Pokeweed mitogen-induced <i>in vitro</i> B-cell differentiation and immunoglobulin production by peripheral blood mononuclear cells from patients with rheumatoid arthritis

Abstract

The peripheral blood mononuclear cells (MNC) were obtained by Conray-Ficoll gradient sedimentation method from 24 patients with rheumatoid arthritis (RA) and 30 healthy donors as controls. The percentage of Bl antigen positive cells in MNC was examined with mouse monoclonal antibody. And MNC were cultured with or without stimulation of Pokeweed mitogen (PWM, 10μl/ml) for 7 days. After culture, immunoglobulin (Ig) secreting cells (ISC) and the amount of Ig secreted in supernatants were simultaneously measured using reverse hemolytic plaque assay and enzyme-linked immunosorbent assay (ELISA) respectively.The percentage of B1-positive cells in MNC of RA patients was not significatnly different from controls. The numbers of both spontaneous and PWM-induced ISC in RA patients were significantly lower than controls. The amounts of IgG and IgA secreted in culture supernatants after 7 day-culture with addition of PWM in RA patients (IgG; Mean±SEM=618±108ng/me, n=20: IgA; 382±104) were not significantly different from controls (IgG; 684±93: IgA; 538±119, n=22). On the other hand, the amount of IgM secreted in RA patients (438±92) was significantly lower than controls (868±100), although the amount of total Ig secreted in RA was not different from controls. In addition, when the amount of Ig secreted in culture supernatants was divided by the number of ISC in stimulation of PWM, measured simultaneously, it became clear that the amount of Ig produced by a single ISC was significantly larger in RA patients than controls.