David H. DeHeer

Influence of the immunoregulatory serum lipoprotein LDL-In on the in vivo proliferation and differentiation of antigen-binding and antibody-secreting lymphocytes during a primary immune response.

Abstract The proliferation and differentiation of selected antigen-specific lymphocyte subpopulations were analyzed in order to gain insight into the cellular targets and the major in vivo biological effects of the immunoregulatory serum lipoprotein, LDL-In. The influence of SRBC antigen dose on the number and relative ratio of splenic B- and T-antigen-binding lymphocytes and plaque-forming cells was characterized in control mice. When LDL-In was administered intravenously 24 hr before antigen, there was a profound reduction throughout the primary response in the number of splenic ABL and PFC, the mean lytic plaque volume, and the serum hemagglutination titer. Kinetic analyses of the cellular events suggest that a simple reduction in effective antigen dose does not account for the suppressive activity. The pattern of suppression points to inhibition of lymphocyte proliferation rather than differentiation as the probable predominant mechanism whereby LDL-In exerts its immuno-suppressive effects in vivo.

Estimation of relative antibody affinity at the level of the antibody-secreting cell during maturation of the immune response

Abstract The relative affinity of specific antibody secreted by mouse spleen cells following primary immunization with SRBC was estimated by competitive inhibition assay of antibody secreted by PFC as well as by inhibition of observed PFC number. Inhibition of direct and of indirect anti-SRBC plaque assays by the addition of specific antigen (SRBC stromata) gave sigmoid inhibition profiles from which the concentration of antigen required to inhibit 50% of the plaques (PI 50 ) was determined, Alternatively, the sum of the cube of individual plaque diameters (Σd 3 ) provided a measure of total anti-SRBC antibody secreted by PFCs from which the concentration of antigen required to inhibit 50% of the antibody (Ab 50 ) was determined. Ab 50 , rather than PI 50 : (a) was a more sensitive measure of inhibition by antigen; (b) decreased following immunization indicating a progressive increase in mean antibody affinity; and (c) correlated with the results of hemolysin transfer experiments, an independent measure of mean affinity of circulating anti-SRBC antibody. From theoretical considerations, estimation of mean antibody affinity requires quantitative analysis of fractional antibody inhibition by antigen. Determination of Ab 50 , rather than PI 50 , provides an estimate of bound and of free antibody and therefore should provide a more valid estimate of the relative antibody affinity at the cellular level. Experimentally, utilizing Ab 50 analysis, the IgM and IgG responses of C3H mice to immunization with SRBC demonstrated a progressive increase in affinity during maturation of the immune response.

Aberrant maturational characteristics of the immune responses of NZB mice to autologous and heterologous erythrocyte antigens

Abstract The maturational characteristics of the humoral immune responses of C3H and NZB mice to autologous and heterologous erythrocyte antigens were investigated. Clonal selection of antibody-secreting B lymphocytes was examined at the plaque-forming cell level of analysis of changes in mean antibody affinity and the heterogeneity of binding affinities. The primary immune response of C3H mice to SRBC exhibited a progressive temporal increase in mean relative antibody affinity and a concomitant restriction in the heterogeneity of binding affinities consistent with clonal selection and restriction of B lymphocytes to high affinity antibody-secreting cells. By contrast, the anti-SRBC immune response of NZB mice displayed aberrant maturational characteristics with a progressive decrease in mean relative antibody affinity but also clonal restriction with selection of clones of cells secreting low affinity antibodies. The spontaneous autoimmune responses of NZB mice to autologous erythrocyte surface autoantigens X and HB were different from the response to heterologous erythrocytes in that there was neither a progressive change in mean relative binding affinity nor evidence of progressive clonal restriction. Although the precise mechanisms responsible for the aberrant selection and derepression of B lymphocyte clones in NZB mice have not been identified, the very nature of the aberration suggests the existence of one or more defects which may be intrinsic to the B lymphocytes of NZB mice.

Aberrant maturation of the immune response to the thymus-independent antigen DNP-Lys-Ficoll by New Zealand black mice

Abstract The maturational features of immune responses to DNP-Lys-Ficoll, a thymus-independent antigen, were compared in immunologically abnormal NZB mice and the normal C3H strain. Competitive inhibition PFC assays, employing DNP-lysine, were used to assess relative antibody affinity and the heterogeneity of antibody affinities at the level of the antibody-secreting B lymphocyte. The primary anti-DNP response of control C3H mice exhibited a progressive temporal increase in relative antibody affinity and a concomitant restriction in the heterogeneity of binding affinities. In the contrasting anti-DNP immune response of NZB mice, mean antibody affinity progressively decreased, a change accompanied by a decrease in the heterogeneity of antibody affinities. These results are consistent with the clonal selection of cells secreting low-affinity antibody in NZB mice. Analyses of plaque diameters in uninhibited and DNP-lysine-inhibited assays demonstrated a preferential neutralization of large PFC in C3H mice, but of small PFC in NZB mice. This maturational defect of NZB mice appears to result from a relative paucity of B cells with high-affinity receptors for antigen and/or the failure of such cells to proliferate after exposure to antigen.

The influence of antigen on a model for analysis of multiple coordinate events associated with the primary immune response.

Abstract The influence of sheep red blood cells (SRBC), a T-dependent antigen, was analyzed in an experimental model of multiple coordinate functions of the immune response. The influence of antigen dose on the kinetics with which splenic B and T antigen-binding lymphocytes (ABL) and plaque-forming cells (PFC) developed, and the relative ratio of each of these cellular subsets were analyzed during the primary immune response to SRBC. As a function of increased antigen dose: 1) the total number of ABL increased, with relative increases on day 3 day 7; 2) the number of direct PFC increased, with relative antigen-dependent increases on day 3 > day 5 > day7; and 3) mean PFC lytic plaque volume progressively decreased. At low antigen doses, T ABL preferentially increased relative to B ABL, whereas at high antigen doses the reverse was observed. Thus, the increased capacity for a secondary response after a primary immunization at low antigen doses could be a consequence of the greater numbers of T ABL induced by small quantities of antigen. Both the rate of proliferation of ABL and their differentiation to PFC were influened positively by increasing doses of antigen; however, the ratio of PFC to ABL was higher at low antigen doses, suggesting that the relative rate of differentiation of ABL to PFC was greater than the rate of proliferation of ABL under conditions of limited antigen dose. Mean lytic plaque volume decreased slightly with time and markedly with increasing antigen dose. This could reflect shifts in binding affinity, mean antibody secreted per antibody-forming cell, or both. Whereas each of these parameters have been separately reported, the coordinate use of each provides a more critical analysis of each of the antigen-specific cell populations that participate in a response and their modification by immunoregulatory factors or events.

SUPPRESSION OF THE IN VIVO HUMORAL IMMUNE RESPONSE BY A SPECIES OF NORMAN HUMAN SERUM LOW DENSITY LIPOPROTEIN

Publisher Summary This chapter focuses on the suppression of the in vivo humoral immune response by a species of normal human serum low-density lipoprotein. A species of normal human serum low density lipoprotein (LDL-In) has been identified with suppressive activity for in vitro human lymphocyte mitogen and allogenic cell stimulations. In a study described in the chapter, LDL-In was isolated in a partially purified form from pooled normal human serum. The effects of in vivo administration of LDL-In also were influenced by antigen dose. Additional data also was obtained on the numbers of plaque-forming cells, the mean plaque diameters and numbers of antigen-binding cells in LDL-In suppressed mice.