Unit: Regulation of Retroviral Infections

Director: Françoise BARRE-SINOUSSI

Our Unit is conducting research on mechanisms involved in innate or induced regulations of HIV/SIV infections within the host and/or host cells and tissues.

I - Regulation of HIV-1 infection at the level of the placenta barrier. Group leader: Elisabeth MENU

Our studies aim to investigate the mechanisms involved in the efficient control of in utero mother-to-child transmission (MTCT) of HIV-1. We have shown that placental trophoblast cells are not permissive to infection with cell-free virus and that the restriction occurs at the early steps of the viral replication cycle before the integration. This restriction is also observed at the level of the placental tissue, according to the lack of detection of productive infection in the absence of any exogenous activation. We are actually studying the mechanisms and the factors involved in this restriction. We have also shown that the passage of the virus through the trophoblast barrier after cell-to-cell infection is regulated by the cytokines of the placental environment, in particular the TNF-α whose expression is increased during co-infections. These findings might explain the increased risk of MTCT observed in a multicentric study, performed in collaboration with the " Centre Pasteur " of Cameroon and the PHPT ("Perinatal HIV Prevention Trial") group in Thailand. In this study, we found that the efficiency of MTCT preventive antiretroviral treatment might be partially explained by the impact of the therapy on placental cytokine expression. These observations are further supported by data showing that AZT is reducing the level of expression of placental TNF-αin vitro.

Overall, our research is contributing to a better understanding of multiple parameters involved in innate and/or therapeutic control of HIV-1 MTCT and, consequently, to the development and evaluation of new therapeutic strategies to prevent HIV-1 MTCT. For example, we are now studying the efficacy of viral entry inhibitors on cell-to-cell infection at the level of the placental barrier in vitro.

II - Regulation of HIV-1 replication and of the survival of infected thymocytes within the thymic environment. Group leader: Nicole ISRAEL.

Defects in both intrathymic and extrathymic pathways have been proposed to explain the progressive decline of CD4+ T cells in HIV-1 infection. Our study aims to identify mechanisms that impact HIV-1 infection and survival of infected cells within the thymic environment. We previously demonstrated that two cytokines (IL-7 and TNF) are critical for HIV-1 replication and survival of thymic target cells in vitro and that IL-7 favors X4 HIV-1 replication over R5 viruses in mature thymocytes and protects them from apoptosis. Immature thymocytes might also participate to the viral spreading in the context of an inflammatory TNF response, but they remain sensitive to virus-induced apoptosis, and consequently, their infection might contribute to a defect in T cell renewal. According to these findings, we studied the beneficial or adverse effects of IL7 therapy in macaques infected by SIV. We recently demonstrated that IL-7 is inducing a renewal of T cells in the animals despite its impact on X4-HIV-1 replication and on the survival of infected thymocytes. Our data are encouraging immunotherapy trials in humans with IL-7.

Recently, we also studied whether an alteration of thymic function is responsible or not for the low CD4+ T cell regeneration observed in some patients with undetectable viral load under antiretroviral therapy, presenting a high frequency of X4-HIV-1. We found that the sustained CD4+ lymphopenia observed in these patients is related to a persisting high level of T cell activation and of apoptosis rather to a defect in thymic function. These findings raise the question of the implication of X4-Gp120 on the persistent T cell activation.

Finally, we also showed recently that HIV-1, in particular R5 viruses are able to replicate at high level in thymic dendritic cells (DC). In addition, we found that HIV-1 is inducing thymic DC death as well as modifications of their phenotypes that might impact their functions, in particular in terms of negative selection of T cells.

III- Early host determinants of protection against AIDS in African Green Monkeys. Group leader: Michaela C.MULLER-TRUTWIN.

We are studying the infection by a simian lentivirus (SIV) in African Green monkeys (AGM) as a model for natural protection against AIDS. Our previous data revealed a high replication of SIVagm in these monkeys, both in blood and the intestine, indicating that virus replication is necessary but not sufficient to induce CD4 T cell depletion. In the same line, data of the literature show that the CD4 T cell depletion in HIV-infected humans is only indirectly associated with viral load, but directly with T cell activation levels. We have shown that chronically infected AGMs do not show increased T cell activation despite high viremia levels. By performing a kinetic study starting from day 1 post-infection in blood and lymph nodes, we demonstrate that AGMs are able to control T cell activation very early on after infection. This control is associated with a preferential induction of anti-inflammatory cytokines (TGF-β, IL-10) during the first week post-infection. The increase of TGF-β correlates with an early augmentation of FoxP3 expression and of CD25+ T cell frequencies in vivo. These findings indicate that regulatory CD25+ T cells might play a role in the control of T cell activation. Early interactions between dendritic cells and T cells might be determinant for these profiles. We are thus presently investigating the maturation profiles and function of dendritic cells and T lymphocytes during SIVagm infection ex vivo. These studies are conducted using classical (flow cytometry, real time PCR...) and global approaches (microarrays).

IV - Factors of natural resistance to HIV-1 infection. Group leader: Gianfranco PANCINO.

Our research aims to identify host factors involved in the control of HIV-1 infection. In collaboration with the Institut Pasteur of Hô Chi Minh City (Vietnam), Cambodia and Bangui (CAR), we studied the mechanisms of resistance to infection in intravenous drug users (IDU) or partners of HIV-1+ individuals who, despite repeated exposures to HIV-1, are not infected (exposed uninfected, EU). We previously reported a decreased susceptibility of EU peripheral blood mononuclear cells to in vitro infection by HIV-1 compared to control individuals. This relative resistance to infection was associated either to an inhibitory activity of CD8 T cells or to restrictions of viral replication in EU CD4 T lymphocytes. We identified three distinct restriction phenotypes that affect HIV-1 entry or post-entry steps of the viral replication cycle. Using an in vitro model for studying HIV-1 replication in primary macrophages, we also demonstrated that macrophage activation through FcγR, mimicking the stimulation by immune complexes, suppresses HIV-1 replication by blocking viral integration into the host genome. FcγR-mediated inhibition also affects other lentiviruses, including HIV-2 and SIV. In the context of these studies, we developed a bioluminescence imaging method allowing real-time quantification of HIV-1 gene expression in infected primary single living cells, in collaboration with the Dynamic Imaging Platform (Figure).

Our present work aims to identify the molecular mechanisms responsible for the restriction of viral replication observed in EU CD4 T cells as well as in activated macrophages. In parallel, we are studying the mechanisms of control of HIV-1 replication in individuals who do not have detectable plasma viral load, in spite of being infected for more than 10 years (HIV-controllers). Altogether these studies might contribute to the identification of new anti-HIV mechanisms and to new therapeutic and vaccine strategies.

V - Regulations of HIV infection by Innate Immunity. Group leader: Daniel SCOTT-ALGARA.

We are studying the role of the innate immunity, in particularly NK cells, in the control of HIV infection. In collaboration with the Institut Pasteur of Ho Chi Minh city, Vietnam and with G. Pancino's team, we previously reported an enhanced NK cell cytotoxic and cytokine in a cohort of Vietnamese intravascular drug users, which remained uninfected by HIV despite their high-risk exposure (EU). We are now analyzing the expression of NK cell markers that may characterize NK cell activation in EUs compared to HIV seronegative individuals and to HIV+ individuals in Vietnam using flow cytometry analysis and real time PCR (collaboration with P. Paul, Marseille). Our results demonstrate that NK cell subsets bearing particular activating and inhibitory receptors are abnormally elevated in some EU, suggesting a selective expansion of NK cell subsets that might represent an advantage for NK cells in better resolving early HIV infection in EU. We are now further investigating the impact of the cross talk between NK cells and dendritic cells (or macrophages) on the expansion of NK cell subsets to gain insights on the role of NK cells in controlling viral spreading and/or disease progression.

Photo

Real-time quantification of HIV-1 gene expression in infected single living cells.

A. Bioluminescent image superimposed with the bright field image showing photon emission of a macrophage after infection with HIV-1 particles bearing the luciferase gene. B. Real-Time imaging (up) and quantification (down) of rescue of viral gene expression in a latently infected primary CD4+ T cell after PHA stimulation (1, blue). HIV-1 replication in a pre-activated CD4+ T cell already emitting photons before stimulation (2, green).

Keywords: HIV/SIV, AIDS, African Green Monkey model, innate regulation, T cell activation, T cell regeneration, restriction factors


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