title: Hypoxia-induced abrogation of contact-dependent inhibition of rheumatoid arthritis synovial fibroblast proliferation.
abstract: OBJECTIVE
  Uncontrolled proliferation of synovial fibroblasts is characteristic of the pathology of rheumatoid arthritis (RA). Since synovial tissues in the rheumatoid joints are hypoxic, we investigated how hypoxia affects RA synovial fibroblast (RASF) proliferation.
  
  
  METHODS
  RASF were cultured at 2000 cells (low density culture) or at 5000 cells (high density, growth-inhibitory confluent culture) per microtiter well under hypoxic (10%, 3%, or 1% O2) or normoxic (21% O2) conditions. Some RASF were treated with recombinant human interleukin 1 receptor antagonist (IL-1ra), anti-tumor necrosis factor-alpha (TNF-alpha)-neutralizing antibodies, anti-N-cadherin-blocking antibodies, or MG132. 3H-labeled thymidine incorporation was quantified to assess their proliferation. Total RNA and cell lysates were prepared for real-time polymerase chain reaction and Western blot analyses.
  
  
  RESULTS
  Hypoxia exerted no effect on proliferation of RASF cultured at low density. At high density, it abrogated contact-dependent growth inhibition of RASF, but not of human dermal fibroblasts. Addition of anti-TNF-alpha antibodies or IL-1ra did not affect the results. Upregulated expression of cyclin-dependent kinase inhibitor p27Kip1 was observed in the cells cultured at high density under normoxic conditions, but not under hypoxic conditions. Hypoxia decreased N-cadherin expression on RASF. Addition of anti-N-cadherin-blocking antibodies mimicked the effects of hypoxic culture; it promoted proliferation of RASF cultured at high density under normoxic conditions. This antibody treatment also downmodulated p27Kip1 expression.
  
  
  CONCLUSIONS
  Hypoxia downregulates N-cadherin expression on RASF, and thus prevents p27Kip1 upregulation for their contact inhibition. It is likely that hypoxia in rheumatoid synovial tissues contributes to rheumatoid pathology by augmenting proliferation of synovial fibroblasts.
authors.affiliation: Department of Medicine and Rheumatology, Graduate School, Tokyo Medical and Dental University, 1-5-45, Yushima, Bunkyo-ku, 113-8519, Tokyo, Japan.
authors.forename: Yoshinori
authors.initials: Y
authors.lastname: Nonomura
authors.forename: Fumitaka
authors.initials: F
authors.lastname: Mizoguchi
authors.forename: Akiko
authors.initials: A
authors.lastname: Suzuki
authors.forename: Toshihiro
authors.initials: T
authors.lastname: Nanki
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Kato
authors.forename: Nobuyuki
authors.initials: N
authors.lastname: Miyasaka
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Kohsaka
mesh_terms: D000818:Animals; D000906:Antibodies / Q000276:immunology; D001172:Arthritis, Rheumatoid / Q000276:immunology / Q000473:pathology* / Q000503:physiopathology*; D015820:Cadherins / Q000276:immunology; D049109:Cell Proliferation; D002478:Cells, Cultured; D003260:Contact Inhibition / Q000502:physiology*; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000276:immunology; D005347:Fibroblasts / Q000166:cytology / Q000502:physiology*; D006801:Humans; D000860:Hypoxia / Q000378:metabolism*; D053590:Interleukin 1 Receptor Antagonist Protein / Q000276:immunology; D014409:Tumor Necrosis Factor-alpha / Q000276:immunology
chemical_list: D000906:Antibodies; D015820:Cadherins; D053590:Interleukin 1 Receptor Antagonist Protein; D014409:Tumor Necrosis Factor-alpha; D050760:Cyclin-Dependent Kinase Inhibitor p27