MTHFR、MTRR基因多态性与不明原因反复妊娠丢失的相关性研究

Correlation Study of MTHFR and MTRR Gene Polymorphisms and Unexplained Recurrent Pregnancy Loss

摘要: 目的：在uRPL (unexplained Recurrent pregnancy loss, uRPL)人群中对MTHFR基因C677T、A1298C位点和MTRR基因A66G位点进行检测，以明确其与uRPL发生及不同流产次数的相关性，为uRPL的病因筛查提供遗传学证据，并指导临床应用。方法：本研究收集了2012~2015年在山东大学629例育龄期夫妇的临床信息和外周血样本，其中既往与同一丈夫连续发生≥2次妊娠28周内妊娠丢失且排除明确病因的患者纳入uRPL组，既往至少有一次活产且没有流产或任何其他不良妊娠结局及并发症病史的患者纳入对照组。根据患者的年龄进行1:1个案匹配，最终有230例uRPL组的女性及230例对照组女性匹配成功。采用高分辨熔融曲线分析技术进行SNP位点基因分型，根据SNP检测结果，对uRPL组和对照组的叶酸代谢风险和叶酸代谢相关基因MTHFR C677T、MTHFR A1298C、MTRR A66G的基因型以及等位基因频率分布进行统计分析，并按照流产次数(2、3、≥4次)分层分析进一步比较。结果：叶酸代谢风险等级在uRPL人群中分布无显著差异，并与既往流产次数无关(P > 0.05)。进一步对三个多态位点的基因型分布进行统计分析，发现：1) MTHFR基因C667T位点在uRPL人群中总体CT (杂合突变)比例增加，而TT (纯合突变)比例降低，亚组分析当既往自然流产次数为3时，纯合突变比例降低，具有显著差异(P < 0.05，Bonferroni法校正后P < 0.0167)，等位基因T的频率也同样显著降低；2) A1298C多态位点在uRPL人群中分布无显著差异(P > 0.05)；3) MTRR基因A66G多态位点在uRPL人群中分布也无显著差异(P > 0.05)，但亚组分析当既往自然流产次数为3时，纯合突变比例降低具有显著差异(P < 0.05，Bonferroni法校正后P < 0.0167)，但等位基因的频率分布未见显著差异。结论：发现叶酸代谢风险等级在uRPL人群中的分布无显著差别，MTHFR基因C667T、A1298C两个多态位点和MTRR基因A66G多态位点突变频率在uRPL总体人群中未见显著增加，并与既往流产次数无显著相关性。

Abstract: Objective: Detect the C677T, A1298C loci of the MTHFR gene, and A66G loci of the MTRR gene in the unexplained recurrent pregnancy loss (uRPL) population to clarify their correlation with the occur-rence of uRPL and different miscarriages, provide genetic evidence for the etiology screening of uRPL, and guide clinical application. Methods: Clinical information and peripheral blood samples were collected from couples of reproductive age attending the Shandong University 2012 to 2015. 629 uRPL patients included those with ≥2 previous consecutive pregnancy losses within 28 weeks of gestation with the same spouse and excluded a clear etiology, while patients in the control group were those with at least one previous live birth and without a history of miscarriage or any other adverse pregnancy outcomes. Pair matching 1:1 was conducted according to female age. 230 pa-tients in the uRPL group and 230 patients in the control group were successfully matched. SNP genotyping technology was performed using high-resolution melting curve analysis. The risk of fo-late metabolism and genotypes of the genes related to folate metabolism, MTHFR C677T, MTHFR A1298C, and MTRR A66G, as well as the allele frequency distribution, were statistically analyzed in the uRPL and control groups and further compared by stratified analysis of the number of miscar-riages (2, 3, ≥4). Results: There was no significant difference in the distribution of folic acid meta-bolic risk grades in the uRPL population, and it was not related to the number of previous miscar-riages (P > 0.05). 1) Further statistical analysis of the genotype distribution of the three polymor-phic sites found that the overall proportion of CT (heterozygous mutation) at the C667T site of the MTHFR gene in the uRPL population increased, while the proportion of TT (homozygous mutation) decreased. When the number of previous spontaneous abortions was 3, the proportion of homozy-gous mutations decreased significantly (P < 0.05, P < 0.0167 after Bonferroni method correction), and the frequency of allele T also decreased significantly; 2) There was no significant difference in the distribution of A1298C (P > 0.05); 3) There was also no significant difference in the distribution of the MTRR gene A66G polymorphism in the uRPL population (P > 0.05), but in subgroup analysis, when the number of previous spontaneous abortions was 3, the proportion of homozygous muta-tions significantly decreased (P < 0.05, P < 0.0167 after Bonferroni method correction), but no sig-nificant difference was found in the frequency distribution of alleles. Conclusion: There was no sig-nificant difference in the distribution of risk level of folate metabolism among the women with uRPL. In the overall uRPL population, the frequencies of polymorphisms of MTHFR (C667T, A1298C), and MTRR (A66G) did not increase significantly, which were also not significantly associated with the number of previous miscarriages.