Characterization of platelet-derived growth factor beta-receptor expressing cells in the vasculature of human rheumatoid synovium

Platelet-derived growth factor (PDGF) beta-receptor expression in normal and rheumatoid synovia was investigated by double immunofluorescence staining of frozen sections and by in situ hybridization. In the inflamed synovia, PDGF beta-receptor mRNA was present in vascular cells, as well as in discrete stromal cells. PDGF beta-receptor expressing cells in rheumatoid synovia were characterized by double immunofluorescence staining using the PDGFR-B2 monoclonal antibody at a concentration at which this antibody merely stained granular accumulations of PDGF beta-receptors. Granular accumulations of PDGF beta-receptors were articulate in blood vessel cells, but also appeared in discrete stromal cells. Thus, the overall distribution of cells having granular accumulations of PDGF beta-receptors was similar to the distribution of cells expressing PDGF beta-receptor mRNA. Double immunofluorescence stainings showed that: (a) a majority (greater than 90%) of resident macrophages did not express granular PDGF beta-receptor staining, but macrophages were often juxtaposed to PDGF beta-receptor-positive cells; (b) T lymphocytes did not express PDGF beta-receptors, but these cells were frequently found in the proximity of cells stained by PDGFR-B2; (c) in some blood vessels both HLA-DR expressing cells and PDGF beta-receptor expressing cells could be visualized, whereas in other blood vessels, cells expressing only one of these activation markers could be detected; (d) smooth muscle cells in blood vessels contained PDGF beta-receptors; and (e) capillary endothelial cells in the inflamed synovia recurrently displayed granular PDGF beta-receptor staining. The granular accumulations of PDGF beta-receptors may reflect internalization of the receptor as a result of paracrine or autocrine ligand stimulation. In support of such a possibility are the findings that elevated levels of PDGF B chain mRNA were detected by in situ hybridization in the inflamed synovia, and that cells expressing PDGF B chain mRNA were distributed similarly to cells expressing PDGF beta-receptor mRNA. Taken together, the results indicate that PDGF has a role in the inflammatory process in rheumatoid synovitis, most likely by stimulating proliferative events in the vasculature

Imatinib mesylate inhibits proliferation of rheumatoid synovial fibroblast-like cells and phosphorylation of Gab adapter proteins activated by platelet-derived growth factor

Receptors for platelet-derived growth factor (PDGF) are abundantly expressed on synovial fibroblast-like (SFL) cells from patients with rheumatoid arthritis (RA), and stimulation with PDGF enhances both the anchorage-dependent and -independent growth of RA–SFL cells. To elucidate the molecular mechanisms responsible for the excessive growth of RA–SFL cells and to seek a novel molecular-targeting therapy for RA, we examined the expression of adapter proteins and the effect of the specific inhibition of PDGF receptor activation by imatinib mesylate. Cultured SFL cells were used in the present study after 2–5 passages. The anchorage-dependent and -independent growth patterns of the SFL cells were evaluated using a tetrazolium-based assay and colony formation in 0·3% agar, respectively. Adapter proteins Gab1 and Gab2 were expressed in RA–SFL cells, and both proteins were rapidly (< 1 min) tyrosine-phosphorylated after the stimulation of RA–SFL cells with 10 ng/ml of PDGF and, to a lesser extent, after stimulation with 100 ng/ml of epidermal growth factor (EGF). The inhibition of PDGF receptor tyrosine kinase activation by 1 µM or less of imatinib mesylate specifically suppressed the PDGF-dependent, but not EGF-dependent, tyrosine phosphorylation of various proteins. Moreover, imatinib mesylate abolished both the anchorage-dependent and -independent proliferation of RA–SFL cells induced by PDGF stimulation. These results suggest that Gab adapter proteins are expressed and likely to be involved in the growth signalling of rheumatoid synovial cells and that imatinib mesylate, a key drug in the treatment of chronic myeloid leukaemia, may also be effective for the treatment of RA