致心律失常性右室心肌病心肌能量代谢研究

发布时间：2018-07-04 08:13

本文选题：致心律失常性右室心肌病 + 扩张型心肌病　；参考：《北京协和医学院》2017年博士论文

【摘要】：背景致心律失常性右室心肌病(ARVC/D)以右室心肌纤维脂肪替换为主要病理特征,以右室心律失常和右室心功能下降为主要临床表现。相比于其他类型心肌病,ARVC/D以右室心肌受累为主,而左室心肌的病变程度较轻。目前对ARVC/D发病机制的研究发现,ARVC/D右室心肌的纤维脂肪替换与心肌脂肪酸代谢调节紊乱有关。这说明心肌的能量代谢重塑在疾病发生过程中发挥重要作用。但是,目前关于ARVC/D心肌的葡萄糖底物利用、氨基酸代谢、三羧酸循环和氧化应激代谢重塑过程,以及与ARVC/D右室心肌受累的关联尚不清楚。目的本研究的目的是探究ARVC/D左室与右室心肌的能量代谢改变,以从代谢角度解析ARVC/D右室心功能下降和右室心律失常的发生机制,以期为ARVC/D的临床治疗提供新的线索。方法马松染色检测非心衰心肌、扩张型心肌病(DCM)和ARVC/D患者的心肌纤维化和心肌细胞体积,并用Image-Pro Plus(IPP)软件进行定量分析。透射电子显微镜观察心肌细胞内脂滴沉积。qPCR检测心衰心肌标志物脑钠肽(BNP)的mRNA表达水平。Spearman相关分析比较超声心动图参数(RVD,LVEDD,LVEF)与心肌BNP的mRNA表达水平高低的相关性。qPCR检测7例非心衰心肌、8例DCM和8例ARVC/D患者的左室与右室心肌中参与脂肪酸底物利用、葡萄糖底物利用、谷氨酸代谢、三羧酸循环和氧化应激相关转运蛋白、转录因子、代谢酶的mRNA表达水平。超高效液相色谱质谱(UPLC-MS)技术检测26例ARVC/D患者的左室与右室心肌中主要代谢中间产物的含量。人线粒体DNA(mtDNA)检测试剂盒检测线粒体DNA拷贝数相对含量。TUNEL凋亡检测试剂盒检测心肌细胞凋亡。结果与DCM患者相比,ARVC/D患者左室扩张(49.50 ± 11.30mm vs.65.50 ±14.50mm,ARVC/D vs.DCM,LVEDD,P0.001)和左室收缩功能障碍(43.50±19.90%vs.22.00±6.90%,ARVC/D vs.DCM,LVEF,P0.001)程度较轻,但右室扩张程度(43.50± 13.00mm vs.27.00±4.30mm,ARVC/D vs.DCM,RVD,P0.001)较重。ARVC/D右室心肌纤维化,肌纤维降解,心肌细胞体积增大,心肌细胞胞浆内可见明显的脂滴沉积。ARVC/D右室心肌BNP的表达水平比DCM右室高12.7倍。ARVC/D和DCM右室BNP的表达水平高低与右室内径(RVD)大小呈正相关关系(r=0.692,P=0.003)。ARVC/D左室心肌与DCM左室心肌BNP的表达水平之间没有显著差异(P=0.247)。ARVC/D和DCM左室BNP的表达水平高低与左室舒张末内径(LVEDD)(P=0.122)和左室射血分数(LVEF)(P=0.138)之间也没有显著相关性。与非心衰对照的左室与右室心肌相比,ARVC/D和DCM的左室与右室心肌中参与脂肪酸底物利用(SLC27A1,SLC27A6,FABP3,CPT1B,ACADVL,ACADS,PPARA)、甘油三酯平衡(GPAT2,PNPLA2,PLIN5)、葡萄糖底物利用(SLC2A4,PFKM,PKM,PDHB)、线粒体谷氨酸氧化(SLC1A5、GLUD1)、三羧酸循环(CS)的重要基因的mRNA表达水平下调或呈下降趋势。与DCM右室心肌相比,ARVC/D右室心肌中参与脂肪酸代谢(SLC27A1,SLC27A6,FABP3,CPT1B,ACADVL,ACADS,PPARA)、甘油三酯平衡(GPAT2,PNPLA2,PLIN5)、葡萄糖代谢(SLC2A4,PFKM,PKM,PDHB)、三羧酸循环(CS)的大部分基因的mRNA表达水平下调或呈下降趋势。与DCM左室心肌相比,ARVC/D左室心肌中上述基因的mRNA表达水平没有显著差异。我们发现,参与谷氨酰胺-谷氨酸代谢的酶类,如谷氨酰胺酶(GLS)、谷氨酸-氨连接酶(GLUL)、氨甲酰基-磷酸合成酶(CPS1),和抗氧化酶谷胱甘肽还原酶(GSR)的基因mRNA表达水平只在ARVC/D和DCM右室下调,而在ARVC/D和DCM左室却没有显著改变。此外,ARVC/D右室心肌中尿素循环代谢中间产物和谷胱甘肽(GSH)含量低于左室(P=0.0013),而氧化型谷胱甘肽(GSSG)含量高于左室(P=0.0039)。与非心衰右室心肌相比,ARVC/D右室心肌线粒体DNA含量下降(P=0.0013)。ARVC/D右室残余心肌细胞存在显著的凋亡改变。结论第一,ARVC/D与DCM左室与右室心肌都存在脂肪酸、葡萄糖、谷氨酸底物利用下降和三羧酸循环障碍。但是,ARVC/D右室心肌能量代谢受损更加严重,产生能量减少,导致ARVC/D右室心功能下降。第二,ARVC/D右室心肌存在谷氨酰胺-谷氨酸代谢障碍和谷胱甘肽代谢稳态失衡,引起右室心肌氧化应激损伤,这可能与ARVC/D右室心肌电生理紊乱和心肌凋亡发生相关。
[Abstract]:Background arrhythmogenic right ventricular cardiomyopathy (ARVC/D) is the main pathological feature of right ventricular myocardial fiber fat replacement, with right ventricular arrhythmia and right ventricular function decline as the main clinical manifestation. Compared with other types of cardiomyopathy, ARVC/D is mainly involved in right ventricular myocardium, and left ventricular myocardium is lighter. The pathogenesis of ARVC/D is present. The study found that the replacement of fibrous fat in the right ventricular myocardium of the ARVC/D is related to the disorder of the metabolic regulation of the myocardial fatty acid. This suggests that the remodeling of the energy metabolism of the myocardium plays an important role in the development of the disease. However, the use of glucose substrates, the metabolism of amino acids, the three carboxylic acid cycle and the remodeling process of oxidative stress metabolism are present in the ARVC/D myocardium. The purpose of this study is to explore the energy metabolism changes in the left ventricular and right ventricular myocardium of the ARVC/D in order to analyze the mechanism of ARVC/D right ventricular dysfunction and right ventricular arrhythmia from the metabolic angle, so as to provide new clues for the clinical treatment of ARVC/D. Method Masson staining was used to detect the clinical treatment of the right ventricular myocardium. Myocardial fibrosis and volume of myocardium in patients with non heart failure, dilated cardiomyopathy (DCM) and ARVC/D and quantitative analysis by Image-Pro Plus (IPP) software. Transmission electron microscopy (TEM) observation of lipid droplet deposition in cardiac myocytes,.QPCR detection of cardiac markers of heart failure, BNP,.Spearman correlation analysis and comparison of echocardiography The correlation between RVD, LVEDD, LVEF and the mRNA expression level of myocardial BNP detected in 7 cases of non heart failure myocardium, 8 cases of DCM and 8 cases of ARVC/D patients involved in the use of fatty acid substrates, glucose substrate utilization, glutamate metabolism, three carboxylic acid cycle and oxidative stress related transporters, transcription factors, metabolic enzymes. MRNA expression level. The content of major metabolic intermediates in left ventricular and right ventricular myocardium in 26 patients with ARVC/D was detected by ultra high performance liquid chromatography mass spectrometry (UPLC-MS). Mitochondrial DNA (mtDNA) detection kit was used to detect the relative content of mitochondrial DNA copy number by.TUNEL apoptosis detection kit to detect cardiomyocyte apoptosis. Compared with DCM patients, the results were compared with those of DCM patients. The left ventricular dilatation of ARVC/D patients (49.50 + 11.30mm vs.65.50 + 14.50mm, ARVC/D vs.DCM, LVEDD, P0.001) and left ventricular systolic dysfunction (43.50 + 19.90%vs.22.00 + 6.90%, ARVC/D vs.DCM, LVEF, etc.) were relatively mild, but the right ventricular dilatation degree was more severe than that of the right ventricular myocardium. Muscle fiber degradation, myocardial cell volume increased, the expression level of BNP in.ARVC/D right ventricular myocardium was 12.7 times higher than that of DCM right ventricle,.ARVC/D and DCM right ventricle BNP expression level were positively correlated with right ventricular diameter (RVD) size (r=0.692, P=0.003).ARVC/D left ventricular myocardium and DCM left ventricular myocardium BNP. There was no significant difference between the levels of.ARVC/D and DCM (P=0.247) and left ventricular BNP expression level with left ventricular end diastolic diameter (LVEDD) and left ventricular ejection fraction (LVEF) (LVEF). Compared with the left ventricular and right ventricular myocardium, the left ventricular and right ventricular myocardium in ARVC/D and DCM were involved in fatty acid bottom. SLC27A1, SLC27A6, FABP3, CPT1B, ACADVL, ACADS, PPARA), triglyceride balance (GPAT2, PNPLA2, PLIN5). The glucose substrate uses (SLC2A4, PFKM), mitochondrial glutamate oxidation, the expression level of the important genes of the three carboxylic acid cycle down or downward trend. The muscles involved in fatty acid metabolism (SLC27A1, SLC27A6, FABP3, CPT1B, ACADVL, ACADS, PPARA), triglyceride balance (GPAT2, PNPLA2, PLIN5), and glucose metabolism (SLC2A4, PFKM, and PLIN5). The expression level of most genes of the three carboxylic acid cycle decreased or declined. There was no significant difference in expression level. We found that the genes involved in glutamine - glutamic acid metabolism, such as glutaminase (GLS), glutamate ammonia ligase (GLUL), aminoformyl phosphate synthetase (CPS1), and antioxidant enzyme glutathione reductase (GSR) gene mRNA expression level only down in the right ventricle of ARVC/D and DCM, and in ARVC/D and DCM left ventricle. In addition, the content of intermediate product of urea metabolism and glutathione (GSH) in ARVC/D right ventricular myocardium was lower than that of left ventricle (P=0.0013), and the content of oxidized glutathione (GSSG) was higher than that of left ventricle (P=0.0039). The mitochondrial DNA content of ARVC/D right ventricular myocardium decreased (P=0.0013).ARVC/D right ventricular remnant myocardium compared with non heart failure right ventricular myocardium. There were significant apoptotic changes in the cell. Conclusion first, both ARVC/D and DCM left ventricular and right ventricular myocardium have fatty acid, glucose, glutamic acid substrate utilization and three carboxylic acid circulation disorder. However, the energy metabolism of ARVC/D right ventricular myocardium is more severely damaged, resulting in reduced energy, resulting in the decrease of ARVC /D right ventricular function. Second, ARVC/D right ventricular myocardium exists. The disorder of glutamine glutamic acid metabolism and the homeostasis of glutathione metabolism may cause oxidative stress injury in the right ventricular myocardium, which may be associated with the electrophysiological disorder of the right ventricular myocardium and the occurrence of myocardial apoptosis in ARVC/D.
【学位授予单位】：北京协和医学院
【学位级别】：博士
【学位授予年份】：2017
【分类号】：R542.2

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