BCAS2在小鼠颗粒细胞和卵母细胞中的定位表达及其对颗粒细胞增殖凋亡的影响

发布时间：2018-04-08 14:04

本文选题：BCAS2　切入点：颗粒细胞　出处：《华中农业大学》2015年硕士论文

【摘要】：BCAS2是雌激素受体的一种辅助转录激活因子,其表达量的变化会影响雌激素受体相关基因的转录活性。雌激素在哺乳动物卵泡发育中有重要的调控作用。目前关于BCAS2的研究主要集中在癌细胞系中,如乳腺癌细胞、Hela细胞等,而关于其在正常体细胞和生殖细胞内的功能研究报道还较少。因此,本研究以小鼠为实验动物模型,利用细胞免疫荧光、Western blot、Real-time PCR等实验技术,检测了BCAS2在小鼠卵巢组织中的定位和表达;利用RNA干扰技术对BCAS2在小鼠卵巢颗粒细胞内的功能做了初步的探索,结果如下:(1)BCAS2在小鼠颗粒细胞和卵母细胞中的表达和定位。细胞免疫荧光结果显示,BCAS2主要定位在颗粒细胞和卵母细胞的细胞核中。Western blot结果显示,BCAS2在小鼠GV-MII期卵母细胞中的表达量逐步升高。在超数排卵处理后,PMSG注射后6 h和12 h卵巢中,BCAS2的表达水平显著高于其它时期(P0.05)。Real-time PCR结果显示,BCAS2 m RNA在腔前卵泡中的表达量显著高于有腔卵泡(P0.05)。FSH可以促进体外培养颗粒细胞中BCAS2 m RNA的表达。(2)RNAi干扰敲低BCAS2的表达对颗粒细胞增殖和凋亡的影响。在颗粒细胞中干扰BCAS2后,能显著性地降低了颗粒细胞的增殖水平(P0.05),同时显著地下调增殖相关基因PCNA的表达(P0.05),干扰BCAS2之后,促进颗粒细胞凋亡,并且抗凋亡基因BCL-2表达量显著下调,促凋亡基因BAX的表达量显著上调(P0.05)。综上,本研究发现BCAS2定位于小鼠颗粒细胞和卵母细胞的细胞核内,同时,BCAS2在不同发育时期的卵泡颗粒细胞中具有特殊的表达规律,颗粒细胞中干扰BCAS2之后,会显著降低细胞的增殖水平,并显著促进细胞凋亡,同时,显著性影响增殖和凋亡相关基因的表达,该结果说明BCAS2可能参与卵泡颗粒细胞增殖和凋亡的生理过程。
[Abstract]:BCAS2 is an assistant transcriptional activator of estrogen receptor, and its expression will affect the transcriptional activity of estrogen receptor related genes.Estrogen plays an important role in the regulation of follicle development in mammals.At present, the study of BCAS2 is mainly focused on cancer cell lines, such as breast cancer cells, such as Hela cells, but its function in normal somatic cells and germ cells is less reported.Therefore, in this study, the localization and expression of BCAS2 in mouse ovarian tissues were detected by using CIFA Western blotreal-time PCR and other experimental techniques.The function of BCAS2 in mouse ovarian granulosa cells was studied by RNA interference technique. The results were as follows: the expression and localization of BCAS2 in mouse granulosa cells and oocytes.The results of cellular immunofluorescence showed that BCAS2 was mainly located in the nucleus of granulosa cells and oocytes. Western blot showed that the expression of BCAS2 in mouse GV-MII oocytes increased gradually.Expression of BCAS2 m RNA in granulosa cells. Effect of RNAi interference knockout on proliferation and apoptosis of granulosa cells.After interfering with BCAS2 in granulosa cells, the proliferation level of granulosa cells was significantly decreased, and the expression of proliferation-related gene PCNA was significantly down-regulated. After interfering with BCAS2, granulosa cells apoptosis was promoted.The expression of anti-apoptotic gene BCL-2 was down-regulated, and the expression of pro-apoptotic gene BAX was significantly up-regulated (P 0.05).In conclusion, we found that BCAS2 was located in the nucleus of mouse granulosa cells and oocytes, and that BCAS2 had a special expression pattern in follicular granulosa cells at different developmental stages, after interfering with BCAS2 in granulosa cells.The results indicated that BCAS2 may be involved in the physiological process of proliferation and apoptosis of follicular granulosa cells.
【学位授予单位】：华中农业大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S814

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