engleski

SYSTEMIC AND LOCAL OSTEOCLASTOGENIC POTENTIAL OF MONONUCLEAR CELLS IN PATIENTS WITH RHEUMATOID ARTHRITIS AND SPONDYLOARTHRITIS

Background: Three major forms of chronic joint diseases are classified: osteoarthritis (OA), rheumatoid arthritis (RA) and spondyloarthritis (SpA). Those forms differ in the osteoclast activity and the intensity of cartilage and bone destruction, with RA as a prototype of "destructive" arthritis and SpA as a prototype of "remodeling" arthritis. Objectives: Although the enhanced osteoclastogenesis and osteoresorption have been confirmed in several forms of arthritis, relative contribution of systemic versus local changes in osteoclast differentiation and activity has not been fully revealed. The aim of our study was to assess the osteoclastogenic potential from peripheral-blood mononuclear cells (PBMC) and local synovial-fluid derived mononuclear cells (SFMC) in patients with RA and SpA, and to analyze the differences in osteoclastogenic responses. Methods: PBMC and SFMC were collected from RA patients (n=7), AS patients (n=2) and PA patients(n=5), after the informed consent. Osteoclast differentiation was stimulated with macrophage colony-stimulating factor (M-CSF) and receptor activator of NF-kB ligand (RANKL). Briefly, cells were seeded in a density of 0.8x106 cells per well of 96-well plate over-night. Non- adherent cells were harvested and plated on glass cover-slips in a density of 0.4x106 cells per well of 96-well plate supplemented with RANKL (60 ng/ml) and M-CSF (40 ng/ml) for the first two days of culture. Further on, cells were cultured with RANKL (40 ng/ml) and M-CSF (25 ng/ml) up to culture day 10. Osteoclasts (OCs) were detected as tartrate resistant acid phosphatase (TRAP)- positive multinucleated cells. Cultures were classified as: no osteoclastogenic potential, low l (less than 50 OC per well) and high (more than 50 OC per well). In addition, RNA was extracted from PBMC/SFMC and osteoclastogenic cultures, reversely trancribed to cDNA, and amplified by quantitative PCR for the expression of osteoclast differentiation genes: RANK (RANKL receptor) and cFms (M-CSF receptor). Results: Multinucleated TRAP-positive OCs were generated from all analyzed PBMC samples, whereas some of the SFMC samples did not contain osteoclastogenic progenitors. PBMC from RA and SpA patients had similar osteoclastogenic potential, whereas SFMC from RA patients generated higher number of OCs compared with SFMC from SpA patients. Additionally, SFMC from RA group showed greater osteoclastogenic potential than PBMC, whereas in SpA group SFMC had lower osteoclastogenic potential than PBMC. Greater osteoclastogenic potential was paralleled with the increased expression of osteoclast differentiation genes. Conclusion: Our results indicate that osteoclast progenitors, capable for differentiation into TRAP-positive multinucleated cells, are contained among both PBMC and SFMC but not with the same frequency in different forms of arthritis and even within the PBMC and SFMC of the same patient. In our further analyses, we will correlate the PBMC and SFMC osteoclastogenic potential with the BP and SF level of proresorptive osteoclastogenic factors in order to assess if the changes in their systemic and local production determine osteoclast differentiation.

arthritis; peripheral blood; synovial fluid; osteoclasts; osteoresortion

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