Genetic variation of the folate metabolic pathway in recurrent pregnancy loss in Azerbaijani women
DOI:
https://doi.org/10.37800/RM.3.2026.670Keywords:
thrombophilia, recurrent pregnancy loss , methylenetetrahydrofolate reductase , methionine synthase , methionine synthase reductaseAbstract
Relevance: Hereditary thrombophilias are a common cause of treatable recurrent miscarriages. The role of genetic thrombophilia in such women has been evaluated, and methylenetetrahydrofolate reductase (MTHFR) polymorphisms have been implicated in RPL and other reproductive complications.
The study aimed to examine the frequency of these molecular alterations in women after 2 or more spontaneous pregnancy losses.
Materials and Methods: Genetic analyses were performed via real-time PCR using the RealBest-D3801 kit (“Vektor-Best” Koltsovo, Russia) on venous blood samples from 384 women with a history of miscarriages and 192 healthy control women with normal, complication-free pregnancies and healthy live births.
Results: Statistical analysis of individual polymorphisms demonstrated a highly significant association between recurrent pregnancy loss (RPL) and the heterozygous variants of MTHFR A1298C (61,7% in RPL vs 43,2 % in controls, P = 0,0003), MTR A2756G (44,5% vs 30,2%, P = 0,0037), and MTRR A66G (63,5% vs 48,1%, P = 0,000027). Conversely, the homozygous mutant genotypes for these genes, as well as both heterozygous and homozygous variants of MTHFR C677T (P = 0,816), showed no statistically significant differences between the studied groups. Regarding combined polymorphisms, the simultaneous carriage of multiple variants did not correlate with an increased risk of RPL (P > 0.05 for all analyzed combinations). The triple heterozygous combination (MTHFR A1298C (AC)+MTR A2756G (AG)+MTRR A66G (AG)) exhibited a minor, non-significant trend toward higher frequency in the RPL group (9,9% vs 5,7%, P = 0,091), further supporting that the co-existence of these thrombophilia-related mutations does not exert a statistically significant cumulative or synergistic effect on RPL susceptibility.
Conclusions: The studied MTHFR, MTR, and MTRR polymorphisms independently act as genetic risk factors for RPL, primarily driven by their heterozygous status. Their combined carriage lacks synergistic risk enhancement, suggesting individual screening holds higher clinical relevance than complex multi-locus evaluation.
References
1. Genovese HG, McQueen DB. The prevalence of sporadic and recurrent pregnancy loss. Fertil Steril. 2023 Nov;120(5):934-936
https://doi.org/10.1016/j.fertnstert.2023.08.954
2. Hyde KJ, Schust DJ. Genetic considerations in recurrent pregnancy loss. Cold Spring Harb Perspect Med. 2015 Feb 6;5(3):a023119.
https://doi.org/10.1101/cshperspect.a023119
3. Shahine L, Lathi R. Recurrent pregnancy loss. Obstet Gynecol Clin. North Am. 2015;42:117–134.
https://doi.org/10.1016/j.ogc.2014.10.002
4. Deng TB, Liao XB, Zhu S. Recent advances in treatment of recurrent spontaneous abortion. Obstet Gynecol Surv. 2022;77:355–366.
https://doi.org/10.1097/OGX.0000000000001033
5. Diao J, Luo L , Li J, Zhang S, Li Y, Qin J, Maternal homocysteine and folate levels and risk of recurrent spontaneous abortion: A meta-analysis of observational studies. 2020 Dec;46(12):2461-2473.
https://doi.org/10.1111/jog.14500
6. dos Santos TB, de Paula HK, Balarin MAS, Silva-Grecco RL, Lima MFP, Rodrigues de Resende EAM, Gomes MKO, Cintra MTR. Can the genetic polymorphisms of the folate metabolism have an influence in the polycystic ovary syndrome? Arch Endocrinol Metab. 2019;63(5):501-508.
https://doi.org/10.20945/2359-3997000000167
7. Xu Y, Ban Y, Ran L, Yu Y, Zhai S, Sun Z, Zhang J, Zhang M, Hong T, Liu R, Ren L, Hu L. Relationship between unexplained recurrent pregnancy loss and 5,10-methylenetetrahydrofolate reductase polymorphisms. Fertil Steril. 2019;111,597-603.
https://doi.org/10.1016/j.fertnstert.2018.11.011
8. Nguyen Ngoc N, Tran Ngoc Thao M, Trieu Tien S, Vu Tung S, Le H, Ho Sy H, Nguyen Thanh T, Trinh The S. Evaluating the Association Between Genetic Polymorphisms Related to Homocysteine Metabolism and Unexplained Recurrent Pregnancy Loss in Women. Appli Clin Genet. 2022:15:55–62.
https://doi.org/10.2147/TACG.S365281
9. Coppedè F. Pregnancy loss and polymorphisms in folic acid genes. In: Molecular Nutrition: Mother and Infant. 2021;317-335.
https://doi.org/10.1016/B978-0-12-813862-5.00013-X
10. Adelekan OO, Uche EI, Balogun TM, Osunkalu VO, Akinbami AA, Ismail KA, Badiru MA, Dosunmu AO, Kamson OR. Methylene tetrahydrofolate reductase gene mutation in sickle cell anaemia patients in Lagos, Nigeria. Pan Afr Med J. 2019 Dec 30;34:213.
https://doi.org/10.11604/pamj.2019.34.213.19524
11. Binia A, Contreras AV, Canizales-Quinteros S, et al. Geographical and ethnic distribution of single nucleotide polymorphisms within genes of the folate/homocysteine pathway metabolism. Genes Nutr. 2014;9:421.
https://doi.org/10.1007/s12263-014-0421-7
12. Clare CE, Brassington AH, Kwong WY, Sinclair KD. One-Carbon Metabolism: Linking Nutritional Biochemistry to Epigenetic Programming of Long-Term Development. Annu Rev Anim Biosci. 2019;7:263–287.
https://doi.org/10.1146/annurev-animal-020518-115206
13. Yıldırmak ZY, Genç DB, Kural A, et al. Assessment of vitamin B12 and homocysteine levels in pregnant women admitted for delivery and cord blood samples of their newborn babies: a multicenter study. Turk J Pediatr 2024; 66: 429-438.
https://doi.org/10.24953/turkjpediatr.2024.4545
14. Akbarova DA. The clinical importance of homocysteine levels during pregnancy: Implications for maternal and fetal health. Int J Med Sci. 2025;5(06):244-249.
https://www.academicpublishers.org/journals/index.php/ijms/article/view/5282/6216
15. Nasri K. Overview of neural tube defects: history, folate-homocysteine cycle, MTHFR C677T polymorphism and preventions. J Pediatr Neonatal Individual Med. 2025;14(2):e140201.
https://doi.org/10.7363/140201
16. Kozłowska-Wytyk M, Kurzawińska G, Ożarowski M, Mrozikiewicz AE, Olbromski P, Karpiński TM, Czerny B, Wolski H. The Importance of Gene Variants Related to Folate Uptake and Distribution in the Etiology of Recurrent Miscarriage and Intrauterine Fetal Demise. Appl Sci. 2024;14(24):11847.
https://doi.org/10.3390/app142411847
17. Practice Committee of the American Society for Reproductive Medicine. Definitions of infertility and recurrent pregnancy loss: a committee opinion. Fertil Steril. 2020; 113(3): 533–535.
https://doi.org/10.1016/j.fertnstert.2019.11.025
18. Feng W, Zhang Y, Pan Y, Zhang Y, Liu M, Huang Y, Xiao Y, Mo W, Jiao J, Wang X, Tian D, Yang L, Ma Y. Association of three missense mutations in the homocysteine-related MTHFR and MTRR gene with risk of polycystic ovary syndrome in Southern Chinese women. Reprod Biol Endocrinol. 2021;19(1):5.
https://doi.org/10.1186/s12958-020-00688-8
19. Lu M, Peng K, Song L, Luo L, Liang P, Liang Y. Association between genetic polymorphisms in methylenetetrahydrofolate reductase and risk of autoimmune diseases: a systematic review and meta-analysis. Dis Markers. 2022; 1: 4568145.
https://doi.org/10.1155/2022/4568145
20. Blanco-Muñoz J, Lacasaña M, Gamboa R, Huesca-Gómez C, Hernández-Mariano JÁ, Aguilar-Garduño C. Interaction between MTHFR 677C>T, PON1 192Q>R and PON1 55L>M polymorphisms and its effect on non-recurrent spontaneous abortion in Mexican women. Gene. 2019;689:69–75.
https://doi.org/10.1016/j.gene.2018.11.093
21. Wu AN, Yu RX, Song KW, Tuo Y. Genetic polymorphisms in folate metabolism and their association with recurrent spontaneous abortion. J Matern Fetal Neonatal Med. 2025;38(1):250576
https://doi.org/10.1080/14767058.2025.2505769
22. Kaye AD, Jeha GM, Pham AD, Fuller MC, Lerner ZI, Sibley GT, Cornett EM, Urits I, Viswanath O, Kevil CG. Folic Acid Supplementation in Patients with Elevated Homocysteine Levels. Adv Ther. 2020;37(10):4149–4164.
https://doi.org/10.1007/s12325-020-01474-z
23. Lino FL, Traina É, Barreto JA, Moron AF, Mattar R. Thrombophilic mutations and polymorphisms, alone or in combination, and recurrent spontaneous abortion. Clin Appl Thromb Hemost. 2015;21(4):365–372. Epub 2014.
https://doi.org/10.1177/1076029613520465
24. Thrombophilia and hyperhomocysteinemia [Internet]. Vector-best.ru. 2025. Available from:
25. Nucleic acids extraction [Internet]. Vector-best.ru. 2025. Available from:
https://en.vector-best.ru/catalog/pcr/kits/nucleic-acids-extraction/
26. Szafarowska M, Segiet A, Jerzak MM. Methylenetetrahydrofolate reductase A1298C and C677T polymorphisms and adverse pregnancy outcome in women with PCOS. Neuro Endocrinol Lett. 2016;37(2):141–146.
https://pubmed.ncbi.nlm.nih.gov/27179578/
27. Zhou J, Zhu Y, Liu Y, Zhan H, Niu P, Chen H, Zhang J. (2024). The association between methionine synthase reductase c.66A>G variant and the risk of recurrent pregnancy loss: A systematic review and meta-analysis. J Gynecol Obstet Hum Reprod. 2024;53(10):102849.
https://doi.org/10.1016/j.jogoh.2024.102849
28. Al-Achkar W, Wafa A, Ammar S, Moassass F, Jarjour RA. Association of methylenetetrahydrofolate reductase C677T and A1298C gene polymorphisms with recurrent pregnancy loss in Syrian women. Reprod Sci. 2017;24(9):1275–1279.
https://doi.org/10.1177/1933719116682874
29. Li S, Wu Y, Gao Y, et al. Maternal hyperhomocysteinemia induces fetal growth restriction by suppressing angiogenesis at the maternal-fetal interface. Cell Biosci 2026;16:17
https://doi.org/10.1186/s13578-025-01529-0
30. Afaq E, Ali A, Jamil R, Waseem HF. Association of plasma folic acid, vitamin-B12 and homocysteine with recurrent pregnancy loss. “A case control study”. Pak J Med Sci. 2023 Sep-Oct;39(5):1280-1285.
https://doi.org/10.12669/pjms.39.5.7432
31. Wan L, Li Y, Zhang Z, Sun Z, He Y, Li R. Methylenetetrahydrofolate reductase and psychiatric diseases. Transl Psychiatry. 2018 Nov 5;8(1):242.
https://doi.org/10.1038/s41398-018-0276-6
32. Zhao L, Li T, Dang M, Li Y, Fan H, Hao Q, Song D, Lu J, Lu Z, Jian Y, Wang H, Wang X, Wu Y and Zhang G. Association of methylenetetrahydrofolate reductase (MTHFR) rs1801133 (677C>T) gene polymorphism with ischemic stroke risk in different populations: An updated meta-analysis. Front Genet. 2023;13:1021423.
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