HtrA 4和GCM1在子痫前期患者胎盘中的表达和意义

发布时间：2018-07-10 11:53

  本文选题：HtrA4 + GCM1　； 参考：《山东大学》2014年硕士论文

【摘要】：研究目的 HtrA4(high temperature reguirement A4, HtrA4)是HtrA家族蛋白中不典型一员,HtrA家族蛋白是丝氨酸蛋白酶,对折叠错误和位置错误的蛋白起质控作用。最近几年,对HtrA1等已经有所报道,主要在调控滋养细胞分化及侵袭中起一定作用,但对HtrA4研究不多。研究表明子痫前期病人的HtrA4在胎盘及血清中表达水平均较正常血压的孕妇水平要低；相较正常绒毛,子痫前期的病人的HtrA4在合体滋养细胞层表达显著增多。已知GCM1(Glial cells missing1)是胎盘形成过程中的一个重要转化因子,调节作用于合体滋养细胞层分化的syncytin融合蛋白的表达,也调节促进胎盘血管生成的胎盘生长因子(PGF)的表达。Liang-Jie Wang[3]等通过染色质免疫沉淀芯片分析,确认HtrA4基因是GCM1的靶基因。本研究就HtrA4和GCM1在妊娠期高血压患者胎盘组织中的表达,探讨两者参与调节滋养细胞侵袭机制的关系,为子痫前期等病理妊娠的发病机制提供理论基础。 实验方法 1.研究对象随机收集山东大学齐鲁医院2013年4月～2013年7月产科住院的产妇胎盘60例,其中妊高症患者40例作为实验组(早发型子痫前期23例,晚发型子痫子痫前期17例),另选同期在本院正常晚期妊娠孕妇20例为对照组。子痫前期的诊断标准参照《妇产科学》(第8版)。两组的孕周、年龄均无统计学差异。所选对象无慢性高血压、糖尿病、心脏病、肾脏病等病史。 2. RT-PCR提取总RNA后,选取合格的mRNA作为模板进行反转录反应,反转录条件严格按照说明书进行操作。PCR引物由北京博爱森生物技术公司合成,GCMl上游：5'-GTTAGAAGCTGAGGCAAGAA-3',下游：5'-TAAAGATTACCCGCGGAT-3'; HtrA4上游：5'-GTCAGCACCAAACAGCG-3',下游：5'-GGAGATTCCATCAGTCACCC-3'; β-actin上游：5'-AACTCCATCATGAAGTGAC-3'下游：5’-GATCCACATCTGCTGGAAGG-3',均按说明操作进行PCR,扩增产物经凝胶电泳后,在D-140图像记录分析系统上进行分析。HtrA4和GCM1的mRNA表达量分别以HtrA4或GCM1扩增DNA条带的灰度值计算。 Western blotting:取100mg组织剪碎进行蛋白提取,凝胶电泳,转膜,杂交,显色,成像仪成像,分析结果 3.统计学方法：应用SPSS17.0软件包进行统计分析,以均数±标准差表示,组间比较采用单因素方差分析,采用t检验,组间相关性分析用spearman检验。 结果 在mRNA水平,HtrA4与GCM1在组中均有表达,HtrA4与GCM1mRNA表达量在研究组中低于健康对照组,差异具有统计学意义(P0.01)；子痫前期组胎盘组织中HtrA4与GCM1的表达均呈正相关(mRNA:r-0.426,蛋白：r=0.686、P0.01)。 讨论 GCM1与子痫前期发病的关系GCM1即GCMa, GCM蛋白是一种具有保守DNA结合区域的转录因子。人类的GCM1主要在胎盘组织,是胎盘发育过程中一个必要的转录因子。在低氧情况下GCM1泛素化降解而活性被抑制,GCM1主要通过促进金合胞素和PGF进而促进滋养细胞融合分化,而HtrA4可以通过PDZ结构域与合胞素的表面亚结构结合,已有报道HtrA4和GCM1交叉表达于母胎界面的间质EVTs中。我们实验证明了在子痫前期的胎盘中GCM1和HtrA4在基因和蛋白水平均降低,且具有相关性,因此,GCM1有可能通过激活HtrA4的表达来调节EVTs的分化,进而促进EVTs侵袭,并同时抵消合胞素的融合活性,但HtrA家族其他蛋白主要通过PDZ结构进行活化作用,所以,HtrA4也有可能促进合胞素表达,还需要进一步实验研究。 结论 HtrA4与GCM1在子痫前期胎盘中均表达异常,而且具有相关性,两者在子痫前期发展过程中可能相互作用,有助于进一步了解子痫前期发病机制。
[Abstract]:research objective
HtrA4 (high temperature reguirement A4, HtrA4) is an untypical member of the HtrA family protein. The HtrA family protein is serine protease, which plays a quality control role in the error of folding and position error. In recent years, some reports have been made on HtrA1 and so on, which mainly play a role in regulating the differentiation and invasion of trophoblastic cells, but for HtrA4 studies. Not much. The expression level of HtrA4 in preeclampsia patients in preeclampsia was lower than that of normal blood pressure pregnant women; compared with normal villi, the expression of HtrA4 in the syncytio cell layer was significantly increased in the patients with preeclampsia. The known GCM1 (Glial cells missing1) is an important transformation factor in the process of fetal disc formation. The expression of syncytin fusion protein and the expression of placental growth factor (PGF), the expression of placental growth factor (PGF), which promote the formation of placenta, and the expression of.Liang-Jie Wang[3] by chromatin immunoprecipitation chip, confirm that the HtrA4 gene is the target gene of GCM1. This study was on HtrA4 and GCM1 in placental group of hypertensive patients with pregnancy. The relationship between the two involved in regulating the invasion mechanism of trophoblastic cells was explored, providing a theoretical basis for the pathogenesis of preeclampsia and other pathological pregnancies.
Experimental method
1. the subjects randomly collected 60 cases of maternal placenta in the Qilu Hospital of Shandong University from April 2013 to July 2013, of which 40 cases of pregnancy induced hypertension were used as the experimental group (23 cases of early onset preeclampsia and 17 cases of late eclampsia), and 20 cases of pregnant women in the same period were selected as the control group. There was no statistical difference in gestational age between the two groups (Eighth Edition). There was no significant difference in age, gestational age, diabetes, heart disease and kidney disease.
2. RT-PCR extract total RNA, select the qualified mRNA as a template for reverse transcription reaction, reverse transcription conditions strictly according to the instructions to operate.PCR primers by Beijing Bo arson Biotech Corp, GCMl upstream: 5'-GTTAGAAGCTGAGGCAAGAA-3', downstream: 5'-TAAAGATTACCCGCGGAT-3'; HtrA4 upstream: 5'-GTCAGCACCAAACAGCG-3', downstream. : 5'-GGAGATTCCATCAGTCACCC-3'; upstream of beta -actin: downstream of 5'-AACTCCATCATGAAGTGAC-3': 5 '-GATCCACATCTGCTGGAAGG-3', all PCR is carried out according to instructions. After gel electrophoresis, the amplified products are analyzed on the D-140 image recording analysis system for the mRNA expression of.HtrA4 and GCM1 in HtrA4 or GCM1 amplification of the DNA strip, respectively. Count.
Western blotting: was taken from 100mg tissue to cut and break for protein extraction, gel electrophoresis, transmembrane, hybridization, color rendering, imaging instrument analysis, and results.
3. statistical method: the statistical analysis was carried out by SPSS17.0 software package, the average number of standard deviation was expressed. The single factor variance analysis was used in the group. The t test was used, and the correlation analysis between groups was tested by Spearman.
Result
At mRNA level, both HtrA4 and GCM1 were expressed in the group, and the expression of HtrA4 and GCM1mRNA in the study group was lower than that in the healthy control group. The difference was statistically significant (P0.01). The expression of HtrA4 and GCM1 in the placental tissue of the preeclampsia group was positively correlated (mRNA:r-0.426, protein: r= 0.686, P0.01).
discuss
The relationship between GCM1 and preeclampsia is GCM1, GCMa, and GCM protein is a transcription factor with conservative DNA binding region. Human GCM1 is mainly in placental tissue. It is a necessary transcription factor in the development of placenta. In hypoxic conditions, the activity of GCM1 is degraded and the activity of GCM1 is inhibited, and GCM1 is mainly through the promotion of syncytin and PGF. Promoting the fusion and differentiation of trophoblastic cells, while HtrA4 can bind to the surface substructure of the PDZ domain with the surface substructure of syncytin, it has been reported that HtrA4 and GCM1 are intersecting in the interstitial EVTs of the maternal fetal interface. Our experiment proved that GCM1 and HtrA4 in the placenta of preeclampsia have an average decrease in gene and protein water and are related, therefore, GCM1 has a possible effect. The expression of HtrA4 can regulate the differentiation of EVTs, and then promote the invasion of EVTs and counteract the fusion activity of the syncytin, but the other proteins of the HtrA family are activated by the PDZ structure. So, HtrA4 may also promote the expression of syncytin, and a further experimental study is needed.
conclusion
HtrA4 and GCM1 are abnormal in the placenta of preeclampsia and are related, and they may interact in the development of preeclampsia, which can help to further understand the pathogenesis of preeclampsia.
【学位授予单位】：山东大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R714.244

【共引文献】

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2 赵海君;张园;崔毓桂;;紧密连接在妊娠生理与病理生理中的作用[J];国际妇产科学杂志;2013年05期

3 陆竹梅;邓姗;邱小菊;;子痫前期患者血清hs-CRP和D-二聚体水平与不良妊娠结局的分析[J];华夏医学;2013年06期

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5 倪华;贲文锐;李文萍;李瑞婷;靳方圆;;TEAD1在小鼠围着床期子宫中的表达研究[J];东北农业大学学报;2014年06期

6 张一鸣;袁小松;蒋雅琴;蒋丽霞;;胎盘生长因子及可溶性血管内皮生长因子受体1的水平变化与子痫前期发病的关系[J];南京医科大学学报(自然科学版);2010年10期

7 余筱琳;刘小萍;胡雪飞;;子痫前期孕妇尿液VEGF、sFlt-1水平的变化及意义[J];南昌大学学报(医学版);2013年08期

8 李莉;郑丹;;Caspase-3在妊娠期高血压疾病胎盘组织中的表达及其意义[J];江西医药;2014年02期

9 刘娜;徐汉卿;崔健;茆灿泉;许雷;;拟除虫菊酯类农药降解酯酶EstA融合蛋白表达质粒的构建和诱导条件优化[J];河南农业科学;2014年02期

10 罗方媛;刘兴会;杨悦;何国琳;陈锰;;NKG2A、NKG2C受体及其配体HLA-E在子痫前期患者蜕膜中的表达[J];四川大学学报(医学版);2014年04期

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1 熊慧娟;CD4~+CD25~+调节性T细胞在妊娠及分娩发动中的作用及机制探讨[D];中南大学;2011年

2 张明;牛IFNτ基因成熟蛋白CDS区的克隆、原核表达及其在妊娠建立过程中的生物学功能研究[D];四川农业大学;2007年

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4 庄学伟;1、子痫前期胎盘syncytin-1甲基化状态的研究 2、Syncytin-1在非小细胞肺癌的初步研究[D];山东大学;2013年

5 余俊;天然化合物S-烯丙基-L-半胱氨酸治疗子痫前期的相关研究[D];华中科技大学;2013年

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7 洪俊平;血管内皮生长因子过表达对肾小管上皮间充质转化的抑制作用及机制探讨[D];北京协和医学院;2012年

8 梁海霞;Ran GTPase在嗜热四膜虫大核无丝分裂和程序性核死亡过程的功能研究[D];山西大学;2013年

9 郭玲;人异常胎盘转录组分析及MAGEL2基因在猪胚胎中的印记研究[D];华中农业大学;2013年

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2 刘德红;正常妊娠母胎界面免疫微环境的研究[D];安徽医科大学;2009年

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