Peter E. Lipsky

Regulation of Human B Lymphocyte Activation, Proliferation, and Differentiation

Publisher Summary This chapter discusses several specific aspects of human B cell activation. The sequence of events and the signals involved in initial B cell activation and the relationship to subsequent proliferation and the generation of Ig-secreting cells have been delineated. The functional and phenotypic heterogeneity found among B cells as well as activation requirements of specific B cell subpopulations have been examined. Finally, the roles of various cytokines including B cell growth factor (BCGF), interleukin 2 (IL-2), interferon-gamma (IFN-γ), and interleukin 1 (IL-1) in B cell responses and their temporal relationship to other B cell activation events are discussed. The nature of the signals transmitted during initial activation determined the capacity of the activated cells to respond subsequently with ongoing proliferation and differentiation. Anatomic site of origin, surface isotype expression, and activation status all play a role in determining the nature of the activation requirements and response potential of B cell subsets. Specific cytokines are presented to provide signals both during and after activation that governed the nature and magnitude of the resultant response. Regulation of human B cell responsiveness is an exquisitely controlled and remarkably complex process.

Immune responses in patients with brain tumors: Factors such as anti-convulsants that may contribute to impaired cell-mediated immunity

The responsiveness of lymphocytes obtained from patients with brain tumors to in vitro stimulation with mitogenic lectins was examined. The previously reported finding of decreased responsiveness was confirmed. To investigate the factors responsible for the hyporesponsiveness, mitogen (phytohemagglutin and pokeweed mitogen) induced lymphocyte activation was evaluated using lymphocytes from 22 patients with brain tumors and 22 normal individuals. Lymphocytes from 13 patients with brain tumors, showed depressed responsiveness when cultured in autologous serum; in eight this was marked and in five moderate. Normal, rather than autologous, serum corrected lymphocyte function from only one of the markedly hyporesponsive patients, suggesting the existence of an intrinsic lymphocyte abnormality in some patients with brain tumors. However, serum from the hyporesponsive patients depressed mitogen‐induced activation of lymphocytes from both tumor patients and normals. The presence of suppressive serum factors could not be related to the nature of the tumor (benign versus malignant, site, cell type or degree of anaplasia). The present studies showed that significant depression of in vitro lymphocyte responsiveness occurred with exposure to two anti‐convulsant agents (phenytoin and phenobarbital) and dexamethasone. Thus, impaired lymphocyte function in patients with brain tumors may have a complex explanation with drug (corticosteroids, anticonvulsants) induced suppression playing a significant role.

Functional heterogeneity of human antigen-presenting cells: Presentation of soluble antigen but not self-Ia by monocytes

These studies were undertaken to examine the phenotype of the antigen-presenting cells (APC) circulating in human peripheral blood. Cells adherent to glass were found to be efficient APC, restoring antigen-induced3H-thymidine incorporation to T4-positive T cells that had been rigorously depleted of contaminating APC. In order to identify the APC within the glass-adherent cells (AC), these cells were stained with a number of monocyte-specific monoclonal antibodies (Mo-Mab) including 3C10, 63D3, and 61D3, and the Mo-Mab-positive and -negative cells were separated with the fluorescence-activated cell sorter. This method of preparation yielded Mo-Mab(+) AC populations that were more than 98% positive for the relevant Mab when reanalyzed with the fluorescence-activated cell sorter. Less than 1% of the Mo-Mab(−) AC populations were positive when reanalyzed with the Mab used for the separation. However, each Mo-Mab(−) AC population was contaminated with variable numbers (4–60%) of Mo as detected by morphologic criteria, histochemical analysis for esterase activity, or staining with a different Mo-Mab. Both Mo-Mab(+) and (−) AC populations were found to be similarly effective APC, with as few as 500 cells/well supporting responses to streptokinase-streptodornase, tetanus toxoid, andCandida albicans antigen. In the absence of antigen, only 3C10(−), 63D3(−), or 61D3(−) AC consistently stimulated3H-thymidine incorporation of autologous T4 cells; large numbers (>5×103/well) of APC were necessary to induce this response. These results support the conclusion that cells identified by Mo-specific Mab are capable of functioning as APC, inducing3H-thymidine incorporation in response to exogenous antigens. However, Mo-Mab(+) AC are not unique in this activity since Mo-Mab(−) AC also appeared to be able to present antigen. These Mo-Mab(−) AC appear to contain the majority of cells inducing autologous T4-cell reactivity.

Regulation of human natural killer (NK) cell function: Induction of killing of an NK-resistant renal carcinoma cell line

Natural killer (NK)-like activity against a renal carcinoma cell line, Cur, was assessed. There was no spontaneous killing of Cur cells by human peripheral blood mononuclear cells in 4-hr assays. Cur killing was observed in 18-hr assays, but the magnitude of killing was variable and always markedly less than that against K562. Cur killing was mediated by a nonadherent, nonphagocytic lymphocyte, the activity of which could be modulated both positively and negatively by monocytes or their products. Preincubation of effectors with monocyte supernatant, interleukin 1 (IL-1), α-interferon (αIFN), or interleukin 2 (IL-2) greatly increased the magnitude of Cur killing and accelerated the kinetics of lysis. The addition of prostaglandin E2 (PGE2) duringin vitro activation of NK by IL-2 profoundly inhibited subsequent Cur lysis, whereas only minimal inhibition of K562 lysis was noted. However, following activation with IL-2, lysis of Cur targets was less sensitive to the inhibitory effects of PGE2. Removal of Leu 11b(+), OKM1(+), orl-leucylleucine methyl ester-sensitive cells markedly decreased both Cur and K562 lysis. Moreover, CD16(+) cells purified with the fluorescence-activated cell sorter were found to mediate Cur killing. Whereas Cur and K562 lysis is mediated by phenotypically similar effector cells, the present studies demonstrate that the cytotoxic functions defined by the ability to lyse these two targets differ in response to a variety of immunoregulatory stimuli.

Low-density lipoprotein (LDL) and lymphocyte responses: direct suppression by native LDL and indirect inhibition from zinc chelation by contaminating EDTA

Low-density lipoproteins (LDL) have been shown to have a number of effects on the function of various cell types. To appreciate whether the in vitro effects of LDL have in vivo relevance, it is necessary to demonstrate that the biologic action described can be accounted for by native LDL and not by an alteration in the molecule or an addition to the preparation occurring during isolation. EDTA is frequently added to LDL during preparation to prevent oxidation. The effect of EDTA dialysis on LDL-mediated inhibition of lymphocyte responses was therefore examined. LDL alone did not inhibit mitogen-induced initial lymphocyte activation but rather suppressed lymphocyte DNA synthesis and subsequent proliferation in a transferrin-reversible manner. LDL dialysed with EDTA also inhibited lymphocyte responsiveness but the inhibition was not reversed by transferrin. Further experiments demonstrated that after dialysis EDTA in the LDL accounted for the change in its inhibitory effects. EDTA did not alter the lipoprotein but itself inhibited lymphocyte responses by chelating zinc necessary for DNA synthesis. These data indicate that LDL preparations may exhibit at least two separate effects on lymphocyte function. LDL is directly suppressive, while small amounts of contaminating EDTA can additionally be suppressive by chelating zinc. Thus, EDTA present in LDL preparations can alter their apparent biologic effects.