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AB0206 Investigation of the inducibility of retroviruses from rheumatoid arthritis synovial fibroblasts (ra-sf) and ra-synovial fluid cells (ra-sfc)
  1. CA Seemayer1,
  2. SA Kolb2,
  3. J Böni3,
  4. M Neidhart1,
  5. B Simmen4,
  6. RE Gay1,
  7. BA Michel1,
  8. J Schüpbach3,
  9. S Gay1
  1. 1Center Exp Rheum
  2. 2Electron Microsc Pathol, University Hospital
  3. 3Nat Swiss Center Retrov, University
  4. 4Orth Surg, Schulthess Clin, Zürich, Switzerland

Abstract

Objectives In many investigations virus like particles (VLP) and especially type-C retroviruses could be activated from tumour cells in culture after stimulation with DNA damaging agents. Co-culture with H9 cells, a lymphatic cell line, has been successfully used to isolate exogenous human retroviruses such as HTLV and HRV-5. The aim of this study was to induce potentially hidden retroviruses from cell cultures from RA patients and to analyse for VLP and reverse transcriptase (RT)-activity.

Methods Rheumatoid arthritis synovial fibroblasts (RA-SF) were derived from synovial biopsies and synovial fluid cells from RA synovial fluids. Normal skin fibroblasts served as controls for the RA-SF and SFC from arthritis urica as well as unclassified oligoarthritis for the RA-SFC. As positive control we used baby hamster kidney cells, murine mast cells and pig fibroblasts, which produce endogenous retroviruses. Subconfluent RA-SF and controls were incubated with azacytidin (AzaC: 2 μg/ml for 24 h), bromodeoxyuridine (BUDR: 25 μg/ml for 24 h) or irradiated with 2 or 8 Gy X-rays, respectively. Supernatants of cell cultures were taken after 24 h or 48 h and analysed for RT-activity by the product enhanced reverse transcriptase (PERT)-assay. The cells were harvested on days 2–6 without trypsinization, fixed in glutaraldehyde and prepared for electron microscopy. Furthermore, RA-SF and RA-SFC were co-cultured with H9 cells for more than 3 weeks, and the supernatant was collected every 3–4 days and analysed by the PERT-assay.

Results In the virus induction and co-culture experiments, no evidence for RT-activity could be detected. In contrast, the positive controls including supernatants of cell cultures from baby hamster kidney cells, murine mast cells and pig fibroblasts revealed a strong positive RT-activity. The electron microscopical investigation of the AzaC, BUDR treated or irradiated cells did not reveal VLP.

Conclusion The current investigation does not support the involvement of infectious retroviruses in RA, but cannot exclude a role of exogenous and/or endogenous retroviruses, or the involvement of endogenous retroviral elements in the pathogenesis of RA.

C. A. Seemayer supported by the DAAD and by the Theodor und Ida Herzog-Egli Stiftung, all others by their institutions.

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