山羊围着床期子宫中ITGB1、ITGAV、ITGB3和ITGA5的表达研究

发布时间：2018-08-04 19:23

【摘要】：山羊属于反刍动物,着床方式为表面着床,胚胎与子宫肉阜上皮细胞相融合、增生,最后形成的胎盘为子叶状胎盘。胚胎着床是成功建立妊娠的关键环节,涉及到胚胎与母体子宫之间复杂的相互作用。整合素作为细胞黏附分子家族成员,是一种细胞表面结合蛋白,主要介导粘附和行使信号传递功能,在胚胎着床过程发挥重要作用。我们通过转录组测序发现:整合素ITGAV(integrinαv,ITGAV)、ITGB3(integrinβ3,ITGB3)和ITGA5(integrinα5,ITGA5)mRNA在山羊围着床期(D6、D16和D19)子宫中表达变化特异。因此本文详细研究整合素亚基ITGB1(integrinβ1,ITGB1)、ITGAV、ITGB3、ITGA5蛋白在山羊围着床子宫中的表达和激素调节,为深入研究整合素在反刍动物胚胎着床中的作用提供实验依据。本实验的目的是检测整合素亚基ITGB1、ITGAV、ITGB3和ITGA5在山羊发情周期、早期妊娠和激素处理子宫中的表达,来确定这些整合素亚基是否在山羊胚胎着床中起作用,是否受类固醇激素调控。本实验材料为黑龙江本地山羊,采用Western blot方法检测ITGB1、ITGAV、ITGB3和ITGA5在发情周期、早期妊娠和激素处理山羊子宫中的表达和类固醇激素调控;通过免疫组织化学技术研究ITGB3和ITGA5蛋白在山羊着床阶段子宫中的定位。本实验所得结果如下:(1)在发情周期中,Western blot检测到ITGB1、ITGAV、ITGB3和ITGA5在山羊整个发情周期的子宫中均表达,ED0表达较高,排卵后(ED6、ED12)表达降低,ED16表达升高。(2)在山羊早期妊娠过程中,Western blot结果显示ITGB1在山羊胚胎粘附和胎盘形成早期(D16-D30)高表达,ITGAV、ITGB3和ITGA5在妊娠期表达基本一致,均在山羊胚胎粘附期(D16)高表达;免疫组织化学结果显示ITGB3在胚胎粘附期(D16、D19)子宫腔上皮和基质中有表达,ITGA5在子宫腔上皮、腺上皮和基质中有表达。(3)在类固醇激素处理模型中,Western blot显示孕酮上调ITGB1的表达,雌激素无明显影响;雌激素上调ITGAV、ITGB3和ITGA5的表达,孕酮无明显影响。本实验结果表明ITGB1的表达在山羊子宫中呈现胚胎着床特异性,在胚胎着床和胎盘形成早期,ITGB1蛋白一直处于高水平。推测ITGB1参与山羊胚胎着床和胎盘形成的过程。孕酮上调ITGB1的表达,雌激素无明显影响。ITGAV、ITGB3和ITGA5在山羊胚胎粘附期(D16)处于高水平,并且ITGB3表达于着床期子宫腔上皮和基质中,ITGA5表达于着床期子宫腔上皮、腺上皮和基质中,推测ITGAV、ITGB3和ITGA5在山羊胚胎粘附过程发挥着重要作用。雌激素上调ITGAV、ITGB3和ITGA5的表达,孕酮无明显影响。山羊胚胎着床过程中,不同的整合素表达方式不同,类固醇调控机制不同,可能发挥不同作用。本实验为研究整合素在反刍动物胚胎着床和胎盘形成中作用机制提供了依据。
[Abstract]:Goat belongs to ruminant, the implantation mode is surface implantation, embryo and uterine caruncle epithelial cells merge and proliferate, the last placenta is cotyledon placenta. Embryo implantation is the key to the successful establishment of pregnancy, involving the complex interaction between embryo and womb. Integrin, as a member of cell adhesion molecule family, is a cell surface binding protein, which mediates adhesion and signal transduction, and plays an important role in embryo implantation. By transcriptome sequencing, we found that the expression of ITGB3 (integrin 尾 3ITGB3) and ITGA5 (integrin 伪 5ITGA5) mRNA were specific in the uterus of goat (D6D16 and D19). Therefore, the expression and hormone regulation of integrin subunit ITGB1 (integrin 尾 1 ITGB1) ITGAVG3ITGA5 protein in goat surrounding bed uterus were studied in detail, which provided experimental basis for further study on the role of integrin in implantation of ruminant embryos. The purpose of this study was to detect the expression of ITGB3 and ITGA5 in goat estrous cycle, early pregnancy and hormone treated uterus to determine whether these integrin subunits play a role in goat embryo implantation and whether they are regulated by steroid hormones. In this study, the expression of ITGB3 and ITGA5 in estrous cycle, early pregnancy and hormone treated goat uterus were detected by Western blot method. The localization of ITGB3 and ITGA5 proteins in goat uterus during implantation was studied by immunohistochemical technique. The results were as follows: (1) in estrous cycle, ITGB1, ITGB1, ITGB3 and ITGA5 were detected to be highly expressed in the uterus of goat estrous cycle. After ovulation (ED6 + ED12), the expression of ED16 was decreased. (2) during early pregnancy, the expression of ITGB1 in goat embryo adhesion and placental formation (D16-D30) was highly expressed. The expression of ITGAV TGB3 and ITGA5 was basically the same during pregnancy, and both expressed in goat embryo adhesion phase (D16). Immunohistochemical results showed that ITGB3 was expressed in uterine cavity epithelium and matrix during embryonic adhesion phase (D16 / D19). (3) in steroid treated model, the expression of ITGB1 was up-regulated by progesterone by Western blot. Estrogen did not affect the expression of ITGB3 and ITGA5, but progesterone did not. The results showed that the expression of ITGB1 was specific to embryo implantation in goat uterus, and the expression of ITGB1 protein was always at a high level in the early stage of embryo implantation and placenta formation. It is speculated that ITGB1 is involved in the process of implantation and placenta formation of goat embryos. Progesterone upregulated the expression of ITGB1, estradiol had no significant effect on the expression of ITGB3 and ITGA5 in goat embryo adhesion phase (D16), and ITGB3 was expressed in uterine cavity epithelium and matrix in implantation stage, glandular epithelium and matrix. It is inferred that ITGAV ITGB3 and ITGA5 play an important role in the process of goat embryo adhesion. Estrogen upregulated the expression of ITGB3 and ITGA5, but progesterone had no effect. Different integrin expression patterns and steroid regulation mechanisms may play different roles in goat embryo implantation. This study provides a basis for the study of the role of integrin in embryo implantation and placental formation in ruminants.
【学位授予单位】：东北农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S827

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