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Effect of Angiotensin Ⅱ on the Expression of Clock Genes in Cultured Neonatal Cardiomyocytes and Myocardial Fibroblasts

Author: HuangXinYan
Tutor: LiQingPing
School: Nanjing Medical University
Course: Cardiovascular Pharmacology
Keywords: Angiotensin-converting enzyme Angiotensin II Myocardial hypertrophy Cardiomyocytes Myocardial fibroblasts Clock gene
CLC: R33
Type: Master's thesis
Year: 2005
Downloads: 153
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Abstract


Blood pressure in healthy persons possesses circadian rhythm. The level of blood pressure is in correlation with Ang II. Angiotensin-converting enzyme acts on Ang I and converts Ang I into Ang II, which is the important source of Ang II. This study observed the circadian rhythm of serum ACE relative activity in mature SHR rat, RHR rat and control SD rat, to analysis the relationship between the circadian rhythm of blood pressure and serum ACE relative activity level.Ang II in blood circulation relates with acute change of blood pressure. But Ang II coming form autocrine ways and paracrine ways relates with chronic regulation of heart structure, and results in myocardial hypertrophy. The histiocyte morphologic transformation of myocardial hypertrophy includes hypertrophy of cardiomyocytes, hyperplasia of myocardial fibroblasts, accumulation of collogen, et al. That is remodeling of cardiovascular system.Cardiomyocytes and myocardial fibroblasts are the important carriers of heart structure and physiology function, such as the development of heart, the contract and diastole of heart. In addition, they are involved in pathology process of cardiovascular diseases, such as myocardial hypertrophy of hypertension, remodeling, trauma and restore. So,investigation of cardiomyocytes and myocardial fibroblasts is of great benefit to comprehending the mechanism of myocardial hypertrophy.All eukaryotes possess endogenous timekeeping mechanisms, which allow an organism to predict cyclical changes in its environment and to adjust their physiology accordingly. Biology clock locates in the hypothalamus of the brain （the suprachiasmatic nucleus SCN） and peripheral tissues. The cultured cells also possess the biology clock. Zeitgebers modulate the expression of clock genes, which affect the translation of the down-stream gene.Angiotensin II, which can induce the development of myocardial hypertrophy, is the important member of RAS system. This subject is to study the expression of clock genes in the cultured cardiomyocytes and myocardial fibroblasts, and to explore the role of circadian clocks in the process of cardiac hypertrophy.This study includes three parts:1. The circadian rhythm of serum ACE relative activity in mature SHR rat, RHR rat and control SD rat.Mature SHR rats, RHR rat and SD rats of normal blood pressure were fed in closed environment for two weeks to build the circadian cycles model. And then we took the serum to measure the activity of ACE. We found that the curves of ACE relative activity in SHR rat, RHR rat and SD rat were different from each other. They are vale shape, the shape ofChinese characters-" 山 " and the shape of English word - " W". The ACE relative activity in SHR rat, RHR rat and SD rat was not different markedly in t test. All these hinted that the activity of ACE in SHR rat and RHR rat was abnormal.2. Effect of Ang II on expression of clock genes in cultured neonatal cardiomyocytesCardiomyocytes were isolated from the ventricles of the SD rat heart by enzymatic dissociation （0.1%trysin） and were purified by different adhesion. The cells were incubated in DMEM nutrient medium. After 72h, we changed the nutrient medium and added Ang II and（or） Tel to a final concentration at 0.1 u mol ? L’1 once a day for three times. The expression of clock genes in cardiomyocytes was detected using RT-PCR. We found that expression of clock genes of cardiomyocytes treated with 0.1 u mol.L-1 Ang II was higher than that of normal cardiomyocytes, and Tel can block the high expression of clock genes induced by Ang II.3. Effect of Ang II on expression of clock genes in cultured neonatal myocardial flbroblasts.Myocardial fibroblasts were isolated from the ventricles of the SD rat heart by enzymatic dissociation （0.1%trysin） and were purified by different adhesion. The cells were incubated in DMEM nutrient medium. After 72h, we changed the nutrient medium and added Ang II and（or） Tel to a final concentration at 0.1 u mol . L-1 once a day for three times. The expression

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