title: Analysis of the antigen-specific T cell response in reactive arthritis by flow cytometry.
abstract: OBJECTIVE
  In reactive arthritis (ReA) a bacteria-specific T cell response to the triggering microbe is detected in synovial fluid (SF), and an impaired Th1 cytokine response has been described. The recent identification of immunodominant bacterial proteins/peptides and new technologies make a more detailed analysis of the immune response possible. The aim of the present study was to use these new techniques to determine the antigen-specific T cell frequency and the cytokine secretion pattern on stimulation with bacteria-derived recombinant proteins in the peripheral blood (PB) and SF from patients with ReA.
  
  
  METHODS
  In 3 patients with Chlamydia-induced ReA and 2 patients with Yersinia-induced ReA, the SF T cell response was investigated after stimulation with the Chlamydia-derived proteins major outer membrane protein (MOMP) and heat-shock protein 60 (Hsp60) and the Yersinia-derived proteins 19-kd protein and Hsp60. In 3 of these patients, the PB T cell response was investigated in parallel. T cells were stimulated in whole blood or whole SF with antigen plus anti-CD28 for 6 hours, brefeldin A was added after 2 hours, and cells were fixed and stained with antibodies against the surface markers CD4 and CD69 and against the cytokines interferon-gamma (IFNgamma), tumor necrosis factor alpha, interleukin-10 (IL-10), and IL-4. Positive cells were quantified by flow cytometry.
  
  
  RESULTS
  In the 3 patients with Chlamydia-induced ReA, the antigen-specific T cell frequency (percentage of IFNgamma CD69 double-positive CD4+ T cells) in response to MOMP (mean +/- SD 1.2 +/- 1.38%) and to Hsp60 (1.21 +/- 1.45%) in SF was about the same. In the 2 patients with Yersinia-induced ReA, the mean +/- SD frequency was 0.66 +/- 0.36% in response to the Hsp60 and 03% +/- 0.22 in response to the 19-kd protein. In the 3 patients whose PB was evaluated, the corresponding T cell response was > or =10 times lower. In 2 patients with Chlamydia-induced ReA, antigen-specific IL-10-positive CD4+ T cells were detected in 0.10-0.23% of the CD4+ T cell subpopulation.
  
  
  CONCLUSIONS
  The frequency of antigen-specific T cells to Chlamydia- and Yersinia-derived antigens in the SF of ReA patients is between 1:200 and 1:50. Both the chlamydial Hsp60 and MOMP are dominant T cell antigens in Chlamydia-induced ReA. In patients with Chlamydia-induced ReA, we detected antigen-specific IL-10 secretion, which might mediate an inhibition of effective bacterial clearance.
authors.affiliation: Benjamin Franklin University Hospital, and German Rheumatology Research Center, Berlin.
authors.forename: A
authors.initials: A
authors.lastname: Thiel
authors.forename: P
authors.initials: P
authors.lastname: Wu
authors.forename: R
authors.initials: R
authors.lastname: Lauster
authors.forename: J
authors.initials: J
authors.lastname: Braun
authors.forename: A
authors.initials: A
authors.lastname: Radbruch
authors.forename: J
authors.initials: J
authors.lastname: Sieper
mesh_terms: D000328:Adult; D000942:Antigens, Bacterial / Q000097:blood; D016918:Arthritis, Reactive / Q000097:blood* / Q000276:immunology / Q000473:pathology; D002689:Chlamydia / Q000276:immunology; D016207:Cytokines / Q000378:metabolism; D000939:Epitopes / Q000032:analysis; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D008213:Lymphocyte Activation / Q000276:immunology; D008297:Male; D008875:Middle Aged; D000091402:Prohibitins; D013582:Synovial Fluid / Q000737:chemistry; D013601:T-Lymphocytes / Q000276:immunology*; D015007:Yersinia / Q000276:immunology
chemical_list: D000942:Antigens, Bacterial; D016207:Cytokines; D000939:Epitopes; C000717331:PHB2 protein, human; D000091402:Prohibitins