兰州大学机构库 >公共卫生学院
β-hCG介导的VEGF-MEK ERK信号通路在早期绒毛血管生成和滋养细胞凋亡中的作用研究
Alternative TitleThe Role of beta hCG-Mediated VEGF-MEK/ERK Signaling Pathway in Early Villi Angiogenesis and Trophoblast Cell Apoptosis
井光壮
Subtype硕士
Thesis Advisor李芝兰
2020-05-24
Degree Grantor兰州大学
Place of Conferral兰州
Degree Name医学硕士
Degree Discipline儿少卫生与妇幼保健学
Keyword稽留流产 绒毛组织 绒毛血管生成 VEGF-MEK/ERK信号通路 滋养细胞凋亡
Abstract目的:通过检测血清&beta-hCG、VEGF含量以及绒毛组织中VEGF-MEK/ERK信号通路和滋养细胞凋亡相关因子的表达水平,探讨稽留流产的发病机制。 方法:(1)收集稽留流产和同孕周、同年龄组因意外妊娠行人工流产术正常早孕妇女血清和绒毛组织标本各12例,所有受试者均无职业病有害因素暴露史。(2)采用放射免疫分析方法检测两组血清&beta-hCG含量。(3)采用酶联免疫吸附实验(ELISA)检测两组血清VEGF含量。(4)制备绒毛组织HE切片,光镜下观察绒毛组织病理学改变。(5)采用免疫组织化学技术CD34定位检测绒毛组织血管生成情况,并计算微血管密度(MVD)。(6)qRT-PCR检测绒毛组织VEGF-MEK/ERK信号通路相关基因VEGF、VEGFR2、RAS、MEK1/2、ERK1/2 mRNA表达水平以及凋亡基因Bcl-2、Bax、Cyt c、Casepase 3 mRNA表达水平。(7)采用Wes全自动蛋白分析技术检测绒毛组织VEGF-MEK/ERK信号通路相关蛋白VEGF、VEGFR2、RAS、MEK1/2、p-MEK1/2、ERK1/2、p-ERK1/2表达水平以及凋亡相关蛋白Bcl-2、Bax、Cyt c、Casepase 3表达水平。 结果:(1)血清&beta-hCG、VEGF含量:稽留流产组血清&beta-hCG、VEGF水平均低于正常早孕组(P<0.05)。(2)绒毛组织病理改变:镜下可见稽留流产组绒毛组织滋养层明显变薄,细胞数量减少,细胞结构紊乱,部分滋养层细胞缺失。(3)绒毛组织血管生成情况:稽留流产组绒毛间质内血管小,管腔狭窄,数量明显减少。微血管计数结果显示:稽留流产组绒毛组织中微血管密度低于正常早孕组(P<0.05)。(4)绒毛组织VEGF-MEK/ERK信号通路相关因子表达情况:稽留流产组绒毛组织中VEGF、VEGFR2、MEK1/2、ERK1/2 mRNA表达水平均低于正常早孕组(P<0.05)稽留流产组绒毛组织中VEGF、RAS、p-MEK1/2、p-ERK1/2蛋白表达水平及p-MEK/MEK、p-ERK/ERK比值均低于正常早孕组(P<0.05)。(5)绒毛组织滋养细胞凋亡相关因子表达情况:稽留流产组绒毛组织中Bcl-2 mRNA表达水平低于正常早孕组(P<0.05)稽留流产组绒毛组织中Bax、Cyt c和Caspase 3 mRNA表达水平均高于正常早孕组(P<0.05)稽留流产组绒毛组织中Bcl-2 蛋白表达水平低于正常早孕组(P<0.05)稽留流产组绒毛组织中Bax/Bcl-2比值、Cyt c和Caspase 3 蛋白表达水平均高于正常早孕组(P<0.05)。(6)绒毛组织VEGF、ERK1/2蛋白表达水平与绒毛组织微血管密度(MVD)的相关性分析:在稽留流产组中,ERK1/2蛋白表达水平与绒毛组织微血管密度(MVD)成正相关(r=0.909,P<0.05)。 结论:孕早期孕妇体内&beta-hCG生成不足,可能使VEGF-MEK/ERK信号通路的表达下调,继而减少绒毛血管生成和激活线粒体凋亡通路,最终导致孕早期绒毛血管生成障碍和滋养细胞功能异常,可能是稽留流产发生的重要机制。
Other AbstractObjective: To explore the mechanism of missed abortion, the contents of serum &beta-hCG, VEGF and the expression levels of VEGF-MEK/ERK signaling pathway and trophoblast cell apoptosis related factors of villous tissues were measured and analyzed. Methods: (1)Serum specimens and villous tissue specimens were collected from 12 cases of missed abortion and 12 termination of pregnancy by induced abortion due to unintended pregnancy in normal pregancy. All subjects were in the same age group, at the same gestational week, and had no history of exposure to occupational hazards. (2)Radioimmunoassay was used to detect &beta-hCG levels in serum of the two groups. (3)Enzyme-linked immunosorbent assay was used to detect VEGF levels in serum. (4)Hematoxylin-eosin sections of villous tissue were prepared and the histopathological changes of villous tissue were observed by light microscopy. (5)Immunohistochemistry CD34 positioning was used to detect the villous angiogenesis. (6)The mRNA expression levels of VEGF, VEGFR2, RAS, MEK1/2 and ERK1/2 related genes in villi, as well as the mRNA expression levels of apoptotic genes Bcl-2, Bax, Cyt c and Casepase 3 was detected by qRT-PCR. (7)The expression levels of VEGF-MEK/ERK signaling pathway related proteins including VEGF, VEGFR2, RAS, MEK1/2, p-MEK1/2, ERK1/2, p-ERK1/2 and the expression levels of apoptosis-related proteins Bcl-2, Bax, Cyt c and Casepase 3 were detected by Wes Simple Western system. Results: (1)The content of &beta-hCG and VEGF in serum: The missed abortion group of serum &beta-hCG, VEGF levels were lower than normal pregnancy group (P <0.05). (2)The pathological changes of villi tissues in the two groups: the trophoblast of the villi tissues in the missed abortion was significantly thinner, the number of cells was reduced, the cell structure was disordered, and part of the trophoblast cells were absent. (3)The angiogenesis of villi: In missed abortion group, the blood vessels in the villi interstitium were small, the vessel lumen was narrow, and the number of vessels was decreased obviously. The microvascular count results showed that the microvascular density in the villi of the missed abortion group was lower than the normal early pregnancy group (P <0.05). (4)Expression of VEGF-MEK/ERK signaling pathway related factors in villi: The expression levels of VEGF, VEGFR2, MEK1/2 and ERK1/2 mRNA in the villi of the missed abortion group were lower than the normal early pregnancy group (P <0.05)The expression levels of VEGF, RAS, p-MEK1/2 and p-ERK1/2 proteins and the p-MEK/MEK and p-ERK/ERK ratios in the villi of the missed abortion group were lower than the normal early pregnancy group (P <0.05). (5)The expression of apoptosis-related factors in trophoblast cells in villi: the expression level of Bcl-2 mRNA in villi of the missed abortion group was lower than the normal early pregnancy group (P <0.05)The mRNA expression levels of Bax, Cyt c and Caspase 3 in the villi of the missed abortion group were higher than the normal early pregnancy group (P <0.05)The expression level of Bcl-2 protein in the villi of the missed abortion group was lower than the normal early pregnancy group (P <0.05)The expression levels of Bax/Bcl-2, Cyt c and Caspase 3 proteins in the villi of the missed abortion group were higher than the normal early pregnancy group (P <0.05). (6)The correlation analysis of VEGF and ERK1/2 protein expression level in villi tissue and villous microvessel density: In missed abortion group, the ERK1/2 protein expression level was positively correlated with villous microvessel density (r=0.909, P <0.05). Conclusion: The &beta-hCG, VEGF levels in the serum were decreased, the expression of the VEGF-MEK/ERK signaling pathway was down-regulated, villi angiogenesis were reduced and mitochondrial apoptotic pathway was activated, villi angiogenesis and trophoblast dysfunction were impaired in early pregnancy, which may be an important mechanism for missed abortion.
Pages52
URL查看原文
Language中文
Document Type学位论文
Identifierhttps://ir.lzu.edu.cn/handle/262010/463457
Collection公共卫生学院
Affiliation
公共卫生学院
First Author AffilicationSchool of Public health
Recommended Citation
GB/T 7714
井光壮. β-hCG介导的VEGF-MEK ERK信号通路在早期绒毛血管生成和滋养细胞凋亡中的作用研究[D]. 兰州. 兰州大学,2020.
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