小鼠卵巢Evi-1表达水平与卵泡发育的相关性研究

发布时间：2018-04-04 10:27

本文选题：Evi-1　切入点：转化生长因子-β超家族　出处：《山东师范大学》2011年硕士论文

【摘要】：Evi-1(the ecotropic viral integration site-1)基因作为一个逆转录病毒常见的插入位点,首先是在由逆转录病毒诱发的急性髓系白血病小鼠(AKXD)中被发现的。正常情况下,Evi-1基因在人的造血细胞上不表达,但在逆转录病毒诱导的伴染色体3q26异常的人类髓系白血病细胞上过度表达,3q26染色体也是Evi-1定位的染色体。研究发现Evi-1是转化生长因子β(TGF-β)超家族信号通路中重要的调节因子,在TGF-β信号通路中发挥抑制作用。缺失Evi-1基因的小鼠将在胚胎发育的前几周因多种生理缺陷而死亡。多项研究证实Evi-1在小鼠的胚胎组织和成体组织中均有表达。但是Evi-1在卵巢中的研究甚少。TGF-β信号通路在卵泡发育过程中的作用以及Evi-1在其通路中的抑制作用使得我们推测Evi-1在卵泡的发育过程中可能发挥某些功能。因此,本实验的目的在于定位Evi-1在卵巢中的位置并进一步研究其在卵泡发育过程中的生物学功能。本实验结果显示: 1. Evi-1蛋白定位于卵母细胞质中,在颗粒细胞,膜细胞,以及卵母细胞核中均未见其表达。 2.早期卵泡发育组,Evi-1呈现出高表达的状态。在出生后1天和4天的小鼠卵巢中,Evi-1的表达量最高。随着卵泡的发育,Evi-1在卵母细胞质中的表达量逐渐降低,在出生10天和16天的卵巢中呈现出一个中间级的表达量水平。最后在23天的小鼠卵巢中表现出一个最低量的表达。由此可见,随着卵泡的发育,Evi-1的表达表现出一个逐级下降的趋势。 3.促性腺激素处理组,Evi-1依旧表达与卵母细胞质中,并且随着卵泡的发育其表达水平继续呈现下降的趋势。在大的有腔卵泡中,Evi-1的表达已经非常低,但是在一些小卵泡中表现出高表达状态,大小卵泡的对比明显。在PMSG24小时处理组中,大的有腔卵泡中可以看到其表达,但是在HCG4小时和HCG11小时的处理组中,大的有腔卵泡中Evi-1的表达已经非常非常弱了。最后,在黄体中未见Evi-1的表达。 4.发情周期检测组,Evi-1主要表达于小卵泡的卵母细胞质中。其中,发现Evi-1在间情期和发情期中的表达量要略高于发情前期和发情后期,提示Evi-1的表达可能受到激素的调节。 5.排卵相关基因的检测,real-time PCR检测Evi-1 mRNA水平和排卵相关基因(Ptgs2, Tnfaip6, Has2, Cd44, C1qbp)mRNA的水平。在HCG处理4小时后检测发现,Evi-1 mRNA的表达水平下调而上述排卵相关基因的水平却上调。总结:Evi-1在卵泡发育的早期过程中呈现高表达状态,随着卵泡的发育表达水平逐渐降低。这种特定的表达方式表明Evi-1分子在早期的卵泡发育过程中可能发挥一定的作用。
[Abstract]:Evi-1(the ecotropic viral integration site-1 (Evi-1(the ecotropic viral integration site-1) gene, as a common insertion site of retrovirus, was first found in mouse acute myeloid leukemia induced by retrovirus (ARV).Under normal conditions, Evi-1 gene was not expressed in human hematopoietic cells, but overexpression of chromosome 3q26 in retrovirus-induced human myeloid leukemia cells with abnormal chromosome 3q26 was also a Evi-1 mapped chromosome.It is found that Evi-1 is an important regulatory factor in TGF- 尾) superfamily signaling pathway and plays an inhibitory role in TGF- 尾 signaling pathway.Mice that lack the Evi-1 gene will die of multiple physiological defects in the first few weeks of embryonic development.Several studies have confirmed the expression of Evi-1 in mouse embryonic and adult tissues.However, there are few studies on the role of TGF- 尾 signaling pathway in follicular development and the inhibition of Evi-1 on follicular development. It is suggested that Evi-1 may play some roles in follicular development.Therefore, the purpose of this study was to locate the position of Evi-1 in ovary and to further study its biological function during follicular development.The results show that:1.Evi-1 protein was localized in oocyte cytoplasm, but not expressed in granulosa cells, membranous cells and oocyte nuclei.2.The expression of Evi-1 in early follicular development group was high.The expression of Evi-1 was the highest in the ovaries of 1 and 4 days postnatal mice.With the development of follicle, the expression of Evi-1 in oocyte cytoplasm decreased gradually, and the expression level of Evi-1 in ovary at 10 and 16 days after birth showed an intermediate level.At last, a minimum expression was observed in the ovaries of 23 day old mice.It can be seen that the expression of Evi-1 in follicular development showed a decreasing trend.3.In gonadotropin treated group, Evi-1 was still expressed in oocyte cytoplasm, and the expression level of Evi-1 continued to decrease with the development of follicles.The expression of Evi-1 in large antral follicles has been very low, but in some small follicles, the expression of Evi-1 is high, and the contrast of large and small follicles is obvious.The expression of Evi-1 was observed in large antral follicles in PMSG24 hour group, but the expression of Evi-1 in large antral follicles was very, very weak in HCG4 hour and HCG11 hour groups.Finally, no expression of Evi-1 was found in corpus luteum.4.Evi-1 was mainly expressed in oocyte cytoplasm of small follicles in estrous cycle detection group.It was found that the expression of Evi-1 in interoestrus and estrus was slightly higher than that in preestrus and late estrus suggesting that the expression of Evi-1 might be regulated by hormones.5.The level of Evi-1 mRNA and the levels of Ptgs2, Tnfaip6, Has2, CD44, C1qbp)mRNA were detected by real-time PCR.After 4 hours of HCG treatment, it was found that the expression of Evi-1 mRNA was down-regulated and the level of ovulation-associated genes was up-regulated.It was concluded that the high expression of Evi-1 was observed in the early stage of follicular development, and the expression level decreased gradually with the development of follicle.This specific expression suggests that Evi-1 may play a role in early follicular development.
【学位授予单位】：山东师范大学
【学位级别】：硕士
【学位授予年份】：2011
【分类号】：R321

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