本文選題：單一畸形精子癥 + 流產(chǎn)率　； 參考：《第四軍醫(yī)大學(xué)》2014年碩士論文

【摘要】：不育癥，是人類現(xiàn)今面臨的重大健康問題之一，影響了10-15%的育齡夫婦，其中男性因素占了大約50%。在臨床上，男性因素不育通常是根據(jù)異常的精子參數(shù)（如少精、弱精、畸形精子等）來診斷的。精子作為配子之一，關(guān)乎著人類的繁衍生息。據(jù)報(bào)道，70%的男性不育癥患者由精液或者精子異常導(dǎo)致。諸多研究顯示，與精液的其他參數(shù)（濃度、活力、存活率等）相比，精子形態(tài)對(duì)胚胎受精潛能和臨床妊娠結(jié)局具有更好的預(yù)測(cè)價(jià)值，這一結(jié)論在體內(nèi)和體外試驗(yàn)中均得到了驗(yàn)證。正是由于精子形態(tài)的重要預(yù)測(cè)價(jià)值和目前男性精液質(zhì)量日益下降的現(xiàn)狀，近年來，WHO關(guān)于畸形精子癥患者的鑒定閾值一直在更新，正常精子形態(tài)百分比由1978年的50%，到1992年的30%，到1999年的15%，尤其是2010年頒布的第五版精液分析標(biāo)準(zhǔn)，將這一閾值定為4%，從而引發(fā)了國內(nèi)外生殖學(xué)家的廣泛關(guān)注和爭議。但目前的研究，絕大部分是關(guān)于精子形態(tài)與輔助生殖技術(shù)后的周期結(jié)局的相關(guān)性分析，而鮮有進(jìn)一步的機(jī)制分析。印記基因是指親本僅一方來源的同源基因表達(dá)，而另一方來源的不表達(dá)。這種差異表達(dá)由DNA序列上印記基因控制區(qū)域的共價(jià)修飾來調(diào)控，主要參與了胚胎的正常生長、發(fā)育及小兒的行為發(fā)展。而精子DNA損傷情況與胚胎發(fā)育、臨床妊娠和流產(chǎn)等均密切相關(guān)。因此，本研究以單一畸形精子癥為研究對(duì)象，從與臨床妊娠結(jié)局相關(guān)性、DNA甲基化修飾和DNA完整性三個(gè)方面進(jìn)行研究。 目的： 回顧性分析單一畸形精子癥與臨床妊娠結(jié)局是否存在相關(guān)性；檢測(cè)精子印記基因（父源H19、母源SNRPN以及Line-1）DNA甲基化改變和精子細(xì)胞凋亡。 方法： 本研究以唐都醫(yī)院生殖醫(yī)學(xué)中心2012年4月-2013年5月期間，因單一畸精癥原因接受輔助受孕治療的患者夫婦為研究對(duì)象，首先回顧性分析了精子形態(tài)與臨床妊娠結(jié)局的相關(guān)性；進(jìn)一步挑選了40例單一畸形精子癥患者精液樣本，通過Isolate法離心洗滌純化精子后提取DNA，采取亞硫酸氫鹽測(cè)序PCR的方法分析父源印記基因H19、母源印記基因SNRPN以及Line-1的DNA甲基化狀態(tài)，并且通過流式細(xì)胞術(shù)檢測(cè)了其精子凋亡（DNA損傷）情況。 結(jié)果： 1、行IVF助孕后，對(duì)照組的胚胎正常受精率、卵裂率、優(yōu)胚率、臨床妊娠率、流產(chǎn)率分別為83.09%、94.52%、49.58%、62%、5.14%，單一畸形精子癥患者的胚胎正常受精率、卵裂率、優(yōu)胚率、臨床妊娠率、流產(chǎn)率分別為83.64%、95.26%、41.88%、64.41%、6.95%，各項(xiàng)指標(biāo)均無顯著差異（P＞0.05）；行ICSI助孕后，對(duì)照組胚胎正常受精率、卵裂率、優(yōu)胚率、臨床妊娠率分別為86.54%、96.80%、43.08%、56%，單一畸形精子癥患者的對(duì)應(yīng)指標(biāo)分別為87.78%、98.13%、41.67%、56.25%，均無統(tǒng)計(jì)學(xué)差異（P＞0.05），但單一畸形精子癥患者的流產(chǎn)率卻顯著升高，差異具有統(tǒng)計(jì)學(xué)意義（6.19%Vs19.5%，P＜0.05）。 2、DNA甲基化的結(jié)果：①父源印記基因H19：畸形精子組的克隆丟失率為36.05±2.86%，CpG島丟失率為3.85±0.41%，CTCF6結(jié)合位點(diǎn)區(qū)域CpG島丟失率為2.43±0.19%，而對(duì)照組則分別為13.41±3.21%，，0.75±0.17%，0.25±0.06%，差異均具有統(tǒng)計(jì)學(xué)意義（P＜0.01）；②母源印記基因SNRPN：DNA甲基化改變?cè)诨尉咏M與對(duì)照組相比，無統(tǒng)計(jì)學(xué)差異（P＞0.05）；③基因Line-1：在畸形精子組和對(duì)照組均呈現(xiàn)高度甲基化的狀態(tài)（對(duì)照組80.54±2.9%Vs畸形精子組79.67±3.2%），且兩組之間甲基化程度無統(tǒng)計(jì)學(xué)差異（P＞0.05）。 3、畸形精子組的細(xì)胞凋亡程度高于對(duì)照組（3.01±0.38Vs0.65±0.19，P＞0.05），且隨著畸形率的升高，細(xì)胞凋亡有增高的趨勢(shì)。 結(jié)論： 1、精子畸形率與臨床輔助助孕方式選擇及妊娠結(jié)局密切相關(guān)。 2、精子畸形率與H19印記控制區(qū)域的低甲基化程度呈顯著正相關(guān)。 3、精子畸形率越高，精子DNA凋亡程度越高，但需大樣本進(jìn)一步確定。
[Abstract]:Infertility is one of the major health problems facing mankind today, affecting the 10-15% couples of childbearing age, in which male factors account for about 50%. in the clinic. Male infertility is usually diagnosed according to abnormal sperm parameters such as oligospermia, weak sperm, abnormal sperm and so on. Sperm is one of the gametes and is related to human reproduction. It is reported that 70% of male infertility are caused by semen or sperm abnormalities. Many studies have shown that sperm morphology has a better predictive value for embryo fertilization potential and clinical pregnancy outcomes than other seminal parameters (concentration, vitality, survival, etc.). This conclusion has been verified in both in vivo and in vitro tests. The important predictive value of sperm morphology and the current decline in the quality of male semen have been updated in WHO in recent years. The percentage of normal sperm morphology has been updated from 50% in 1978 to 30% in 1992, to 15% in 1999, especially in 2010, the fifth version of semen analysis, the threshold of this threshold. It is defined as 4%, which leads to widespread concern and controversy about reproductive scientists at home and abroad. But most of the current studies are about the correlation analysis of the periodic outcome of sperm morphology and assisted reproductive technology, but few mechanism analyses. This differential expression is regulated by covalent modification of the imprinted gene control region on the DNA sequence. It is mainly involved in the normal growth of the embryo and the development of the child's behavior. The DNA damage of the sperm is closely related to the development of the embryo and the clinical pregnancy and abortion. Therefore, this study is based on the single malformed spermatozoa. The correlation between clinical pregnancy outcome, DNA methylation and DNA integrity were studied in three aspects.
Objective:
A retrospective analysis of the correlation between single malformed spermatozoa and clinical pregnancy outcome, and detection of DNA methylation changes and sperm cell apoptosis in sperm imprinting genes (parent source H19, parent SNRPN and Line-1).
Method錛

本文編號(hào)：1846297