Activation of antigen-specific B cells: role of T cells, cytokines, and antigen in induction of growth and differentiation.

Abstract

T cells and cytokines were used to activate highly enriched populations of 2,4,6-trinitrophenyl (TNP)-binding B cells (TNP-ABC). TNP-ABC did not proliferate or differentiate when they were cultured with thymus-dependent (TD) antigen, even in the presence of supernatants known to contain B-cell growth and differentiation factors. However, purified TNP-ABC did proliferate and differentiate when they were cultured with TD antigen in the presence of carrier-primed T cells and antigen (TNP-keyhole limpet hemocyanin)-i.e., linked recognition. TNP The induction of maximal B-cell plaque-forming cell (PFC) responses to certain soluble antigens requires the participation of H-2-restricted antigen-specific T helper cells [hence the antigens are called "thymus-dependent" (TD)] and the appropriate hapten-carrier conjugates (linked recognition) (1-5). The role of cytokines in inducing or sustaining the proliferation and differentiation of B cells responding to TD antigens has not been extensively studied. In general, supernatants containing T cellderived lymphokines that act on B cells have been described in polyclonal systems (6-18) or in systems using erythrocytebound antigens (7,8).In this study we have utilized a population of purified 2,4,6-trinitrophenyl (TNP)-antigen-binding B cells (TNP-ABC) in a TD antigen-driven system to analyze the effect(s) of T cells and cytokines. The major advantage of this system, compared to the use of non-antigen-enriched B-cell populations, is that B-cell growth as well as differentiation can be assessed. In addition, because the B cells are >80% TNP specific, responding TNPspecific B-cell blasts can be visualized and isolated. As described previously (19), when T cells primed to keyhole limpet hemocyanin (KLH) are cultured with antigen (TNP-KLH) and TNP-binding B cells from unprimed mice, B-cell proliferation and differentiation occur.The data presented in this communication indicate that cytokines (with or without antigen) do not induce the differentiation or proliferation of "resting" TNP-ABC. However, when the TNP-ABC are first activated by means of linked recognition, the resultant blasts both proliferate and differentiate in response to cytokines; when activation occurs by means of nonlinked recognition, the blasts proliferate but do not differentiate in response to the same cytokines. Preparation of Cells. The methods used for the isolation of TNP-specific B cells have been described (20). T cells were obtained from the popliteal lymph nodes of mice immunized 5-8 days previously in the hind footpads with 50-75 Ag of KLH (Calbiochem-Behring). Between 90% and 95% of the B cells were removed by adherence to Petri dishes coated with affinity-purified rabbit anti-mouse Ig (RAMIg) (21). Immediately prior to use, the T cells were irradiated (1,500 rads; 1 rad = 0.01 gray).Culture Conditions. Culture conditions have been described (19).Generation of Activated TNP-ABC (TNP-ABC*). TNP-ABC were cultured with primed T cells (3-4 X 105 cells per well in 0.2 ml) and antigen [TNP-KLH or f...

Induction of the c-myc protooncogene after antigen binding to hapten-specific B cells.

“…Such populations of cells have previously been shown to be resting B cells (>95% in G o by acridine orange analysis) (13) that are only activated to expansion and differentiation by TD antigens in the presence of carrier-specific Th cells (6,14) . TNP-ABC were divided into two groups, each consisting of two sets of cultures (2 X 106 cells/ml) ; Group I served as a control, while group 2 received 10 ttg/ ml of TNP-KLH (hapten carrier) .…”

“…In our view, the sum total of all these cellular changes, though insufficient by themselves to initiate the synthesis of ribosomal RNA and DNA, serve to prepare the cell for a major growth stimulus. In the case of the TD antigenic stimulation of B cells, this stimulus will be delivered through the direct interaction between Th cells and the antigen-binding B cells (6,14,26) and will result in the commitment of the B cell to DNA synthesis. Experimental observations from both the plateletderived growth factor stimulation of 3T3 fibroblasts (16,17,20) and anti-A, stimulation of human B cells (18) support this proposed model.…”

B-cell growth factor (B-cell growth factor I or B-cell-stimulating factor, provisional 1) is a differentiation factor for resting B cells and may not induce cell growth.

“…Hence, the results are incompatible with models in which BSFp1 receptors are only expressed after the cells have been induced by anti-,u to enlarge and enter G, (1,5 Cell Culture Conditions. Sorted B cells were cultured as described (12). In addition, small Thy-1.2-cells were cultured at 50,000 per well for 24 hr (phase 1) with either PK 7.1 SN, BSFpl, or anti-8-Sepharose.…”

Alcohol Dehydrogenase: An Autoantibody Target in Patients with Alcoholic Liver Disease

“…This notion has been challenged, however, by studies showing that antibodies directed to intracellular components, such as nuclear antigens, are capable of inflicting damage in vivo (25)(26)(27)(28). Offurther relevance in understanding the pathogenic process is the fact that anti-ADH antibodies belong to the IgG isotype, implicating T-cell 'help' in their generation (29)(30)(31)(32). This raises the possibility that ADH tolerance breakdown occurs not only at B cell level, but also T cell level, a possibility requiring future study.…”