懷孕婦女血中葉酸代謝物分析與其代謝途徑相關基因多型性之探討

Abstract

研究背景 葉酸是維生素B群的一種，從流行病學針對懷孕併發症與新生兒神經管缺損等與葉酸之間關係的研究，目前在臨床上皆建議婦女在懷孕初期攝取高劑量的補充，以降低疾病罹患風險。同時利用代謝質體學與藥物基因體學的研究，我們試以代謝物的角度來觀察懷孕婦女接受葉酸補充的情形和探討其與基因間的關係。 研究方法 本研究共收集96年1月至96年4月惠心婦幼診所之門診孕婦，於例行抽血檢驗時採取血樣進行LC/MS-MS與HPLC葉酸代謝物分析，並且以問卷方式收集孕婦葉酸補充劑之使用情形、基本資料、飲食狀況等資訊，以SNPstream鑑定孕婦的MTHFR、MTR、MTRR、MTHFD1、ALDH1L1、AMT、FOLH1、BHMT等基因型，研究懷孕婦女體內葉酸代謝物分布與基因型的相關性。 研究結果 89位受試者中包含7位一般健康女性、26位未服用葉酸之孕婦和56位服用葉酸的懷孕婦女，成功測定出FA、5MeTHF、5FoTHF、5,10CHTHF四種葉酸代謝物和同半胱胺酸濃度，於曾補充葉酸的孕婦中血球5MeTHF則呈現逐漸增加的狀況，未補充葉酸者的血漿葉酸和血球葉酸雖呈逐漸降低之趨勢但都未達統計上顯著。 懷孕婦女體內血漿同半胱胺酸濃度於每週期比較中，對於使用葉酸與否並無明顯差異，然而血漿同半胱胺酸濃度與血漿葉酸代謝物濃度有顯著反比關係。 成功鑑定18個基因型中有12個基因包括MTHFR、MTR、MTRR、ALDH1L1、AMT、FOLH1、BHMT都分別發現不同基因型與葉酸代謝物濃度之間的關聯性。在未服用葉酸的懷孕婦女中，5FoTHF/5,10CHTHF比值依MTHFR、MTRR、ALDH1L1的基因型而有高低不同的分布。 結論 我們發現受到葉酸補充影響代謝物濃度最為顯著的是5MeTHF，能即時提高血漿5MeTHF濃度，而在曾持續補充葉酸的孕婦中，也觀察到血球5MeTHF濃度隨週期而提升，可能是藉由荷爾蒙調節而提高體內葉酸儲存量所致。在未服用葉酸的受試者中，葉酸代謝途徑的MTHFR、MTR、MTRR、ALDH1L1、AMT等基因型對於葉酸各代謝物的濃度均有顯著性影響，而5FoTHF/5,10CHTHF在MTHFR、MTRR、ALDH1L1的5個SNP中發現具變異型配子的受試者有比值較低的趨勢，可以作為單純評估個體受到來自於基因型層面影響的依據。

Background Folate is one of vitamin B complex. Epidemiological studies reveal the incidence of pregnancy complications and newborn neural tube defect in folate-insufficient pregnant women. Thereby, it is recommended to boost higher dose of folic acid during the first trimester of pregnancy for the prevention from unwilling complications. Combining the knowledge of metabolomics and pharmacogenomics, we tried to study the effect of folate intake in pregnant women and genetic impact from the view point of folate metabolites. Methods Healthy and pregnant women in outpatient service of Eugene Clinic were enrolled in our study. We collected blood samples during their routine blood drawn for LC/MS-MS and HPLC detection of folate metabolites. The use of folate supplement, basic demographic data and dietary preference were surveyed by a questionary. Genotypes of the selected genes including MTHFR, MTR, MTRR, MTHFD1, ALDH1L1, AMT, FOLH1and BHMT were determined using SNPstream system. Results The 89 participants include 7 healthy non-pregnant women, 26 pregnant women without folate supplement and 56 pregnant women with regular folate intake. We successfully detected FA, 5MeTHF, 5FoTHF, 5,10CHTHF, and plasma homocysteine concentration. In pregnant women receiving daily folate supplement, erythrocyte 5MeTHF concentration increases with the gestation age. Although plasma and erythrocyte folate concentration decrease with time in pregnant women without folate supplement, it is not statistically significant (p> 0.05). Folate supplement did not make difference in plasma homocysteine levels of pregnant women. However there’s an inverse relation between plasma folate metabolites and homocysteine. Among18 successfully determined genes, genotypes of MTHFR, MTR, MTRR, ALDH1L1, AMT, FOLH1, BHMT genes were correlated to the concentrations of folate metabolites. The 5FoTHF/5,10CHTHF ratio is correlated to the distribution of genotypes of MTHFR, MTRR, and ALDH1L1. Conclusion Folate supplement importantly changed 5MeTHF concentration compared to the other folate metabolites and elevated plasma 5MeTHF immediately. In the pregnant women with regular folate supplement, erythrocyte 5MeTHF level increases with gestational age. It might be due to the increase during pregnancy in folate uptake capacity via altered hormone regulation. In pregnant women without folate supplement, levels of folate metabolites varied, which was associated with genotypes of MTHFR, MTR, MTRR, ALDH1L1, and AMT. The 5FoTHF/5,10CHTHF ratio is relatively low in pregnant women carrying heterozygotic SNPs of MTHFR, MTRR, ALDH1L1 genes. It may be as a maker for evaluating genetic effects on folate maintenance.