic aorta was separated,and other treatments were the same as the TAC group in the sham group.After 8 weeks,the three groups of mice were tested as follows:(1)Cardiac ultrasonography was used to detect left ventricular chamber and wall thickness.(2)The body weight of the mice was measured.Subsequently,the heart and lung tissues were extracted from the death mice by cervical dislocation,and the heart weight/body weight(HW/BW)and lung weight/body weight(LW/BW)were detected.(3)The shape of the mice heart was observed by stereoscope,and the morphological changes of myocardial cells was observed by htoxylin-eosin(H&E)staining.(4)Colorimetry was used to detect HATs activity.(5)The expression of histone H3K9 ac,p300-HAT,PCAF-HAT and the myocardial hypertrophy markers ANP and ?-MHC were assayed by Western blot.(6)Real-time quantitative polymerase chain reaction(Real-Time PCR)was used to detect the mRNA expression in the heart nucleus transcription factor MEF2 A.Results:(1)The results of stereoscopy showed that the heart of mice in TAC group was significantly larger than that of the normal group,and no significant change in thesham group.(2)The cardiac mass index(CMI)of the TAC group was significantly higher than that of the normal group(P < 0.05),while the lung mass index(LMI)was not statistically different from that of the normal group(P > 0.05).(3)H&E staining results showed that compared with the normal group,the myocardial cells were significantly enlarged,disordered and fibroticin the TAC group.(4)Cardiac ultrasonography results showed that the left ventricular cavity was significantly smaller than that of the normal group,and the ventricular wall thickness was significantly increased in the TAC group,while that of the sham group was not significantly changed.(5)The data of colorimetry showed that HATs activity was significantly increased compared with that of the normal groupin the myocardial tissue of the TAC group(P < 0.05),while there was no apparently increase in the sham group(P >0.05).(6)Western blot results showed that the acetylation level of histone H3K9 ac was significantly higher than that of the normal group in the TAC group(P < 0.05),and the protein expression levels of p300-HAT,PCAF-HAT,myocardial hypertrophy marker ANP and ?-MHC were significantly higher than those of the normal group in the TAC group(P < 0.05).(7)Real-time PCR results showed that compared with the normal group,the mRNA expression of cardiac nucleus transcription factor MEF2 A was significantly increased in the myocardial tissue of the TAC group,and the difference was statistical significance(P < 0.05).Conclusions:(1)The model of cardiac hypertrophy in mice was successfully constructed by constricting the thoracic aorta.(2)Hyperacetylation of histone H3K9 ac mediated by p300-HAT and PCAF-HAT was involved the overexpression of the cardiac nucleus transcription factor MEF2 A,and myocardial hypertrophy markers ANP and ?-MHC in cardiac hypertrophy miceinduced by pressure overload.Part?Anacardic Acid Attenuates Pressure-Overload Cardiac Hypertrophy Through Regulating Histone Acetylation ModificationObjective:To investigate the regulation of histone acetylation modification on histone acetylases(HATs)inhibitor anacardic acid(AA)attenuates myocardial hypertrophy in mice caused by pressure overload.Methods:Kunming mice old 8 weeks were selected as the research object,and were randomly divided into normal group,thoracic aortic constriction(TAC)group,TAC +AA group,TAC + dimethy sulfoxide(DMSO)group,normal + AA group according to random number table method.Part of the thoracic aorta(about 70% of the thoracic aorta diameter)was ligated to 8 weeks after thoracic aortic constriction in the TAC group.AA(5 mg/kg)was given 3 weeks and 3 times a week to 8 weeks afterthoracic aortic constriction by intraperitoneal injection in the TAC + AA group.DMSO was intraperitoneally injected at the same dose and times 3 weeks after thoracic aortic constriction in TAC + DMSO group.Mice were given AA at the same dose and times by intraperitoneal injection for 8 weeks in the normal + AA group.And the myocardial tissues in each group were collected for the following analysis:(1)Cardiac ultrasonography was used to detect left ventricularanterior wall thickness(LVAWT),interventricular septal thickness(IVS),left ventricular end-diastolic diameter(LVEDD),left ventricular end-systolic diameter(LVESD)and left ventricular volume(LVV)at the end of left ventricular diastole and end systolic phase.(2)Htoxylin-eosin(H&E)staining was used to observe the cardiac morphology and myocardial cell surface area in the mice.(3)CHIP-Q-PCR was used to detect the acetylation level of hisotne H3K9 ac and the binding of p300-HAT and PCAF-HAT in the promoter region of cardiac nucleus transcription factor MEF2 A,and to investigate the regulatory relationship of cardiac nucleus transcription factor MEF2 A on myocardialhypertrophy relevant markers ANP and ?-MHC.(4)Real-time quantitative polymerase chain reaction(real-time PCR)was used to detect the mRNA expression level of heart nucleus transcription factor MEF2 A.(5)The expression level of histone H3K9 ac,p300-HAT,PCAF-HAT and myocardial hypertrophy markers ANP and?-MHC were tested by Western blot.(6)The effects of HATs inhibitor anacardic acid on the survival rate and cardiac ejection fraction in thoracic aortic constriction mice were observed.Results:(1)Cardiac ultrasonography results showed that compared with the TAC group,LVAWT,IVS,LVEDD,LVESD and LVV were significantly reduced in TAC +AA group(all P < 0.05).There was no significant difference between TAC + DMSO group and TAC group(all P > 0.05).(2)H&E staining results showed that the degree of myocardial hypertrophy and the surface area of myocardial cells were obviously getting better,compared with that of TAC group in TAC + AA group(P < 0.05),while there was no significant difference between TAC + DMSO group and TAC group(P > 0.05).(3)CHIP-Q-PCR results showed that the binding of p300-HAT and PCAF-HAT in the promoter region of MEF2 A were significantly higher than those ofnormal group in the TAC group mice(P < 0.05).The acetylation level of histone H3K9 ac in the promoter region of MEF2 A was aslo significantly higher than those ofnormal group in the TAC group mice(P < 0.05).However,the binding amount of p300-HAT and PCAF-HAT and the acetylation level of histone H3K9 ac in the promoter region of cardiac nucleus transcription factor MEF2 A were significantly lower than those in the TAC group in the TAC + AA group(all P < 0.05),while the differences between the TAC + DMSO group and the TAC group were not statistical significance(all P >0.05).(4)Real-time PCR results showed that anacardic acid could significantly down-regulate the mRNA expression level of the heart nucleus transcription factor MEF2 A in TAC mice(P < 0.05),while the difference between TAC + DMSO group and TAC group was not statistical significance(P > 0.05).(5)Western blot results showed that compared with the TAC group,the acetylation level of H3K9 ac was significantly down-regulated in the TAC + AA group(P < 0.05),while there was not significantly changed between the TAC + DMSO group and the TAC group(P > 0.05).At the same time,the expression levels of p300-HAT,PCAF-HAT and myocardial hypertrophy markers ANP and ?-MHC were significantly down-regulated compared with those of the TAC groupin the TAC + AA group(all P < 0.05),while the differences between the TAC + DMSO group and the TAC group were not statistically significant(all P > 0.05).Conclusions:(1)The hyperacetylation of histone H3K9 ac mediated by p300-HAT and PCAF-HAT is involved cardiac hypertrophy of mice caused by pressure overload.(2)HATs inhibitor anacardic acid can attenuate the pressure-overload myocardial hypertrophy through inhibiting hyperacetylation of histone H3K9 ac mediated by p300-HAT and PCAF-HAT to regulate the expression level of cardiac hypertrophy related genes.