title: Suppression of type I collagen gene expression by prostaglandins in fibroblasts is mediated at the transcriptional level.
abstract: 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. Interleukin-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.
  
  
  METHODS
  BALBc/3T3 fibroblasts were treated with IL-1 and prostaglandins in the absence and presence of indomethacin to inhibit endogenous prostaglandin biosynthesis. Collagen synthesis was analyzed by SDS-PAGE as [3H]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-1-stimulated synthesis of PGE2 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 PGF2alpha > PGE2 > PGE1 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 PGE2- or PGE1-treated BALBc/3T3 cells. EMSA analysis showed zinc-dependent differences in the binding of nuclear factors from untreated 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.
authors.affiliation: Rheumatology Division, Beth Israel Deaconess Medical Center, New England Baptist Bone & Joint Institute, Harvard Institutes of Medicine, Boston, Massachusetts, USA.
authors.forename: F B
authors.initials: FB
authors.lastname: Riquet
authors.forename: W F
authors.initials: WF
authors.lastname: Lai
authors.forename: J R
authors.initials: JR
authors.lastname: Birkhead
authors.forename: L F
authors.initials: LF
authors.lastname: Suen
authors.forename: G
authors.initials: G
authors.lastname: Karsenty
authors.forename: M B
authors.initials: MB
authors.lastname: Goldring
mesh_terms: D000527:Alprostadil / Q000494:pharmacology; D000818:Animals; D002478:Cells, Cultured; D003094:Collagen / Q000235:genetics*; D004247:DNA / Q000235:genetics / Q000378:metabolism; D018983:DNA Footprinting; D004268:DNA-Binding Proteins / Q000032:analysis; D003850:Deoxyribonuclease I / Q000378:metabolism; D015232:Dinoprostone / Q000096:biosynthesis / Q000494:pharmacology; D015536:Down-Regulation / Q000187:drug effects; D005347:Fibroblasts; D017930:Genes, Reporter / Q000235:genetics; D007213:Indomethacin / Q000494:pharmacology; D007375:Interleukin-1 / Q000494:pharmacology; D051379:Mice; D008807:Mice, Inbred BALB C; D016595:Misoprostol / Q000494:pharmacology; D011401:Promoter Regions, Genetic / Q000235:genetics; D011453:Prostaglandins / Q000096:biosynthesis / Q000494:pharmacology*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects*; D014162:Transfection
chemical_list: D004268:DNA-Binding Proteins; D007375:Interleukin-1; D011453:Prostaglandins; D012333:RNA, Messenger; D016595:Misoprostol; D003094:Collagen; D004247:DNA; D003850:Deoxyribonuclease I; D000527:Alprostadil; D015232:Dinoprostone; D007213:Indomethacin