Cell-mediated immune response of synovial fluid lymphocytes to ureaplasma antigen in Reiter's syndrome

Abstract

INTRODUCTION Reiter's syndrome (RS) is an seronegative arthritis that occurs after urogenital or enteric infection which in addition with occular and/or mucocutaneous manifestations presents complete form of disease. According to previous understanding arthritis in the RS is the reactive one, which means that it is impossible to isolate its causative agent. However, there are the more and more authors suggesting that arthritis in the urogenital form of disease is caused by the infective agent in the affected joint. This suggestion is based on numerous studies on the presence of Chlmaydia trachomatis and Ureaplasma urealyticum in the inflamed joint by using new diagnostic methods in molecular biology published in the recent literature [1-3]. Besides, numerous studies of the humoral and cell-mediated immune response to "triggering" bacteria in the affected joint have supported previous suggestions [4-7]. Aim of the study was to determine whether synovial fluid T-cells specifically recognize the "triggering" bacteria presumably responsible for the Reiter's syndrome. METHOD The 3H-thymidine uptake procedure for measuring lymphocyte responses was applied to lymphocytes derived concurrently from synovial fluid (SF) and from peripheral blood (PB) [8]. Ureaplasma antigen and mitogen PHA stimulated lymphocytes in 24 RS patients (24 PB samples, 9 SF samples) and the results were compared with those found in 10 patients with rheumatoid arthritis (RA) (10 PB samples, 5 SF samples). Preparation of ureaplasma antigen. Ureaplasma was cultured on cell-free liquid medium [9]. Sample of 8 ml was heat-inactivated for 15 minutes at 601C and permanently stirred with magnetic mixer. The sample was centrifuged at 2000 x g for 40 minutes and than deposits carefully carried to other sterile glass tubes (Corex) and recentrifuged at 9000 x g for 30 minutes. The deposit was washed 3 times in sterile 0.9% NaCl, and final sediment was resuspended in 1.2 ml sterile 0.9% NaCl. Bacteriology: Chlamydia trachomatis was isolated by cell culture using cycloheximide-treated McCoy cells [10], while Ureaplasma urealyticum was identified according to its biochemical properties grown on cell-free liquid medium [9]. RESULTS Proliferative response of the PB lymphocytes to stimulation by mitogen and ureaplasma antigen did not differ between RS and RA patients. Also, there was no difference in proliferative response of SF lymphocytes to mitogen stimulation between RS and RA patients (Figure 1). However, proliferation of SF lymphocytes stimulated by ureaplasma antigen was significantly elevated in RS patients compared with the control group. This difference is statistically significant (p<0.05) (Figure 2). Difference in proliferative response of the PB and SF lymphocytes stimulated by the ureaplasma antigen was not found in RS patients. DISCUSSION It was found that SF lymphocytes of RS patients showed significantly elevated proliferative response to stimulation by the ureaplasma antigen compared with SF lymphocytes of the control group. There was no difference when the lymphocytes were stimulated by the mitogen. Our findings suggest that elevated proliferative response of lymphocytes is the sign of stimulation cell-mediated immunity to antigen present in inflamed joint. Hence, the main immune response to Ureaplasma is on the cell-mediated level in the affected joint. This confirms the earlier finding reported by Ford et all. who concluded that synovial rather than peripheral blood lymphocytes indicate the microbiological cause of arthritis [11,12]. Horowitz etal. demonstrated the correlation between clinical remission after antibiotic therapy and eradication of Ureaplasma, together with a decrease in cellular immune response synovial fluid lymphocytes to ureaplasma antigen stimulation [13]. In that study Horowitz did not find statisticaly significant difference of ureaplasma proliferative response between PB and SF lymphocytes in patients with RS. We obtained the same results. Than we concluded that sensibilization of immune system exist in the presence of foreign antigen in RS patients. The other authors demonstrated higher stimulation indices than the ones we found in our patients [11-15]. This difference may be the result of different preparation of antigens, in other words selection of serotype of Ureaplasma for antigen preparation different conditions of lymphocyte cultivation. We concluded that the presence of antigen, antigen-specific T cells and efficient antigen-presenting cells (CD4+T cells) in the joint of RS patients strongly suggests that a T-cell-mediated response to bacteria has the central role in the pathogenesis of Reiter's syndrom.

Keywords

• Reiter's syndrome
• ureaplasma antigen
• proliferative lym­phocyte response
• synovial fluid