Thrombin stimulates cell proliferation in human fibroblast-like synoviocytes in nuclear factor-kappaB activation and protein kinase C mediated pathway.
OBJECTIVE
To examine the effect of thrombin on nuclear factor (NF)-kappaB activation and cell proliferation in synovial cells from patients with rheumatoid arthritis (RA).


METHODS
Using cultured human synovial cells from patients with RA, electrophoretic mobility shift assay, [3H]thymidine incorporation assay, and MTT assay were performed. We tested the upregulatory effects of thrombin on NF-kappaB activation and cell proliferation. The effect of thrombin on degradation of IkappaB was analyzed by Western blot.


RESULTS
Thrombin transiently induced DNA-binding activity of NF-kappaB, followed by degradation of IkappaBalpha, but not IkappaBbeta1. Moreover, synovial cell proliferation was stimulated by thrombin in a dose dependent manner. The kinetics of synovial cell proliferation induced by thrombin were almost parallel to those of NF-kappaB activation. Supershift analysis revealed that thrombin induced DNA-binding complexes were made up principally of the p65 and p50 Rel family members. Further, protein kinase C inhibitor calphostin C repressed thrombin induced NF-kappaB activation and cell proliferation in synovial cells.


CONCLUSIONS
Thrombin stimulates synovial cell proliferation involved in NF-kappaB activation, at least in part, through a protein kinase C mediated pathway, possibly indicating that thrombin plays an important role in synovial hyperplasia in RA.
Maruyama
N
N
Second Department of Internal Medicine, Asahikawa Medical College, Japan.
Hirano
F
F
Yoshikawa
N
N
Migita
K
K
Eguchi
K
K
Tanaka
H
H
D000991:Antithrombins / Q000494:pharmacology; D001172:Arthritis, Rheumatoid / Q000378:metabolism / Q000473:pathology*; D002455:Cell Division / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D004247:DNA / Q000187:drug effects / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D020672:I-kappa B Proteins; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D018179:Receptors, Thrombin / Q000235:genetics / Q000378:metabolism; D013583:Synovial Membrane / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism* / Q000473:pathology; D013917:Thrombin / Q000378:metabolism / Q000502:physiology*
D000991:Antithrombins; D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; C083487:I kappa B beta protein; D020672:I-kappa B Proteins; D016328:NF-kappa B; C000606204:NFKBIA protein, human; D012333:RNA, Messenger; D018179:Receptors, Thrombin; D000072000:NF-KappaB Inhibitor alpha; D004247:DNA; D011493:Protein Kinase C; D013917:Thrombin