糖尿病双生子肾虚血瘀证的表观遗传分析-epigenetic analysis of kidney deficiency and blood stasis syndrome in diabetic twins.docx

成都中医药大学博士学位论文糖尿病双生子肾虚血瘀证的表观遗传研究 成都中医药大学博士学位论文 糖尿病双生子肾虚血瘀证的表观遗传研究 PAGE PAGE 1 中文摘要 目的： 1. 从理论上探讨表观遗传学研究方法引入中医证候研究的可行性及合理性。 2. 精选一对典型的糖尿病双生子，筛选糖尿病肾虚血瘀证差异表达基因 mRNA 及差异表达 miRNA。 方法： 1. 理论研究法：对比表观遗传学与中医遗传学的概念、内容，指出引入表观 遗传学研究方法研究中医证候是恰当的拓展。 2. 证候调查法：用四诊、肾虚、血瘀等中医量表对糖尿病双生子家系进行证 候调查，宏观辨析其家系证候及其规律。 3. 实验分析法：联合运用 mRNA 表达谱芯片和 miRNA 芯片，筛选糖尿病肾虚血 瘀证的差异表达 mRNA、miRNA。 结果： 1. 引入表观遗传学方法揭示糖尿病的证候微观实质，能更好地诊断、预防和 治疗疾病。 2.糖尿病双生子家系调查显示，肾虚血瘀证有一定的先天影响和家族性因素， 糖尿病与肾虚血瘀证相关。 3. 筛选出一批代谢、免疫、转录调控类基因作为糖尿病肾虚血瘀证的候选选 基因。其中糖尿病候选差异基因 48 条，肾虚血瘀证的候选差异基因 27 条。 4. 筛选出糖尿病肾虚血瘀证差异表达 miRNA，并与候选差异表达基因相互验 证。筛选出糖尿病候选差异 miRNA 3 条，肾虚血瘀证的候选差异基因 miRNA 2 条 结论： 表观遗传学是当前研究的一个热点，是连接基因型和表型的纽带，中医证候、 疗效及中医遗传等很多问题都难以通过简单的单基因突变之类的经典遗传学内容 解释，需要引入表观遗传学的理论进行研究。而双生子是研究表观遗传的良好模型。 通过一个糖尿病肾虚血瘀双生子家系的证候调查，显示肾虚血瘀证有一定的遗 传影响，且糖尿病与肾虚血瘀证密切相关。联合运用表达谱芯片和 miRNA 芯片实验， 筛选出一批代谢、转录调控、免疫类基因作为糖尿病肾虚血瘀证的候选差异 mRNA， 并发现 miRNA-32 可能是糖尿病的差异表达 miRNA。 关键词：糖尿病，肾虚血瘀证，双生子，表观遗传，miRNA Objective: Abstract To Discusses the rationality and feasibility of using the theory and means of epigenetics to TCM research. To investigate microcosmic essentials of kidney deficiency with blood stasis syndrome at the gene level in diabetes. Method: Compare the concept and content of epigenetics with TCM genetics to get syndrome study need to introduce the theory and means of epigenetics. Clinical investigations with scaling tables on four diagnosis, kidney-deficiency, and blood stasis. mRNA and miRNNA microarray tests to screen out potential differentially expressed genes for type 2 diabetes with kidney-deficiency and blood stasis syndrome. Results: With applying the concepts and methods of epigenetics to the field of Chinese medicine research, it will provide new ideas, new methods and new technology in many aspects such as viscera state heredity, endowment, combination of disease and syndrome, and lay the foundation for prevention and treatment of disease. A pair of diabetes twins and their families were systematically investigated during the last consecutive two year period