中国汉族女性多囊卵巢综合征和卵巢早衰的相关遗传学研究

发布时间：2018-03-15 11:40

本文选题：多囊卵巢综合征　切入点：ANRIL　出处：《安徽医科大学》2014年硕士论文　论文类型：学位论文

【摘要】：目的本研究目的是探究ANRIL多态性与中国汉族人群多囊卵巢综合征的易感性。方法在中国汉族女性中收集139例多囊卵巢综合征患者和141例对照者，提取DNA，设计探针引物通过实时荧光PCR TaqMan探针方法对INK4区域反义非编码RNA(ANRIL)的一个SNPrs1333049进行基因分型分析。结果在PCOS患者中，各项生化指标和临床表现在各基因型之间差异无统计学意义。通过分析TaqMan探针基因分型的结果，rs1333049位可以分为3种基因型：CC，CG,GG。通过统计分析，PCOS患者和对照者间基因型分布和等位基因频率差异无统计学意义(2=1.87, P=0.17;2=3.25, P=0.20)。结论染色体9p21上rs1333049多态性与PCOS高雄激素血症无关。目的为探究CITED2基因在卵巢早衰的疾病发生过程中的作用，本研究将在卵巢早衰病人中筛查CITED2基因潜在的突变，以确认CITED2基因与该疾病的关系。方法收集189例特发性卵巢早衰患者和200例健康女性对照的外周血样，提取DNA后，PCR扩增直接测序探查CITED2基因启动子区和编码区的变异。随后，构建相应质粒，通过哺乳动物双杂交,双萤光素酶报告实验,亚细胞定位等功能试验检验CITED2基因中突变位点的作用。结果对189例卵巢早衰患者和200例健康对照女性的CITED2基因编码区和启动子区测序峰图，进行chromas软件解读比对，一位POF患者的CITED2基因编码区和启动子区各发现一个突变，分别为c.634AG Met212Val和T-339G，这两个突变均未在正常对照中发现,且这两个突变均不在研究热点区域。通过体外功能实验，，发现野生型CITED2可以通过调节HIF1-a来调控VEGF的表达。而实验发现的编码区突变在双荧光实验和哺乳动物双杂交试验中均可通过竞争性抑制HIF1-a与CBP/300结合，来恢复VEGF的表达。亚细胞定位可见野生型和突变型CITED2对于细胞定位并无明显影响。启动子中的突变相较于野生型可以升高基因的表达。结论首次在中国汉族特发性POF患者中发现CITED2基因中的突变，并通过功能实验进一步明确了突变可能的作用，并由此推断对于其可能与POF的发生相关。
[Abstract]:Objective to explore the relationship between ANRIL polymorphism and susceptibility to polycystic ovary syndrome (PCOS) in Chinese Han population. Methods A total of 139 polycystic ovary syndrome (PCOS) patients and 141 controls were collected from Han women in China, and a probe primer was designed to analyze a SNPrs1333049 of INK4 region antisense noncoding RNAs by real-time fluorescence PCR TaqMan probe method. Results in PCOS patients, There was no significant difference in biochemical indexes and clinical manifestations among genotypes. The results of TaqMan probe genotyping could be divided into three genotypes: 1: CCG GG. By statistical analysis of genotypes between PCOS patients and controls, the genotypes of rs1333049 could be divided into 3 genotypes. There was no significant difference in distribution and allele frequency (P = 1.87, P = 0.17, P = 0.20, P = 0.20). Conclusion rs1333049 polymorphism on chromosome 9p21 is not associated with PCOS hyperandrogenemia. Objective to investigate the role of CITED2 gene in the pathogenesis of premature ovarian failure (POF). In this study, potential mutations of CITED2 gene were screened in premature ovarian failure patients to confirm the relationship between CITED2 gene and the disease. Methods Peripheral blood samples of 189 patients with idiopathic premature ovarian failure and 200 healthy female controls were collected. After DNA was extracted, the mutation of promoter and coding region of CITED2 gene was detected by direct sequencing. Then the corresponding plasmids were constructed. The function of mutation site in CITED2 gene was tested by mammalian two-hybrid, double-luciferase report and subcellular localization. Results the CITED2 gene coding region and promoter region were sequenced from 189 preterm ovarian failure patients and 200 healthy women, and chromas software was used to analyze and compare. One mutation was found in the CITED2 gene coding region and one in the promoter region in one POF patient, and one mutation was found in the CITED2 gene coding region and promoter region in one POF patient. C. 634AG Met212Val and T-339G. these two mutations were not found in normal control, and neither of them was found in the hot research area. It was found that wild-type CITED2 could regulate the expression of VEGF by regulating HIF1-a, and the coding region mutation could be inhibited by competitive inhibition of HIF1-a binding to CBP/300 in both bifluorescence and mammalian two-hybrid experiments. Subcellular localization showed that wild type and mutant CITED2 had no significant effect on cellular localization. Mutations in promoter could increase gene expression compared with wild type. Conclusion the mutation of CITED2 gene was first found in Chinese Han patients with idiopathic POF, and the possible role of the mutation was further clarified by functional experiments, and it was deduced that the mutation might be related to the occurrence of POF.
【学位授予单位】：安徽医科大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R711.75

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