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International audience; Objective: Mucosae are the gateway for many pathogens. However, few vaccines have been developed to specifically target mucosal immunity. Recent studies in the laboratory evidenced local expression of IL-7 in the acutely SIV-infected intestinal mucosa and IAV-infected lung in macaques or mice, respectively. This overexpression led to chemokine production and infiltration of immune cells into these mucosae. In addition, systemic injection of IL-7 to macaques rapidly stimulates both the production of chemokines and immune cells homing into the mucosa. These results prompted us to study if IL-7 could be used as a mucosal vaccine adjuvant.Methods: Mice were intratracheally treated with IL-7 or PBS, then immunized by the same route against diphtheria toxoid (DT) or inactivated influenza virus (IAVi) two days later. DT immunized-mice were sacrificed at day 14 while those immunized with IAVi were infected with virulent IAV at day 14 and sacrificed at day 29. We measured antigen-specific antibody responses (anti-DT & anti-IAV) in both bronchoalveolar lavages (BAL) and sera, as well as chemokine expressions in lung tissue, by ELISA. The immune cell infiltration into the pulmonary mucosa was evaluated by immunochemistry on lung sections. The effectiveness of IL-7-adjuvanted vaccine in providing protection against IAV pathology was assessed by daily monitoring of animal body weight.Results: Intratracheal administration of IL-7 stimulated the production of pro-inflammatory chemokines in the lung parenchyma, leading to massive infiltration of immune cells found to be mainly organized in lymphoid aggregates. Moreover, intratracheal administration of IL-7 before primo-immunization allowed antigens-specific IgAs and IgGs production in the pulmonary mucosa. These effects were not observed in control mice vaccinated without IL-7 administration. In addition, only IL-7-treated immunized mice were protected against influenza pathology. This protection seems to be related to a high level of IAV-specific antibodies in BAL and lymphoid aggregate appearance in the pulmonary mucosa.Conclusion: By attracting immune cells into mucosae, local IL-7 administration prepares the mucosal immune system, gathering conditions that result in enhanced antigen-specific pulmonary immune responses upon antigenic stimulation. Hence, IL-7 appears as a mucosal adjuvant able to increase mucosal antibody responses, an important immune arm implicated in the protection against most mucosal infections.

International audience; Objective: HIV-1 and HIV-2 differ notably in their epidemiology, with worldwide HIV-1 spread and HIV-2 mainly confined to West Africa. Natural killer (NK) cells are critical antiviral effectors of the immune system; however, limited information is available about these innate effector cells during HIV-2 infection.Method: In this study, 24 untreated HIV-2-infected patients were analyzed and compared with 21 long-term nonprogressor and 10 controller HIV-1 patients, and healthy donors. Extensive phenotype and functional NK-cell characteristics, as well as ligands of activating NK receptors involved in NK lysis were determined by flow cytometry.Results: We report in HIV-2 patients a very significant reduced expression of the activating NKp30 receptor (P < 0.0001) on NK cells, much higher than observed in HIV-1 patients. The impaired expression of NKp30 is correlated negatively with HLA-DR (r = -0.5970; P = 0.0002), and positively with both NKG2A (r = 0.5324; P < 0.0001) and Siglec-7 (r = 0.5621; P = 0.0004). HIV-2 patients with NKp30 NK cells displayed overproduction of IFN-γ (P < 0.0001) associated with impaired cytolytic function when tested against target cells expressing surface B7-H6. This cellular ligand of NKp30 is strongly detectable as a surface molecule on CD4 T cells infected by HIV-2.Conclusion: Altogether, our data suggested that the defective expression of NKp30 may be induced by the chronic engagement of this receptor by B7-H6 expressed on HIV-2-infected target cells. This represents a novel mechanism by which the chronic ligand exposure by the viral environment may subvert NK-cell-mediated function to establish persistent HIV-2 infection.

The low pathogenicity and replicative potential of HIV-2 are still poorly understood. We investigated whether HIV-2 reservoirs might follow the peculiar distribution reported in models of attenuated HIV-1/SIV infections, i.e. limited infection of central-memory CD4 T lymphocytes (TCM). Antiretroviral-naive HIV-2 infected individuals from the ANRS-CO5 (12 non-progressors, 2 progressors) were prospectively included. Peripheral blood mononuclear cells (PBMCs) were sorted into monocytes and resting CD4 T-cell subsets (naive [TN], central- [TCM], transitional- [TTM] and effector-memory [TEM]). Reactivation of HIV-2 was tested in 30-day cultures of CD8-depleted PBMCs. HIV-2 DNA was quantified by real-time PCR. Cell surface markers, co-receptors and restriction factors were analyzed by flow-cytometry and multiplex transcriptomic study. HIV-2 DNA was undetectable in monocytes from all individuals and was quantifiable in TTM from 4 individuals (median: 2.25 log10 copies/106 cells [IQR: 1.99-2.94]) but in TCM from only 1 individual (1.75 log10 copies/106 cells). HIV-2 DNA levels in PBMCs (median: 1.94 log10 copies/106 PBMC [IQR = 1.53-2.13]) positively correlated with those in TTM (r = 0.66, p = 0.01) but not TCM. HIV-2 reactivation was observed in the cells from only 3 individuals. The CCR5 co-receptor was distributed similarly in cell populations from individuals and donors. TCM had a lower expression of CXCR6 transcripts (p = 0.002) than TTM confirmed by FACS analysis, and a higher expression of TRIM5 transcripts (p = 0.004). Thus the low HIV-2 reservoirs differ from HIV-1 reservoirs by the lack of monocytic infection and a limited infection of TCM associated to a lower expression of a potential alternative HIV-2 co-receptor, CXCR6 and a higher expression of a restriction factor, TRIM5. These findings shed new light on the low pathogenicity of HIV-2 infection suggesting mechanisms close to those reported in other models of attenuated HIV/SIV infection models.

International audience; Background: Despite considerable research efforts, mucosal immunity remains particularly difficult to stimulate through vaccines. Systemic injection of IL-7 stimulates chemokines-induced recruitment of circulating T-cells into mucosae.Methods: The optimal dose of IL-7 to be sprayed on mucosal surface was defined on 14 rhesus macaques. On mucosal biopsies collected after IL-7 administration, we quantified local transcription of 19 chemokines by qRT-PCR and cell infiltration by immunochemistry plus image analysis. Six macaques were immunized with antigens (DT and the HIV-1 gp41-P1 peptide) applied directly on the vaginal mucosa, two days after either IL-7 or PBS administration. The immunizations were repeated thrice, four months apart, and the macaques were euthanized 2 weeks after the last immunization. Antigen-specific IgA and IgG productions were quantified in vaginal secretions by ELISA. Antigen-specific plasma cells were detected by reverse immunohistochemistry in tissue, and by B-cell ELISPOT in PBMCs.Results: A significant overexpression of twelve chemokines was observed 48 hours after mucosal administration of 10µg of IL-7. Subsequently, mDC, macrophage, NK, B- and T-cell numbers significantly raised in the IL-7-treated mucosae, suggesting massive chemokine-driven infiltration. Administration of antigens led to a stronger mucosal immune response in the IL-7-treated macaques as compared to animals immunized with antigens alone. Robust production of antigen-specific IgAs and IgGs was detected in vaginal secretions. The immunizations repeated thrice sustained mucosal specific immune responses. Antigen-specific-antibody secreting cells were recovered from PBMC and more DT-specific plasma cells were found in the vaginal mucosae (IgA isotype) and the draining lymph nodes (IgG isotype) of IL-7-treated macaques. Tertiary lymphoid organs were observed in vaginal mucosae from IL-7-treated macaques only.Conclusions: Pre-treatment by non-traumatic vaginal administration of IL-7 (10µg by spray), allows for the development of a strong mucosal immune response in macaques following subsequent mucosal vaccination, through local chemokine expression and the recruitment of immune cells in the vaginal mucosa. The mucosal localization of IgA-specific plasma cells argues for their main contribution in the high levels of specific-IgAs evidenced in the vaginal secretions. These data suggest that IL-7 could be used as a mucosal adjuvant to elicit vaginal antibody response, the most promising way to confer protection to numerous STD.

© 2012 Albuquerque et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited., Forkhead box N1 (FOXN1) is a transcription factor crucial for thymic epithelium development and prevention of its involution. Investigation of a patient with a rare homozygous FOXN1 mutation (R255X), leading to alopecia universalis and thymus aplasia, unexpectedly revealed non-maternal circulating T-cells, and, strikingly, large numbers of aberrant doublenegative ab T-cells (CD4negCD8neg, DN) and regulatory-like T-cells. These data raise the possibility that a thymic rudiment persisted, allowing T-cell development, albeit with disturbances in positive/negative selection, as suggested by DN and FoxP3+ cell expansions. Although regulatory-like T-cell numbers normalized following HLA-mismatched thymic transplantation, the abDN subset persisted 5 years post-transplantation. Involution of thymus allograft likely occurred 3 years post-transplantation based on sj/bTREC ratio, which estimates intrathymic precursor T-cell divisions and, consequently, thymic explant output. Nevertheless, functional immune-competence was sustained, providing new insights for the design of immunological reconstitution strategies based on thymic transplantation, with potential applications in other clinical settings., This work was supported by grants from ‘‘Fundação para a Ciência e a Tecnologia’’ (FCT) and by ‘‘Programa Operacional Ciência e Inovação 2010’’ (POCI2010) to RMMV and AES. ASA received a scholarship from FCT. Thymus transplantation performed at the Duke University Medical Center, Durham, USA was financially supported by the Portuguese National Health Services. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Human immunodeficiency virus (HIV) replication and T cell proliferation was investigated in situ by a PCR based analysis of individual microdissected splenic white pulps. Founder effects, revealed by an exquisite compartmentalization of HIV genotypes and T cells, indicated the recruitment of latently infected CD4+ T cells through highly localized antigen presentation, rather than the infection of CD4+ T lymphoblasts by blood borne virus or immune complexes. HIV infected white pulps could be infiltrated by HIV specific cytotoxic T lymphocytes, so implicating them in CD4+ T cell destruction in vivo. Together these data describe an iterative and deleterious mechanism of antigen driven T cell recruitment and activation, HIV replication and spread, with consequent destruction of the newly infected cells.

Thymic function decreases progressively with age but may be boosted in certain circumstances. We questioned whether heart transplantation was such a situation and whether thymic function was related to the onset of rejection. Twenty-eight antithymocyte globulin-treated heart transplant recipients were included. Patients diagnosed for an antibody-mediated rejection on endomyocardial biopsy had a higher proportion of circulating recent thymic emigrant CD4+ T cells and T cell receptor excision circle levels than other transplanted subjects. Thymus volume and density, assessed by computed tomography in a subset of patients, was also higher in patients experiencing antibody-mediated rejection. We demonstrate that thymic function is a major determinant of onset of antibody-mediated rejection and question whether thymectomy could be a prophylactic strategy to prevent alloimmune humoral responses.

Forkhead box N1 (FOXN1) is a transcription factor crucial for thymic epithelium development and prevention of its involution. Investigation of a patient with a rare homozygous FOXN1 mutation (R255X), leading to alopecia universalis and thymus aplasia, unexpectedly revealed non-maternal circulating T-cells, and, strikingly, large numbers of aberrant double-negative αβ T-cells (CD4negCD8neg, DN) and regulatory-like T-cells. These data raise the possibility that a thymic rudiment persisted, allowing T-cell development, albeit with disturbances in positive/negative selection, as suggested by DN and FoxP3+ cell expansions. Although regulatory-like T-cell numbers normalized following HLA-mismatched thymic transplantation, the αβDN subset persisted 5 years post-transplantation. Involution of thymus allograft likely occurred 3 years post-transplantation based on sj/βTREC ratio, which estimates intrathymic precursor T-cell divisions and, consequently, thymic explant output. Nevertheless, functional immune-competence was sustained, providing new insights for the design of immunological reconstitution strategies based on thymic transplantation, with potential applications in other clinical settings.