EGCG对cTnIR193H限制型心肌病小鼠cTnI表达的影响

发布时间：2018-06-24 02:18

  本文选题：EGCG + 心脏舒张功能障碍　； 参考：《重庆医科大学》2017年硕士论文

【摘要】：表没食子儿茶素没食子酸酯(EGCG)源于茶多酚,具有多种生物活性。EGCG可以降低组蛋白去乙酰化酶(HDACs)的活性,从而调控基因的表达。本课题组前期对cTnI R193H限制型心肌病模型小鼠研究发现,正常cTnI表达降低、而突变cTnI异常表达是限制性心肌病(RCM)发病机制之一。研究还发现,EGCG干预R193H模型小鼠,可以降低模型小鼠心肌细胞对钙的敏感性,改善其舒张功能。本课题组前期对老年小鼠的研究发现,老年小鼠存在衰老性舒张功能障碍并同时伴有cTnI表达降低,给予EGCG可明显升高老年小鼠cTnI的表达,改善其衰老性舒张功能障碍。为此提出科学假设:EGCG还可能通过上调R193H小鼠正常cTnI的表达,参与改善舒张功能障碍。目的本实验利用EGCG对cTnI R193H模型小鼠进行3个月干预,以期探寻EGCG升高模型小鼠心脏正常cTn I的表达量,及其组蛋白乙酰化的调控机理。方法选择8周龄的健康spf级c57bl/6小鼠和ctnir193h雄性小鼠各30只,两种小鼠均分别分成egcg干预组、dmso干预组以及未干预组。egcg干预采用腹腔注射溶于dmso的egcg,按照各个小鼠体重,每1kg体重给予egcg50mg,dmso干预采用腹腔注射与egcg干预组溶剂容积一致的dmso,未干预组仅进行腹腔注射行为,但不注射。每周干预5天,干预3个月后收集各组小鼠心脏组织。分别用westernblot、rt-pcr、chip-q-pcr检测ctni的表达量;ctni、gata4及hdac1的mrna表达水平;ctni基因启动子区hdac1、gata4的结合水平及组蛋白h3k9乙酰化水平。结果r193h小鼠心脏egcg干预组与dmso干预组和未干预组比较结果显示:1.r193h小鼠egcg干预组正常ctni的表达量在mrna及蛋白水平均升高(p0.05),而正常的野生型c57小鼠则无此变化。2.r193h小鼠egcg干预组gata4的mrna表达水平升高,hdac1的mrna表达水平降低(p0.05)。3.r193h小鼠egcg干预组组蛋白h3k9乙酰化水平升高,ctni启动子区域hdac1结合水平降低,gata4的结合水平升高(p0.05)。结论egcg可通过抑制hdac1的表达及hdac1在ctni启动子区域的结合,促进组蛋白h3k9的乙酰化、心脏核心转录因子gata4的表达及gata4在ctni启动子区域的结合,上调限制型心肌病心肌正常ctni的表达。
[Abstract]:Epigallocatechin gallate (EGCG) is derived from tea polyphenols. EGCG can reduce the activity of histone deacetylase (HDACs) and regulate gene expression. The previous study of mice with cTnI R193H restricted cardiomyopathy showed that the expression of normal cTnI was decreased, and the abnormal expression of mutant cTnI was one of the pathogenesis of restrictive cardiomyopathy (RCM). It was also found that EGCG could decrease the sensitivity of cardiac myocytes to calcium and improve the diastolic function of R193H model mice. In the early stage of the study, we found that senile diastolic dysfunction and decreased cTnI expression were found in the aged mice. EGCG could significantly increase the expression of cTnI and improve the aging diastolic dysfunction in the aged mice. Therefore, the scientific hypothesis is that EGCG may be involved in improving diastolic dysfunction by up-regulating the expression of normal cTnI in R193H mice. Objective in this study, EGCG was used to intervene cTnI R193H model mice for 3 months, in order to explore the expression of normal cTnI and the regulation mechanism of histone acetylation. Methods A total of 30 healthy spf grade c57bl/6 mice and 30 ctnir193h male mice were randomly divided into two groups: the egcg intervention group and the non-intervention group. The mice were injected intraperitoneally with dmso soluble egcg according to the weight of each mouse. Each 1kg was treated with egcg 50mg / dmso by intraperitoneal injection with the same solvent volume as that in the egcg intervention group, but not by intraperitoneal injection in the non-intervention group. Heart tissues were collected after intervention for 5 days a week for 3 months. The expression level of ctni and mrna of hdac1 were detected by Western blottl rt-pcrtip-q-PCR, respectively. The binding level of hdac1 and gata4 in promoter region of CTNI gene and the acetylation level of histone h3k9 were detected. Results compared with dmso intervention group and dmso intervention group, the expression of normal ctni in egcg intervention group was higher than that in dmso intervention group (p0.05), while that in normal wild-type c57 mice was not. 2. R193h mice. The level of mrna expression of gata4 in egcg intervention group was increased (p0.05). The mrna expression of hdac1 was decreased (p0.05). 3. The level of h3k9 acetylation of histone was increased in egcg intervention group. The level of hdac1 binding in the promoter region of ctni decreased significantly (p0.05). Conclusion egcg can promote the acetylation of histone h3k9, the expression of heart core transcription factor gata4 and the binding of gata4 to ctni promoter by inhibiting the expression of hdac1 and the binding of hdac1 in ctni promoter region. To upregulate the expression of normal ctni in myocardium of restricted cardiomyopathy.
【学位授予单位】：重庆医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R725.4

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