title: Inflammation-induced up-regulation of TLR2 expression in human endothelial cells is independent of differential methylation in the TLR2 promoter CpG island.
abstract: Toll-like receptors play an important role in endothelial inflammation; however, little is known on the mechanisms regulating their expression. Differential promoter DNA methylation is an increasingly recognized mechanism that determines a switch between gene silencing and gene transcription. We hypothesized that epigenetic mechanisms are involved in the regulation of endothelial TLR2 expression because of the localization of the TLR2 promoter on a CpG-island. Resting human umbilical vein endothelial cells (HUVECs) displayed rather low TLR2 mRNA expression, while a strong expression increase occurred under inflammatory conditions. We examined the TLR2 promoter methylation pattern in resting HUVECs and compared it to cells treated either with the inflammatory cytokine TNF-α or the DNA-demethylating agent 5-azacytidine. DNA bisulfite conversion was followed by either genomic sequencing or single nucleotide primer extension (SNuPE) HPLC. Results of both techniques showed a low- or non-methylated TLR2 promoter in resting HUVECs and no alteration of the methylation pattern under inflammatory conditions. Whereas 5-azacytidine significantly increased the mRNA expression of the epigenetically regulated gene H19, TLR2 expression was not affected. Taken together, employing different methodological approaches, our data show no implication of methylation pattern changes in inflammatory induction of TLR2 expression in human endothelial cells.
authors.affiliation: Department of Pharmaceutical Biology, Saarland University, Saarbrücken, Germany.
authors.forename: Britta
authors.initials: B
authors.lastname: Diesel
authors.forename: Nadège
authors.initials: N
authors.lastname: Ripoche
authors.forename: Rebecca T
authors.initials: RT
authors.lastname: Risch
authors.forename: Sascha
authors.initials: S
authors.lastname: Tierling
authors.forename: Jörn
authors.initials: J
authors.lastname: Walter
authors.forename: Alexandra K
authors.initials: AK
authors.lastname: Kiemer
mesh_terms: D001374:Azacitidine / Q000494:pharmacology; D002460:Cell Line; D018899:CpG Islands / Q000235:genetics; D019175:DNA Methylation / Q000187:drug effects; D042783:Endothelial Cells / Q000187:drug effects / Q000276:immunology / Q000378:metabolism* / Q000473:pathology; D044127:Epigenesis, Genetic; D005786:Gene Expression Regulation; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D007113:Immunity, Innate; D007249:Inflammation / Q000235:genetics / Q000276:immunology*; D011401:Promoter Regions, Genetic / Q000235:genetics; D017422:Sequence Analysis, DNA; D051195:Toll-Like Receptor 2 / Q000235:genetics*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
chemical_list: C495342:TLR2 protein, human; D051195:Toll-Like Receptor 2; D014409:Tumor Necrosis Factor-alpha; D001374:Azacitidine