ABSTRACT
Zika virus (ZIKV) is a significant threat to pregnant women and their fetuses as it can cause severe birth defects and congenital neurodevelopmental disorders, referred to as congenital Zika syndrome (CZS). Thus, a safe and effective ZIKV vaccine for pregnant women to prevent in utero ZIKV infection is of utmost importance. Murine models of ZIKV infection are limited by the fact that immunocompetent mice are resistant to ZIKV infection. As such, interferon-deficient mice have been used in some preclinical studies to test the efficacy of ZIKV vaccine candidates against lethal virus challenge. However, interferon-deficient mouse models have limitations in assessing the immunogenicity of vaccines, necessitating the use of immunocompetent mouse pregnancy models. Using the human stat2 knock-in (hSTAT2KI) mouse pregnancy model, we show that vaccination with a purified formalin-inactivated Zika virus (ZPIV) vaccine prior to pregnancy successfully prevented vertical transmission. In addition, maternal immunity protected offspring against postnatal challenge for up to 28 days. Furthermore, passive transfer of human IgG purified from hyper-immune sera of ZPIV vaccinees prevented maternal and fetal ZIKV infection, providing strong evidence that the neutralizing antibody response may serve as a meaningful correlate of protection.
ABSTRACT
Zika virus (ZIKV) infection during pregnancy poses significant threats to maternal and fetal health, leading to intrauterine fetal demise and severe developmental malformations that constitute congenital Zika syndrome (CZS). As such, the development of a safe and effective ZIKV vaccine is a critical public health priority. However, the safety and efficacy of such a vaccine during pregnancy remain uncertain. Historically, the conduct of clinical trials in pregnant women has been challenging. Therefore, clinically relevant animal pregnancy models are in high demand for testing vaccine efficacy. We previously reported that a marmoset pregnancy model of ZIKV infection consistently demonstrated vertical transmission from mother to fetus during pregnancy. Using this marmoset model, we also showed that vertical transmission could be prevented by pre-pregnancy vaccination with Zika purified inactivated virus (ZPIV) vaccine. Here, we further examined the efficacy of ZPIV vaccination during pregnancy. Vaccination during pregnancy elicited virus neutralizing antibody responses that were comparable to those elicited by pre-pregnancy vaccination. Vaccination also reduced placental pathology, viral burden and vertical transmission of ZIKV during pregnancy, without causing adverse effects. These results provide key insights into the safety and efficacy of ZPIV vaccination during pregnancy and demonstrate positive effects of vaccination on the reduction of ZIKV infection, an important advance in preparedness for future ZIKV outbreaks.
ABSTRACT
Zika virus (ZIKV) infection during pregnancy causes severe developmental defects in newborns, termed congenital Zika syndrome (CZS). Factors contributing to a surge in ZIKV-associated CZS are poorly understood. One possibility is that ZIKV may exploit the antibody-dependent enhancement of infection mechanism, mediated by cross-reactive antibodies from prior dengue virus (DENV) infection, which may exacerbate ZIKV infection during pregnancy. In this study, we investigated the impact of prior DENV infection or no DENV infection on ZIKV pathogenesis during pregnancy in a total of four female common marmosets with five or six fetuses per group. The results showed that negative-sense viral RNA copies increased in the placental and fetal tissues of DENV-immune dams but not in DENV-naïve dams. In addition, viral proteins were prevalent in endothelial cells, macrophages, and neonatal Fc receptor-expressing cells in the placental trabeculae and in neuronal cells in the brains of fetuses from DENV-immune dams. DENV-immune marmosets maintained high titers of cross-reactive ZIKV-binding antibodies that were poorly neutralizing, raising the possibility that these antibodies might be involved in the exacerbation of ZIKV infection. These findings need to be verified in a larger study, and the mechanism involved in the exacerbation of ZIKV infection in DENV-immune marmosets needs further investigation. However, the results suggest a potential negative impact of preexisting DENV immunity on subsequent ZIKV infection during pregnancy in vivo.
Subject(s)
Dengue Virus , Dengue , Zika Virus Infection , Zika Virus , Animals , Female , Pregnancy , Callithrix , Antibodies, Neutralizing , Antibodies, Viral , Endothelial Cells , Placenta , Cross ReactionsABSTRACT
Zika virus (ZIKV) is a mosquito-borne arbovirus that can cause severe congenital birth defects. The utmost goal of ZIKV vaccines is to prevent both maternal-fetal infection and congenital Zika syndrome. A Zika purified inactivated virus (ZPIV) was previously shown to be protective in non-pregnant mice and rhesus macaques. In this study, we further examined the efficacy of ZPIV against ZIKV infection during pregnancy in immunocompetent C57BL6 mice and common marmoset monkeys (Callithrix jacchus). We showed that, in C57BL/6 mice, ZPIV significantly reduced ZIKV-induced fetal malformations. Protection of fetuses was positively correlated with virus-neutralizing antibody levels. In marmosets, the vaccine prevented vertical transmission of ZIKV and elicited neutralizing antibodies that remained above a previously determined threshold of protection for up to 18 months. These proof-of-concept studies demonstrate ZPIV's protective efficacy is both potent and durable and has the potential to prevent the harmful consequence of ZIKV infection during pregnancy.
ABSTRACT
The herpesviruses varicella-zoster virus (VZV) and human cytomegalovirus (HCMV) are endemic to humans. VZV causes varicella (chicken pox) and herpes zoster (shingles), while HCMV causes serious disease in immunocompromised patients and neonates. More effective, less toxic antivirals are needed, necessitating better models to study these viruses and evaluate antivirals. Previously, VZV and HCMV models used fetal tissue; here, we developed an adult human skin model to study VZV and HCMV in culture and in vivo While VZV is known to grow in skin, it was unknown whether skin could support an HCMV infection. We used TB40/E HCMV and POka VZV strains to evaluate virus tropism in skin organ culture (SOC) and skin xenograft mouse models. Adult human skin from reduction mammoplasties was prepared for culture on NetWells or mouse implantation. In SOC, VZV infected the epidermis and HCMV infected the dermis. Specifically, HCMV infected fibroblasts, endothelial cells, and hematopoietic cells, with some infected cells able to transfer infection. VZV and HCMV mouse models were developed by subcutaneous transplantation of skin into SCID/beige or athymic nude mice at 2 independent sites. Viruses were inoculated directly into one xenograft, and widespread infection was observed for VZV and HCMV. Notably, we detected VZV- and HCMV-infected cells in the contralateral, uninoculated xenografts, suggesting dissemination from infected xenografts occurred. For the first time, we showed HCMV successfully grows in adult human skin, as does VZV. Thus, this novel system may provide a much-needed preclinical small-animal model for HCMV and VZV and, potentially, other human-restricted viruses.IMPORTANCE Varicella-zoster virus and human cytomegalovirus infect a majority of the global population. While they often cause mild disease, serious illness and complications can arise. Unfortunately, there are few effective drugs to treat these viruses, and many are toxic. To complicate this, these viruses are restricted to replication in human cells and tissues, making them difficult to study in traditional animal models. Current models rely heavily on fetal tissues, can be prohibitively expensive, and are often complicated to generate. While fetal tissue models provide helpful insights, it is necessary to study human viruses in human tissue systems to fully understand these viruses and adequately evaluate novel antivirals. Adult human skin is an appropriate model for these viruses because many target cells are present, including basal keratinocytes, fibroblasts, dendritic cells, and lymphocytes. Skin models, in culture and xenografts in immunodeficient mice, have potential for research on viral pathogenesis, tissue tropism, dissemination, and therapy.
Subject(s)
Chickenpox/virology , Cytomegalovirus/physiology , Herpes Zoster/virology , Herpesvirus 3, Human/physiology , Skin/virology , Animals , Antiviral Agents/pharmacology , Chickenpox/pathology , Cytomegalovirus/drug effects , Disease Models, Animal , Endothelial Cells , Fibroblasts/pathology , Fibroblasts/virology , Herpes Zoster/pathology , Herpesvirus 3, Human/drug effects , Heterografts , Humans , Male , Mice , Mice, Nude , Mice, SCID , Organ Culture Techniques , Skin/pathologyABSTRACT
Zika virus (ZIKV) infection during human pregnancy may cause diverse and serious congenital defects in the developing fetus. Previous efforts to generate animal models of human ZIKV infection and clinical symptoms often involved manipulating mice to impair their Type I interferon (IFN) signaling, thereby allowing enhanced infection and vertical transmission of virus to the embryo. Here, we show that even pregnant mice competent to generate Type I IFN responses that can limit ZIKV infection nonetheless develop profound placental pathology and high frequency of fetal demise. We consistently found that maternal ZIKV exposure led to placental pathology and that ZIKV RNA levels measured in maternal, placental or embryonic tissues were not predictive of the pathological effects seen in the embryos. Placental pathology included trophoblast hyperplasia in the labyrinth, trophoblast giant cell necrosis in the junctional zone, and loss of embryonic vessels. Our findings suggest that, in this context of limited infection, placental pathology rather than embryonic/fetal viral infection may be a stronger contributor to adverse pregnancy outcomes in mice. Our finding demonstrates that in immunocompetent mice, direct viral infection of the embryo is not essential for fetal demise. Our immunologically unmanipulated pregnancy mouse model provides a consistent and easily measurable congenital abnormality readout to assess fetal outcome, and may serve as an additional model to test prophylactic and therapeutic interventions to protect the fetus during pregnancy, and for studying the mechanisms of ZIKV congenital immunopathogenesis.
Subject(s)
Disease Models, Animal , Fetal Diseases/pathology , Placenta Diseases/pathology , Pregnancy Complications, Infectious/pathology , Zika Virus Infection/pathology , Zika Virus/physiology , Animals , Female , Fetal Diseases/virology , Infectious Disease Transmission, Vertical , Mice , Mice, Inbred C57BL , Placenta Diseases/virology , Pregnancy , Pregnancy Complications, Infectious/virology , Pregnancy Outcome , RNA, Viral , Zika Virus Infection/virologyABSTRACT
CD4+ T cells mediate protection against Mycobacterium tuberculosis (Mtb); however, the phenotype of protective T cells is undefined, thereby confounding vaccination efforts. IL-27 is highly expressed during human tuberculosis (TB), and absence of IL-27R (Il27ra) specifically on T cells results in increased protection. IL-27R deficiency during chronic Mtb infection does not impact antigen-specific CD4+ T cell number but maintains programmed death-1 (PD-1), CD69, and CD127 expression while reducing T-bet and killer cell lectin-like receptor G1 (KLRG1) expression. Furthermore, T-bet haploinsufficiency results in failure to generate KLRG1+, antigen-specific CD4+ T cells, and in improved protection. T cells in Il27ra(-/-) mice accumulate preferentially in the lung parenchyma within close proximity to Mtb, and antigen-specific CD4+ T cells lacking IL-27R are intrinsically more fit than intact T cells and maintain IL-2 production. Improved fitness of IL-27R-deficient T cells is not associated with increased proliferation but with decreased expression of cell death-associated markers. Therefore, during Mtb infection, IL-27R acts intrinsically on T cells to limit protection and reduce fitness, whereas the IL-27R-deficient environment alters the phenotype and location of T cells. The significant expression of IL-27 in TB and the negative influence of IL-27R on T cell function demonstrate the pathway by which this cytokine/receptor pair is detrimental in TB.
Subject(s)
CD4-Positive T-Lymphocytes/immunology , Mycobacterium tuberculosis/immunology , Receptors, Cytokine/immunology , Receptors, Interleukin/immunology , Tuberculosis/immunology , Adult , Animals , Antigens, CD/genetics , Antigens, CD/immunology , Antigens, Differentiation, T-Lymphocyte/genetics , Antigens, Differentiation, T-Lymphocyte/immunology , CD4-Positive T-Lymphocytes/pathology , Female , Humans , Interleukin-7 Receptor alpha Subunit/genetics , Interleukin-7 Receptor alpha Subunit/immunology , Interleukins/genetics , Interleukins/immunology , Lectins, C-Type/genetics , Lectins, C-Type/immunology , Male , Mice , Mice, Knockout , Programmed Cell Death 1 Receptor/genetics , Programmed Cell Death 1 Receptor/immunology , Receptors, Cytokine/genetics , Receptors, Immunologic/genetics , Receptors, Immunologic/immunology , Receptors, Interleukin/genetics , Trans-Activators/genetics , Trans-Activators/immunology , Tuberculosis/genetics , Tuberculosis/pathologyABSTRACT
Immunomodulatory agents potentially represent a new class of broad-spectrum antimicrobials. Here, we demonstrate that prophylaxis with immunomodulatory cytosine-phosphate-guanidine (CpG) oligodeoxynucleotide (ODN), a toll-like receptor 9 (TLR9) agonist, confers protection against Yersinia pestis, the etiologic agent of plague. The data establish that intranasal administration of CpG ODN 1 day prior to lethal pulmonary exposure to Y. pestis strain KIM D27 significantly improves survival of C57BL/6 mice and reduces bacterial growth in hepatic tissue, despite paradoxically increasing bacterial growth in the lung. All of these CpG ODN-mediated impacts, including the increased pulmonary burden, are TLR9 dependent, as they are not observed in TLR9-deficient mice. The capacity of prophylactic intranasal CpG ODN to enhance survival does not require adaptive immunity, as it is evident in mice lacking B and/or T cells; however, the presence of T cells improves long-term survival. The prophylactic regimen also improves survival and reduces hepatic bacterial burden in mice challenged intraperitoneally with KIM D27, indicating that intranasal delivery of CpG ODN has systemic impacts. Indeed, intranasal prophylaxis with CpG ODN provides significant protection against subcutaneous challenge with Y. pestis strain CO92 even though it fails to protect mice from intranasal challenge with that fully virulent strain.
Subject(s)
Adjuvants, Immunologic/administration & dosage , Oligodeoxyribonucleotides/administration & dosage , Plague/prevention & control , Yersinia pestis , Adjuvants, Immunologic/pharmacology , Administration, Intranasal , Animals , Gene Expression Regulation/immunology , Liver/microbiology , Lung/cytology , Lung/microbiology , Mice , Mice, Inbred C57BL , Oligodeoxyribonucleotides/pharmacology , Specific Pathogen-Free Organisms , Toll-Like Receptor 9/agonists , Toll-Like Receptor 9/genetics , Toll-Like Receptor 9/metabolism , Virulence , Yersinia pestis/immunology , Yersinia pestis/pathogenicityABSTRACT
Cell-mediated immune responses are known to be critical for control of mycobacterial infections whereas the role of B cells and humoral immunity is unclear. B cells can modulate immune responses by secretion of immunoglobulin, production of cytokines and antigen-presentation. To define the impact of B cells in the absence of secreted immunoglobulin, we analyzed the progression of Mycobacterium tuberculosis (Mtb) infection in mice that have B cells but which lack secretory immunoglobulin (AID(-/-)µS(-/-)mice). AID(-/-)µS(-/-) mice accumulated a population of activated B cells in the lungs when infected and were more susceptible to aerosol Mtb when compared to wild type (C57BL/6) mice or indeed mice that totally lack B cells. The enhanced susceptibility of AID(-/-)µS(-/-) mice was not associated with defective T cell activation or expression of a type 1 immune response. While delivery of normal serum to AID(-/-)µS(-/-) mice did not reverse susceptibility, susceptibility in the spleen was dependent upon the presence of B cells and susceptibility in the lungs of AID(-/-)µS(-/-)mice was associated with elevated expression of the cytokines IL-6, GM-CSF, IL-10 and molecules made by alternatively activated macrophages. Blocking of IL-10 signaling resulted in reversal of susceptibility in the spleens and lungs of AID(-/-)µS(-/-) mice. These data support the hypothesis that B cells can modulate immunity to Mtb in an organ specific manner via the modulation of cytokine production and macrophage activation.
Subject(s)
B-Lymphocytes/immunology , Mycobacterium tuberculosis/immunology , Tuberculosis/immunology , Animals , B-Lymphocytes/metabolism , Granulocyte-Macrophage Colony-Stimulating Factor/metabolism , Interleukin-10/metabolism , Interleukin-6/metabolism , MiceABSTRACT
The Gram-negative bacterium Yersinia pestis causes plague, a rapidly progressing and often fatal disease. The formation of fibrin at sites of Y. pestis infection supports innate host defense against plague, perhaps by providing a nondiffusible spatial cue that promotes the accumulation of inflammatory cells expressing fibrin-binding integrins. This report demonstrates that fibrin is an essential component of T cell-mediated defense against plague but can be dispensable for Ab-mediated defense. Genetic or pharmacologic depletion of fibrin abrogated innate and T cell-mediated defense in mice challenged intranasally with Y. pestis. The fibrin-deficient mice displayed reduced survival, increased bacterial burden, and exacerbated hemorrhagic pathology. They also showed fewer neutrophils within infected lung tissue and reduced neutrophil viability at sites of liver infection. Depletion of neutrophils from wild-type mice weakened T cell-mediated defense against plague. The data suggest that T cells combat plague in conjunction with neutrophils, which require help from fibrin to withstand Y. pestis encounters and effectively clear bacteria.
Subject(s)
Fibrin/physiology , Immunity, Innate , T-Lymphocyte Subsets/immunology , T-Lymphocyte Subsets/microbiology , Yersinia pestis/immunology , Animals , Bacterial Proteins/physiology , Fibrinogen/physiology , Mice , Mice, Inbred C57BL , Mice, Knockout , Plague/immunology , Plague/metabolism , Plasminogen Activators/physiologyABSTRACT
Animals lacking the inducible nitric oxide synthase gene (nos2(-/-)) are less susceptible to Mycobacterium avium strain 25291 and lack nitric oxide-mediated immunomodulation of CD4(+) T cells. Here we show that the absence of nos2 results in increased accumulation of neutrophils and both CD4(+) and CD8(+) T cells within the M. avium containing granuloma. Examination of the T-cell phenotype in M. avium infected mice demonstrated that CD4(+)CD44(hi) effector T cells expressing the Th1 transcriptional regulator T-bet (T-bet(+)) were specifically reduced by the presence of nitric oxide. Importantly, the T-bet(+) effector population could be separated into CD69(hi) and CD69(lo) populations, with the CD69(lo) population only able to accumulate during chronic infection within infected nos2(-/-) mice. Transcriptomic comparison between CD4(+)CD44(hi)CD69(hi) and CD4(+)CD44(hi)CD69(lo) populations revealed that CD4(+)CD44(hi)CD69(lo) cells had higher expression of the integrin itgb1/itga4 (VLA-4, CD49d/CD29). Inhibition of Nos2 activity allowed increased accumulation of the CD4(+) CD44(hi)T-bet(+)CD69(lo) population in WT mice as well as increased expression of VLA-4. These data support the hypothesis that effector T cells in mycobacterial granulomata are not a uniform effector population but exist in distinct subsets with differential susceptibility to the regulatory effects of nitric oxide.
Subject(s)
Mycobacterium avium/immunology , Nitric Oxide Synthase Type II/immunology , Nitric Oxide/immunology , Th1 Cells/immunology , Tuberculosis, Pulmonary/immunology , Animals , Antigens, CD/genetics , Antigens, CD/immunology , Antigens, Differentiation, T-Lymphocyte/genetics , Antigens, Differentiation, T-Lymphocyte/immunology , CD8-Positive T-Lymphocytes/immunology , Cell Adhesion Molecules/genetics , Cell Adhesion Molecules/immunology , Granuloma, Respiratory Tract/genetics , Granuloma, Respiratory Tract/immunology , Granuloma, Respiratory Tract/microbiology , Granuloma, Respiratory Tract/veterinary , Lectins, C-Type/genetics , Lectins, C-Type/immunology , Mice , Mice, Knockout , Nitric Oxide/genetics , Nitric Oxide Synthase Type II/genetics , T-Box Domain Proteins/genetics , T-Box Domain Proteins/immunology , Tuberculosis, Pulmonary/genetics , Tuberculosis, Pulmonary/veterinaryABSTRACT
CD4 T cells, and especially T follicular helper cells, are critical for the generation of a robust humoral response to an infection or vaccination. Importantly, immunosenescence affects CD4 T-cell function, and the accumulation of intrinsic defects decreases the cognate helper functions of these cells. However, much less is known about the contribution of the aged microenvironment to this impaired CD4 T-cell response. In this study, we have employed a preclinical model to determine whether the aged environment contributes to the defects in CD4 T-cell functions with aging. Using an adoptive transfer model in mice, we demonstrate for the first time that the aged microenvironment negatively impacts at least three steps of the CD4 T-cell response to antigenic stimulation. First, the recruitment of CD4 T cells to the spleen is reduced in aged compared to young hosts, which correlates with dysregulated chemokine expression in the aged organ. Second, the priming of CD4 T cells by DCs is reduced in aged compared to young mice. Finally, naïve CD4 T cells show a reduced transition to a T follicular helper cell phenotype in the aged environment, which impairs the subsequent generation of germinal centers. These studies have provided new insights into how aging impacts the immune system and how these changes influence the development of immunity to infections or vaccinations.
Subject(s)
Aging/immunology , CD4-Positive T-Lymphocytes/immunology , Animals , CD4-Positive T-Lymphocytes/cytology , CD4-Positive T-Lymphocytes/metabolism , Cell Differentiation/immunology , Cell Growth Processes/immunology , Chemokine CCL21/immunology , Chemokine CXCL13/immunology , Dendritic Cells/cytology , Dendritic Cells/immunology , Lymphocyte Activation , Lymphoid Tissue/cytology , Lymphoid Tissue/immunology , Mice , Mice, Inbred C57BL , Mice, Knockout , Spleen/cytology , Spleen/immunologyABSTRACT
Mouse invariant natural killer T cells (iNKT cells) provide cognate and noncognate help for lipid and protein-specific B cells, respectively. However, the long-term outcome for B cells after cognate help is provided by iNKT cells is unknown at present. Here we found that cognate iNKT cell help resulted in a B cell differentiation program characterized by extrafollicular plasmablasts, germinal-center formation, affinity maturation and a robust primary immunoglobulin G (IgG) antibody response that was uniquely dependent on iNKT cell-derived interleukin 21 (IL-21). However, cognate help from iNKT cells did not generate an enhanced humoral memory response. Thus, cognate iNKT cell help for lipid-specific B cells induces a unique signature that is a hybrid of classic T cell-dependent and T cell-independent type 2 B cell responses.
Subject(s)
Antigens/immunology , B-Lymphocytes/immunology , Interleukins/physiology , Lipids/immunology , Natural Killer T-Cells/immunology , Animals , Germinal Center/immunology , Immunity, Humoral , Immunologic Memory , Mice , Mice, Inbred C57BL , Mice, Transgenic , Receptors, Antigen, B-Cell/immunology , Signal Transduction/immunology , Spleen/immunologyABSTRACT
IL-23 is required for the IL-17 response to infection with Mycobacterium tuberculosis, but is not required for the early control of bacterial growth. However, mice deficient for the p19 component of IL-23 (Il23a(-/-)) exhibit increased bacterial growth late in infection that is temporally associated with smaller B cell follicles in the lungs. Cxcl13 is required for B cell follicle formation and immunity during tuberculosis. The absence of IL-23 results in decreased expression of Cxcl13 within M. tuberculosis-induced lymphocyte follicles in the lungs, and this deficiency was associated with increased cuffing of T cells around the vessels in the lungs of these mice. Il23a(-/-) mice also poorly expressed IL-17A and IL-22 mRNA. These cytokines were able to induce Cxcl13 in mouse primary lung fibroblasts, suggesting that these cytokines are likely involved in B cell follicle formation. Indeed, IL-17RA-deficient mice generated smaller B cell follicles early in the response, whereas IL-22-deficient mice had smaller B cell follicles at an intermediate time postinfection; however, only Il23a(-/-) mice had a sustained deficiency in B cell follicle formation and reduced immunity. We propose that in the absence of IL-23, expression of long-term immunity to tuberculosis is compromised due to reduced expression of Cxcl13 in B cell follicles and reduced ability of T cells to migrate from the vessels and into the lesion. Further, although IL-17 and IL-22 can both contribute to Cxcl13 production and B cell follicle formation, it is IL-23 that is critical in this regard.
Subject(s)
Germinal Center/immunology , Germinal Center/pathology , Interleukin-23/physiology , Mycobacterium tuberculosis/immunology , Tuberculosis, Pulmonary/immunology , Tuberculosis, Pulmonary/pathology , Animals , Cells, Cultured , Chemokine CXCL13/biosynthesis , Germinal Center/microbiology , Interleukin-23/deficiency , Interleukin-23/genetics , Lung/immunology , Lung/microbiology , Lung/pathology , Mice , Mice, Inbred C57BL , Mice, Knockout , Mycobacterium tuberculosis/growth & development , Time Factors , Tuberculosis, Pulmonary/microbiologyABSTRACT
Survival of mature B cells is regulated by B cell receptor and BAFFR-dependent signals. We show that B cells from mice lacking the G(alphaq) subunit of trimeric G proteins (Gnaq(-/-) mice) have an intrinsic survival advantage over normal B cells, even in the absence of BAFF. Gnaq(-/-) B cells develop normally in the bone marrow but inappropriately survive peripheral tolerance checkpoints, leading to the accumulation of transitional, marginal zone, and follicular B cells, many of which are autoreactive. Gnaq(-/-) chimeric mice rapidly develop arthritis as well as other manifestations of systemic autoimmune disease. Importantly, we demonstrate that the development of the autoreactive B cell compartment is the result of an intrinsic defect in Gnaq(-/-) B cells, resulting in the aberrant activation of the prosurvival factor Akt. Together, these data show for the first time that signaling through trimeric G proteins is critically important for maintaining control of peripheral B cell tolerance induction and repressing autoimmunity.
Subject(s)
Autoimmunity/immunology , B-Lymphocytes/cytology , B-Lymphocytes/immunology , Cell Differentiation/immunology , GTP-Binding Protein alpha Subunits, Gq-G11/genetics , Heterotrimeric GTP-Binding Proteins/physiology , Immune Tolerance , Anemia/blood , Anemia/immunology , Animals , Antibodies, Anti-Idiotypic/immunology , Antibodies, Antinuclear/blood , Antibodies, Antinuclear/immunology , Antibodies, Monoclonal/immunology , Antibodies, Monoclonal/pharmacology , Antigen-Antibody Complex/metabolism , Arthritis/immunology , Arthritis/pathology , Autoantigens/immunology , Autoimmune Diseases/genetics , Autoimmune Diseases/immunology , Autoimmune Diseases/mortality , Autoimmune Diseases/pathology , Autoimmunity/genetics , B-Cell Activating Factor/immunology , B-Cell Activating Factor/pharmacology , B-Lymphocyte Subsets/cytology , B-Lymphocytes/drug effects , B-Lymphocytes/metabolism , B-Lymphocytes/transplantation , Cell Differentiation/genetics , Cell Movement/genetics , Cell Movement/immunology , Cell Survival/genetics , Cell Survival/immunology , Homeostasis/immunology , Kidney/metabolism , Kidney/pathology , Lymphocyte Activation/genetics , Lymphocyte Activation/immunology , Mice , Mice, Inbred C57BL , Mice, Knockout , Proto-Oncogene Proteins c-akt/metabolism , Radiation Chimera/immunology , Receptors, Antigen, B-Cell/immunology , Spleen/cytology , Spleen/drug effects , T-Lymphocytes/cytologyABSTRACT
In vitro generated OVA-specific IL-17-producing CD8 T effector cells (Tc17) from OT-1 mice, adoptively transferred into B16-OVA tumor-bearing mice, controlled tumor growth in early and late stage melanoma. IL-17, TNF, and IFN-gamma from the Tc17 effectors all played a role in an enhanced recruitment of T cells, neutrophils, and macrophages to the tumor. In addition, Tc17 cells and recently recruited, activated neutrophils produced further chemokines, including CCL3, CCL4, CCL5, CXCL9, and CXCL10, responsible for the attraction of type 1 lymphocytes (Th1 and Tc1) and additional neutrophils. Neutrophils were rapidly attracted to the tumor site by an IL-17 dependent mechanism, but at later stages the induction of the chemokine CXCL2 by Tc17-derived TNF and IFN-gamma contributed to sustain neutrophil recruitment. Approximately 10-50 times as many Tc17 effectors were required compared with Tc1 effectors to exert the same level of control over tumor growth. The recruitment of neutrophils was more prominent when Tc17 rather than Tc1 were used to control tumor and depletion of neutrophils resulted in a diminished capacity to control tumor growth.
Subject(s)
CD8-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/transplantation , Immunotherapy, Adoptive/methods , Interleukin-17/biosynthesis , Melanoma, Experimental/immunology , Melanoma, Experimental/therapy , Animals , CD8-Positive T-Lymphocytes/pathology , Cell Line, Tumor , Cytotoxicity Tests, Immunologic , Disease Models, Animal , Female , Interleukin-17/physiology , Lymphocytes, Tumor-Infiltrating/immunology , Lymphocytes, Tumor-Infiltrating/pathology , Melanoma, Experimental/pathology , Mice , Mice, Inbred C57BL , Mice, Knockout , Mice, Transgenic , Neoplasm Staging , Ovalbumin/immunology , T-Lymphocytes, Cytotoxic/immunology , T-Lymphocytes, Cytotoxic/pathology , T-Lymphocytes, Cytotoxic/transplantationABSTRACT
We examined the expression and influence of IL-10 during influenza infection. We found that IL-10 does not impact sublethal infection, heterosubtypic immunity, or the maintenance of long-lived influenza Ag depots. However, IL-10-deficient mice display dramatically increased survival compared with wild-type mice when challenged with lethal doses of virus, correlating with increased expression of several Th17-associated cytokines in the lungs of IL-10-deficient mice during the peak of infection, but not with unchecked inflammation or with increased cellular responses. Foxp3(-) CD4 T cell effectors at the site of infection represent the most abundant source of IL-10 in wild-type mice during high-dose influenza infection, and the majority of these cells coproduce IFN-gamma. Finally, compared with predominant Th1 responses in wild-type mice, virus-specific T cell responses in the absence of IL-10 display a strong Th17 component in addition to a strong Th1 response and we show that Th17-polarized CD4 T cell effectors can protect naive mice against an otherwise lethal influenza challenge and utilize unique mechanisms to do so. Our results show that IL-10 expression inhibits development of Th17 responses during influenza infection and that this is correlated with compromised protection during high-dose primary, but not secondary, challenge.
Subject(s)
Influenza A Virus, H1N1 Subtype/immunology , Interleukin-10/deficiency , Interleukin-10/immunology , Interleukin-17/immunology , Orthomyxoviridae Infections/immunology , T-Lymphocytes, Helper-Inducer/immunology , Animals , Antigens, Viral/immunology , Forkhead Transcription Factors/immunology , Interleukin-10/genetics , Interleukin-10/metabolism , Mice , Orthomyxoviridae Infections/genetics , Orthomyxoviridae Infections/metabolism , Survival RateABSTRACT
CD38 controls the chemotaxis of leukocytes to some, but not all, chemokines, suggesting that chemokine receptor signaling in leukocytes is more diverse than previously appreciated. To determine the basis for this signaling heterogeneity, we examined the chemokine receptors that signal in a CD38-dependent manner and identified a novel "alternative" chemokine receptor signaling pathway. Similar to the "classical" signaling pathway, the alternative chemokine receptor pathway is activated by Galpha(i2)-containing Gi proteins. However, unlike the classical pathway, the alternative pathway is also dependent on the Gq class of G proteins. We show that Galpha(q)-deficient neutrophils and dendritic cells (DCs) make defective calcium and chemotactic responses upon stimulation with N-formyl methionyl leucyl phenylalanine and CC chemokine ligand (CCL) 3 (neutrophils), or upon stimulation with CCL2, CCL19, CCL21, and CXC chemokine ligand (CXCL) 12 (DCs). In contrast, Galpha(q)-deficient T cell responses to CXCL12 and CCL19 remain intact. Thus, the alternative chemokine receptor pathway controls the migration of only a subset of cells. Regardless, the novel alternative chemokine receptor signaling pathway appears to be critically important for the initiation of inflammatory responses, as Galpha(q) is required for the migration of DCs from the skin to draining lymph nodes after fluorescein isothiocyanate sensitization and the emigration of monocytes from the bone marrow into inflamed skin after contact sensitization.
Subject(s)
Dendritic Cells/physiology , GTP-Binding Protein alpha Subunits, Gq-G11/physiology , Granulocytes/physiology , Receptors, Chemokine/physiology , Signal Transduction , ADP-ribosyl Cyclase 1/deficiency , ADP-ribosyl Cyclase 1/genetics , ADP-ribosyl Cyclase 1/physiology , Animals , Antigens, CD/physiology , CD4-Positive T-Lymphocytes/immunology , Chemotaxis, Leukocyte , Mice , Mice, KnockoutABSTRACT
Secondary lymphoid organs (SLOs) promote primary immune responses by recruiting naive lymphocytes and activated APCs. However, their role in the persistence or responsiveness of memory lymphocytes is unclear. We tested whether memory cells were maintained and could respond to challenge in the absence of SLOs. We found that influenza-specific CD8 cells in the lung acquired a memory phenotype, underwent homeostatic proliferation, recirculated through nonlymphoid tissues, and responded to and cleared a challenge infection in the complete absence of SLOs. Similarly, influenza-specific virus-neutralizing antibody was generated and maintained in the absence of SLOs. Inducible bronchus-associated lymphoid tissue (iBALT) was also formed in the lungs of previously infected mice and may provide a niche for the maintenance of memory cells at the local level. These data show that SLOs are dispensable for the maintenance of immunologic memory and directly demonstrate the utility of local tissues, such as iBALT, in secondary immune responses.