Differential influences of bucillamine and methotrexate on the generation of fibroblast-like cells from bone marrow CD34+ cells of rheumatoid arthritis patients

We have recently demonstrated that bone marrow CD34+ cells from rheumatoid arthritis (RA) patients displayed abnormal capacities to respond to TNF-α and to differentiate into fibroblast-like cells producing MMP-1 (type B synoviocyte -like cells). The current study examined the effects of representative potent disease-modifying antirheumatic drugs, including bucillamine (BUC) and methotrexate (MTX) on the in vitro generation of fibroblast-like cells from RA bone marrow CD34+ cells. CD34+ cells purified from bone marrow specimens of 8 patients with active RA were cultured in the presence or absence of pharmacologically attainable concentrations of intramolecular disulfide form of bucillamine (BUC-ID, 3 μM), a major metabolite of BUC or MTX (20 nM). After incubation for 28 days, the generation of fibroblast-like cells was assessed under phase-contrast light microscopy and the concentrations of MMP-1 and VEGF in the culture supernatants were measured by ELISA. BUC-ID, but not MTX, significantly suppressed the generation of fibroblast-like cells from RA bone marrow CD34+ cells stimulated with SCF, GM-CSF and TNF-α (p = 0.024 as determined by Wilcoxon signed rank test). Accordingly, BUC-ID, but not MTX, significantly suppressed the production of MMP-1 (p = 0.017) and VEGF (p = 0.017) by RA bone marrow CD34+ cells, without inhibition of β2-microglobulin production. These results demonstrate that BUC-ID, but not MTX, is a potent inhibitor of differentiation of fibroblast-like cells from RA bone marrow CD34+ cells. Since MTX, but not BUC, has been previously shown to influence on type A synoviocytes, the data provide rationale of combination of BUC and MTX in the treatment of RA.