正常及虚寒状态大鼠肝脏的DNA甲基化水平差异
发布时间：2018-08-05 12:15
【摘要】： 本研究是国家“973”课题《基于“药性构成三要素”的中药药性实质研究》(课题编号2007CB512605)的一部分,课题着重考察中药药性形成和药性表达的诸种因素,提出“药性是药物作用于机体状态的、与化学成分相关的生物学效应的归纳和概括”工作假说,认为机体状态、化学成分及生物学效应是构成药性的三要素。其中,机体是药物发挥作用的载体,是功效产出的依托,是构成药性的重要要素。不同的机体状态如何用各种生理指标进行表征,以及不同状态如何影响药物生物效应的表达是研究的内容之一。 前期研究已证实不同机体状态肝脏的代谢酶活性水平存在差异,本研究试图以DNA甲基化水平作为研究的切入点,寻求差异产生的可能原因。所谓DNA甲基化是指在DNA甲基转移酶(DNA methyltransferase,DNMTs)的催化下,以S-腺苷甲硫氨酸(S-adenosylmethionine,SAM)为甲基供体,将甲基转移到胞嘧啶的5'位置上。它作为哺乳动物基因组发生的最为常见的一种表观遗传学事件,可以通过对基因表达的影响和染色质结构的调节来制约细胞和组织的能力,并表现出机体的各种生理功能的差别。 本研究主要通过对虚寒状态大鼠和正常状态大鼠肝组织DNA甲基化水平的检测,观察不同机体状态DNA甲基化水平的差别,从而为进一步探寻DNA甲基化对机体状态可能产生影响的分子机制奠定基础。 主要进行了以下2个方面的研究: 一建立测定大鼠肝组织DNA甲基化水平的高效液相色谱分析方法;用酚氯仿抽提法从大鼠肝组织提取基因组DNA,通过酸水解将DNA水解为单个碱基,最终用HPLC测定碱基的含量。 二以氢化可的松为造模剂塑造大鼠虚寒模型,对比研究虚寒状态和正常状态大鼠肝组织DNA甲基化水平的差异,结果分析表明,与正常状态相比阳虚状态组大鼠肝组织DNA甲基化水平明显降低,具有显著性差异(P＜0.01)。 分析虚寒和正常大鼠肝组织DNA甲基化水平存在差异的原因可能是氢化可的松通过与细胞质中的糖皮质激素受体结合,转移到细胞核,调节基因的转录,对DNA甲基化水平产生了影响,表现出DNA甲基化水平的差异,而这种甲基化水平的差异影响了遗传信息的表达,最终通过蛋白质大分子的功能活动表现出机体的各种生理功能和状态差别。以上推测有待实验证实。
[Abstract]:This research is part of the national "973" project "the essential study of traditional Chinese Medicine based on" three elements of Drug composition "(subject number 2007CB512605), which focuses on the factors of the formation and expression of traditional Chinese medicine. This paper puts forward the working hypothesis that "the drug acts on the state of the body and the biological effects related to the chemical composition". It is considered that the state of the body, the chemical composition and the biological effect are the three elements that constitute the drug. Among them, the body is the carrier of the drug, the support of the efficacy and the important factor of the drug. One of the contents of the study is how different organism states are characterized by various physiological indexes and how different states affect the expression of drug biological effects. Previous studies have confirmed that there are differences in the activity of metabolic enzymes in different body states. This study attempts to use the level of DNA methylation as the starting point to explore the possible causes of the difference. The so-called DNA methylation refers to the transfer of methyl to the 5 'position of cytosine with S-adenosylmethionine (S-adenosylmethionine) as methyl donor catalyzed by DNA methyltransferase (DNA). As one of the most common epigenetic events occurring in the mammalian genome, it restricts the ability of cells and tissues to regulate gene expression and chromatin structure. It also shows the difference of various physiological functions of the body. The purpose of this study was to observe the difference of DNA methylation level in liver tissues of rats with deficiency and cold and normal rats, in order to observe the difference of DNA methylation levels in different organism states. The results provide a basis for further exploring the molecular mechanism of DNA methylation affecting the state of organism. The following two aspects were studied: one was to establish a high performance liquid chromatography (HPLC) method for the determination of DNA methylation in rat liver tissue. Genomic DNA was extracted from rat liver tissue by phenol chloroform extraction method. DNA was hydrolyzed into a single base by acid hydrolysis, and the content of base was determined by HPLC. Secondly, hydrocortisone was used as a model to establish rat model of deficiency cold. The difference of DNA methylation level in liver tissue between rats with deficiency cold state and normal state was studied. Compared with the normal state, the DNA methylation level of liver tissue in the Yang deficiency group was significantly lower than that in the normal group (P < 0. 01). The reason for the difference in DNA methylation between deficiency cold and normal rat liver may be that hydrocortisone is transferred to the nucleus by binding with glucocorticoid receptor in cytoplasm to regulate gene transcription. It has an effect on DNA methylation level, showing the difference of DNA methylation level, and the difference of this methylation level affects the expression of genetic information. Finally, the functional activities of protein macromolecules show various physiological functions and state differences. The above conjecture needs to be verified by experiments.
【学位授予单位】：北京中医药大学
【学位级别】：硕士
【学位授予年份】：2009
【分类号】：R341
本文编号：2165766
[Abstract]:This research is part of the national "973" project "the essential study of traditional Chinese Medicine based on" three elements of Drug composition "(subject number 2007CB512605), which focuses on the factors of the formation and expression of traditional Chinese medicine. This paper puts forward the working hypothesis that "the drug acts on the state of the body and the biological effects related to the chemical composition". It is considered that the state of the body, the chemical composition and the biological effect are the three elements that constitute the drug. Among them, the body is the carrier of the drug, the support of the efficacy and the important factor of the drug. One of the contents of the study is how different organism states are characterized by various physiological indexes and how different states affect the expression of drug biological effects. Previous studies have confirmed that there are differences in the activity of metabolic enzymes in different body states. This study attempts to use the level of DNA methylation as the starting point to explore the possible causes of the difference. The so-called DNA methylation refers to the transfer of methyl to the 5 'position of cytosine with S-adenosylmethionine (S-adenosylmethionine) as methyl donor catalyzed by DNA methyltransferase (DNA). As one of the most common epigenetic events occurring in the mammalian genome, it restricts the ability of cells and tissues to regulate gene expression and chromatin structure. It also shows the difference of various physiological functions of the body. The purpose of this study was to observe the difference of DNA methylation level in liver tissues of rats with deficiency and cold and normal rats, in order to observe the difference of DNA methylation levels in different organism states. The results provide a basis for further exploring the molecular mechanism of DNA methylation affecting the state of organism. The following two aspects were studied: one was to establish a high performance liquid chromatography (HPLC) method for the determination of DNA methylation in rat liver tissue. Genomic DNA was extracted from rat liver tissue by phenol chloroform extraction method. DNA was hydrolyzed into a single base by acid hydrolysis, and the content of base was determined by HPLC. Secondly, hydrocortisone was used as a model to establish rat model of deficiency cold. The difference of DNA methylation level in liver tissue between rats with deficiency cold state and normal state was studied. Compared with the normal state, the DNA methylation level of liver tissue in the Yang deficiency group was significantly lower than that in the normal group (P < 0. 01). The reason for the difference in DNA methylation between deficiency cold and normal rat liver may be that hydrocortisone is transferred to the nucleus by binding with glucocorticoid receptor in cytoplasm to regulate gene transcription. It has an effect on DNA methylation level, showing the difference of DNA methylation level, and the difference of this methylation level affects the expression of genetic information. Finally, the functional activities of protein macromolecules show various physiological functions and state differences. The above conjecture needs to be verified by experiments.
【学位授予单位】：北京中医药大学
【学位级别】：硕士
【学位授予年份】：2009
【分类号】：R341
【引证文献】
相关硕士学位论文 前2条
1 郑飞;虚寒、虚热状态大鼠肝组织DNA甲基化水平的研究[D];北京中医药大学;2010年
2 刘一洋;附子及其乙酸乙酯提取物对虚寒证模型大鼠的影响及机制研究[D];山东中医药大学;2012年
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