Interaction of Brucella abortus lipopolysaccharide with major histocompatibility complex class II molecules in B lymphocytes.

Lipopolysaccharide (LPS), a major amphiphilic molecule located at the outer membrane of gram-negative bacteria, is a potent antigen known to induce specific humoral immune responses in infected mammals. LPS has been described as a polyclonal activator of B lymphocytes, triggering the secretion of antibodies directed against distinct sugar epitopes of the LPS chain. But, how LPS is handled by B cells remains to be fully understood. This task appears to be essential for a better knowledge of the anti-LPS humoral immune response. In this study, we examine the internalization of LPS and its interaction with antigen-presenting major histocompatibility complex (MHC) class II molecules in murine and human B-cell lines. By use of immunofluorescence, we observe that structurally different LPSs from Brucella and Shigella strains accumulate in an intracellular compartment enriched in MHC class II molecules. By use of immunoprecipitation, we illustrate that only Brucella abortus LPS associates with MHC class II molecules in a haplotype-independent manner. Taken together, these results raise the possibility that B. abortus LPS may play a role in T-cell activation.     

Lipopolysaccharide augments HLA-A,B,C molecule expression but inhibits interferon-gamma-induced HLA-DR molecule expression on cultured human endothelial cells.

The effect of bacterial lipopolysaccharide (LPS) on the expression of class I and II major histocompatibility complex (MHC) molecules on the surface of cultured human umbilical vein endothelial cells (HUVEC) was determined by indirect immunofluorescent staining followed by flow cytometric analysis. LPS at concentrations higher than 0.01 micrograms/ml augmented class I MHC (HLA-A,B,C) expression on HUVEC in a concentration-dependent manner. Optimal augmentation, approximately sixfold compared with control, was seen with 10 micrograms/ml of LPS. Time-course experiments indicated that the augmentation was maximal on Day 4. In contrast, LPS had no effect on the induction of class II MHC (HLA-DR) molecules and at concentrations higher than 0.01 micrograms/ml inhibited the interferon-gamma(IFN-gamma)-induced class II MHC expression. The inhibition was about 60% at the concentration of 100 micrograms/ml of LPS. Interleukin-1 (IL-1) had a similar effect as LPS on class I and II MHC expression. However, LPS appeared to affect MHC expression directly and not through production of IL-1 or cyclo-oxygenase pathway products, since anti-IL-1 antibodies or an inhibitor of cyclo-oxygenase pathway products, indomethacin, failed to reverse the effects of LPS. These data stress the role of LPS as a direct modulatory factor of class I and II MHC expression on endothelial cells during the development of immune and inflammatory response against Gram-negative bacteria.     

Synergistic effect between CD40 and class II signals overcome the requirement for class II dimerization in superantigen-induced cytokine gene expression

Although staphylococcal enterotoxin A (SEA), B (SEB), and toxic shock syndrome toxin 1 (TSST-1) bind to major histocompatibility complex (MHC) class II molecules, they differ in their mode of binding. Signaling induced by these toxins via MHC class II molecules seems to be largely mediated by their mode of interaction. In the present study, we have demonstrated that contrary to SEA, stimulation of the human monocytic cell line THP-1 with SEB or TSST-1 failed to induce interleukin-1β or tumor necrosis factor-α gene expression. Treatment of THP-1 cells with interferon-γ increased the level of MHC class II expression but did not enhance the SEB and TSST-1 response. However, cross-linking of SEB or TSST-1 bound to MHC class II molecules with specific antibodies leads to cytokine gene expression, indicating that dimerization of class II molecules is a requirement for this superantigen-induced response. The presence of anti-CD40 antibodies in the course of SEB or TSST-1 stimulation overcomes this requirement, indicating that certain signal(s) induced via CD40 molecules can replace those induced by dimerization of class II molecules. Pretreatment with anti-lymphocyte functional antigen-1 (LFA-1) antibodies completely inhibited SEA-induced response as well as that induced by SEB or TSST-1 in the presence of CD40 antibodies, supporting the involvement of LFA-1 intercellular adhesion molecule system in these responses. The entirety of these results demonstrate clearly that dimerization of class II molecules is a prerequisite for superantigen-induced T cell-independent cytokine gene expression which can be replaced by signaling via CD40 in an LFA-1-dependent system.     

Inhibition of cell surface MHC class II expression by Salmonella

Peptide presentation by MHC molecules is an essential component of the adaptive immune response. To persist in a host, many pathogens have evolved strategies that interfere with MHC antigen-presentation. We show that in human cells harboring intracellular Salmonella, MHC class II cell surface expression was substantially reduced. The effect was specific for MHC class II as expression of additional surface receptors remained unchanged. We investigated the underlying mechanism and showed that class II biosynthesis and peptide loading were unaffected by the presence of Salmonella; however, infection led to an intracellular accumulation of mature molecules. The intracellular class II colocalized with lysosome-associated membrane protein-1 and HLA-DM but not with the Salmonella-containing vacuole. Using Salmonella mutants defective in different components and effectors of the Salmonella pathogenicity island-2 type-III secretion system, we traced the effect on class II to the sifA locus. SifA has been shown to be involved in recruiting membrane for the Salmonella-containing vacuoles. Our data suggest an additional role for SifA in interfering with MHC class II antigen-presentation.     

Phenotype of Resting and Activated Monocyte-Derived Dendritic Cells Grown from Peripheral Blood of Patients with Ankylosing Spondylitis

Decreased levels of class II major histocompatibility complex (MHC) expression and impaired formation of immunological synapse by dendritic cells (DCs) of HLA-B27 transgenic rats have been recently demonstrated. The resulting dysfunction of DCs may be implicated in the pathogenesis of the HLA-B27-related disease in transgenic animals. The phenotype of DCs in patients with ankylosing spondylitis (AS) has not been evaluated. Monocyte-derived DCs (MDDCs) were grown from patients with active AS and age-matched healthy volunteers. Surface expression of HLA-DR, co-stimulation molecules CD80, CD86 and CD40, as well as CD83 was assessed by flow cytometry and compared between the groups under 3 conditions: in resting state, after stimulation by lipopolysaccharide (LPS) and after stimulation by LPS in the presence of etanercept, a soluble receptor of tumor necrosis factor α. Lower baseline expression of class II MHC molecules (HLA-DR) was observed by MDDCs grown from AS patients, as compared to healthy subjects. Post-stimulated levels of HLA-DR were comparable in both groups, suggesting greater up-regulation of class II MHC molecules by MDDCs from AS in response to LPS. No difference between groups in the levels of expression of co-stimulation molecules and CD83 was observed. Lower basic expression of class II MHC by the MDDCs grown from patients with AS may be associated with impaired regulation of their activity. Functional studies on DCs from patients with AS are needed to evaluate the integrity of their antigen-presenting function.     

Immune complex-FcgammaR interaction modulates monocyte/macrophage molecules involved in inflammation and immune response

The interaction between receptors for the Fc portion of IgG (FcgammaRs) from monocytes/macrophages and immune complexes (IC) triggers regulatory and effector functions. Recently, we have demonstrated that IC exert a drastic inhibition of basal and IFN-gamma-induced expression of MHC class II on human monocytes. Taking into account that the regulation of MHC class II molecules is a crucial event in the immune response, in this report we extend our previous studies analysing the effect of STAT-1 phosphorylation in the down-regulatory process, the fate of the intracellular pool of MHC class II molecules and the effect of complement on MHC class II down-regulation induced by IC. We also studied the effect of IC on the expression of MHC class II (I-A(d)) in macrophages using a mouse model of chronic inflammation. We demonstrate that IC induce a depletion not only on surface expressed but also on intracellular MHC class II content and that IC-induced down-regulation of MHC class II is not mediated by the inhibition of STAT-1 phosphorylation. On the other hand, the effect of IC is not specific for the down-regulation of MHC class II, for it could be restricted to other molecules involved in inflammatory processes. Our experiments also show that the activation of the complement system could be a crucial step on the regulation of the effect of IC on MHC class II expression. In agreement with our in vitro experiments using human monocytes, IC treatment reduces the expression of MHC class II in a mouse model of chronic inflammation.     

Human CD1a molecule expressed on monocytes plays an accessory role in the superantigen-induced activation of T lymphocytes

The CD1 molecules exhibit characteristics of the MHC class I and class II molecules. They are expressed on cortical thymocytes and, similarly to MHC class II molecules, on antigen-presenting cells. In the present study, we investigated the role of the CD1 molecules in the T-cell response to bacterial superantigens. Indeed, we have observed that CD1 molecules could be detected on the CD14-positive population of some healthy donors (14% of donors tested). The CD1 expression on monocytes is correlated with an activation state of the donors as demonstrated by the increased expression of the CD25, CD38, CD45R0, and MHC class II molecules on their lymphocytes. On these donors, CD1a mAbs induced a clear inhibition (65%) of lymphocyte proliferation induced by either staphylococcal enterotoxin A or toxic shock syndrome toxin-1, whereas this proliferation was constantly unaffected by the addition of mAbs directed against CD1b or CD1c. Moreover, an intracellular calcium flux was induced in monocytes following CD1a engagement, and this calcium flux was partially inhibited by preincubation of these cells with the superantigen. These results attribute to the CD1a molecule expressed by monocytes a role in the transduction of signal(s) involved in superantigen-induced activation.     

Dendritic cell function in cytomegalovirus-infected patients with mononucleosis

Dendritic cells (DCs) are important target cells for human cytomegalovirus (HCMV) infection, and the virus has been shown to hamper the differentiation and maturation pathways of these cells in vitro. In the present study, we examined the function of monocyte-derived DCs obtained from immunocompetent individuals undergoing symptomatic HCMV infection in terms of immunophenotypic characteristics, pinocytosis, lymphocyte stimulation capacity, and cyto-chemokine secretion in comparison with DCs obtained from healthy controls. Immature and lipopolysaccharide (LPS)-stimulated DCs obtained from patients actively infected with HCMV expressed significantly lower levels of major histocompatibility complex (MHC) class II molecules. The inhibition of expression of MHC class II molecules by HCMV appeared to be functionally relevant, as mature DCs obtained from patients with HCMV mononucleosis were inefficient in stimulating proliferation of allogenic lymphocytes. Finally, the pattern of cyto-chemokines secreted by DCs obtained from patients with HCMV mononucleosis was characterized by a proinflammatory profile with an increased production of interleukin (IL)-1beta, tumor necrosis factor alpha, CC chemokine ligand 2 (CCL2) and CCL3, and reduced secretion of IL-10 upon LPS stimulation. During symptomatic HCMV infection in the immunocompetent host, DCs exhibit an impaired immunophenotype and function. These effects may contribute to the viral-induced immunomodulation, which is often observed in HCMV-infected patients.     

MHC class II molecules in tumour immunology: prognostic marker and target for immune modulation

MHC class II molecules presenting MHC class II restricted antigens play an important role in the activation of CD4+ T cells, which are the central orchestrating cells of an immune response. This review focuses on the particular role of MHC class II molecules in tumour immunology. The MHC class II antigen presentation pathway and the expression of MHC class II molecules on tumour cells related to clinical outcome is discussed. Improving the MHC class II tumour antigen presentation pathway, for instance by downregulation of the invariant chain or modulation of HLA-DO expression, offers many opportunities for developing new modalities of immunotherapy.     

Forcing Tumor Cells to Present Their Own Tumor Antigens to the Immune System： a Necessary Design for an Efficient Tumor Immunotherapy

The general principle for tumor cells to escape from immune surveillance is to prevent tumor antigens from being recognized by the immune system. Many methods have been developed to increase the immunogenecity of the tumor cells. The most efficient methods are able to force tumor cells to present their own tumor antigens to the immune system. Stimulating Th cells by converting tumor cells into MHC class II /Ii- antigen presenting cells is one of the most efficient technologies. Using antisense methods, we suppress the expression of the Ii protein that normally co-expresses with MHC class II molecules and blocks the antigenic peptide binding site of MHC class II molecules during synthesis in the endoplasmic reticulum. In such tumor cells, the“unprotected“ MHC class II molecules pick up endogenous tumor antigenic peptides, which have been transported into the ER for binding to MHC class I molecules. Simultaneous presentation of tumor antigens by both MHC class I and II molecules generates a robust and long-lasting anti-tumor immune response. MHC class II /Ii- tumor cells are potent tumor cell vaccines and also cure a significant number of animals with renal and prostate tumors. We have developed analogous human gene vectors that are suitable for most patients and cancers.     

MHC class II antigen expression is not induced on murine epidermal keratinocytes by interferon-gamma alone or in combination with tumour necrosis factor-alpha

Epidermal keratinocytes are induced to express MHC class II molecules in a variety of disease states associated with immune activity. To investigate the mechanism of this process we have exposed murine and rat keratinocytes to a variety of lymphokines and monitored changes in their MHC molecule expression. Murine cultured keratinocytes were treated with recombinant interferon-gamma (IFN-gamma), and MHC antigen expression quantified by flow cytometry. IFN pretreatment resulted in the up-regulation of class I molecule expression, but no class II expression was detected. In addition, cultured murine keratinocytes exposed to a combination of recombinant tumour necrosis factor (TNF)-alpha and IFN-gamma, or crude lymphocyte supernatants, failed to show positive membrane staining for class II molecules. However, rat keratinocytes cultured under conditions identical to murine cells were induced to express class II molecules after IFN-gamma pretreatment. The inability of IFN to induce class II expression on murine keratinocytes appears not to result from cell culture, as subcutaneous injection of IFN fails to induce epidermal class II antigen expression. However, class II expression can be induced on rat epidermis in vivo. Thus, the response of epidermal keratinocytes to IFN-gamma appears to show species variation.     

Tumor Growth Alters Macrophage Responsiveness to Macrophage Colony-Stimulating Factor During Reactivity Against Allogeneic and Syngeneic Mhc Class II Molecules

Tumor-induced changes in macrophage (M?)² accessory activities significantly suppress T-cell recognition of allogeneic and syngeneic major histocompatibility complex (MHC) class II molecules. Because these changes are often associated with altered responses to stimulatory and inhibitory cytokines, we investigated the possibility that tumor growth alters the contribution of a macrophage regulatory cytokine, macrophage colony-stimulating factor (M-CSF), during reactivity against allogeneic and syngeneic MHC class II molecules. T-cell reactivity against allogeneic MHC class II molecules was significantly suppressed by tumor-bearing host (TBH) M? in the presence of M-CSF. M-CSF-induced suppression was independent of TBH M? prostaglandin E₂ (PGE₂) synthesis. T-cell reactivity against syngeneic MHC class II molecules increased in the presence of M-CSF when normal host (NH) M? served as the source of syngeneic molecules. However, T-cell reactivity against syngeneic MHC class II molecules in the presence of M-CSF did not change when TBH M? served as stimulator/accessory cells. Although T-cell reactivity against NH syngeneic MHC class II molecules was additively increased by M-CSF and indomethacin (a PGE₂ synthesis inhibitor) treatment, reactivity against TBH syngeneic MHC class II molecules increased solely through PGE₂ synthesis inhibition. Admixtures of both NH and TBH M? in the absence or presence of M-CSF suggest that tumor-induced suppression was not strictly due to decreased expression of MHC class II molecules. Collectively, these data suggest that TBH M? are partly suppressive through altered responsiveness to M-CSF.