TLR2和TLR4的表达与自然流产的相关性研究

发布时间：2018-09-15 20:09

【摘要】：目的:流产(Abortion)是指怀孕未满28周、胎儿体重未满1000g而使怀孕受到终止,早期流产是指发生在怀孕12周以前者,晚期流产是指发生在满12周及其之后者。自然流产是早孕期间比较多见的病症,其发病原因和机制十分繁杂,除去常见的解剖结构异常(如子宫畸形、宫颈功能不全等)、染色体异常、遗传、内分泌、理化环境及感染等因素外,近年来生殖免疫成为研究热点之一,也有相关研究表明了免疫因素与自然流产密切相关。该理论认为,从免疫移植学角度而言,之所以怀孕期间孕囊作为带有“外来”抗原的同种异体移植物可以存在于母亲体内,是因为母体胎儿面免疫系统的精细和复杂的调节。很多因素可以经过活化母亲体内Toll样受体(Toll like receptors,TLRs),改变母体胎儿面多种活性因子的分布水平,破坏了母体胎儿面的免疫均衡,使得孕囊被母亲体内的免疫反应所排斥,引发流产。TLRs是一类能辨识病原微生物的模式分子,有调控天然免疫和获得性免疫的作用,并且参与母体胎儿面的抗病原微生物的天然免疫应答,TLRs既表达在免疫活性细胞,也在其他部位表达,如绒毛滋养细胞、蜕膜组织等,在维系健康妊娠和自然流产发病中起到不可忽视的作用。目前关于与妊娠流产的相关TLRs中,对Toll样受体2和4的研究相对较多,它们是体内非常关键的自然免疫蛋白,也是TLRs家族亚类特征性的两个,在大部分细胞系的细胞膜上都有分布,骨髓中单核细胞分布最多,在中性粒细胞、巨噬细胞、树突状细胞等细胞膜上也分布较多,另外,还分布于心肌细胞、微血管内皮细胞、气道上皮细胞、脂肪细胞、消化道上皮细胞、人胚胎肾细胞、齿龈纤维母细胞、人表皮和脐静脉内皮细胞以及胎盘滋养细胞[1]。Toll样受体2辨识肽聚糖,Toll样受体4辨识脂多糖、内毒素等,经过髓样分化蛋白88(MyD88)依赖性和非依赖性反应转导路径引起母体胎儿面的免疫活性因子Th1/Th2失去平衡。Th1分泌的趋化因子产生过多,对滋养层细胞产生细胞毒性作用而形成免疫损伤,直接或间接影响孕囊的着床和成长发育,而Th2则起到了维持母体胎儿面的免疫保护作用。故而,当Th1/Th2偏向Th1时发生病理性怀孕的风险增加。如果对自然流产的患者盲目给予保胎,而不是对因治疗,往往达不到医生和患者所期待的效果。因此尽最大可能进行分析和寻找病因变的重要之至。文章就Toll样受体2和4在绒毛和外周血中的分布情况与自然流产的相关性进行探索。方法:选择2014年06月-2014年12月在解放军第105医院妇产科门诊患者(年龄35岁、孕周12周)作为本次实验的研究对象:本次怀孕结局为自然流产者、因意外怀孕自愿行人流终止妊娠的健康的早孕妇女。纳入的研究对象均由专科医师对病史进行问询,包括既往史、月经史、婚育史、家族史等,并核实填写项目是否真实、准确。体格检查包括:一般体格检查:血压、身高、体重、甲状腺、乳腺等,及常规妇科检查。所有对象均接受基础性激素检查、b超检查、夫妇染色体检查、有无急慢性传染病、妇科炎症及与怀孕相关联的免疫抗体(如抗磷脂抗体、抗β2糖蛋白抗体、抗核抗体、抗甲状腺抗体、狼疮抗凝血因子等为阳性者)等辅助检查,尽可能排除遗传、染色体、解剖等相关因素。本次怀孕结局为自然流产患者列为研究组;将曾有怀孕和生育史,本次因意外怀孕自愿行人流终止妊娠的健康孕妇列为对照组,每组各20例。采用免疫组化法检查每例患者绒毛滋养细胞中toll样受体2和4的分布水平(每个样本随机抽选一张切片,以观察绒毛滋养细胞胞浆内是否含有棕黄色颗粒为判断标准)。采用spss17.0统计软件进行分析处理。用平均值加减标准差分析。若p0.05,则有统计学差异;采用流式细胞仪检测tlr2和tlr4分别在外周血淋巴细胞群、单核细胞群和粒细胞群的分布比值进行分析。结果:1、自然流产组与妊娠对照组之间的年龄、孕龄和孕次的差异等均无统计学意义。2、tlr2和tlr4在自然流产组和妊娠对照组的绒毛滋养细胞均见分布,主要分布在合体滋养细胞的胞浆中。3、tlr2在自然流产组和妊娠对照组的绒毛、外周血中单核细胞群和粒细胞群的分布水平差异无统计学意义(p0.05),tlr2在自然流产组和妊娠对照组的外周血中淋巴细胞群的分布水平差异具有统计学意义(p0.05)。4、自然流产组和妊娠对照组相比,tlr4在两组绒毛及外周血淋巴细胞群、单核细胞群和粒细胞群的分布水平差异有统计学意义(p0.05)。结论:1.tlr2和tlr4在自然流产组和妊娠对照组绒毛滋养细胞、外周血淋巴细胞群、单核细胞群和粒细胞群中都有表达。2.tlr2在自然流产组外周血中淋巴细胞群的表达明显升高,tlr4在自然流产组绒毛滋养细胞的表达明显上调。3.tlr4在自然流产组绒毛组织、外周血中淋巴细胞群、单核细胞群和粒细胞群中的分布均升高明显。通过对早孕期妇女进行外周血的tlr2和tlr4的检测,有利于预测孕妇可能发生的自然流产的风险,以期预防或干预,降低流产的发生率。
[Abstract]:Objective: Abortion refers to the termination of pregnancy before 28 weeks of gestation and under 1000g of fetal weight. Early abortion refers to those who occur before 12 weeks of gestation. Late abortion refers to those who occur after 12 weeks of gestation. In recent years, reproductive immunity has become one of the hotspots of research, and some related studies have shown that immune factors are closely related to spontaneous abortion. During this period, gestational sac, as an allograft with "foreign" antigen, can exist in the mother because of the fine and complicated regulation of the maternal fetal immune system. TLRs are a kind of model molecule which can identify pathogenic microorganisms, regulate innate immunity and acquired immunity, and participate in the innate immune response against pathogenic microorganisms on the maternal and fetal surfaces. TLRs are expressed in immune activity. Sex cells, also expressed in other parts of the body, such as choriotrophoblasts and decidual tissue, play an important role in maintaining healthy pregnancy and spontaneous abortion. Two characteristic subclasses are distributed on the cell membranes of most cell lines. Monocytes are most distributed in bone marrow. They are also distributed on the cell membranes of neutrophils, macrophages, dendritic cells and so on. In addition, they are also distributed in cardiomyocytes, microvascular endothelial cells, airway epithelial cells, adipocytes, gastrointestinal epithelial cells, and human beings. Embryonic kidney cells, gingival fibroblasts, human epidermal and umbilical vein endothelial cells and placental trophoblasts [1]. Toll-like receptor 2 recognizing peptidoglycans, Toll-like receptor 4 recognizing lipopolysaccharides, endotoxins, etc., through myeloid differentiation protein 88 (MyD88) dependent and independent response transduction pathways cause the loss of maternal and fetal immune activity factor Th1/Th2. Th1 produces too many chemokines, which produce cytotoxic effects on trophoblast cells and cause immune damage, directly or indirectly affecting implantation and growth of gestational sacs, while Th2 protects the immune function of maintaining maternal and fetal surface. Therefore, the risk of pathological pregnancy increases when Th1/Th2 is biased towards Th1. The distribution of Toll-like receptors 2 and 4 in villi and peripheral blood and the relationship between the distribution of Toll-like receptors 2 and 4 in villi and peripheral blood and spontaneous abortion were explored. From June 14 to December 2014, patients aged 35 and 12 weeks of gestation in the outpatient department of gynecology and obstetrics of the 105th Hospital of the PLA were selected as the subjects of this study. Physical examinations include general physical examinations: blood pressure, height, weight, thyroid gland, breast gland, and routine gynecological examinations. All subjects were examined for basic sex hormones, ultrasound, couple chromosomes, acute and chronic infectious diseases, gynecological inflammation. The pregnant women with spontaneous abortion were enrolled in the study group. The pregnant women with previous pregnancy and childbearing history were enrolled in the study group. The distribution of Toll-like receptors 2 and 4 in trophoblastic cells of each patient was examined by immunohistochemistry. A section was randomly selected from each sample to observe whether there were brown granules in cytoplasm of trophoblastic cells. SPSS 17.0 statistical software was used for analysis and processing.The standard deviation analysis was used for average addition and subtraction.If p0.05, there was statistical difference.Flow cytometry was used to detect the distribution ratio of TLR2 and TLR4 in peripheral blood lymphocyte population, monocyte population and granulocyte population respectively.Results:1. Natural abortion group and pregnancy control group. TLR2 and TLR4 were distributed in the cytoplasm of syncytiotrophoblast. 3. the distribution of TLR2 in the villi of spontaneous abortion group and pregnancy control group was different. There was no significant difference between the two groups (p0.05). The distribution of TLR2 in peripheral blood lymphocyte group was significantly different between the spontaneous abortion group and the pregnancy control group (p0.05). 4. Compared with the pregnancy control group, the distribution of TLR4 in villi and peripheral blood lymphocyte group, monocyte group and granulocyte group were statistically different between the two groups. Conclusion: 1. The expression of TLR2 and TLR4 in choriotrophoblasts, peripheral blood lymphocyte, monocyte and granulocyte in spontaneous abortion group and pregnancy control group were significantly increased. 2. The expression of TLR2 in peripheral blood lymphocyte was significantly increased in spontaneous abortion group, and the expression of TLR4 in choriotrophocyte in spontaneous abortion group was significantly increased. The distribution of TLR4 in villi, peripheral blood lymphocyte, monocyte and granulocyte increased significantly in spontaneous abortion group. The detection of TLR2 and TLR4 in peripheral blood of early pregnant women is helpful to predict the risk of spontaneous abortion in order to prevent or intervene and reduce the incidence of abortion.
【学位授予单位】：蚌埠医学院
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R714.21

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