title: Downregulation of a rheumatoid arthritis-related antigen (RA-A47) by ra-a47 antisense oligonucleotides induces inflammatory factors in chondrocytes.
abstract: Previously we have shown that the expression of RA-A47 (rheumatoid arthritis-related antigen) which is identical to HSP47, a collagen-binding chaperon, is downregulated in chondrocytes by tumor necrosis factor alpha (TNFalpha). RA-A47 was also found on the surface of chondrocytes where it is recognized as an antigen in the serum of rheumatoid arthritis (RA) patients. Its translocation to the cell surface from endoplasmic reticulum membrane where it is normally located was also enhanced by TNFalpha. To understand the significance of RA-A47 downregulation in chondrocytes independent from other effects of TNFalpha, we used an antisense oligonucleotide approach and investigated the effect of this treatment on the expression of molecules related to matrix degradation and production of growth factors for chondrocytic, endothelial, and synovial cells. Here we show that treatment of rabbit chondrocyes and human chondrosarcoma cells HCS-2/8 by ra-a47 antisense S-oligonucleotides significantly reduced the expression of ra-a47 both at mRNA and protein level. Interestingly, this TNFalpha-independent RA-A47 downregulation was associated with a strong induction of matrix metalloproteinase (MMP)-9 mRNA and inducible NO synthase (iNOS) mRNA. The induction of active-type MMP-9 was further detected by gelatin zymography. Under the same conditions, the release of basic fibroblast growth factor (bFGF) and connective tissue growth factor (CTGF) from HCS-2/8 cells into the conditioned medium (CM) was strongly enhanced. These effects were not a result of TNFalpha upregulation, since the ra-a47 antisense oligonucleotide treatment did not enhance TNFalpha synthesis. These observations indicate that downregulation of RA-A47 induces TNFalpha-independent cartilage-degrading pathways involving iNOS and MMP-9. Furthermore, the stimulation of bFGF and CTGF release from chondrocytes may stimulate the proliferation of adjacent endothelial and/or synovial cells.
authors.affiliation: Department of Biochemistry and Molecular Dentistry, Okayama University Graduate School of Medicine and Dentistry, Okayama, Japan.
authors.forename: Takako
authors.initials: T
authors.lastname: Hattori
authors.forename: Harumi
authors.initials: H
authors.lastname: Kawaki
authors.forename: Satoshi
authors.initials: S
authors.lastname: Kubota
authors.forename: Yasutaka
authors.initials: Y
authors.lastname: Yutani
authors.forename: Benoit
authors.initials: B
authors.lastname: de Crombrugghe
authors.forename: Klaus
authors.initials: K
authors.lastname: von der Mark
authors.forename: Masaharu
authors.initials: M
authors.lastname: Takigawa
mesh_terms: D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D019902:Chondrocytes / Q000378:metabolism*; D002813:Chondrosarcoma / Q000378:metabolism; D055513:Connective Tissue Growth Factor; D017077:Culture Media, Conditioned / Q000737:chemistry; D015536:Down-Regulation; D004790:Enzyme Induction; D004797:Enzyme-Linked Immunosorbent Assay; D005109:Extracellular Matrix / Q000737:chemistry; D016222:Fibroblast Growth Factor 2 / Q000378:metabolism; D050958:HSP47 Heat-Shock Proteins; D006360:Heat-Shock Proteins / Q000096:biosynthesis*; D006801:Humans; D017874:Immediate-Early Proteins / Q000378:metabolism; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism; D020780:Matrix Metalloproteinase 9 / Q000096:biosynthesis; D019001:Nitric Oxide Synthase / Q000096:biosynthesis; D052247:Nitric Oxide Synthase Type II; D016376:Oligonucleotides, Antisense; D011233:Precipitin Tests; D012333:RNA, Messenger / Q000032:analysis; D011817:Rabbits; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis
chemical_list: C526317:CCN2 protein, human; D017077:Culture Media, Conditioned; D050958:HSP47 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D017874:Immediate-Early Proteins; D036341:Intercellular Signaling Peptides and Proteins; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; C494660:SERPINH1 protein, human; D014409:Tumor Necrosis Factor-alpha; D016222:Fibroblast Growth Factor 2; D055513:Connective Tissue Growth Factor; C496319:NOS2 protein, human; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; D020780:Matrix Metalloproteinase 9