title: Induction of proliferation and pro-inflammatory cytokine production in rheumatoid arthritis peripheral blood mononuclear cells by a 65 KDa chondrocyte membrane-specific, constitutive target autoantigen (CH65).
abstract: OBJECTIVE
  To analyze proliferation and pro-inflammatory cytokine production of peripheral blood mononuclear cells (PBMC) from rheumatoid arthritis (RA) patients following stimulation with a purified chondrocyte membrane-associated autoantigen (CH65).
  
  
  METHODS
  CH65 was highly purified from bovine chondrocyte membranes by solubilization and ion exchange chromatography. PBMC of RA patients (n = 37; 28 seropositive, nine seronegative) and non-arthritic donors (n = 20) were isolated by ficoll centrifugation and used in cell proliferation assays. The levels of interleukin (IL)-1, tumo necrosis factor (TNF) and IL-6 produced after stimulation with CH65 were determined by enzyme-linked immunosorbent assay (ELISA). Statistical analysis was performed using Mann-Whitney U-test and Spearman rank test and the software SPSS 13.0TM(SPSS Inc.; Chicago, IL, USA).
  
  
  RESULTS
  Peripheral blood mononuclear cells exhibited a strong proliferative response to purified CH65 in approximately 50% of the RA patients (seropositive > seronegative), with a maximum reactivity at 0.15 or 0.30 μg/mL culture medium. In contrast, PBMC from normal donors did not show a proliferative response to CH65 at any dose. The proliferative response in RA patients peaked at days 7-9 and returned to control levels at day 13, indicating an antigen-driven process. CH65-stimulated RA PBMC produced moderate to high amounts of IL-1, TNF and IL-6. This was comparable to the response after exposure to isolated whole chondrocyte membranes or purified collagen type II.
  
  
  CONCLUSIONS
  These results demonstrate a significant cellular immune response to CH65 protein in RA patients. Given the high similarity between bovine and human CH65, the results suggest a pathogenetic involvement of this molecule as a cartilage-specific potential target autoantigen in RA.
authors.affiliation: Institute of Clinical Microbiology and Hospital Hygiene, University Hospital Erlangen, University of Erlangen-Nuremberg, Erlangen, Germany.
authors.forename: Rayya J
authors.initials: RJ
authors.lastname: Alsalameh
authors.affiliation: Department of Biochemistry, Rush University Medical Center, Chicago, Illinois, USA.
authors.forename: Rachael C
authors.initials: RC
authors.lastname: Casey
authors.affiliation: Department of Biochemistry, Rush University Medical Center, Chicago, Illinois, USA.
authors.forename: Jürgen
authors.initials: J
authors.lastname: Mollenhauer
authors.affiliation: Department of Medicine III, Institute for Clinical Immunology and Rheumatology, University of Erlangen-Nuremberg, Erlangen, Germany.
authors.forename: Joachim R
authors.initials: JR
authors.lastname: Kalden
authors.affiliation: Department of Internal Medicine, Rheumatology and Clinical Immunology, Charite Centre for Internal Medicine and Dermatology, Charite-Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: Gerd R
authors.initials: GR
authors.lastname: Burmester
authors.affiliation: Department of Medicine III, Institute for Clinical Immunology and Rheumatology, University of Erlangen-Nuremberg, Erlangen, Germany.|Division of Rheumatology and Immunology, Department of Medicine, Tawam Hospital, Al Ain, Abu Dhabi, UAE.
authors.forename: Saifeddin M
authors.initials: SM
authors.lastname: Alsalameh
mesh_terms: D000328:Adult; D000368:Aged; D000818:Animals; D001172:Arthritis, Rheumatoid / Q000097:blood / Q000276:immunology*; D001324:Autoantigens / Q000276:immunology* / Q000378:metabolism; D002417:Cattle; D049109:Cell Proliferation; D002478:Cells, Cultured; D019902:Chondrocytes / Q000276:immunology* / Q000378:metabolism; D016207:Cytokines / Q000276:immunology* / Q000378:metabolism; D005260:Female; D006801:Humans; D007111:Immunity, Cellular; D018836:Inflammation Mediators / Q000276:immunology* / Q000378:metabolism; D007963:Leukocytes, Mononuclear / Q000276:immunology* / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D013997:Time Factors
chemical_list: D001324:Autoantigens; D016207:Cytokines; D018836:Inflammation Mediators
keywords: CH65; T-cells; autoimmunity to cartilage; chondrocytic autoantigen; pro-inflammatory cytokines; rheumatoid arthritis