Suppression of type I collagen gene expression by prostaglandins in fibroblasts is mediated at the transcriptional level

Background: Tissues undergoing a chronic inflammatory process, such as the synovium in rheumatoid arthritis, are characterized by the infiltration of lymphocytes of different subsets and activation of monocyte/macrophages. Interleultin-1 (IL-1), a monocyte/macrophage product that stimulates synovial fibroblasts to produce matrix metalloproteinases (MMPs), prostaglandins, and other cytokines, also has profound effects on the synthesis of extracellular matrix components such as type I collagen. In previous studies, we have shown that synovial fibroblasts and chondrocytes isolated from human joint tissues are particularly sensitive to prostaglandins, which modulate the effects of IL-1 on collagen gene expression in an autocrine manner. Materials and Methods: BALBc/3T3 fibroblasts were treated with IL-I and prostaglandins in the absence and presence of indomethacin to inhibit endogenous prostaglandin biosynthesis. Collagen synthesis was analyzed by SDS-PAGE as [H-3]proline-labeled, secreted proteins, and prostaglandin production and cyclic adenosine 3',5'-cyclic monophosphate (camp) content were assayed. The expression of type I collagen gene (Col1a1) promoter-reporter gene constructs was examined in transient transfection experiments, and the binding of nuclear factors to the Col1a1 promoter region spanning -222 bp/+ 116 bp was analyzed by DNase I footprinting and electrophoretic mobility shift (EMSA) assays. Results: IL-1 increased the synthesis of type I and type III collagens in BALBc/3T3 fibroblasts; greater increases were observed when IL-l-stimulated synthesis of PGE, was blocked by indomethacin. Transient transfection experiments demonstrated dose-dependent inhibition of the -222 bp Col1a1 promoter by exogenously added prostaglandins with the order of potency of PGF(2)alpha > PGE(2) > PGE(1). DNase I footprinting showed increased protection, which extended from the region immediately upstream of the TATA box, owing to the binding of nuclear factors from PGE(2)- or PGE(1)-treated BALBc/3T3 cells. EMSA analysis showed zinc-dependent differences in the binding of nuclear factors from unheated and prostaglandin-treated cells to the -84 bp/-29 bp region of the Col1a1 promoter. conclusions: These results show that the inhibition of Col1a1 expression by IL-1 in fibroblasts is mediated by prostaglandins at the transcriptional level and suggest that PGE-responsive factors may interact directly or indirectly with basal regulatory elements in the proximal promoter region of the Col1a1 gene.