Optimizing examination of methylenetetrahydrofolate reductase gene promoter methylation in cleft lip with or without cleft palate non-syndromic patients using the pyrosequencing method
Downloads
Background: Cleft lip with or without cleft palate (CL/P) is the most common congenital anomaly found in Indonesia. CL/P is caused by hereditary (genetic) and environmental factors. Environmental factors can result in methylation in the promoter of the methylenetetrahydrofolate reductase (MTHFR) gene, affecting its expression. Methylation takes place at the CpG site found at chromosome 1, coordinates 11,805,406–11,806,509. Pyrosequencing technology can detect the percentage methylation of a gene quickly, simply, and accurately.Purpose: The aim of the study is to optimize detection of methylation of the MTHFR gene using the pyrosequencing method.Methods: Methods used in this study were DNA extraction from blood, DNA bisulfite conversion, polymerase chain reaction (PCR), and methylation detection using CpG pyrosequencing assay. Samples were taken from 20 CL/P patients (C) and 44 normal patients (N).Results:The pyrosequencing method was successful in detecting methylation at three MTHFR gene sites at coordinates 11,805,507–11,805,529. The methylation level at the third site was higher in group C than in group N, while at the first and second positions, group C had a lower methylation level than group N. In general, the percentage of methylation for both groups was low or hypomethylated (less than 5%).Conclusion:The pyrosequencing method can be used to determine methylation levels in the MTHFR gene with the results presented as percentages (quantitative data). Hypomethylation occurs in groups C and N at the coordinates 11,805,507–11,805,529 of the MTHFR gene promoter.
Downloads
Nugraha AP, Yang H, Chen J, Yang K, Kraisintu P, Zaww K, Ma A, Wang R, Alhadi NEAM, Vanegas Sáenz JR, Hong G. β-tricalcium phosphate as alveolar bone grafting in cleft lip/palate: A systematic review. Dent J. 2023; 11(10): 234. doi: https://doi.org/10.3390/dj11100234
Lesmana S, Auerkari EI. Genes contributing in cleft lip and cleft palate: A literature review. J Int Dent Med Res. 2016; 9(Special Issue): 441–8. web: http://www.jidmr.com/journal/DENTISTRY/2016/Special_Issue/30_D16_60_Stella.pdf
Centre for Disease Control and Prevention (CDC). Facts about cleft lip and cleft palate. Birth defects. 2020. Available from: https://www.cdc.gov/ncbddd/birthdefects/cleftlip.html. Accessed 2022 Aug 22.
Ittiwut R, Siriwan P, Suphapeetiporn K, Shotelersuk V. Epidemiology of cleft lip with or without cleft palate in Thais. Asian Biomed. 2017; 10(4): 335–8. doi: https://doi.org/10.5372/1905-7415.1004.495
Priyono GP, Rafiyah I, Nurhidayah I. Parents' self esteem of children with cleft lip and palate. J Nurs Care. 2018; 1(2): 164–71. doi: https://doi.org/10.24198/jnc.v1i2.17129
Taslim T, Joenoes H, Sulistyani LD, Latief BS, Auerkari EI. MTHFR C677T polymorphism in Indonesian patients with oral cleft. J Int Dent Med Res. 2017; 10(Special Issue): 723–8. web: http://www.jidmr.com/journal/wp-content/uploads/2017/12/57.76Tania-Taslim-et-al-rev.pdf
Sosiawan A, Kurniati M, Danudiningrat CP, Wahjuningrum DA, Mulyawan I. The role of family history as a risk factor for non-syndromic cleft lip and/or palate with multifactorial inheritance. Dent J. 2021; 54(2): 108–12. doi: https://doi.org/10.20473/j.djmkg.v54.i2.p108-112
Fitrie RNI, Hidayat M, Dahliana L. Incidence of cleft lip with or without cleft palate at Yayasan Pembina Penderita Celah Bibir dan Langit-Langit (YPPCBL) in 2016-2019. J Med Heal. 2022; 4(1): 12. doi: https://doi.org/10.28932/jmh.v4i1.3396
Arif Harahap W. Role of DNA methylation as a diagnostic biomarker of sporadic breast cancer. J thee Med Sci (Berkala Ilmu Kedokteran). 2016; 48(4 Suplement): 42–3. doi: https://doi.org/10.19106/JMedScieSup004804201636
Ferrazzi E, Tiso G, Di Martino D. Folic acid versus 5- methyl tetrahydrofolate supplementation in pregnancy. Eur J Obstet Gynecol Reprod Biol. 2020; 253: 312–9. doi: https://doi.org/10.1016/j.ejogrb.2020.06.012
Coppedè, Stoccoro, Tannorella, Gallo, Nicolí¬, Migliore. Association of polymorphisms in genes involved in one-carbon metabolism with MTHFR methylation levels. Int J Mol Sci. 2019; 20(15): 3754. doi: https://doi.org/10.3390/ijms20153754
Fleckhaus J, Schneider PM. Novel multiplex strategy for DNA methylation-based age prediction from small amounts of DNA via pyrosequencing. Forensic Sci Int Genet. 2020; 44: 102189. doi: https://doi.org/10.1016/j.fsigen.2019.102189
Wan L, Li Y, Zhang Z, Sun Z, He Y, Li R. Methylenetetrahydrofolate reductase and psychiatric diseases. Transl Psychiatry. 2018; 8(1): 242. doi: https://doi.org/10.1038/s41398-018-0276-6
Dell'edera D, Tinelli A, Milazzo GN, Malvasi A, Domenico C, Pacella E, Pierluigi C, Giuseppe T, Marcello G, Francesco L, Epifania AA. Effect of multivitamins on plasma homocysteine in patients with the 5,10 methylenetetrahydrofolate reductase C677T homozygous state. Mol Med Rep. 2013; 8(2): 609–12. doi: https://doi.org/10.3892/mmr.2013.1563
Nasroen SL, Maskoen AM, Soedjana H, Soeria Soemantri ES, Hilmanto D. The effects of IRF6 rs2235373 polymorphism on mRNA expression changes in non-syndromic cleft lip and palate with various phenotypes. Padjadjaran J Dent. 2018; 30(3): 221–30. doi: https://doi.org/10.24198/pjd.vol30no3.17949
Kedar R, Chandel D. MTHFR gene polymorphism and associated nutritional deficiency in the etiology and pathogenesis of Down syndrome. Egypt J Med Hum Genet. 2019; 20(1): 12. doi: https://doi.org/10.1186/s43042-019-0010-9
Huang W-J, Lu X-L, Li J-T, Zhang J-M. Effects of folic acid on oligozoospermia with MTHFR polymorphisms in term of seminal parameters, DNA fragmentation, and live birth rate: a double"blind, randomized, placebo"controlled trial. Andrology. 2020; 8(1): 110–6. doi: https://doi.org/10.1111/andr.12652
Loscalzo J, Handy DE. Epigenetic modifications: basic mechanisms and role in cardiovascular disease (2013 Grover Conference series). Pulm Circ. 2014; 4(2): 169–74. doi: https://doi.org/10.1086/675979
Liu H-Y, Liu S-M, Zhang Y-Z. Maternal folic acid supplementation mediates offspring health via DNA methylation. Reprod Sci. 2020; 27(4): 963–76. doi: https://doi.org/10.1007/s43032-020-00161-2
Santana Bezerra H, Severo de Assis C, dos Santos Nunes MK, Wanderley de Queiroga Evangelista I, Modesto Filho J, Alves Pegado Gomes CN, Ferreira do Nascimento RA, Pordeus Luna RC, de Carvalho Costa MJ, de Oliveira NFP, Camati Persuhn D. The MTHFR promoter hypermethylation pattern associated with the A1298C polymorphism influences lipid parameters and glycemic control in diabetic patients. Diabetol Metab Syndr. 2019; 11(1): 4. doi: https://doi.org/10.1186/s13098-019-0399-9
Charoenvicha C, Sirimaharaj W, Khwanngern K, Chattipakorn N, Chattipakorn S. Alterations in DNA methylation in orofacial clefts. Int J Mol Sci. 2022; 23(21): 12727. doi: https://doi.org/10.3390/ijms232112727
Copyright (c) 2024 Dental Journal
This work is licensed under a Creative Commons Attribution-ShareAlike 4.0 International License.
- Every manuscript submitted to must observe the policy and terms set by the Dental Journal (Majalah Kedokteran Gigi).
- Publication rights to manuscript content published by the Dental Journal (Majalah Kedokteran Gigi) is owned by the journal with the consent and approval of the author(s) concerned.
- Full texts of electronically published manuscripts can be accessed free of charge and used according to the license shown below.
- The Dental Journal (Majalah Kedokteran Gigi) is licensed under a Creative Commons Attribution-ShareAlike 4.0 International License
