CD40 ligation of rheumatoid synovial fibroblasts regulates RANKL-mediated osteoclastogenesis: evidence of NF-kappaB-dependent, CD40-mediated bone destruction in rheumatoid arthritis.
OBJECTIVE
To determine whether CD40 ligation of rheumatoid arthritis synovial fibroblasts (RASFs) is able to induce RANKL expression and osteoclastogenesis in RASFs, and to identify its mechanism of action in patients with RA.


METHODS
CD40 of RASFs was ligated with CD40 ligand (CD40L)-transfected L cells or activated T cells. The formation of osteoclasts in cocultures of CD40-ligated RASFs and T lymphocyte-depleted peripheral blood mononuclear cells was evaluated by tartrate-resistant acid phosphatase staining, detection of calcitonin receptor, and resorption pit formation assay. The expression of NF-kappaB, IkappaB alpha, ERK-1/2, phospho-ERK-1/2, p38, phospho-p38, and RANKL was examined by immunoblotting and/or semiquantitative reverse transcription-polymerase chain reaction.


RESULTS
CD40 ligation of RASFs by CD40L-transfected L cells or activated T cells induced RANKL expression and enhanced osteoclastogenesis. CD40 ligation of RASFs also induced activation of ERK-1/2, p38 MAPK, and NF-kappaB and up-regulation of CD40 ligation-induced RANKL expression, whereas osteoclastogenesis was reduced in RASFs transfected with a dominant-negative mutant of IkappaB alpha or by an NF-kappaB inhibitor. However, specific inhibitors of MAPK/ERK-1/2 and p38 MAPK partially blocked the induction of RANKL expression and osteoclastogenesis. Monoclonal antibodies against interleukin-1 and tumor necrosis factor alpha partially inhibited CD40 ligation-mediated osteoclastogenesis.


CONCLUSIONS
These results indicate that CD40 ligation of RASFs induces RANKL expression mainly via NF-kappaB activation and also results in enhanced osteoclast formation, both of which might play important roles in bone and cartilage destruction in RA. Inhibition of the CD40-CD40L interaction is a potential strategy for the prevention of bone damage in RA.
Lee
Hak-Yong
HY
Chonbuk National University Medical School and Research Institute of Clinical Medicine, Jeonju, Korea.
Jeon
Hyun-Soon
HS
Song
Eun-Kyung
EK
Han
Myung-Kwan
MK
Park
Sung-Il
SI
Lee
Sang-Il
SI
Yun
Hee-Jin
HJ
Kim
Jung-Ryul
JR
Kim
Jong-Suk
JS
Lee
Yong-Chul
YC
Kim
Sung-Il
SI
Kim
Hang-Rae
HR
Choi
Jin-Young
JY
Kang
Insoo
I
Kim
Ho-Youn
HY
Yoo
Wan-Hee
WH
D001172:Arthritis, Rheumatoid / Q000473:pathology / Q000503:physiopathology*; D019013:CD40 Antigens / Q000276:immunology / Q000502:physiology*; D002352:Carrier Proteins / Q000032:analysis / Q000502:physiology*; D002478:Cells, Cultured; D005347:Fibroblasts / Q000502:physiology*; D006801:Humans; D051550:I-kappa B Kinase / Q000032:analysis; D020672:I-kappa B Proteins / Q000032:analysis; D008562:Membrane Glycoproteins / Q000032:analysis / Q000502:physiology*; D019950:Mitogen-Activated Protein Kinase 1 / Q000032:analysis; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000032:analysis / Q000502:physiology*; D010010:Osteoclasts / Q000502:physiology*; D053245:RANK Ligand; D053246:Receptor Activator of Nuclear Factor-kappa B; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013583:Synovial Membrane / Q000166:cytology*; D048051:p38 Mitogen-Activated Protein Kinases / Q000032:analysis
D019013:CD40 Antigens; D002352:Carrier Proteins; D020672:I-kappa B Proteins; D008562:Membrane Glycoproteins; D016328:NF-kappa B; C000606204:NFKBIA protein, human; D053245:RANK Ligand; D053246:Receptor Activator of Nuclear Factor-kappa B; C505686:TNFRSF11A protein, human; C505688:TNFSF11 protein, human; D000072000:NF-KappaB Inhibitor alpha; D051550:I-kappa B Kinase; D019950:Mitogen-Activated Protein Kinase 1; D048051:p38 Mitogen-Activated Protein Kinases