Alain Lamarre

IgA production without |[mu]| or |[delta]| chain expression in developing B cells

Surface, membrane-bound, immunoglobulin M (IgM) or IgD expression early in B cell ontogeny is considered essential for the differentiation of antibody-producing cells in mammals; only in IgM+ B cells is the heavy chain locus rearranged to express antibodies of other classes. We show here that IgA is selectively expressed in μMT mice, which lack IgM or IgD expression and have a pro-B cell developmental block. μMT IgA binds proteins of commensal intestinal bacteria and is weakly induced by Salmonella infection, although not through conventional immunization. This μMT IgA pathway requires extrasplenic peripheral lymphoid tissues and may be an evolutionarily primitive system in which immature B cells switch to IgA production at peripheral sites.

IgA responses in the intestinal mucosa against pathogenic and non-pathogenic microorganisms.

IgA is the most abundant immunoglobulin produced in mammals; most is secreted as a dimer across mucous membranes. This review discusses the different mechanisms of induction of IgA, and its role in protecting mucosal surfaces against pathogenic and non-pathogenic microorganisms.

Mechanisms of Neonatal Mucosal Antibody Protection

Following an abrupt transition at birth from the sterile uterus to an environment with abundant commensal and pathogenic microbes, neonatal mammals are protected by maternal Abs at mucosal surfaces. We show in mice that different Ab isotypes work in distinct ways to protect the neonatal mucosal surface. Secretory IgA acts to limit penetration of commensal intestinal bacteria through the neonatal intestinal epithelium: an apparently primitive process that does not require diversification of the primary natural Ab repertoire. In contrast, neonatal protection against the exclusively luminal parasite Heligmosomoides polygyrus required IgG from primed females. This immune IgG could either be delivered directly in milk or retrotransported via neonatal Fc receptor from the neonatal serum into the intestinal lumen to exert its protective effect.

Neutralizing antiviral antibody responses.

Publisher Summary Neutralizing antibodies are evolutionarily important effectors of immunity against viruses. Their evaluation has revealed a number of basic insights into specificity, rules of reactivity (tolerance), and memory—namely, (1) Specificity of neutralizing antibodies is defined by their capacity to distinguish between virus serotypes; (2) B cell reactivity is determined by antigen structure, concentration, and time of availability in secondary lymphoid organs; and (3) B cell memory is provided by elevated protective antibody titers in serum that are depending on antigen stimulation. These perhaps slightly overstated rules are simple, correlate with in vivo evidence as well as clinical observations, and appear to largely demystify many speculations about antibodies and B cell physiology. The chapter also considers successful vaccines and compares them with those infectious diseases where efficient protective vaccines are lacking, it is striking to note that all successful vaccines induce high levels of neutralizing antibodies (nAbs) that are both necessary and sufficient to protect the host from disease. Successful vaccination against infectious diseases such as tuberculosis, leprosy, or HIV would require induction of additional long-lasting T cell responses to control infection.

Novel Plant Virus-Based Vaccine Induces Protective Cytotoxic T-Lymphocyte-Mediated Antiviral Immunity through Dendritic Cell Maturation

ABSTRACT Currently used vaccines protect mainly through the production of neutralizing antibodies. However, antibodies confer little or no protection for a majority of chronic viral infections that require active involvement of cytotoxic T lymphocytes (CTLs). Virus-like particles (VLPs) have been shown to be efficient inducers of cell-mediated immune responses, but administration of an adjuvant is generally required. We recently reported the generation of a novel VLP system exploiting the self-assembly property of the papaya mosaic virus (PapMV) coat protein. We show here that uptake of PapMV-like particles by murine splenic dendritic cells (DCs) in vivo leads to their maturation, suggesting that they possess intrinsic adjuvant-like properties. DCs pulsed with PapMV-like particles displaying the lymphocytic choriomeningitis virus (LCMV) p33 immunodominant CTL epitope (PapMV-p33) efficiently process and cross-present the viral epitope to p33-specific transgenic T cells. Importantly, the CTL epitope is also properly processed and presented in vivo, since immunization of p33-specific T-cell receptor transgenic mice with PapMV-p33 induces the activation of large numbers of specific CTLs. C57BL/6 mice immunized with PapMV-p33 VLPs in the absence of adjuvant develop p33-specific effector CTLs that rapidly expand following LCMV challenge and protect vaccinated mice against LCMV infection in a dose-dependent manner. These results demonstrate the efficiency of this novel plant virus-based vaccination platform in inducing DC maturation leading to protective CTL responses.

Regulation of activation-induced deaminase stability and antibody gene diversification by Hsp90.

Hsp90 stabilizes and prevents degradation of cytoplasmic activation-induced deaminase.

Systems analysis of a RIG-I agonist inducing broad spectrum inhibition of virus infectivity.

The RIG-I like receptor pathway is stimulated during RNA virus infection by interaction between cytosolic RIG-I and viral RNA structures that contain short hairpin dsRNA and 5′ triphosphate (5′ppp) terminal structure. In the present study, an RNA agonist of RIG-I was synthesized in vitro and shown to stimulate RIG-I-dependent antiviral responses at concentrations in the picomolar range. In human lung epithelial A549 cells, 5′pppRNA specifically stimulated multiple parameters of the innate antiviral response, including IRF3, IRF7 and STAT1 activation, and induction of inflammatory and interferon stimulated genes - hallmarks of a fully functional antiviral response. Evaluation of the magnitude and duration of gene expression by transcriptional profiling identified a robust, sustained and diversified antiviral and inflammatory response characterized by enhanced pathogen recognition and interferon (IFN) signaling. Bioinformatics analysis further identified a transcriptional signature uniquely induced by 5′pppRNA, and not by IFNα-2b, that included a constellation of IRF7 and NF-kB target genes capable of mobilizing multiple arms of the innate and adaptive immune response. Treatment of primary PBMCs or lung epithelial A549 cells with 5′pppRNA provided significant protection against a spectrum of RNA and DNA viruses. In C57Bl/6 mice, intravenous administration of 5′pppRNA protected animals from a lethal challenge with H1N1 Influenza, reduced virus titers in mouse lungs and protected animals from virus-induced pneumonia. Strikingly, the RIG-I-specific transcriptional response afforded partial protection from influenza challenge, even in the absence of type I interferon signaling. This systems approach provides transcriptional, biochemical, and in vivo analysis of the antiviral efficacy of 5′pppRNA and highlights the therapeutic potential associated with the use of RIG-I agonists as broad spectrum antiviral agents.

Proteasome-Independent Major Histocompatibility Complex Class I Cross-Presentation Mediated by Papaya Mosaic Virus-Like Particles Leads to Expansion of Specific Human T Cells

ABSTRACT The development of versatile vaccine platforms is a priority that is recognized by health authorities worldwide; such platforms should induce both arms of the immune system, the humoral and cytotoxic-T-lymphocyte responses. In this study, we have established that a vaccine platform based on the coat protein of papaya mosaic virus (PapMV CP), previously shown to induce a humoral response, can induce major histocompatibility complex (MHC) class I cross-presentation of HLA-A*0201 epitopes from gp100, a melanoma antigen, and from influenza virus M1 matrix protein. PapMV proteins were able to assemble into stable virus-like particles (VLPs) in a crystalline and repetitive structure. When we pulsed HLA-A*0201+ antigen-presenting cells (APCs) with the recombinant PapMV FLU or gp100, we noted that antigen-specific CD8+ T cells were highly reactive to these APCs, demonstrating that the epitope from the VLPs were processed and loaded on the MHC class I complex. APCs were preincubated with two different proteasome inhibitors, which did not affect the efficiency of peptide presentation on MHC class I. Classical presentation from an endogenous antigen was abolished in the same conditions. Clearly, antigen presentation mediated by the PapMV system was proteasome independent. Finally, PapMV-pulsed APCs had the capacity to expand highly avid antigen-specific T cells against the influenza virus M1 HLA-A*0201 epitope when cocultured with autologous peripheral blood mononuclear cells. This study demonstrates the potential of PapMV for MHC class I cross-presentation and for the expansion of human antigen-specific T cells. It makes VLPs from PapMV CP a very attractive platform to trigger cellular responses for vaccine development against chronic infectious diseases and cancers.

Efnb1 and Efnb2 Proteins Regulate Thymocyte Development, Peripheral T Cell Differentiation, and Antiviral Immune Responses and Are Essential for Interleukin-6 (IL-6) Signaling

Background: Ephrins (Efn) are the ligands of Eph kinases. The roles of Efn in the T cell compartment are studied. Results: Efnb1 and Efnb2 double knock-out mice showed compromised thymocyte development, Th1 and Th17 function, IL-6 receptor signaling, and antivirus responses. Conclusion: Efnb1 and Efnb2 are involved in the T cell development and function. Significance: This study has revealed novel biological roles of Efns. Erythropoietin-producing hepatocellular kinases (Eph kinases) constitute the largest family of cell membrane receptor tyrosine kinases, and their ligand ephrins are also cell surface molecules. Because of promiscuous interaction between Ephs and ephrins, there is considerable redundancy in this system, reflecting the essential roles of these molecules in the biological system through evolution. In this study, both Efnb1 and Efnb2 were null-mutated in the T cell compartment of mice through loxP-mediated gene deletion. Mice with this double conditional mutation (double KO mice) showed reduced thymus and spleen size and cellularity. There was a significant decrease in the DN4, double positive, and single positive thymocyte subpopulations and mature CD4 and CD8 cells in the periphery. dKO thymocytes and peripheral T cells failed to compete with their WT counterparts in irradiated recipients, and the T cells showed compromised ability of homeostatic expansion. dKO naive T cells were inferior in differentiating into Th1 and Th17 effectors in vitro. The dKO mice showed diminished immune response against LCMV infection. Mechanistic studies revealed that IL-6 signaling in dKO T cells was compromised, in terms of abated induction of STAT3 phosphorylation upon IL-6 stimulation. This defect likely contributed to the observed in vitro and in vivo phenotype in dKO mice. This study revealed novel roles of Efnb1 and Efnb2 in T cell development and function.

Efficient T cell repertoire selection in tetraparental chimeric mice independent of thymic epithelial MHC

Nonthymic epithelial cells were compared with thymic epithelial cells for their role in T cell repertoire selection. Tetraparental aggregation chimeras were generated from T and B cell-deficient mice (H-2d SCID or H-2b Rag−/−) and thymus-deficient nude mice (H-2b or H-2d). These tetraparental mice showed primary protective CD8+ T cell responses, after lymphocytic choriomeningitis virus infection, that were peptide-specifically restricted to either thymic or nonthymic epithelial MHC at comparable levels. These chimeras also mounted neutralizing IgG responses dependent on cognate CD4+ T helper cell activity restricted to nonthymic epithelial MHC. Therefore, in contrast to earlier results with irradiation or thymus chimeras, these relatively undisturbed tetraparental mice reveal that the MHC of nonthymic epithelial cells efficiently selects a functional T cell repertoire.