title: Frequency of human B cells that differentiate in response to anti-CD3-activated T cells.
abstract: Activation of T cells by mAb to the CD3 molecular complex induces the differentiation of many more Ig-secreting cells (ISC) from resting human B cells in bulk cultures than do other modes of polyclonal B cell activation. In the current experiments, a limiting dilution assay was used to demonstrate that this increase in ISC generation reflects an increased frequency of responding B cells. Highly purified B cells were cultured at densities of between 1000 cells and 0.5 cell per microwell with fresh, mitomycin C-treated T cells (T mito) or T cell clones stimulated by immobilized mAb to CD3. After 5 days in culture, the number of wells containing ISC was determined, and the frequency of responding B cells was calculated. The proportion of B cells responding to anti-CD3-stimulated T cells was very large (10.7 +/- 2.8%) and greatly surpassed that induced by other polyclonal activators. B cells cultured with anti-CD3-stimulated T cell clones responded better than did those cultured with T mito. The addition of exogenous IL-2 or IL-6 to cultures supported by activated T mito enhanced the frequency of responding B cells, whereas IL-4 did not increase the generation of ISC and inhibited the augmentation of B cell responses induced by IL-2. Supplementation of cultures with mitomycin C-treated B cells as accessory cells had less of an effect. The addition of both accessory cells and IL-2 markedly increased B cell responsiveness, with precursor frequencies of 60 to 80% noted. In some experiments, cultures were carried out for 7 to 14 days and supernatants were analyzed for IgM, IgG, and IgA secretion. B cells activated by anti-CD3-stimulated T cells produced all three Ig isotypes. When the classes of Ig produced by single B cells were examined, it was observed that the stimulation of individual B cell precursors led to the production of multiple Ig isotypes, suggesting that isotype switching occurs in these cultures. These results demonstrate that under optimum culture conditions, T cells stimulated with immobilized anti-CD3 can activate the majority of human peripheral blood B cells to produce Ig and induce isotype switching by many.
authors: Harold C. Simmons Arthritis Research Center, University of Texas Southwestern Medical Center, Dallas 75235.
authors: K
authors: K
authors: Amoroso
authors: P E
authors: PE
authors: Lipsky
mesh_terms: D000328:Adult; D000921:Antibody-Producing Cells / Q000502:physiology; D000938:Antigen-Presenting Cells / Q000502:physiology; D000945:Antigens, Differentiation, T-Lymphocyte / Q000276:immunology*; D001402:B-Lymphocytes / Q000276:immunology / Q000502:physiology*; D017252:CD3 Complex; D002454:Cell Differentiation; D006801:Humans; D007132:Immunoglobulin Isotypes / Q000096:biosynthesis; D007136:Immunoglobulins / Q000096:biosynthesis; D007376:Interleukin-2 / Q000494:pharmacology; D015847:Interleukin-4 / Q000494:pharmacology; D015850:Interleukin-6 / Q000494:pharmacology; D008213:Lymphocyte Activation; D011948:Receptors, Antigen, T-Cell / Q000276:immunology*; D013601:T-Lymphocytes / Q000276:immunology*
chemical_list: D000945:Antigens, Differentiation, T-Lymphocyte; D017252:CD3 Complex; D007132:Immunoglobulin Isotypes; D007136:Immunoglobulins; D007376:Interleukin-2; D015850:Interleukin-6; D011948:Receptors, Antigen, T-Cell; D015847:Interleukin-4