The Effect Of SAHA,a Hdac Inhibitor,on Diastolic Function In Rcm Mouse

cTnI was a key point of diastolic dysfunction.In the present study,we demonstrated that the expression of ssTnI,fetal type of troponin I,and cTnI were influented by Histone acetylation.So,we question whether the expression of ssTnI or cTnI would increase by a higher Histone acetylation level.SAHA,a PAN-histone deacetylase inhibitor,increased the Histone H3 acetylation level exactly.In the study,we try to increased the expression of ssTnI and cTnI in restrictive cardiomyopathy mice to improve its diastolic dysfunction by SAHA.PART I: THE OPTIMUM DOSE OF SAHA TREATMENT IN C57BL/6 MICEObjective The optimum dose of SAHA,a PAN-histone deacetylase inhibitor,in vivo treatment is still unclear.It is essential that finding out the optimum dose of SAHA in vivo treatment.Methods Eighteen male adult C57BL/6 mouse were employed and divided into6 groups randomly,NORMAL group,DMSO group,SAHA-treatment group(10 ?g/g,25? g/g,50? g/g,100 ?g/g).Mice received subcutaneous injection with normal saline 1?l/g,DMSO 1?l/g,10?g/g SAHA,25?g/g SAHA,50 ? g/g SAHA,100 ? g/g SAHA for 7 days respectively.The acetylation level of Histone H3 was detected by Western Blotting.Results The level of acetylated Histone H3 of 50?g/g and 100?g/g SAHA group was incresased compared to Normal group,DMSO group,10?g/g SAHA group,25?g/g SAHA group(P<0.05).And there is no significance between 50?g/g SAHA group and 100?g/g SAHA group.Conclusion The optimum dose of SAHA would be 50?g per gram.PART II: ANIMAL REPRODUCTION: CTNI R193 H MICE Objective cTnI R193 H mice were useful for diastolic dysfunction study.In this part we aim to reproduce the RCM model,cTnI R193 H mice,and to make sure the diastolic dysfunction.Methods Raise the male and female cTnI R193 H mice in a 1/2 ratio together and then observe the vaginal plug closely the day after mating.Divided the pregnant mice into a single cage.The tail tissue were collected from the infant mice 10 days after birth.Then extracted the DNA for genotyping.Raise the infant cTnI R193 H mice for Experiment and Reproduce.Four male C57BL/6 mice as NORMAL group.All the mice were 3 month old.Detect the diastolic function of the two groups by Echocardiography.Results Compared to NORMAL group,IVRT,DT and Tei increased in cTnI R193 H group(P <0.05).No significance in E,A,E/A,LVFS,LVEF,SV,CO,LVIDd and LVVd between the two groups.Conclusions cTnI R193 H mice show a impaired diastolic function compared to normal mice.PART III: EFFECT OF SAHA ON DIASTOLIC DYSFUNCTION/DHFObjective The etiology of Restrictive Cardiomyopathy(RCM)is still unclear and some study showed that mutation troponin would lead a RCM presentation.The acetylated H3 in promoter of ssTnI/cTnI were down-regulated during the ssTnI silencing after birth and cTnI decreasing in aging.The efficiency of SAHA on influence of ssTnI/cTnI and diastolic dysfunction is unknown.This study aim to learn if the cardiac diastolic function in cTnI R193 H mice changes after SAHA treated.Methods Sixteen male cTnI R193 H mice were employed and divided into the following three groups: 1)CTRL group,2)DMSO group,3)SAHA group.All the mice were 3 month old.Mice received subcutaneous injection with normal saline 1?l/g,DMSO 1?l/g,50?g/g SAHA for 56 days respectively.The data of diastolic function were collected by Echocardiography.The level of acetylated Histone H3 and expression of cTnI as well as ssTnI were detected by Western Blotting,the following m RNA were detected by RT-q PCR: TNNI1,TNNI3,ATP2A2,HDAC1,HDAC2,HDAC3,the level of ac H3 in cTnI promoter were detected by CHIP.Results (1)The acetylated Histone H3 and ac H3 level of cTnI promoter in SAHA group was increased compared to CTRL group(P<0.05).(2)Compared to CTRL group,IVRT,DT,Tei increased in SAHA group(P <0.05),E and A decreased(P <0.05)as E/A have no significantly difference.LVFS,LVEF,SV and CO decreased in SAHA group(P <0.05).(3)Expression of cTnI show no significance among three groups as ssTnI was undetected.Relative expression of m RNA of TNNI1,TNNI3,ATP2A2,HDAC1,HDAC2,HDAC3 show no significance among three groups.Conclusions (1)Maybe SAHA treatment increased the acetylation level of Histone H3 and ac H3 level of c Tni promoter in cTnI R193 H mice.(2)SAHA may not do help to diastolic dysfunction of RCM as even make the diastolic function and systolic function worse.(3)SAHA may not influent the expression of cTnI nor ssTnI as the m RNA expression of TNNI1,TNNI3,ATP2A2,HDAC1,HDAC2,HDAC3.