Pancreatic expression of B7 co-stimulatory molecules in the non-obese diabetic mouse

Expression of the co-stimulatory molecule B7-1 (CD80) on pancreaticß cells can overcome peripheral T cell tolerance Intransgenic models of autoimmune disease. This study aimed todetermine if aberrant B7-1 or B7-2 (CD86) expression on pancreaticß cells is involved in the pathogenesis of autoimmunediabetes in non-obese diabetic (NOD) mice. Immunohistochemlcalanalysis of NOD pancreas sections revealed no evidence of B7-1or B7-2 expression on pancreatic ß cells at any stageprior to the onset of either spontaneously arising or cyclophosphamldeaccelerateddiabetes. Likewise, the NOD-derived NIT-1 ß cell linedid not express surface B7 orB7-1 mRNA either constitutivelyor following exposure to IFN- and TNF-, two cytokines knownto be present in the Insulitis lesion of NOD mice, or cAMP whichcan induce B7-1 expression on B cells. Both B7-1 and B7-2 were,however, highly expressed on the majority of islet-infiltratinginflammatory cells in NOD mice between days 7 and 12 after theadministration of cyclophosphamide which results in acceleratedß cell destruction. Likewise B7-1 and B7-2 were extensivelyexpressed on islet-infiltrating cells present at the time ofdiabetes onset in NOD SCID mice with adoptively transferreddiabetes. By immunohistochemistry and flow cytometry, it wasdetermined that the phenotype of B7⁺ cells in the pancreas ofNOD mice 9 days after cyclophosphamide included a mixture ofmacrophages and both CD4⁺ and CD8⁺ T cells. B7-2 was also expressedon islet-infiltrating cells in the spontaneously occurring diabetesof female NOD mice, but the levels of B7-1 expression were lowin comparison with the accelerated models of diabetes. RIP-IL-2transgenic mice, which have extensive islet infiltration butno autoimmune ß cell destruction, also had virtuallyno B7-1 expression and a minority of B7-2-expressing inflammatorycells. Thus, the activation of ß cell-specific T cellsin NOD mice does not appear to be a result of aberrant expressionof B7 on the ß cells. Expression of B7-1 and B7-2on islet-infiltrating cells is, however, associated with autoimmuneß cell destruction, suggesting a role for the B7-CD28interaction in this process.     

B7-H1协同刺激淋巴细胞增殖并诱导产生抑制性T细胞

目的：探讨B7-H1分子在抗原诱导的免疫反应中的作用。方法：在可溶性抗原PPD诱导淋巴细胞增殖体系中,加入中和抗体检测B7-H1的协同刺激作用。转染表达B7-H1的ECV304细胞,观测其对PPD诱导的淋巴细胞增殖、细胞因子分泌及淋巴细胞亚群比例的影响。将刺激后的淋巴细胞过继至自体或同种异体增殖体系中,检测其功能。结果：抗B7-H1中和抗体可降低PPD诱导的淋巴细胞增殖反应和细胞因子分泌;转染B7-H1的ECV304细胞可协同刺激抗原诱导的淋巴细胞增殖,促进细胞因子IL-10的表达,并改变T细胞亚群分布,增加CD4^＋T细胞亚群比例;自体或同种异体过继实验显示,B7-H1刺激后的淋巴细胞具有显著的免疫抑制功能。结论：B7-H1分子可协同刺激淋巴细胞增殖并诱导产生具有免疫抑制功能的T细胞亚群。     

Prevention and amelioration of collagen-induced arthritis by blockade of the CD28 co-stimulatory pathway: requirement for both B7-1 and B7-2

Collagen type II-induced arthritis (CIA) is an experimental model of arthritis that has been successfully used to dissect the pathogenesis of human rheumatoid arthritis and to identify potential therapeutic targets. We have used this model to evaluate the role of T cell co-stimulation in both disease development and progression. T cell co-stimulation is provided by ligation of CD28 with either B7-1 or B7-2 present on antigen-presenting cells and can be prevented by a soluble form of CTLA-4 (CTLA-4Ig) which binds with high affinity to both B7-1 and B7-2. We found that administration of CTLA-4Ig at the time of immunization prevented the development of CIA and was associated with lack of lymphocyte expansion within the draining lymph node and failure to produce anti-collagen IgG1 or IgG2a antibodies. To determine which CD28 ligand plays a more dominant role in CIA, we treated mice with monoclonal antibodies (mAb) against either B7-1 or B7-2. Neither anti-B7-1 nor anti-B7-2 had any effect on the course of CIA when given alone, but resulted in reduced incidence and clinical scores when given together. Interestingly, when treatment was delayed until after the onset of clinical disease, both CTLA-4Ig or anti-B7-1 plus anti-B7-2 mAb still ameliorated disease. Effective treatment was associated with a reduction in interferon-γ production by lymph node cells following stimulation in vitro, suggesting that Th1 responses were diminished. This study points to a critical role of CD28 co-stimulation in the development and perpetuation of CIA in DBA/1 mice. Interestingly, it demonstrates an active role for T cells in the later stages of this disease and implicates both B7-1 and B7-2-mediated co-stimulation in the pathogenesis of CIA.     

Interaction of B7RP-1 with ICOS Negatively Regulates Antigen Presentation by B Cells

Stimulation of T cells through the T cell receptor is insufficient for optimal T cell activation. A second activation signal is necessary, being usually provided by the costimulatory molecule CD28. Recently, additional costimulatory pathways have been identified, including inducible costimulator (ICOS) and its ligand B7RP-1. We have examined the role of the B7RP-1/ICOS costimulatory pathway on antigen presentation by B cells, using the I-A^k and I-E^k-positive CH27 B cell line and several different T cell lines. We found that CH27 expressed B7RP-1 and PD-L1 whereas the T cell lines expressed ICOS and PD-1. In the presence of HEL, the T cell hybridomas C10 and 3A9 released IL-2, which is indicative of antigen-specific T cell activation by the CH27 cells. Unexpectedly, blocking antibodies for B7RP-1 and ICOS enhanced the IL-2 response in both T cells. As expected, an increase in the production of IL-2 was seen when blocking antibodies for PD-1 were used. Blocking with antibodies for I-A^k, CD28, B7.1, and B7.2 lead to a decrease in IL-2 production. Additionally we tested a Th1 and a Th2 T cell clone. Blockade of B7RP-1/ICOS lead to an increased IFN- response in Th1 cells (A.E7) and an increased IL-4 response in Th2 cells (D10.G4.1). Intracellular staining also showed an increase in cytokine production when the B7RP-1/ICOS pathway was blocked. In conclusion, the B7RP-1/ICOS pathway is negatively regulating T cell activation by B cells and may play a role similar to that of the PD-L1/PD-1 pathway.     

Interleukin-10 differentially regulates B7-1 (CD80) and B7-2 (CD86) expression on human peripheral blood dendritic cells

Most of the immunosuppressive effects of interleukin-10 (IL-10) are related to functional inhibition of antigen-presenting cells (APC). Herein, we investigate the influence of recombinant (r)IL-10 on human dendritic cells (DC) purified from peripheral blood of healthy volunteers. First, we found that rIL-10 inhibited in a dose-dependent manner the proliferative responses as well as the production of IL-2 and interferon-γ (IFN-γ) in mixed lymphocyte reaction (MLR) between purified T cells and DC. This rIL-10 effect could be attributed to a direct effect on DC, as DC preincubated with rIL-10 were found to be deficient in the induction of alloreactive T cells even when anti-IL-10 neutralizing mAb was added at the time of MLR. Flow cytometric analysis indicated that rIL-10 did not modify the expression of ICAM-1 (CD54) and B7-1 (CD80), but decreased HLA-DR and B7-2 (CD86) expression at the DC surface. We conclude that the inhibitory effect of rIL-10 on primary alloreactive T cell responses involves down-regulation of class II MHC and B7-2 expression at the DC surface.     

调节性B淋巴细胞在自身免疫性疾病中的研究进展

自身免疫性疾病是指机体对自身成分产生免疫应答所致的一种疾病状态,这种自身免疫反应会影响相应器官的功能.在大多数自身免疫性疾病中,B淋巴细胞（简称B细胞）能产生自身抗体,因此一般被认为是自身免疫性疾病的”罪魁祸首”.最近大量研究发现,一类特殊的B细胞亚群即调节性B细胞（Bregs）能下调免疫反应,很好地维持免疫稳态,它们的丢失或缺乏会导致更加严重的自身免疫性疾病,且这种免疫调节功能主要通过自身分泌白细胞介素-10（IL-10）来实现.Bregs的起源、与其分化和功能相关的分子及其在自身免疫性疾病的作用将被进一步讨论.     

The role of B lymphocytes as antigen-presenting cells

B lymphocytes are regarded as professional antigen-presenting cells (APCs) despite their primary role in humoral immunity. Over the last two decades, studies designed to define the role of the B cells as APCs have generated discrepant results, showing that B cells are either unnecessary or required for T cell priming and either immunogenic or tolerogenic to T cells. The reasons for these discrepancies are not clear. Here we review mechanisms regulating B cell antigen presentation and the data derived from the major studies conducted by different groups representing each school of thought. In general it is clear that B cells process and present specific and nonspecific antigens differently. The presentation of specific antigen through the B cell antigen receptor occurs with very high efficiency and is associated with B cell activation, resulting in the activation of cognate T cells. In contrast, the presentation of nonspecific antigen by B cells is minimized and dissociated from B cell activation. As a result, B cells inactivate T cells that recognize nonspecific antigenic epitopes presented by B cells, or they induce regulatory T cell differentiation or expansion. These mechanisms serve to ensure effective production of high-affinity antigen-specific antibodies but minimize the production of nonspecific antibodies and autoantibodies.     

Nitric oxide limits the expansion of antigen-specific T cells in mice infected with the microfilariae of Brugia pahangi

Infection of BALB/c mice with the microfilariae (Mf) of the filarial nematode Brugia pahangi results in an antigen-specific proliferative defect that is induced by high levels of NO. Using carboxyfluorescein diacetate succinimydl ester and cell surface labeling, it was possible to identify a population of antigen-specific T cells from Mf-infected BALB/c mice that expressed particularly high levels of CD4 (CD4(hi)). These cells proliferated in culture only when inducible NO synthase was inhibited and accounted for almost all of the antigen-specific proliferative response under those conditions. CD4(hi) cells also expressed high levels of CD44, consistent with their status as activated T cells. A similar population of CD4(hi) cells was observed in cultures from Mf-infected gamma interferon receptor knockout (IFN-gammaR(-/-)) mice. Terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling staining revealed that the CD4(+) T cells from Mf-infected wild-type mice were preferentially susceptible to apoptosis compared to CD4(+) T cells from IFN-gammaR(-/-) mice. These studies suggest that the expansion of antigen-specific T cells in Mf-infected mice is limited by NO.     

Kinetics of chemokine secretion in human macrophages infected with various strains of Mycobacterium tuberculosis

Background and Purpose: It has been shown that chemokine secretion upon infection with Mycobacterium tuberculosis is influenced by the virulence of the strain, and it is suggested that virulence-associated differences in chemokine secretion contribute to the failure in containing the infection due to poor granuloma formation. Materials and Methods: In this study, we used prevalent M tuberculosis clinical strains (S7 and S10) to study the chemokine secretion profile in infected THP-1 cells and monocyte-derived macrophages (MDM) and compared this with the chemokine secretion induced by laboratory strains. Results: This study showed that comparatively lower levels of IP-10 were induced by clinical strains than by laboratory strains in both differentiated THP-1 and MDMs. The secretion of MIP-1α was also depressed but only in the THP-1 cells infected with clinical strains. This depressed chemokine secretion may hinder the movement of Th-1 cells from the periphery into the infection foci to control the infection. Correlation between IP-10 and IL-12p40 showed a negative relationship in control MDMs, while there was a positive correlation in all the infected strains, indicating their cooperative role in attracting and activating Th1 cells for a protective immune response at the site. This relationship was strain dependent, with avirulent H37Ra showing higher correlation, followed by the clinical strains and the virulent H37Rv. A positive correlation of IP-10 with IFN-γ (S7 and H37Ra) and with IL-10 (H37Ra and H37Rv) suggested a definitive interplay of these molecules in infection. Conclusions: The chemokines secretion by infected THP-1 cells and MDMs was strain dependant and the lower induction by the clinical strains may indicate that the clinical strains maintain a quiescent nature to mislead the host immune system for their benefit.     

Reciprocal expression of co-stimulatory molecules,B7-1 and B7-2, on murine T cells following activation

The co-stimulatory B7 molecules (B7-1 and B7-2) are expressed on professional antigen-presenting cells in mice. In this study, we demonstrate that B7-1 (CD80) and B7-2 (CD86) are also expressed on murine T cells in the absence of major histocompatibility complex class II molecules. The temporal expression of these two molecules on T cells varies with the state of activation where resting T cells express B7-2 but show little or no expression of B7-1. Following activation, B7-2 expression is down-regulated and there is a concomitant increase in the expression of B7-1 on the cell surface which peaks at about 72 h. Thus these two co-stimulatory molecules are reciprocally expressed on the T cell surface. This pattern of expression of B7-1 and B7-2 on T cells suggests that these two molecules may have different roles in the generation and regulation of immune responses.     

Functional role of regulatory T cells in B cell lymphoma and related mechanisms

B cell lymphoma (BCL) has a higher degree of malignancy and complicated pathogenic mechanism. Regulatory T cells (Treg cells) are known to exert certain immune suppression functions, in addition to immune mediating effects. Recent studies have revealed the role of Treg cells in pathogenesis and progression of multiple malignant tumors. This study therefore investigated the functional role and related mechanism of Treg cells in BCL. A cohort of thirty patients who were diagnosed with BCL in our hospital between January 2013 and December 2014. Another thirty healthy individuals were recruited. Peripheral blood mononuclear cells (PBMCs) were separated and analyzed for the ratio of CD4+/CD25+ Treg cells. The mRNA expression levels of Foxp3, transforming growth factor (TGF)-β1 and interleukin (IL)-10 genes were quantified by real-time PCR, while their serum levels were determined by enzyme-linked immunosorbent assay (ELISA). Meanwhile all laboratory indexes for patients were monitored during the complete remission (CR) stage. BCL patients significantly elevated ratio of CD4+/CD25+ Treg cells, which were decreased at CR stage. mRNA levels of Foxp3, TGF-β1 and IL-10, in addition to protein levels of TGF-β1 and IL-10 were potentiated in lymphoma patients but decreased in CR patients (P<0.05 in all cases). CD4+/CD25+ Treg cells exert immune suppressing functions in BCL via regulating cytokines, thereby facilitating the pathogenesis and progression of lymphoma.     

Comparative analysis of B7-1 and B7-2 expression in Langerhans cells: differential regulation by T helper type 1 and T helper type 2 cytokines

Epidermal Langerhans cells (LC) are Ia-bearing potent antigen-presenting cells (APC) of dendritic cell lineage that play a crucial role in primary and secondary T cell-dependent immune responses. LC express several costimulatory molecules such as B7, which has been implicated as one of the important determinants of professional APC. Recently, B7 antigens have been shown to include three distinct molecules termed B7-1, B7-2, and B7-3, and the expression of B7-1 and B7-2 in LC has been already confirmed. However, little is known of the regulation of B7-1 and B7-2 expression in LC. We demonstrated that LC do not express B7-1 and B7-2 in situ; however, the expression of both molecules is rapidly induced during the first 3 days of culture, and high levels of expression are maintained at least until day 6. We show that the expression of B7-2 in LC is much higher than that of B7-1 in each experiment, and that B7-1 and B7-2 expression is reproducibly augmented by interleukin (IL)-4 in a dose-dependent manner; however, IL-2 affected expression very little. Finally, B7-1 expression is significantly and dose-dependently down-regulated by interferon (IFN)-γ or IL-10, and B7-2 expression is consistently inhibited by IL-10, but not by IFN-γ. The effects of these cytokines are active only in the induction phase (during first 3 days of culture) of B7 expression: the modulatory effects of cytokines are hardly detected in the plateau phase (days 4 to 6 of culture) of B7 expression in LC. These findings suggest that B7-1 and B7-2 expression are indeed selectively and differentially regulated by these T cell-derived cytokines, and that the cytokines may modulate the synthesis of B7 molecules rather than the degradation of already-expressed B7 molecules.     

Therapeutic targeting of gut-originating regulatory B cells in neuroinflammatory diseases

Regulatory B cells (Bregs) are immunosuppressive cells that support immunological tolerance by the production of IL-10, IL-35, and TGF-β. Bregs arise from different developmental stages in response to inflammatory stimuli. In that regard, mounting evidence points towards a direct influence of gut microbiota on mucosal B cell development, activation, and regulation in health and disease. While an increasing number of diseases are associated with alterations in gut microbiome (dysbiosis), little is known about the role of microbiota on Breg development and induction in neuroinflammatory disorders. Notably, gut-originating, IL-10- and IgA-producing regulatory plasma cells have recently been demonstrated to egress from the gut to suppress inflammation in the CNS raising fundamental questions about the triggers and functions of mucosal-originating Bregs in systemic inflammation. Advancing our understanding of Bregs in neuroinflammatory diseases could lead to novel therapeutic approaches. Here, we summarize the main aspects of Breg differentiation and functions and evidence about their involvement in neuroinflammatory diseases. Further, we highlight current data of gut-originating Bregs and their microbial interactions and discuss future microbiota-regulatory B cell-targeted therapies in immune-mediated diseases.     

IL-10-producing B cells involved in the pathogenesis of Coxsackie virus B3-induced acute viral myocarditis

Background: Interleukin-10 (IL-10)-producing B cells, a subset of regulatory B cells, play critical roles in autoimmune and infectious diseases. However, the role of IL-10-producing B cells in acute viral myocarditis (AVMC) remains unknown. Methods: BALB/c mice were intraperitoneally (i. p.) infected with coxsackievirus B3 (CVB3) to establish AVMC models (AVMC group), while control mice (control group) were treated with phosphate-buffered saline (PBS) i. p. According to the time after injection, the AVMC group mice or control group mice were randomly separated into 1 week and 2 week subgroup. Myocardial histopathological changes were observed by hematoxylin and eosin staining and the frequency of splenic IL-10-producing B cells was measured by flow cytometry. Results: Histopathologic examination of heart tissues showed that mice infected with CVB3 developed AVMC. Compared with control group, the frequency of splenic IL-10-producing B cells was increased significantly in the AVMC group, with the 1 week AVMC subgroup (3.58 ± 0.47%) higher than the 2 week AVMC subgroup (2.50 ± 0.42%) (all P < 0.05). Conclusions: IL-10-producing B cells are increased in CVB3-induced AVMC, indicating that IL-10-producing B cells may play an important role in the pathogenesis of CVB3-induced AVMC.     

The ontogeny of B cells in the thymus of normal, CD3 epsilon knockout (KO), RAG-2 KO and IL-7 transgenic mice.

The ontogeny of thymic B cells was determined by three-color flow cytometry and the presence or absence of B cell progenitors confirmed by cell culture experiments. In the thymus of young normal mice, CD117(+), B220(low) pro- and pre-B cells are present but disappear with age. B220(low), CD5(+), B-1 B cells are present in the thymus of older animals following the appearance of similar cells in the peritoneal cavity and blood. In CD3 epsilon gene-deleted mice, the phenotypic progression and number of thymic B cells remains unaltered, showing that blocking T cell development does not automatically result in an increase of thymic B lymphopoiesis. Pro-B cells in RAG-2 knockout mice are found in the fetal and neonatal blood, spleen and thymus, but with increasing age are only found in the bone marrow. B lymphopoiesis in adult IL-7 transgenic mice is dramatically altered with CD117(+) pro- and pre-B cells present in spleen, lymph node and blood. In the thymus of adult IL-7 transgenic mice, the fraction of CD117(+) thymic B cells is significantly increased. These results show that in the steady state, the phenotype of thymic B cells is critically dependent on both mouse age and the phenotype of circulating B cells.     

鼠感染Babesia microti和 Babesia rodhaini后早期 IL-12、IL-10诱生的研究

目的 研究Babesia microti 和Babesia rodhaini感染后早期IL-12和IL-10诱生情况,以探讨B. microti和 B. rodhaini不同种属感染的发病机制与免疫应答效应.方法 使用ELISA方法分别检测Babesia microti 和 Babesia rodhaini 感染鼠后的0、3、6、9、12、18、24、36、48、72、96 h血清中IL-12及IL-10的浓度.结果 B. microti感染后3 h,小鼠血液中IL-12产生达一个高峰,至感染后24 h达最高峰值,与对照组比较差异显著,同组鼠在感染的早期(96 h前)血液中IL-10水平无明显变化.而B. rodhaini感染鼠,在感染后3～72 h,血清中IL-10和IL-12水平与对照组比较均无明显差异.在感染后96 h,血清中IL-10和IL-12水平开始下降,与对照组比较差异显著.结论 B. microti 感染鼠早期诱生的细胞因子主要为IL-12,而B. rodhaini感染后早期IL-12和IL-10水平无明显变化.     

Interleukin-10 and antigen-presenting cells actively suppress Th1 cells in BALB/c mice infected with the filarial parasite Brugia pahangi

Infection with the third-stage larvae (L3) of the filarial nematode Brugia results in a Th2-biased immune response in mice and humans. Previously we have shown that the production of interleukin 4 (IL-4) is critical for down-regulating polyclonal Th1 responses in L3-infected mice. However, the in vitro neutralization of IL-4 did not fully recover the defective polyclonal Th1 responses, nor did it result in the production of any antigen (Ag)-specific Th1 cytokines, suggesting that perhaps infection with L3 does not result in priming of Th1 cells in vivo. In this study, we analyzed the role of IL-10 and Ag-presenting cells (APCs) in the spleen as additional factors controlling the Th2 bias in infected mice. Our data show that IL-10 and APCs also contribute to the suppression of mitogen-driven Th1 responses of spleen cells from infected mice. In addition, the neutralization of IL-10 or the replacement of the resident APC population from spleen cell cultures resulted in the production of Ag-specific Th1 cytokines. Irradiated spleen cells from either L3-infected or uninfected mice were able to restore Ag-specific Th1 responses in vitro. Therefore, it appears that Brugia-reactive Th1 cells are primed following infection with L3, but are actively suppressed in vivo by a mechanism that involves IL-10 and the resident APC population, but not IL-4. These results indicate that a complex interplay of cytokines and cell populations underscores the Th2-polarized response in L3-infected mice.     

TNFR2 expression is a hallmark of human memory B cells with suppressive function

Tumor Necrosis Factor Receptor 2 (TNFR2) expression is increasingly being linked to tolerogenic immune reactions and cells with suppressor function including a subset of T-regulatory cells. B-regulatory cells play an important role in control of T-cell responses and inflammation. Recently, we described TNFR2 as a marker for IL-10-producing B cells, a hallmark of this cell subset. Here, we demonstrate that proliferation of T cells is reduced in the presence of TNFR2 positive human memory B cells generated with TLR9 ligand, while TNFR2- and TNFR2+CD27- B cells display costimulatory activity. Our data further reveal that IL-10 secretion is characteristic of IgM+ naïve and memory B cells but suppressive activity is not restricted to IL-10: (i) the inhibitory effect of TNFR2+ switched memory B cells was comparable to that exerted by TNFR2+ IgM+ memory B cells although IL-10 secretion levels in the cocultures were lower; (ii) supernatants from TNFR2+ memory B cells failed to suppress T-cell proliferation. Based on our findings, we propose that formation of Breg is a specific characteristic of human memory B cells undergoing terminal differentiation. Our data further corroborate that TNFR2 represents a viable marker for identification of memory B cells with regulatory function.     

Co-transfer of B cells converts resistance into susceptibility in T cell-reconstituted, Leishmania major-resistant C.B-17 scid mice by a non-congnate mechanism

Resistance to infection of mice with Leishmania major parasitesis dependent on the production of IFN- by CD4⁺ T helper cells.C.B-17 scid mice, lacking both T and B cells, succumb very quicklyto the infection, but develop resistance if reconstituted withappropriate numbers of T cells from BALB/c mice. In this model,we studied the role of B cells with regard to their abilityto influence disease outcome and to function as antigen-presentingcells for T cells. For this purpose, we reconstituted scid mice(H-2^d) with either T cells or with T and B cells obtained from(BALB/c x BALB.B)F₁ mice (H-2^d x ^b), and infected them withL. major parasites 1 day after reconstitution. Mice reconstitutedwith T cells alone cured the disease, whereas additional B cellreconstitution led to susceptibility. Healing was associatedwith a predominant T_h1-type response. In all mice, L. mayor-specificT cell proliferation was restricted to the MHC phenotype ofthe recipient (H-2^d) but not to that of the donor (H-2^d x ^b),indicating that there was no detectable contribution of donorB cells in the priming of a T cell response. Furthermore, Bcells, when purified from infected BALB/c mice, were unableto stimulate a L. mayor-specific CD4⁺ T cell clone (L1/1) withoutaddition of exogenous antigen, in contrast to macrophages fromthe same animal. These data suggest that B cells, in vivo, donot carry L. major antigen in a form capable of activating specificCD4⁺ T cells. Therefore, B cells promote disease by means otherthan cognate interaction with CD4⁺ T cells.