The requirement for interferon-gamma (IFN gamma) in resolution of an HSV-2 vaginal infection and the cellular sources of this cytokine in the vaginal mucosa were assessed. IFN gamma levels in vaginal secretions peaked on Days 2 and 5 following HSV-2 inoculation. Natural killer (NK) cell depletion greatly diminished the early production of IFN gamma but had no significant effect on the rate of virus clearance. CD4+ T cells were primarily responsible for the second peak of IFN gamma levels and neutralization of this IFN gamma beginning 3 days after virus inoculation delayed, but did not prevent, virus clearance from the vagina. HSV-2 persisted in mice depleted of both CD4+ and CD8+ T cells while clearance was delayed in CD4+, but not CD8+ T cell-depleted mice, demonstrating the T cell dependence and predominant role of CD4+ T cells in resolution of the infection. Together, these data suggest that IFN gamma is not essential for virus clearance but plays an important role in enhancing T cell-mediated clearance mechanisms. The implication of these results is that IFN gamma produced locally in the genital tract enhances virus clearance and may ultimately be important for reducing the amount of virus available to infect sensory ganglia.

Human metapneumovirus (hMPV), a member of the family Paramyxoviridae, is a leading cause of lower respiratory tract infections in children, the elderly, and immunocompromised patients. Virus-and host-specific mechanisms of pathogenesis and immune protection are not fully understood. By an intranasal inoculation model, we show that hMPV-infected BALB/c mice developed clinical disease, including airway obstruction and hyperresponsiveness (AHR), along with histopathologic evidence of lung inflammation and viral replication. hMPV infection protected mice against subsequent viral challenge, as demonstrated by undetectable viral titers, lack of body weight loss, and a significant reduction in the level of lung inflammation. No crossprotection with other paramyxoviruses, such as respiratory syncytial virus, was observed. T-lymphocyte depletion studies showed that CD4 ؉ and CD8 ؉ T cells cooperate synergistically in hMPV eradication during primary infection, but CD4؉ more than CD8 ؉ T cells also enhanced clinical disease and lung pathology. Concurrent depletion of CD4؉ and CD8 ؉ T cells completely blocked airway obstruction as well as AHR. Despite impaired generation of neutralizing anti-hMPV antibodies in the absence of CD4؉ T cells, mice had undetectable viral replication after hMPV challenge and were protected from clinical disease, suggesting that protection can be provided by an intact CD8 ؉ T-cell compartment. Whether these findings have implications for naturally acquired human infections remains to be determined.

Dengue (DEN) is a mosquito-borne disease caused by four DENV serotypes (DENV-1, -2, -3, and -4) that have no treatments or vaccines. Primary infection with one DENV usually leads to acute illness followed by lifelong homotypic immunity, but susceptibility to infection by the other three DENVs remains. Therefore, a vaccine needs to protect from all four DENVs simultaneously. To date a suitable animal model to mimic systemic human illness exists only for DENV-2 in immunocompromised mice using passaged viruses; however, models are still needed for the remaining serotypes. This study describes establishment of a lethal systemic DENV-3 infection model with a human isolate in immunocompromised mice and is the first report of lethal infection by a nonadapted clinical DENV isolate without evidence of neurological disease. Our DENV-3 model provides a relevant platform to test DEN vaccines and antivirals.

The effects of 17β-estradiol (E2) on immune function have been extensively reported. The effects are dependent on concentration and duration of exposure and potential differences in signaling between the known E2 receptors, estrogen receptors (ER) α and ERβ. Through the use of ER-deficient mice, we and others have begun to demonstrate the role of the two known receptors in modulating immune functional activities. Previous studies have shown that cells of the innate immune system have altered function (bactericidal capacity) and patterns of cytokine expression (increased proinflammatory cytokine expression) through amelioration of ERα signaling. In this study, we extend these studies to analysis of T cell differentiation and proliferation in APC-dependent and APC-independent in vitro assay systems. Our results demonstrate that ERα deficiency in splenic macrophages, but not CD11c+ splenic dendritic cells pulsed with OVA significantly enhances proliferative responses and IFN-γ production by transgenic OVA peptide-specific (OT-II) CD4+ T cells when compared with Ag-pulsed APC from wild-type littermates. The addition of E2 in this culture system did not significantly affect the production of IFN-γ. In addition, when purified CD4+ T cells from ERα-deficient and wild-type littermates were stimulated with anti-CD3/CD28 Ab in the absence of E2, there were no significant differences in IFN-γ or IL-4 production. However, the addition of E2 significantly increased IL-4 secretion, as well as increased GATA-3 mRNA levels from ERα-replete CD4+ T cells, while this effect was abrogated in ERα-deficient CD4+ T cells.

Large numbers of polymorphonuclear leukocytes (PMNs) infiltrated the murine vaginal mucosa within 24 h after intravaginal inoculation with an attenuated strain of herpes simplex virus type 2 (HSV-2). The role of these cells in resolution of a primary genital infection and in protection of HSV-immune animals against challenge with a fully virulent HSV-2 strain was investigated. Depletion of greater than 95% of the PMNs at the vaginal mucosal surface prior to intravaginal inoculation with an attenuated HSV-2 strain resulted in significantly higher virus titers on days 3 to 7 but only slightly delayed resolution of the primary genital infection. These results suggest that neutrophils helped control the infection but that other immune mechanisms ultimately cleared the virus. Interestingly, depletion of PMNs from HSV-immune mice prior to challenge with a fully virulent HSV-2 strain resulted in a rise in virus titers to levels comparable to those of nonimmune mice and a more pronounced diminution of virus clearance from the vaginal mucosa despite the presence of HSV-specific B and T cells. Levels of gamma interferon (IFN-γ) and HSV-specific antibody were comparable in neutrophil-depleted and control-treated immune mice following HSV-2 challenge, suggesting that RB6-8C5 treatment did not impair T- and B-cell function. Therefore, these results suggest that neutrophils play a role in limiting and clearing HSV-2 vaginal infections and that they are, in association with HSV-specific B and T cells, an important component in immune protection of the vaginal mucosa.

The T-cell-mediated resolution of herpes simplex virus type 2 (HSV-2) genital infections is not fully understood. In these studies, the mechanisms by which CD8 ؉ T cells clear virus from the genital epithelium were examined. Ovalbumin (OVA)-specific CD8 ؉ T cells from OT-I transgenic mice cleared a thymidine kinase-deficient, ovalbumin-expressing HSV-2 virus (HSV-2 tk ؊ OVA) from the genital epithelium of recipient mice, and clearance was abrogated by in vivo neutralization of gamma interferon (IFN-␥). Further, CD8؉ OT-I T cells deficient in IFN-␥ were unable to clear HSV-2 tk ؊ OVA from the vaginal epithelium. The requirement for cytolytic mechanisms in HSV-2 tk ؊ OVA clearance was tested in radiation chimeras by adoptive transfer of wild-type or perforin-deficient OT-I T cells to irradiated Fas-defective or wild-type recipients. Although a dramatic decrease in viral load was observed early after challenge with HSV-2 tk ؊ OVA, full resolution of the infection was not achieved in recipients lacking both perforin-and Fas-mediated cytolytic pathways. These results suggest that IFN-␥ was responsible for an early rapid decrease in HSV-2 virus titer. However, either perforin-or Fas-mediated cytolytic mechanisms were required to achieve complete clearance of HSV-2 from the genital epithelium.Herpes simplex virus type 2 (HSV-2) infects epithelial cells in the genital mucosa, spreads to the sensory ganglia via retrograde transport, and establishes a lifelong latent infection in sensory neurons (50). The virus periodically reactivates and descends sensory neurons via anterograde transport, resulting in development of recurrent lesions at or near the site of primary infection or in shedding of infectious virus in the absence of disease symptoms. The primary and recurrent lesions of immunocompetent individuals are generally self limiting and are resolved primarily by cell-mediated immune mechanisms. Recurrent disease is less well controlled in immunocompromised individuals, resulting in more frequent recurrences and sometimes severe mucocutaneous disease manifestations. Studies of HSV infection in human immunodeficiency virus (HIV)-infected individuals suggested that the severity of HSV disease could be inversely correlated with the number of HSV-specific CD8 ϩ T cells (39). Studies of recurrent HSV lesions in immunocompetent humans have demonstrated the early infiltration of CD4 ϩ T cells and macrophages, local production of IFN-␥, and late arrival of CD8ϩ T cells at the site of HSV infection. Both CD4 ϩ and CD8 ϩ T lymphocytes capable of IFN-␥ secretion and HSVspecific cytolysis have been isolated from human herpetic lesions (10) and clearance of infectious virus, and resolution of lesions has been correlated with the detection of HSV-specific cytolytic T-lymphocyte activity (10, 22-23). However, the role for these cytolytic and noncytolytic immune mechanisms in resolution of HSV-2 genital infections is not well understood.Murine models of HSV-2 genital infection have also demonstrated the importance of cell-mediated imm...

A murine model of genital infection with a thymidine kinase-deficient (tk-) strain of herpes simplex virus type 2 (HSV-2) was utilized to examine the development of the local T cell response in the genital mucosa and draining genital lymph nodes (gLN). HSV-specific cytokine-secreting T cells were detected in the gLN 4 days postintravaginal inoculation but not in the urogenital tract or spleen until 5 days postinoculation, suggesting the cellular immune response originates in the gLN. More CD4+ than CD8+ gLN T cells were detected by flow cytometric analysis following primary vaginal inoculation and the majority of HSV-specific gLN T cells detected by ELISPOT were CD4+ and Th1-like based on secretion of IFN gamma and not IL-4 or IL-5. A similar population of HSV-specific memory T cells persisted in the genital tract 2 months following HSV-2 tk- genital inoculation. These data suggest that the urogenital cellular immune response elicited in mice following genital inoculation with HSV-2 tk- is predominantly CD4+ and Th1-like, resembling that observed in humans. The results of this study are important for the rational design of vaccines capable of inducing protective immunity in the genital tract.

Lassa fever (LF) is a zoonotic disease associated with acute and potentially fatal hemorrhagic illness caused by the Lassa virus (LASV), a member of the family Arenaviridae. It is generally assumed that a single infection with LASV will produce life-long protective immunity. This suggests that protective immunity induced by vaccination is an achievable goal and that cell-mediated immunity may play a more important role in protection, at least following natural infection. Seropositive individuals in endemic regions have been shown to have LASV-specific T cells recognizing epitopes for nucleocapsid protein (NP) and glycoprotein precursor (GPC), suggesting that these will be important vaccine immunogens. The role of neutralizing antibodies in protective immunity is still equivocal as recent studies suggest a role for neutralizing antibodies. There is extensive genetic heterogeneity among LASV strains that is of concern in the development of assays to detect and identify all four LASV lineages. Furthermore, the gene disparity may complicate the synthesis of effective vaccines that will provide protection across multiple lineages. Non-human primate models of LASV infection are considered the gold standard for recapitulation of human LF. The most promising vaccine candidates to date are the ML29 (a live attenuated reassortant of Mopeia and LASV), vesicular stomatitis virus (VSV) and vaccinia-vectored platforms based on their ability to induce protection following single doses, high rates of survival following challenge, and the use of live virus platforms. To date no LASV vaccine candidates have undergone clinical evaluation.