胚胎延后植入对小鼠妊娠结局的影响

发布时间：2018-07-03 04:33

本文选题：延后植入 + 妊娠结局　；参考：《黑龙江八一农垦大学》2017年硕士论文

【摘要】：根据世界卫生组织的报告,不孕不育将发展成为续肿瘤和心脑血管疾病之后的第三大疾病。目前,尽管辅助生殖技术已经成为治疗不孕不育的重要手段,但它的成功率仍然仅维持在40%~50%左右的水平上,在动物中的体细胞核移植技术方面也是同样的,该技术的最终产仔率仅仅只有1%~5%。限制这些技术应用的关键环节是胚泡植入失败(Implantation Failure)。胚泡植入既需要胚泡由休眠状态转变为激活状态,也需要子宫从非接受态变为受容状态,且二者必须同步进行方能完成胚泡在子宫内膜的定位、粘附、侵入、发育及分化的生理过程即成功植入。成功植入是决定胚胎继续发育和妊娠结局的关键步骤。在哺乳动物中也包括人类,植入失败是造成妊娠率低的一个主要因素。医学临床研究表明正常育龄女性在一个月经周期内平均妊娠成功率大约为30%,在成功排卵受孕的妇女中仅有50%~60%的早期妊娠胚胎能够发育到20周以后,而其中妊娠失败的70%左右是由于植入失败所导致的。此外,植入失败也是造成动物屡配不孕的主要原因之一。因此,本试验聚焦于胚胎延后植入对妊娠结局的影响,探寻决定胚胎植入时间分子基础和妊娠维持的生理调控机制,为解决胚胎移植后高流产率、低产仔率提供理论基础,为提高动物的繁殖效率、增加经济效益以及濒危动物的保护提供技术支撑。本研究利用胚胎移植技术将妊娠第4天(d 4)的胚泡分别移植入受体鼠d 4、d 5、d 6的子宫内,观察其妊娠状况并统计其妊娠期长、窝产仔数。在移植后的24 h、48 h、72 h和8 d分别检查其植入位点数、胚胎吸收数以及其胎盘发育情况。此外,用人工诱导蜕膜化检查其蜕膜状况。采用Microarray和real-time RT-PCR方法筛选并验证d 4和d 5子宫基因。结果显示,小鼠的胚胎在延后1天(即d 5)也能着床并能诱导蜕膜化的发生,而延后2天则不能发生植入。与正常组(d 4移植)相比,延后着床组的胚胎吸收率明显增加、窝产仔数明显减少且妊娠期延长。Microarray筛选出了284个差异表达的子宫基因。研究结果表明了小鼠胚胎在超出传统认为的子宫植入窗口期亦可着床,但会导致植入位点间隙不均和胎盘发育异常进而产生妊娠不良的结局。子宫方面的调控可能是造成间隙不均和胎盘异常的主要因素。此外,本研究成功地创建了一个可靠的接受期与非接受期的子宫基因数据库。
[Abstract]:Infertility will develop into the third largest disease after cancer and cardiovascular and cerebrovascular diseases, according to the World Health Organization. At present, although assisted reproductive technology has become an important method for the treatment of infertility, its success rate is still only about 40%, and the same is true of somatic cell nuclear transfer in animals. The final birth rate of this technique is only 1 / 5. The key to limiting the application of these technologies is implantation failure. The implantation of blastocyst requires that the blastocyst change from dormancy state to active state, and that the uterus changes from non-accepted state to capacitive state. Both of them must be simultaneously carried out to complete the location, adhesion and invasion of the blastocyst in the endometrium. The physiological process of development and differentiation was successfully implanted. Successful implantation is a crucial step in determining the continued development of the embryo and the outcome of the pregnancy. Failure to implant is a major factor in low pregnancy rates in mammals, as well as in humans. Medical clinical studies have shown that the average pregnancy success rate for normal women of childbearing age is about 30 percent in one menstrual cycle, and that only 50 percent of the women who successfully ovulate and conceive have the ability to develop their early pregnancy embryos after 20 weeks. About 70% of pregnancy failure is due to implantation failure. In addition, the failure of implantation is also one of the main reasons for repeated infertility. Therefore, this study focused on the effect of delayed implantation on the outcome of pregnancy, explored the molecular basis of embryo implantation time and physiological regulation mechanism of pregnancy maintenance, and provided the theoretical basis for solving the problem of high abortion rate and low birth rate after embryo transfer. It provides technical support for improving animal reproduction efficiency, increasing economic benefit and protecting endangered animals. In this study, the blastocysts of the 4th day of pregnancy (d 4) were transplanted into the uterus of the recipient mouse d 4 d 5 d 6 by embryo transfer technique. The pregnancy status was observed and the gestational period was long and litter size was counted. The implantation sites, embryo resorption and placenta development were examined at 24 h, 48 h, 72 h and 8 d after transplantation, respectively. In addition, the decidua status was examined by artificial induced decidualization. D 4 and d 5 genes were screened and verified by microarray and real-time RT-PCR. The results showed that mouse embryos could also implant and induce decidualization 1 day later (i.e. d 5), but delayed 2 days could not induce implantation. Compared with the normal group (d 4 transplantation), the embryo absorption rate of the delayed implantation group was significantly increased, the litter size was significantly decreased and the gestational period prolonged. Microarray screened 284 differentially expressed uterine genes. The results showed that mouse embryos could also be implanted during the implantation window beyond the conventional belief, but it would lead to uneven implantation sites and abnormal placental development, which would lead to the adverse outcome of pregnancy. Uterine regulation may be a major cause of uneven gaps and placental abnormalities. In addition, we successfully created a reliable uterine gene database for both receptive and non-receptive periods.
【学位授予单位】：黑龙江八一农垦大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R714.8

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