Age-associated changes in DNA methylation and expression of the TNFα gene in pigs

DNA methylation is an important mediator of gene expression regulation and has been shown to be closely linked to aging. Immune-related genes tend to be influenced by DNA methylation at different ages. To explore DNA methylation changes in the porcine TNFα gene and analyze their potential effects on gene expression, we measured the methylation level of the TNFα promoter and TNFα mRNA expression in the spleen of Meishan piglets at six developmental stages (1, 7, 14, 21, 28 and 35 days old) by bisulfite sequencing PCR and quantitative PCR. The results revealed a trend for TNFα promoter methylation level to increase and mRNA expression to decrease with age. Correlation analysis showed a significant negative association between methylation level and mRNA expression (Pearson's r = -0.775, P = 4.87E-07). In addition, the transcription factor Sp1 was revealed to bind with the TNFα promoter and regulate TNFα expression. DNA methylation in the TNFα promoter was found to decrease the promoter's activity, and methylation inhibition could enhance the expression level of TNFα, providing functional evidence that promoter methylation controls TNFα expression. Together, our data provide insights into age-associated changes in promoter methylation of the TNFα gene in the spleen and contribute to our understanding of regulatory mechanisms for TNFα expression in the immune system of pigs.