Dna methylation change in genes involved in alcohol metabolism (aldh2) and one-carbon metabolism (mthfr) pathways among people with alcohol use disorder with withdrawal seizure

Alcohol use disorder (AUD) is a serious public health concern. It is influenced by both genetic predisposition and environmental factors, such as the availability of alcohol. studies have also examined genetic loci associated with specific alcohol related phenotypes such as age of initiation of drinking, subjective response to alcohol and consequences of heavy alcohol use, such as complicated withdrawal. Aldehyde dehydrogenase (ALDH2) is the alcohol metabolizing gene which is involved in conversion of acetaldehyde to acetate. MTHFR is a gene involved in one-carbon metabolism, both genes are known to be associated with AUD. Single Nucleotide Variants of MTHFR are associated with alcohol withdrawal seizure. Hyperhomocysteinemia in alcohol dependent subjects has been shown to be associated with alcohol withdrawal seizures and atrophic brain changes. There are evidence suggests that epigenetics may play a role in mechanism underlying withdrawal seizure. It is unclear why only some individuals get withdrawal seizure. We aim to estimate DNA methylation levels in AUD with seizure. Study included male patients Alcohol Use Disorder (N=89) based on ICD 10 criteria, recruited from the clinical services of St John's Medical College Hospital (SJMCH) (Gastroenterology and Psychiatry) and NIMHANS (Centre for Addiction Medicine) after informed consent. Out of 89 AUD cases N=12 had a seizure (SZ+ve) and N=77 had no seizure (SZ-ve). Genotyping was done for variants ALDH2 (rs2238151) and MTHFR (rs1801133, rs1801131) and plasma homocysteine levels were estimated for all the samples. We have Genomic DNA that was bisulfite converted (Zymo gold Bisulfite Kit) for ALDH2 and MTHFR loci. DNA methylation was done by pyrosequencing using Pyromark Q24 (Qiagen). We used Mann-Whitney U tests to analyze differences in both ALDH2 and MTHFR methylation (Individual CpG sites) between groups. Following this we did a logistic regression analysis to study association between DNA methylation levels and SZ+ve while correcting for amount of alcohol consumption and duration of drinking. We studied 3 CpG sites at MTHFR locus and we observed significant hypomethylation at all 3 CpG sites (CpG1, p= < 0.01; CpG2, p= < 0.01; CpG3, p=0.01) in SZ+ve group compared to SZ-ve group. The association remained significant even after correcting for the quantity of alcohol consumption and duration of drinking. At ALDH2 locus we observed decreased methylation levels in SZ+ve group compared to SZ-ve group but the difference did not reach statistical significance. No association was observed between homocysteine levels and alcohol seizure. We did not observe any association between variants analyzed and alcohol withdrawal seizure. Alcohol interacts with methionine synthase (MS), which is responsible for the conversion of homocysteine to methionine. Inhibition of MS leads to disturbance in S-adenosyl methionine (SAM) and S-adenosylhomocysteine (SAH). SAM to SAH conversion is controlled by DNA-methyltransferases (DNMTs) and MTHFR is the key enzyme involved in this process and changes in methylation levels of MTHFR may disturbance of this pathway which will affect the brain function. Studies have reported the interplay between alcohol use, homocysteine, and DNA methylation. In the present study, we have studied DNA methylation from blood samples and not from brain tissue. Altered methylation in blood DNA at candidate loci might serve as a biomarker for prolonged severe alcohol abuse.