The requirement for surface Ig signaling as a prerequisite for T cell:B cell interactions. A possible role for desialylation

Models for T cell:B cell collaboration suggest that activated B cells process and present Ag to Th cells which subsequently induce B cell proliferation and differentiation. In contrast to activated B cells, resting B cells have generally been shown to be less efficient APC. If this model of T:B collaboration is physiologically correct, then resting B cells must undergo a phenotypic change that permits effective interaction with T cells. In this report, the requirement for rapid signaling through surface Ig on resting B cells for the induction of T:B interaction was investigated with an in vitro clustering assay. Resting splenic B cells were unable to form specific conjugates with T cell clones, unless the B cells were first treated with neuraminidase to remove sialic acid. In contrast, LPS-activated B cells were able to form conjugates without prior treatment. The ability of antibody against LFA-1 or L3T4 to inhibit cluster formation depended on the state of B cell activation in that anti-LFA-1 and anti-L3T4 mAb inhibited cluster formation by neuraminidase-treated resting B cells, but not by LPS-activated B cells. In addition, Ag-specific B cells which were isolated by their capacity to bind specific Ag were able to form clusters without any additional treatment. Moreover, treatment of resting splenic B cells with anti-mu-antibody induced clustering potential in B cells in as little as 10 min, suggesting that signaling through surface Ig was sufficient to induce this phenotypic change in B cells. Furthermore, activation of protein kinase C and Ca2+ mobilization were shown to be involved in that PMA and ionomycin treatment were also able to induce clustering potential in resting B cells. The rapid induction of clustering potential in resting B cells after signaling through surface Ig may represent a fundamental change in B cell physiology which occurs after recognition of specific Ag and may be required for effective cognate recognition between resting hapten-specific B cells and carrier-specific T cells. The potential role of desialylation for the induction of T:B interaction is discussed.     

Modified MHC restriction of donor-origin T cells in humans with severe combined immunodeficiency transplanted with haploidentical bone marrow stem cells

The choice of class II MHC determinants that serve as self-recognition elements for murine CD4+ T cells is thought to be determined by the environment in which T cells mature rather than their genotype. Patients with severe combined immunodeficiency (SCID) reconstituted with T cell depleted haploidentical parental stem cells provide an excellent model for studying this phenomenon in humans. After engraftment, the T cells that develop in these infants are all of donor origin. We sought to determine whether the successful immune reconstitution observed in two such SCID chimeras involved modification of the MHC restriction of Ag recognition by the genetically donor T cells as they matured to become competent T cells in the infants' microenvironment. A tetanus toxoid (TT)-specific T cell line and TT-specific T cell clones were established from the blood of two reconstituted SCID patients and from their maternal donors. T cell responsiveness was determined by [3H]thymidine incorporation after TT presentation by EBV-transformed B cell lines (EBV-B) from various donors. The TT-specific T cell line from patient 1 proliferated when presented Ag by patient, maternal donor, and paternal APC. A CD4+ donor origin clone that proliferated when presented TT by patient and paternal EBV-B, but not by maternal donor EBV-B, was isolated from each patient. TT recognition by these clones was shown to be restricted by the HLA DR determinant shared by patient and father, but not present in the donor. Four TT-specific clones isolated from maternal donors failed to proliferate when presented TT by the appropriate paternal EBV-B. These studies demonstrate that, in these human SCID bone marrow chimeras, engrafted donor-origin stem cells maturing to competent T cells in the recipient microenvironment are capable of utilizing recipient HLA determinants as restriction elements for Ag recognition. This suggests that human, as well as murine, MHC restriction patterns for Ag recognition by CD4+ T cells are environmentally determined.     

A human CD4- T cell leukemia subclone with contact-dependent helper function.

Helper T lymphocytes provide a contact dependent signal to resting B cells that is required for optimal differentiation into Ig-secreting cells. The surface structure(s) on T cells that mediate helper function have not been identified but are known to be induced by T cell activation. A CD4- subclone of the Jurkat leukemic T cell line (D1.1) was isolated and found to be distinct from CD4+ Jurkat clones and a variety of other T and non-T cell leukemic lines in that coculture of D1.1 with resting B cells induced B cells to specifically express surface CD23 molecules, a marker of B cell activation. Furthermore, Jurkat D1.1 induced B cell proliferation and terminal B cell differentiation into IgG-secreting cells in the presence of T cell-dependent B cell mitogens. Similar to the helper effector function of activated T cells, the effects of Jurkat D1.1 were neither Ag nor MHC restricted. Paraformaldehyde fixed Jurkat D1.1 cells remained competent to activate B cells while D1.1 supernatants and diffusible factors were inactive. The effect of Jurkat D1.1 on B cell activation was distinct from that of rIL-4 and was not inhibited by antibodies to IL-4. Together these observations suggested that surface structures on D1.1 and not secreted factors, mediated contact-dependent helper effector function.     

Pre-exposure of human B cells to recombinant IL-1 enhances subsequent proliferation

The reported effects of the monocyte-derived cytokine IL-1 on human B lymphocytes are both varied and controversial. IL-1 has been reported to augment both proliferation and Ig secretion of previously activated human B cells. In the present study highly purified splenic B cells were cultured with rIL-1 before, simultaneously with, and after the addition of the polyclonal B cell mitogen, anti-Ig. rIL-1 had no significant effect on B cell proliferation when added simultaneously with or after B cell activation with anti-Ig. However, incubation of splenic B cells with rIL-1 for 24 h before stimulation with anti-Ig appeared to enhance mitogenesis. With the observation that rIL-1 exerted effects on resting B cells, the effect of rIL-1 on several events which accompany B cell activation was examined. rIL-1 failed to stimulate RNA synthesis, effect increases in cell size or intracellular Ca2+ levels, or lead to the hyperexpression of MHC class II or B cell activation Ag. These studies suggest that rIL-1 does not activate B cells but primes them to respond to subsequent activation.     

NK cell function in severe combined immunodeficiency (SCID): evidence of a common T and NK cell defect in some but not all SCID patients

The immunologic work-up of eight infants with the clinical diagnosis of severe combined immunodeficiency (SCID) was performed with special emphasis on natural killer (NK) cell function and ontogeny. Contrary to previous reports, our study shows that not all SCID patients lack NK activity; some may even express very high NK- and antibody-dependent cellular cytotoxicity (ADCC). The present group of eight SCID infants was homogeneous with respect to normal levels of the purine metabolism enzymes adenosine deaminase (ADA) and purine nucleoside phosphorylase (PNP). They all had low serum Ig levels and were defective for specific antibody formation against BSA and diphtheria toxin (DiT). None of the infants' peripheral blood mononuclear cells (PBMC) proliferated significantly upon in vitro stimulation with PHA, concanavalin A (Con A), pokeweed mitogen (PWM), and irradiated allogeneic lymphocytes. Seven of eight patients, however, responded significantly to mitogenic factors present in a lectin-free interleukin 2 (IL 2) preparation, and two exhibited a positive costimulation as well with simultaneous exposure to IL 2 + Con A. The lymphocyte marker analysis revealed high percentages of OKT10+ cells in seven of eight infants, whereas peripheral T cells (OKT3+) with suppressor/killer (OKT8+) or helper/inducer (OKT4+) phenotypes were abnormally low in all infants with one exception. The PBMC of two patients formed low to normal percentages of E rosettes but expressed no B cell markers (B-/SCID). The six other infants had high percentages of B cells (B+/SCID) but lacked E rosette-forming cells. High NK and ADCC activity was found in the two B-/SCID patients. The B+/SCID infants either totally lacked NK and ADCC function (four of six) or expressed low to normal NK activity together with some T cell markers as revealed by monoclonal antibody staining but not by E rosette formation (two of six). From the data presented, an ontogenic model is proposed that assumes the status of an independent cell lineage in between T cells and monocytes for human NK cells, or that places these cells in close proximity to early differentiation steps of the T cell lineage. In any case, NK cell function clearly constitutes an additional parameter of heterogeneity in the immunologic analysis of SCID.     

Enhancement of human B cell proliferation and differentiation by tumor necrosis factor-alpha and interleukin 1

The role of tumor necrosis factor-alpha (TNF-alpha) in human B cell responses was examined and compared with that of interleukin (IL) 1 by assessing the ability of each cytokine to support proliferation and differentiation. Recombinant TNF-alpha (rTNF-alpha) and recombinant IL-1 (rIL-1) each enhanced the generation of immunoglobulin-secreting cells (ISC) in cultures of pokeweed mitogen-stimulated B cells incubated with T cells. To examine the direct effect of rTNF-alpha and rIL-1 on the responding B cell, highly purified peripheral blood B cells were stimulated with Cowan I Staphylococcus aureus (SA). In the absence of T cell factors, proliferation was minimal and there was no generation of ISC. Recombinant IL-2 (rIL-2) supported both responses. Although rTNF-alpha alone did not support SA-stimulated generation of ISC, it did increase SA-stimulated B cell DNA synthesis by two- to eightfold. In addition, rTNF-alpha augmented B cell proliferation in rIL-2 supported SA-stimulated cultures. Moreover, rTNF-alpha enhanced the generation of ISC stimulated by rIL-2 alone or rIL-2 and SA. rIL-1 also augmented DNA synthesis and generation of ISC by B cells stimulated with SA and rIL-2. However, rTNF-alpha enhanced proliferation and ISC generation in SA + rIL-2-stimulated cultures even when they were supplemented with saturating concentrations of rIL-1. Utilizing a two-stage culture system, it was found that the major effect of rTNF-alpha was to enhance responsiveness of SA-activated B cells to rIL-2, whereas it exerted only a minimal effect during initial stimulation. These results indicate that TNF-alpha as well as IL-1 augment B cell responsiveness. Moreover, the ability of rTNF-alpha to enhance B cell responsiveness was not an indirect effect resulting from the induction of Il-1 production by contaminating monocytes, but rather resulted from the delivery of a signal by rTNF-alpha directly to the responding B cell that promoted both proliferation and differentiation after initial activation. The data therefore indicate that human B cell responsiveness can be independently regulated by the action of two separate monocyte-derived cytokines.     

Effect of gamma radiation on resting B lymphocytes. II. Functional characterization of the antigen-presentation defect

The effect of radiation on three discrete Ag-presentation functions in resting B cells was examined: 1) Ag uptake and processing, 2) expression of processed Ag in the context of functional class II molecules, and 3) provision of necessary co-stimulatory, or "second," signals. Analysis of radiation's effect on B cell presentation of intact vs fragmented Ag or its effect on presentation by Ag-pulsed B cells indicated that damage to Ag uptake and processing could not account for the bulk of the radiation-induced Ag-presentation defect. Experiments with phosphatidylinositol hydrolysis as an indirect measure of TCR occupancy suggested that irradiation caused a fairly rapid (within 1 to 2 h) decrease in the ability of the B cell APC to display a stimulatory combination of Ag and class II molecule. Ag dose-response analyses demonstrated that when presenting a fragment of the Ag pigeon cytochrome c to a T cell clone, 3000 rad-treated B cell APC were able to stimulate approximately 50% as much phosphatidylinositol turnover as unirradiated B cells. It was also found that, in contrast to their inability to initiate T cell proliferation, and similarly to chemically cross-linked splenocytes, heavily irradiated resting B cells plus Ag induced a state of Ag hyporesponsiveness in T cell clones. This effect on T cells had the same Ag- and MHC-specificity as did receptor occupancy required for proliferation, indicating that heavily irradiated resting B cells bear functional class II molecules. Co-culture of T cells with allogeneic B cells and syngeneic heavily irradiated B cells or chemically cross-linked splenic APC plus Ag resulted in T cell proliferation and interfered with the induction of the hyporesponsive state. This co-stimulatory function was radiosensitive in resting allogeneic B cells. Together, these data support the hypothesis that the major functional consequences of radiation to resting B cell APC are a reduction in the effective display of Ag plus class II molecules and, probably what is more important, a loss in the ability to provide APC-derived co-stimulatory signals.     

Inhibitory influence of IL-4 on human B cell responsiveness

The role of IL-4 in human B cell activation, proliferation, and differentiation was examined. rIL-2, but not rIL-4, was able to promote maximum proliferation and generation of Ig-secreting cells in cultures of highly purified B cells stimulated with Cowan I Staphylococcus aureus (SA). Addition of rIL-4 to rIL-2-supported cultures of SA-stimulated peripheral blood, spleen, or lymph node B cells dramatically suppressed both proliferation and differentiation. Results from experiments in which rIL-4 was added to culture at progressively later times indicated a requirement for rIL-4 to be present during the first 2 days of a 5-day incubation to cause inhibition of responsiveness. When a two-stage culture system was utilized, rIL-4 was found to support proliferation or differentiation of B cells initially activated with SA for 2 days only minimally. However, rIL-4 did not inhibit responses of SA preactivated B cells supported by IL-2. The presence of rIL-4 during the initial 48-h activation of B cells with SA and rIL-2 resulted in a profound inhibition of the ability of the activated B cells to respond subsequently to rIL-2 or lymphokine-rich T cell supernatants. A similar 48-h incubation with rIL-4 alone without SA had no effect on subsequent B cell responsiveness. The presence of rIFN-gamma during B cell activation decreased the inhibitory effect of IL-4. Other cytokines including IFN-alpha, IL-1, and commercially available low m.w. B cell growth factor also diminished the inhibitory effect of IL-4. These results indicate that IL-4 inhibits the capacity of human B cells to be activated maximally by SA and rIL-2 and therefore suggest a new immunomodulatory role for this cytokine.     

Can resting B cells present antigen to T cells?

Antigen stimulation of T lymphocytes can occur only in the presence of an antigen-presenting cell (APC). An ever-increasing number of cell types have been found to act as APCs; these include macrophages, splenic and lymph node dendritic cells, and Langerhans' cells of the skin. Although activated B lymphocytes and B cell lymphomas are known to serve as APCs, it has been generally believed that resting B cells cannot perform this function. However, in recent studies we have found that resting B cells can indeed present soluble antigen to T cell clones as well as to antigen-primed T cells. The previous difficulty in demonstrating this activity can be explained by the finding that, in contrast to macrophages and dendritic cells, the antigen-presenting ability of resting B cells is very radiosensitive. Macrophages are usually irradiated with 2000-3300 rads to prevent them from incorporating [3H]thymidine in the T cell proliferation assay. Resting B cells, however, begin to lose presenting function at 1500 rads and have completely lost this activity at 3300 rads. It was also possible to distinguish two distinct T cell clonal phenotypes when resting B cells were used as APCs on the basis of two different assays (T cell proliferation, and B cell proliferation resulting from T cell activation). The majority of T cell clones tested were capable of both proliferating themselves and inducing the proliferation of B cells. Some T cells clones, however, could not proliferate in the presence of antigen and B cell APCs, although they were very good at inducing the proliferation of B cells. This suggests that there are two distinct pathways of T cell activation, one leading to T cell proliferation and the other leading only to the release of lymphokines (as measured by the polyclonal activation of B cells).     

Hairy cell leukemia-associated antigen (HC2) is an activation antigen of several hemopoietic cell lineages, inducible on monocytes by IFN-gamma

The HC2 Ag is defined by a mAb raised against leukemic B lymphocytes from a patient with hairy cell leukemia (HCL). This 60 to 70-kDa Ag was immunoprecipitated from EBV-transformed B lymphoblastoid cell lines, from HCL-B cells, from the HUT-102 T cell line infected with HTLVI, and from activated monocytes. A binding assay with radioiodinated Fab' anti-HC2 confirmed this cellular distribution of the Ag and demonstrated 500 to 3000 binding sites on resting T cells, 300 to 11,000 binding sites on non-T cells, less than 3000 binding sites on chronic lymphocytic leukemia B cells, and 29,000 to 223,000 binding sites on HCL-B cells. PMA plus anti-CD3 up-regulated HC2 expression on T cells and IFN-gamma up-regulated expression on monocytes. On B cells, EBV transformation may result in HC2 expression, and antibody to HC2 has been found to inhibit B cell differentiation and proliferation. The combined results suggest an important role for the HC2 membrane-associated Ag on cells responsible for the immune response.     

Characterization of a T4+/Leu-8+ T cell clone that directly helps B cell Ig production by secreting B cell differentiation factor

A human T4+/Leu-8+ T cell clone (YA2) was established by phytohemagglutinin activation and interleukin 2 (IL 2) propagation. Functional characterization of this clone demonstrated that it provided potent help towards Ig production by pokeweed mitogen-stimulated B cells in the presence of small numbers of autologous T cells or by Staphylococcus aureus Cowan I (SAC)-activated B cells in the presence of B cell growth factor (BCGF). YA2 provided no help to resting B cells and minimal help to either unactivated B cells cultured with BCGF or SAC-activated B cells. Supernatant generated from clone YA2 by IL 2 stimulation had significant B cell differentiation activity but no BCGF or IL 2 activity. Thus, YA2 is a T4+/Leu-8+ potent direct helper only to B cells that are activated and proliferating due to its selective secretion of a differentiation factor, and not an activation and growth factor. The availability of phenotypically defined cloned populations of T cells with restricted functional helper activity related to the secretion of selected B cell tropic factors should prove useful in the dissection of the role of individual T cell subsets in the regulation of the human B cell cycle.     

Interleukin 2 induces proliferation of normal "resting" human T cells in the absence of other known external stimulation

Interleukin 2(IL-2) is known to stimulate the progression of activated T cells from G1 through the rest of the cell cycle. We have demonstrated that addition of purified recombinant human IL-2 (rIL-2) to fresh normal human peripheral blood mononuclear cells (PBM), which were IL-2 receptor (Tac) negative by FACS analysis, stimulated marked proliferation of the PBM. IL-2-induced proliferation was also observed with umbilical cord blood mononuclear cells. Monocyte depletion of PBM resulted in a marked reduction of rIL-2-induced proliferative response which could be restored by adding back autologous irradiated monocytes but not by interleukin 1. The T cells preincubated with rIL-2 showed a five to six times enhanced autologous mixed-lymphocyte reaction (AMLR) compared to controls. The rIL-2-induced proliferative response of PBM was inhibited in a concentration-dependent fashion by preincubation of PBM with an anti-HLA-DR framework monoclonal antibody. The proliferating cells were shown by two-color flow cytometric analysis to be primarily Leu-1+ and Leu-4+ T cells (both leu-3+ and Leu-2+ subsets); however, 6 to 19% of responding cells had surface markers for B cells or NK cells. The data demonstrate that rIL-2 can induce proliferation of "resting" human T cells. The phenomenon may be related to a monocyte-dependent AMLR which induces IL-2 receptors and IL-2 responsiveness in a subset of T cells.     

Resting B lymphocytes as APC for naive T lymphocytes: dependence on CD40 ligand/CD40

Although resting B cells as APC are tolerogenic for naive T cells in vivo, we show here that they can provide all the costimulatory signals necessary for naive T cell proliferation in vivo and in vitro. In the absence of an activating signal through the B cell Ag receptor, T cell proliferation after Ag recognition on resting B cells depends on CD40 expression on the B cells, implying that naive T cells use the membrane-bound cytokine, CD40 ligand (CD154), to induce the costimulatory signals that they need. Induction of B7-1 (CD80) and increased or sustained expression of CD44H, ICAM-1 (CD54), and B7-2 (CD86) are dependent on the interaction of CD40 ligand with CD40. Transient expression (12 h) of B7-2 is T cell- and peptide Ag-dependent, but CD40-independent. Only sustained (>/=24 h) expression of B7-2 and perhaps increased expression of ICAM-1 could be shown to be functionally important in this system. T cells cultured with CD40-deficient B cells and peptide remain about as responsive as fresh naive cells upon secondary culture with whole splenic APC. Therefore, B cells, and perhaps other APC, may be tolerogenic not because they fail to provide sufficient costimulation for T cell proliferation, but because they are deficient in some later functions necessary for a productive T cell response.     

Monoclonal antibody against the human peripheral lymph node homing receptor homologue (Leu 8) inhibits B cell differentiation but not B cell proliferation.

Previous studies have shown that a subpopulation of circulating human B cells expresses the Leu 8 peripheral lymph node homing receptor homologue and that these B cells are capable of producing Ig in response to staphylococcus A Cowan I (SAC). In the present study the effect of a signal delivered via the Leu 8 molecule (using anti-Leu 8 mAb) on B cells was examined. Initially, it was shown that immobilized anti-Leu 8 suppressed IgM and IgG secretion of B cells activated by SAC + IL-2 but not that by PWM-prestimulated B cells or B cells stimulated with PWM in the presence of CD4+, Leu 8- T cells (a source of helper cells). It was also shown that anti-Leu 8 did not suppress SAC + IL-2-stimulated B cell proliferation or expression of IL-2R alpha-chain or c-myc mRNA in B cells. The addition of T cells, monocytes, purified IL-2, rIL-1, rIL-6, or human B cell growth factor did not overcome the inhibitory effect of anti-Leu 8 on SAC-stimulated B cell Ig production, and the inhibitory effect of anti-Leu 8 was not blocked by anti-TGF-beta. Finally, inhibition of B cell differentiation occurred even when anti-Leu 8 was added up to 72 hrs after initiation of cell culture. Thus, anti-Leu 8 is unique among inhibitors of B cell function in that it can down-regulate immunoglobulin synthesis without affecting B cell proliferation. These findings suggest that a natural ligand for Leu 8 could affect not only homing of B cells, but also B cell differentiation.     

Interleukin-2 reverses T cell unresponsiveness to Plasmodium falciparum-antigen in malaria immune subjects

Peripheral blood mononuclear cells (PBMC) from a large proportion of 34 healthy adult native residents in a malaria endemic area showed null or marginal proliferative response (low-responders) to schizont-enriched Plasmodium falciparum malaria antigen (M.Ag) but good response to pokeweed mitogen. In contrast, substantial proliferative response to M.Ag was observed in 8/8 adult temporary residents with a history of one to three acute malaria episodes. Purified CD4+ T cells preferentially responded to M.Ag, however in low-responders CD4+ T cell proliferation was poor. Moreover, no inhibition of CD4+ T cell proliferation was observed when graded numbers of CD8+ T cells were added in culture. The addition of recombinant interleukin 2 (rIL-2) to M.Ag restored the proliferative response of low-responders' PBMC. This response was M.Ag-specific when CD4+ T cells grown in M.Ag plus rIL-2, but not in rIL-2 alone, were tested in secondary cultures.     

Human T cell activation. Synergy between CD73 (ecto-5'-nucleotidase) and signals delivered through CD3 and CD2 molecules

Interaction of the glycosyl phosphatidylinositol-linked differentiation Ag CD73 (ecto-5'-nucleotidase) with the CD73-specific mAb 1E9 generates agonistic signals that strongly synergize with T cell activation induced by CD3 and CD2 mAb. This synergy is observed only when 1E9 is immobilized on plastic and occurs in the absence of accessory cells or exogenous lymphokines. 1E9 induces a rapid (though transient) increase in [Ca2+]i in a minor proportion (20 to 30%) of unfractionated T lymphocytes (presumably CD73+ cells). However, this [Ca2+]i mobilization is not sufficient to fully activate CD73+ T cells, as shown by the requirement of additional signals such as CD3 or CD2 stimulation to initiate T cell proliferation. These signals cannot be substituted by the exogenous lymphokines, rIL-1, rIL-2, or rIL-4, or PMA (when T cells are rigorously depleted of monocytes). These data indicate that CD73 may behave as an accessory molecule regulating interactions between T cells and antigens or APC. A comparison was carried out with mAb 9.3 to the differentiation Ag CD28, another agonistic molecule with activating properties similar to CD73. Despite their lower percentage, the ability of CD73+ T cells to amplify the proliferation induced by CD3 or CD2 mAb was equivalent or even greater than that of CD28+ T cells. Once activated, CD73+ cells may recruit the remaining (CD73-) cells primed by CD3 or CD2 stimulation. Based on these data, we suggest that CD73+ T lymphocytes may be a specialized subset to amplify immune responses originated by the CD3 and CD2 activation pathways. Finally, the functional association between CD73 and integral membrane molecules like CD3 and CD2 suggests that GPI-anchored molecules may play a role in transmembrane signaling mediated by conventional second messenger systems.     

Failure to detect IL-3-binding sites on human mast cells

IL-3, a pleiotropic lymphokine, has been termed mast cell growth factor because it promotes growth and differentiation of murine mast cells. Murine mast cells, in turn, express cell surface receptors for IL-3. Human rIL-3 has been shown to induce proliferation and differentiation of human basophils and to activate basophils via high affinity binding sites. To investigate whether human mast cells express IL-3R, binding studies with 125I-radiolabeled human rIL-3 were performed on HMC-1, a novel human mast cell line, and on pure populations (i.e., 93 to 99% purity) of human tissue mast cells obtained with mAb and C from dispersed lung (n = 2). Unexpectedly, neither enriched human lung mast cells nor HMC-1 cells bound radiolabeled human rIL-3 specifically. Moreover, human rIL-3 failed to promote uptake of [3H]thymidine, synthesis of histamine, histamine releasability, or changes in expression of mast cell differentiation Ag (YB5B8, CD54/ICAM-1, CD9/p24, CD33/gp67) on either human lung mast cells or HMC-1 cells. It is hypothesized that the fundamental difference in the biologic response to IL-3 between human and murine mast cells is due to a loss during evolution of mast cell high affinity IL-3 binding sites.     

A monoclonal anti-mouse LFA-1 alpha antibody mimics the biological effects of B cell stimulatory factor-1 (BSF-1)

This study describes the generation of a monoclonal mouse x rat antibody (G-48) that recognizes a determinant on the serologically defined LFA-1 alpha-chain. It immunoprecipitates two noncovalently associated polypeptides of 176,000 and 95,000 Mr respectively from lysates of radioiodinated BCL1 cells, T cells, and B cells. G-48 mimics the biological actions of BSF-1 by inducing increased levels of Ia antigen expression on resting B cells, augmenting the proliferation of anti-delta-stimulated B cells, and in insolubilized form, inducing IgG1 secretion by LPS-activated B cells. G-48 does not have BCDF mu, BCGF II, nor IL 2 activity. These results demonstrate that LFA-1 plays an important role in B cell activation, proliferation, and differentiation.     

Phenotypic and functional analysis of the cellular response in regional lymphoid tissue during an acute virus infection

A phenotypic and functional analysis has been made of the cellular response in regional lymphoid tissue of C57BL/6J mice infected with lymphocytic choriomeningitis virus. Massive recruitment of nondividing cells occurred from 3 days after infection, with total numbers of CD8+ T lymphocytes, B220+ B cells, and Thy-1- B220- null cells being high from day 4 to day 6. In contrast, the peak counts for CD4+ T cells were recorded on day 4 and declined dramatically thereafter. Enhanced expression of IL-2R and Ly-24, both of which can be regarded as T cell activation markers, was found for both the CD4+ and the CD8+ subsets, being most prominent for the CD8+ T cells on day 6. Evidence of T cell proliferation was not recognized until days 5 and 6, coincident with enhanced responsiveness of the lymphocytes to rIL-2 and the development of virus-specific cytotoxic activity. Elimination of the CD4+ T cells by treatment of mice with mAb did not modify either the pathogenesis of lymphocytic choriomeningitis, or the expression of activation markers on the CD8+ T cells which are known to be the key effectors in this disease. Thus, the pattern of responsiveness for the CD8+ population is of recruitment to the lymph node, progressive increase in the expression of activation markers and enhanced sensitivity to rIL-2, with late proliferation and generation of cytotoxic activity. This model provides a system for the rigorous in vivo analysis of parameters influencing lymphocyte differentiation and activation in a virus infection.     

Characterization of the effector mechanism of help for antigen-presenting and bystander resting B cell growth mediated by Ia-restricted Th2 helper T cell lines

The mechanism of help for resting B cell growth in MHC-restricted T-B collaboration was investigated using an in vitro polyclonal model for these T cell-B cell interactions. In the presence of rabbit anti-mouse Ig, small, size-selected B cells elicit help from syngeneic Ia-restricted Th2 cell lines specific for F(ab')2 rabbit globulin. Both Ag-presenting and bystander B cells receive signals from stimulated Th cells that lead to B cell proliferation. The results suggest that the direct activation of resting Ag-presenting and bystander B cells by Th2 cells is mediated by a similar effector mechanism. Although proliferative responses by Ag-presenting B cells are of greater magnitude, help for both Ag-presenting and bystander B cell populations is characterized by the lack of a requirement for membrane Ig cross-linking, by identical kinetics, and by the necessity for direct cell contact or close proximity with Th cells. B cell proliferation is not induced by exposure to the sequence of diffusable mediators released from a synchronized Ag-specific T-B interaction. The T cell-dependent proliferation by both B cell populations can be inhibited by excess mitomycin C-treated syngeneic "cold target" B cells, demonstrating a requirement for a short-range T cell-B cell interaction. mAb inhibition experiments fail to identify a role for class II, LFA-1, or CD4 membrane molecules in the delivery of help to bystander B cells. Antibody against H2d bystander class II molecules has no effect on bystander B cell proliferation at concentrations that completely block Ag presentation by H2d B cells to an H2d-restricted Th cell line. Antibodies against the cell adhesion molecule LFA-1 or the Th cell molecule CD4 do inhibit bystander B cell proliferation, but only to the extent that they block T cell activation and the induction of help. The inductive stimulus leading to resting B cell growth results from an early, short-range interaction with Th cells. B cell proliferation is supported by T cell soluble mediators as a consequence of this interaction, which is required for at least 8 hr after T cell recognition of Ag/Ia on the surface of Ag-presenting B cells.