【摘要】： 本研究是國家“973”課題《基于“藥性構(gòu)成三要素”的中藥藥性實(shí)質(zhì)研究》(課題編號2007CB512605)的一部分,課題著重考察中藥藥性形成和藥性表達(dá)的諸種因素,提出“藥性是藥物作用于機(jī)體狀態(tài)的、與化學(xué)成分相關(guān)的生物學(xué)效應(yīng)的歸納和概括”工作假說,認(rèn)為機(jī)體狀態(tài)、化學(xué)成分及生物學(xué)效應(yīng)是構(gòu)成藥性的三要素。其中,機(jī)體是藥物發(fā)揮作用的載體,是功效產(chǎn)出的依托,是構(gòu)成藥性的重要要素。不同的機(jī)體狀態(tài)如何用各種生理指標(biāo)進(jìn)行表征,以及不同狀態(tài)如何影響藥物生物效應(yīng)的表達(dá)是研究的內(nèi)容之一。 前期研究已證實(shí)不同機(jī)體狀態(tài)肝臟的代謝酶活性水平存在差異,本研究試圖以DNA甲基化水平作為研究的切入點(diǎn),尋求差異產(chǎn)生的可能原因。所謂DNA甲基化是指在DNA甲基轉(zhuǎn)移酶(DNA methyltransferase,DNMTs)的催化下,以S-腺苷甲硫氨酸(S-adenosylmethionine,SAM)為甲基供體,將甲基轉(zhuǎn)移到胞嘧啶的5'位置上。它作為哺乳動物基因組發(fā)生的最為常見的一種表觀遺傳學(xué)事件,可以通過對基因表達(dá)的影響和染色質(zhì)結(jié)構(gòu)的調(diào)節(jié)來制約細(xì)胞和組織的能力,并表現(xiàn)出機(jī)體的各種生理功能的差別。 本研究主要通過對虛寒狀態(tài)大鼠和正常狀態(tài)大鼠肝組織DNA甲基化水平的檢測,觀察不同機(jī)體狀態(tài)DNA甲基化水平的差別,從而為進(jìn)一步探尋DNA甲基化對機(jī)體狀態(tài)可能產(chǎn)生影響的分子機(jī)制奠定基礎(chǔ)。 主要進(jìn)行了以下2個方面的研究: 一建立測定大鼠肝組織DNA甲基化水平的高效液相色譜分析方法;用酚氯仿抽提法從大鼠肝組織提取基因組DNA,通過酸水解將DNA水解為單個堿基,最終用HPLC測定堿基的含量。 二以氫化可的松為造模劑塑造大鼠虛寒模型,對比研究虛寒狀態(tài)和正常狀態(tài)大鼠肝組織DNA甲基化水平的差異,結(jié)果分析表明,與正常狀態(tài)相比陽虛狀態(tài)組大鼠肝組織DNA甲基化水平明顯降低,具有顯著性差異(P＜0.01)。 分析虛寒和正常大鼠肝組織DNA甲基化水平存在差異的原因可能是氫化可的松通過與細(xì)胞質(zhì)中的糖皮質(zhì)激素受體結(jié)合,轉(zhuǎn)移到細(xì)胞核,調(diào)節(jié)基因的轉(zhuǎn)錄,對DNA甲基化水平產(chǎn)生了影響,表現(xiàn)出DNA甲基化水平的差異,而這種甲基化水平的差異影響了遺傳信息的表達(dá),最終通過蛋白質(zhì)大分子的功能活動表現(xiàn)出機(jī)體的各種生理功能和狀態(tài)差別。以上推測有待實(shí)驗(yàn)證實(shí)。
[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.
【學(xué)位授予單位】：北京中醫(yī)藥大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2009
【分類號】：R341

【引證文獻(xiàn)】

相關(guān)碩士學(xué)位論文 前2條

1 鄭飛;虛寒、虛熱狀態(tài)大鼠肝組織DNA甲基化水平的研究[D];北京中醫(yī)藥大學(xué);2010年

2 劉一洋;附子及其乙酸乙酯提取物對虛寒證模型大鼠的影響及機(jī)制研究[D];山東中醫(yī)藥大學(xué);2012年

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本文編號：2165766