Cytokine-induced Chromatin Modifications of the Type I Collagen Alpha 2 Gene during Intestinal Endothelial-to-Mesenchymal Transition

Abstract

Fibrosis of the intestine is currently an irreversible complication of inflammatory bowel disease; yet, little is understood of the underlying pathogenesis and antifibrotic strategies remain elusive. To develop effective therapies, knowledge of the mechanism of transcription and excessive deposition of type I collagen, a hallmark of fibrosis, is needed. We have shown previously that endothelial-to-mesenchymal transition (EndoMT) contributes to the pool of intestinal fibrotic cells and that a cytokine cocktail (interleukin 1-β, tumor necrosis factor α, and transforming growth factor β) induces collagen I alpha 2 (COL1A2) mRNA and protein. Chromatin immunoprecipitation assays on pure cultures of human intestinal mucosal endothelial cells undergoing EndoMT were performed with antibodies to specific histone modifications and RNA polymerase II. Reverse transcriptase-PCR was used to quantify the levels of Col1A2 and endothelial-specific von Willebrand factor (vWF) mRNA. We showed that cytokines induce selective chromatin modifications (histone 4 hyperacetylation, and hypermethylation of histone 3) and phosphorylated RNA polymerase II at the COL1A2 promoter. Hypoacetylated and hypomethylated histone 3 was detected on the repressed vWF gene. Prolonged exposure to cytokines (16 days) retained hyperacetylation of select lysines in H4 on the COL1A2 promoter. Removal of cytokines after 16 days and continued culture for 10 days showed persistent hyperacetylation at lysine 16 in histone H4. This is the first study to show that COL1A2 gene expression is associated with cytokine-induced, temporally ordered, and persistent chromatin modifications and suggests that these are important determinants of gene expression in EndoMT and intestinal fibrosis.