Géraldine Arrode-Brusés

Maternal immune activation by poly(I:C) induces expression of cytokines IL-1β and IL-13, chemokine MCP-1 and colony stimulating factor VEGF in fetal mouse brain

BackgroundMaternal viral infection during pregnancy is associated with an increase in the incidence of psychiatric disorders with presumed neurodevelopmental origin, including autism spectrum disorders and schizophrenia. The enhanced risk for developing mental illness appears to be caused by deleterious effects of innate immune response-associated factors on the development of the central nervous system, which predispose the offspring to pathological behaviors in adolescence and adulthood. To identify the immune response-associated soluble factors that may affect central nervous system development, we examined the effect of innate immune response activation by polyriboinosinic-polyribocytidylic acid (poly(I:C)), a synthetic analogue of viral double-stranded RNA, on the expression levels of pro- and anti-inflammatory cytokines, chemokines and colony stimulating factors in fetal and postnatal mouse brain 6 h and 24 h after treatment.MethodsC57BL/6J pregnant mice (gestational day 16) or newborn mice (postnatal day 4) received a single intraperitoneal injection of the synthetic analogue of viral double-stranded RNA poly(I:C) (20 mg/kg). Thirty-two immune response-associated soluble factors, including pro- and anti-inflammatory cytokines, chemokines and colony stimulating factors, were assayed 6 h and 24 h after poly(I:C) injection using multiplexed bead-based immunoassay (Milliplex Map) and processed in a Luminex 100 IS instrument.ResultsMaternal exposure to poly(I:C) at gestational day 16 induced a significant increase in cytokines interleukin (IL)-1β, IL-7 and IL-13; chemokines monocyte chemoattractant protein 1 (MCP-1), macrophage inflammatory protein (MIP)-1α, interferon gamma-induced protein (IP)-10 and monokine induced by IFN-gamma (MIG); and in the colony stimulating factor vascular endothelial growth factor (VEGF) in the fetal brain. IL-1β showed the highest concentration levels in fetal brains and was the only cytokine significantly up-regulated 24 h after maternal poly(I:C) injection, suggesting that IL-1β may have a deleterious impact on central nervous system development. In contrast, poly(I:C) treatment of postnatal day 4 pups induced a pronounced rise in chemokines and colony stimulating factors in their brains instead of the pro-inflammatory cytokine IL-1β.ConclusionsThis study identified a significant increase in the concentration levels of the cytokines IL-1β and IL-13, the chemokine MCP-1 and the colony stimulating factor VEGF in the developing central nervous system during activation of an innate immune response, suggesting that these factors are mediators of the noxious effects of maternal immune activation on central nervous system development, with potential long-lasting effects on animal behavior.

Characterization of T-Cell Responses in Macaques Immunized with a Single Dose of HIV DNA Vaccine

ABSTRACT The optimization of immune responses (IR) induced by HIV DNA vaccines in humans is one of the great challenges in the development of an effective vaccine against AIDS. Ideally, this vaccine should be delivered in a single dose to immunize humans. We recently demonstrated that the immunization of mice with a single dose of a DNA vaccine derived from pathogenic SHIVKU2 (Δ4SHIVKU2) induced long-lasting, potent, and polyfunctional HIV-specific CD8+ T-cell responses (G. Arrode, R. Hegde, A. Mani, Y. Jin, Y. Chebloune, and O. Narayan, J. Immunol. 178:2318-2327, 2007). In the present work, we expanded the characterization of the IR induced by this DNA immunization protocol to rhesus macaques. Animals immunized with a single high dose of Δ4SHIVKU2 DNA vaccine were monitored longitudinally for vaccine-induced IR using multiparametric flow cytometry-based assays. Interestingly, all five immunized macaques developed broad and polyfunctional HIV-specific T-cell IR that persisted for months, with an unusual reemergence in the blood following an initial decline but in the absence of antibody responses. The majority of vaccine-specific CD4+ and CD8+ T cells lacked gamma interferon production but showed high antigen-specific proliferation capacities. Proliferative CD8+ T cells expressed the lytic molecule granzyme B. No integrated viral vector could be detected in mononuclear cells from immunized animals, and this high dose of DNA did not induce any detectable autoimmune responses against DNA. Taken together, our comprehensive analysis demonstrated for the first time the capacity of a single high dose of HIV DNA vaccine alone to induce long-lasting and polyfunctional T-cell responses in the nonhuman primate model, bringing new insights for the design of future HIV vaccines.

Immunogenicity of a lentiviral-based DNA vaccine driven by the 5 LTR of the naturally attenuated caprine arthritis encephalitis virus (CAEV) in mice and macaques

Increasing the safety and the efficacy of existing HIV vaccines is one of the strategies that could help to promote the development of a vaccine for human use. We developed a HIV DNA vaccine (Δ4-SHIVKU2) that has been shown to induce potent polyfunctional HIV-specific T cell responses following a single dose immunization of mice and macaques. Δ4-SHIVKU2 also induced protection when immunized macaques were challenged with homologous pathogenic viruses. In the present study, our aim was to examine whether a chimeric HIV DNA vaccine (CAL-Δ4-SHIVKU2) whose genome is driven by the LTR of the goat lentivirus, caprine arthritis encephalitis (CAEV) expresses efficiently the vaccine antigens and induces potent immune responses in animal models for HIV vaccine. Data of radioimmunoprecipitation assays clearly show that this chimeric genome drives efficient expression of all HIV antigens in the construct. In addition, evaluation of the p24 Gag protein in the supernatant of HEK-293-T cells transfected in parallel with Δ4-SHIVKU2 and CAL-Δ4-SHIVKU2 showed no difference suggesting that these two LTRs are inducing equally the expression of the viral genes. Immunization of mice and macaques using our single dose immunization regimen resulted in induction of similar IFN-γ ELISPOT responses in Δ4-SHIVKU2- and CAL-Δ4-SHIVKU2-treated mice. Similar profiles of T cell responses were also detected both in mice and macaques when multiparametric flow cytometry analyses were performed. Since CAEV LTR is not dependent of Tat to drive viral gene expression and is not functional for integration with HIV integrase, this new vector increases the safety and efficacy of our vaccine vectors and vaccination strategy.

Changes of biological properties and pathogenesis of CAEV chimeras expressing Nef and Vpx/Vpr accessory proteins in infected goats

Background Caprine Arthritis Encephalitis Virus (CAEV) is a goat lentivirus closely related to HIV that is naturally attenuated with only a minor proportion of infected goats that develop inflammatory diseases, in absence of progression in AIDS. In contrast, to HIV CAEV does not infect productively the CD4+ T cells and has a simpler genome organization. Indeed, CAEV lacks 3 out of 6 regulatory/accessory genes found in HIV. We thought that CAEV/goat is an excellent model to study the functions and the implication of primate lentiviral accessory proteins in the biology and the pathogenesis of lentiviruses.

Cowpox Helped Against Smallpox; Will the Goat Lentivirus (Caprine Arthritis Encephalitis Virus) Help Against HIV-1?

Unlike HIV-1 in humans, the natural goat lentivirus caprine arthritis encephalitis virus (CAEV) does not cause AIDS-like disease in infected goats and some virus strains are totally nonpathogenic. In most naturally or experimentally CAEV-infected healthy goats the virus replicates only transiently in the periphery. CAEV replicates productively in macrophages but not in CD4+ T cells. Unlike HIV-1, we have demonstrated that the CAEV genome is driven by Tat-independent constitutive LTR promoters.1 We have shown that experimental CAEV infection of calves resulted in the total clearance of the virus following 2–4 months of productive replication.2 From these data and those from numerous experimental and natural studies of CAEV infection, we hypothesized that CAEV LTR driving constitutive expression of viral proteins might be one of the key factors of T cell-mediated virus clearance. We generated a simian human immunodeficiency virus (SHIV)-based replication-defective lentivector DNA vaccine whose antigen expression was driven by the 5′ CAEV LTR and found that this DNA vaccine induces potent immune responses both in mice and macaques.3 To augment the immunogenicity we engineered a novel nonintegrative one-cycle SHIV-based lentivector vaccine driven by both 5′ and 3′ LTRs of CAEV (Fig. 1). This vaccine was designed to be delivered as DNA whose expression in in vivo-transfected cells produces viral proteins used either directly as antigen for immune stimulation or for assembly into released virions, and also used as another source of antigen or in mature infectious virions that infect only target cells without integration of the vaccine genome to amplify the vaccines antigen further. This design synergizes the advantages of both DNA and live-attenuated vaccines but lacks their associated inconvenience. FIG. 1. Development of a caprine arthritis encephalitis virus (CAEV)/HIV chimera as an anti-HIV vaccine for humans. The naturally attenuated genome of CAEV is used as a source to isolate key sequences to generate a CAEV/HIV chimeric genome used as a lentivector ... This novel lentivector vaccine was found to be highly immunogenic in PBL hu-SCID mice and induced potent and persistent CD4+ and CD8+ T cells in macaques immunized with a single dose (5 mg) of DNA in the absence of any boost. T cell responses contained effector, memory effector, and central and stem memory T cells with a high recall capacity. Antibody to the major capsid Gag and Env proteins was also repeatedly detected.4 Repeated low-dose challenge of vaccine and control animals with the heterologous highly pathogenic SIVmac251 showed efficient control of virus replication in all (6/6) vaccinated macaques. The absence of vaccine-induced neutralizing antibody responses resulted in a lack of protection against virus acquisition. These data clearly demonstrate that CAEV may provide key elements in the design of safe and efficacious HIV vaccine prototypes for the protection of human populations. The recent findings in the development of HIV broadly neutralizing antibodies with increased breadth and potency in immunized Llamas may provide winning anti-HIV vaccine combination strategies against virus replication and acquisition as well.

P17-01. HIV-specific immune responses induced by a single dose of HIV DNA vaccine in Rhesus macaques

Background DNA vaccine is one of the most promising strategies for development of an efficacious HIV vaccine. Current HIV DNA vaccines induce potent immune responses (IR) in rodents but their immunogenicity remains weak in human and non-human primates (NHP). On the other hand, the MRKAd-5 based HIV vaccine that induces strong HIV-specific T cell responses failed to control HIV infection in human clinical trial. Therefore, strategies that enhance the immunogenicity of HIV DNA vaccines together with a strong commitment to better characterize the vaccine-induced IR are needed. Such comprehensive studies are critical in defining correlates of immune protection.

P03-04. Increased virulence of CAEV chimeras expressing Nef and Vpx/Vpr accessory proteins in infected goats

Background Caprine Arthritis Encephalitis Virus (CAEV) is a goat lentivirus closely related to HIV except that CAEV never causes AIDS-like disease in goat, in addition, only a minor proportion of infected goats develop inflammatory diseases. One of the main differences that distinguishes CAEV and HIV is the simplicity of CAEV genome organization. CAEV lacks 3 out of 6 regulatory/accessory genes found in HIV. We thought that CAEV/goat is an excellent model to study the functions and the implication of primate lentiviral accessory proteins in the pathogenicity of lentiviruses.