(6S)-5-Methyltetrahydrofolate Dicholine Salt Modulates Methyl-Donor Metabolites and Repetitive-Element DNA Methylation in HepaRG Cells

BACKGROUND: One-carbon metabolism supports methyl-donor availability for cellular methylation reactions. (6S)-5-methyltetrahydrofolate dicholine salt (5-MTHF-DCh) is a source of bioactive folate and choline, and a potential modifier of one-carbon metabolism and DNA methylation. However, despite the potential relevance of this dual folate-choline formulation to dietary supplement science, its effects on methyl-donor metabolism and downstream epigenetic endpoints remain insufficiently characterized. STUDY DESIGN: Here, we tested whether 5-MTHF-DCh modifies one-carbon metabolism endpoints and DNA methylation of selected repetitive elements (RE) in HepaRG cells. Differentiated HepaRG cells were treated with 5-MTHF-DCh (0, 1, 10, or 40 nM) for 96 h. Cellular S-adenosylmethionine (SAM), S-adenosylhomocysteine (SAH), and methionine were quantified by liquid chromatography-tandem mass spectrometry (LC-MS/MS). DNA methylation was assessed for five RE (L1PA2, L1PA13, L1PREC1, Alu, and HERVK11I long terminal repeat, LTR). Transcript levels were measured by quantitative real time PCR (qRT-PCR) for DNA methylation machinery and one-carbon enzymes as well as for selected RE. Dose effects were evaluated by ANOVA with Holm-adjusted comparisons. RESULTS: = 0.074), while the mRNA levels of the DNA methylation machinery remained unchanged. In HepaRG cells, 5-MTHF-DCh increased SAM and was associated with selective increases in DNA methylation at specific RE, without detectable changes in core DNA methylation machinery transcripts. CONCLUSIONS: These findings support 5-MTHF-DCh as a modulator of methyl-donor status with measurable epigenetic endpoints in a human hepatic cell model, and provide mechanistic evidence relevant to further evaluation of this folate-choline ingredient.