体外受精胎儿出生性别失衡机制的研究

发布时间：2018-05-27 16:30

  本文选题：体外受精 + 性别失衡　； 参考：《中国农业大学》2016年博士论文

【摘要】：体外受精(in vitro fertilization, IVF)是一种关键的辅助生殖技术,已广泛应用于人类不育治疗及家畜优良种质资源高效利用。尽管绝大部分IVF后代是健康的,但是越来越多的大样本人流行病学调查数据及小鼠、家畜等动物模型实验证据表明,IVF存在一系列潜在的短期和长期健康风险,例如：流产、早产、出生缺陷、性别比例失衡、较高的代谢疾病甚至癌症风险。目前,已有观点认为IVF引发的表观修饰异常是造成这些健康问题的主要因素。然而,这些表观修饰异常的性质以及其在IVF相关健康问题中的作用知之甚少,这使得IVF相关的问题始终不能从根本上解决。在这些健康问题中,IVF引发的性别失衡问题受到广泛的关注。自1991以年,这一现象在牛、猪和人上得到反复证实,但是具体机制一直没能得到解析。作为生殖健康的一个非常重要的指标,性别比例出现失衡会对社会稳定和群体行为造成长期的严重影响。IVF引发的性别失衡意味着IVF胚胎发育存在性别倾向性的发育缺陷。考虑到IVF引发的表观修饰错误可能是造成这些健康问题的主要原因,我们推测IVF可能扰乱了性别特异性的表观修饰事件,进而引起性别倾向性的发育缺陷及出生性别比例失衡。在此,利用小鼠、兔子、牛为研究模型,通过足够的样本数量(每组n500)首次证实小鼠IVF胎儿存在出生性别比例失衡问题(57.17%vs.50.89%),这是由于围植入期雌性胚胎倾向性的发育异常所致；此外,发现雌性IVF附植前胚胎存在Xist表达抑制和印迹X染色体失活(imprinted X-chromosome inactivation, iXCI)不足的表观修饰错误。通过功能性补偿实验校正了IVF出生性别比例,证实IVF附植前胚胎中Rnf12/Xist基因低表达引发iXCI不足,造成围植入期雌性倾向性地发育异常,导致出生性别失衡。最终,发现在胚胎体外培养液中适时添加适量的视黄酸可以校正IVF引发的iXCI不足问题,并有效地校正出生性别比例。该研究表明IVF引起的表观错误在IVF引起的健康问题中具有重要作用,揭示这些表观错误有助于预防IVF健康问题。此外,比较E3.5、E7.5和E10.5三个重要发育阶段的体内胚胎(in vivo, IVO)和IVF胚胎的microRNA-seq数据,发现失调的microRNAs主要通过调控与癌症发生、遗传信息加工、葡萄糖代谢、细胞骨架组成及神经发生等过程的基因。进一步,发现IVF组胚胎中仅有miR-199a-5p从E3.5到E10.5持续低表达。通过功能性缺失与补偿性实验,证实IVF引发的miR-199a-5p低表达是造成IVF胚胎糖酵解率高、发育潜能降低、附植后胚胎存活率低的主要原因。综上,预防miR-199a-5p低表达可能是改善IVF围植入期胚胎发育的有效策略。本研究表明IVF操作及环境引发的表观修饰错误是导致IVF胚胎及出生后代健康问题的主要原因,发现并校正这些表观错误可有效预防这些健康问题。这一思想对于该领域的研究具有指引作用。将来针对其他的IVF健康问题,仍需要大量研究探索是由哪些具体的表观修饰错误引发的,进而提出针对性的解决方案。最终,实现IVF更加安全地应用。
[Abstract]:In vitro fertilization (IVF) is a key assisted reproductive technology. It has been widely used in the treatment of human infertility and the efficient utilization of fine germplasm resources of domestic animals. Although the vast majority of the offspring of IVF are healthy, more and more large samples of epidemiological survey data and experimental evidence of animal models, such as mice and livestock, are used. IVF has a series of potential short-term and long-term health risks, such as abortion, preterm birth, birth defects, gender imbalance, higher metabolic disease and even cancer risk. At present, there is a view that apparent modification abnormality caused by IVF is the main factor causing these health problems. However, the nature of these apparent abnormalities and the nature of these apparent modifications, There is little knowledge about the role of IVF related health problems, which makes IVF related problems never fundamentally solved. In these health problems, the problem of gender imbalance caused by IVF is widely concerned. Since 1991, this phenomenon has been repeatedly proved in cattle, pigs and people, but the specific mechanism has not been resolved. As a very important indicator of reproductive health, the gender imbalance in the long-term serious impact of.IVF on social stability and group behavior is a gender imbalance in the development of IVF embryos. Considering the apparent mistaken errors caused by IVF may be the main cause of these health problems. For reasons, we speculate that IVF may disrupt sexual specific apparent modification events, resulting in sexual developmental defects and the imbalance of birth sex ratio. Here, mice, rabbits and cattle are used to study models with sufficient samples (each group of N500) for the existence of sex ratio imbalance in the IVF fetus (57.17% Vs.50.89%), this is due to the tendency of abnormal development of female embryos in the peri implantation period. In addition, the apparent modification errors of Xist expression inhibition and imprinting of X chromosome inactivation (imprinted X-chromosome inactivation, iXCI) in female preimplantation embryos were found. The sex ratio of IVF birth was corrected by functional compensation experiments. The low expression of Rnf12/Xist gene in IVF preimplantation embryos leads to the deficiency of iXCI, which causes the female tendency to develop abnormality in the peri implantation period, leading to the birth sex imbalance. Finally, it is found that proper addition of retinoic acid in the embryo culture medium can correct the problem of iXCI deficiency caused by IVF and effectively correct the sex ratio of birth. The apparent error caused by the apparent IVF plays an important role in the health problems caused by IVF. It is revealed that these apparent errors contribute to the prevention of IVF health problems. In addition, the microRNA-seq data of the three important developmental stages of the body embryos (in vivo, IVO) and IVF embryos of E3.5, E7.5 and E10.5 are compared. Gene processing, genetic information processing, glucose metabolism, cytoskeleton composition and neurogenesis. Further, it was found that only miR-199a-5p from E3.5 to E10.5 remained low in IVF embryos. Through functional deletion and compensatory experiments, the low expression of miR-199a-5p caused by IVF was the high glycolysis rate and developmental potential of IVF embryos. The main reason for lower survival rate of implanted embryos is that the prevention of low expression of miR-199a-5p may be an effective strategy to improve the development of IVF peri implantation embryos. This study shows that the apparent modification errors caused by IVF operation and environment are the main causes of the health problems of IVF embryos and offspring, and the apparent errors can be found and corrected. Effective prevention of these health problems. This idea has a guiding role in the field of research. In the future, there is still a lot of research and Exploration on other IVF health problems, which are caused by specific apparent modification errors, and then put forward targeted solutions. Finally, the implementation of IVF is more secure.
【学位授予单位】：中国农业大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：R714.8;S814.8
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本文编号：1942947