Fig. 2.

Involvement of DNA methylation in TNF-α-inducible EC-SOD down-regulation in fibroblasts. (A) DNA methylation of cytosine by DNMT and demethylation of 5mC by TETs. (B) Down-regulation of 5hmC by TNF-α. Fibroblasts were treated with or without 10 ng/ml TNF-α for 72 h. Isolated genomic DNA (80 or 40 ng) was subjected to a DNA dot blot analysis with an anti-5hmC antibody. Protein concentrations in samples were confirmed by staining with methylene blue (MB). (C) Fibroblasts were treated with or without 10 ng/ml TNF-α for 72 h, followed by bisulfite-MSP amplification with methylation (M) and unmethylation (U) site primers. (D) Change in 5hmC on the EC-SOD promoter by TNF-α. Fibroblasts were treated with or without 10 ng/ml TNF-α for 72 h, followed by a MeDIP analysis with a 5hmC antibody. *p<0.05 vs control (Ct).