MECHANISMS OF CLONAL ABORTION TOLEROGENESIS .2. CLONAL BEHAVIOR OF IMMATURE B-CELLS FOLLOWING EXPOSURE TO ANTI-MU CHAIN ANTIBODY

  • 1 January 1979
    • journal article
    • research article
    • Vol. 37  (1) , 203-215
Abstract
B[bone marrow-derived]-lymphocyte cloning methods were to quantify the effects of anti-.mu. chain antibody on immature and mature B cells. Nude mouse spleen lymphocytes were incubated with various concentrations of sheep anti-mouse .mu. chain antibody for 10 min- 24 h. They were then washed and plated in the agar B cell colony formation assay. Five to 6 days later, control B cells had developed into colonies with a plating efficiency of about 5%. B cells from newborn mice pretreated with anti-.mu. yielded fewer colonies. Remarkably low concentrations sufficed to inhibit subsequent mitogenesis; i.e., 3 .mu.g/ml acting for 1 h or 0.1 .mu.g/ml acting for 24 h gave > 50% inhibition. Adult B cells were about 30-fold more resistant to negative signaling. Immature cells became more profoundly inhibited as anti-.mu. treatment was prolonged. Anti-Ia or anti-H2 antibodies in the absence of complement, did not deliver a negative signal. Anti-.mu. pretreatment also reduced the capacity of immature B cells to form clones of anti-hapten antibody-forming cells in a liquid microculture system where the triggering stimulus was a T[thymus-derived]-cell independent antigen. Mature T-independent B cells were not inhibited. Populations of hapten-specific B cells prepared by the hapten-gelatin method were investigated in the agar cloning system. Pretreatment of immature cells with anti-.mu. reduced their capacity to form colonies, this subpopulation of cells behaving like unfractionated B cells. Hapten-HGG [human .gamma. globulin] also delivered a negative signal. Mature hapten-specific cells or unfractionated immature spleen cells formed normal numbers of colonies following hapten-HGG treatment. Anti-.mu. antibody and hapten-HGG apparently deliver strong negative signals to immature but not mature cells with appropriate receptors. The value of anti-.mu. as a model, universal tolerogen was supported. Fluorescence-activated cell sorter (FACS) analysis was performed to study the relationships between functional inhibition and Ig [immunoglobulin] receptor modulation. The IgM receptors of immature B cells are more readily modulated by anti-.mu. antibody than those of mature cells. The receptor regeneration could be partially inhibited amongst immature but not mature B cells. There was not a close quantitative relationship between the degree of modulation and the degree of functional inhibition. Irreversible receptor modulation as such was probably not the cause of functional inhibiton.