Effects of disease-modifying antirheumatic drugs and antiinflammatory cytokines on human osteoclastogenesis through interaction with receptor activator of nuclear factor kappaB, osteoprotegerin, and receptor activator of nuclear factor kappaB ligand.
OBJECTIVE
To demonstrate the effects of disease-modifying antirheumatic drugs and antiinflammatory cytokines on human osteoclastogenesis through their effects on receptor activator of nuclear factor kappaB (RANK), osteoprotegerin (OPG), and RANK ligand (RANKL).


METHODS
Peripheral blood mononuclear cells (PBMCs) and rheumatoid arthritis (RA) fibroblast-like synoviocytes (FLS) were cocultured in the presence of macrophage colony-stimulating factor, 1,25-dihydroxyvitamin D(3), and various concentrations of methotrexate (MTX), sulfasalazine (SSZ), hydroxychloroquine (HCQ), anti-tumor necrosis factor alpha monoclonal antibody (infliximab), interleukin-4 (IL-4), and IL-10. Osteoclast formation was assayed by counting cells after staining for tartrate-resistant acid phosphatase. RANKL expression in RA FLS and RANK expression in PBMCs were assayed by Western blotting, reverse transcription-polymerase chain reaction (RT-PCR), and real-time PCR. OPG expression was measured by enzyme-linked immunosorbent assay, RT-PCR, and real-time PCR in cultures of RA FLS.


RESULTS
MTX, SSZ, infliximab, and IL-4, but not IL-10 and HCQ, each inhibited osteoclast formation in a dose-dependent manner. We observed no evidence of synergistic inhibition of osteoclast formation by IL-4 and IL-10. High doses of infliximab suppressed the expression of RANK in PBMCs. MTX, SSZ, infliximab, and IL-4 each inhibited the expression of RANKL in RA FLS in a dose-dependent manner, and also increased the secretion of OPG in RA FLS supernatants.


CONCLUSIONS
MTX, SSZ, infliximab, and IL-4 inhibit human osteoclastogenesis by modulating the interaction of RANKL, RANK, and OPG. These results are indicative of the underlying mechanisms of the antiresorptive effects of these 4 agents.
Lee
Chang-Keun
CK
University of Ulsan College of Medicine, Asan Medical Center, Seoul, Korea.
Lee
Eun Young
EY
Chung
Son Mi
SM
Mun
Se Hwan
SH
Yoo
Bin
B
Moon
Hee-Bom
HB
D000135:Acid Phosphatase / Q000378:metabolism; D018501:Antirheumatic Agents / Q000494:pharmacology*; D001172:Arthritis, Rheumatoid / Q000473:pathology; D015153:Blotting, Western; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002470:Cell Survival / Q000187:drug effects; D018920:Coculture Techniques; D016207:Cytokines / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D006023:Glycoproteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism; D007963:Leukocytes, Mononuclear / Q000187:drug effects / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D010010:Osteoclasts / Q000187:drug effects* / Q000378:metabolism; D010012:Osteogenesis / Q000187:drug effects / Q000235:genetics; D053244:Osteoprotegerin; D053245:RANK Ligand; D012333:RNA, Messenger / Q000378:metabolism; D053246:Receptor Activator of Nuclear Factor-kappa B; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism*; D018124:Receptors, Tumor Necrosis Factor; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013583:Synovial Membrane / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D000071681:Tartrate-Resistant Acid Phosphatase
D018501:Antirheumatic Agents; D002352:Carrier Proteins; D016207:Cytokines; D006023:Glycoproteins; D007527:Isoenzymes; D008562:Membrane Glycoproteins; D053244:Osteoprotegerin; D053245:RANK Ligand; D012333:RNA, Messenger; D053246:Receptor Activator of Nuclear Factor-kappa B; D018160:Receptors, Cytoplasmic and Nuclear; D018124:Receptors, Tumor Necrosis Factor; C505686:TNFRSF11A protein, human; C506030:TNFRSF11B protein, human; C505688:TNFSF11 protein, human; D000135:Acid Phosphatase; D000071681:Tartrate-Resistant Acid Phosphatase