RESUMEN
The induction of local T helper type 1 (Th1)-mediated cellular immunity is crucial for resistance of mice to genital infection by the obligate intracellular bacterium Chlamydia trachomatis. We tested the hypothesis that the route of immunization that elicits relatively high numbers of chlamydia-specific, gamma interferon (IFN-gamma)-secreting T lymphocytes (ISTLs) in the genital tract would induce optimal protective immunity against reinfection. Female BALB/c mice were infected intravaginally (i.v.), intranasally (i.n.), orally (p.o.), or subcutaneously (s.c.) with C. trachomatis. At days 7, 14, 21, and 28 postinfection, T cells isolated from the genital tract tissues were restimulated with chlamydial antigen in vitro, and the amounts of IFN-gamma induced were measured by a sandwiched enzyme-linked immunosorbent assay method. At day 7 postinfection, i.n.- and i.v.-immunized mice had high levels of chlamydia-specific ISTLs in their genital tracts (203.58 +/- 68.1 and 225.5 +/- 12.1 pg/ml, respectively). However, there were no detectable ISTLs in the genital tracts of p.o.- or s.c.-infected mice. When preinfected mice were challenged i.v. 70 days later, animals preexposed by the i.n. route were highly resistant to reinfection, with greatly reduced chlamydial burden, and suffered an attenuated infection that resolved by day 6 postchallenge. Animals preexposed by the i.v. route were modestly protected, whereas p.o. and s.c. groups were indistinguishable in this regard from control mice. The resistance of i.n.-immunized mice (and to some extent the i.v.-exposed mice) to reinfection was associated with early appearance (within 24 h) of high levels of genital ISTLs compared with mice preinfected by other routes. Furthermore, although i.n. and i.v.-immunized mice had comparable levels of chlamydia-specific immunoglobulin A (IgA) antibodies in their vaginal washes, the levels of IgG2a were four- sixfold higher in i.n.-immunized mice than in any of the other groups. The results suggested that immunization routes that foster rapid induction of vigorous genital mucosal cell-mediated immune (CMI) effectors (e.g., IFN-gamma), the CMI-associated humoral effector, IgG2a, and to some extent secretory IgA produce protective immunity against chlamydial genital infection. Therefore, i.n. immunization is a potential delivery route of choice in the development of a vaccine against Chlamydia.
Asunto(s)
Infecciones por Chlamydia/inmunología , Chlamydia trachomatis/inmunología , Enfermedades de los Genitales Femeninos/inmunología , Interferón gamma/metabolismo , Linfocitos T/inmunología , Administración Intranasal , Administración Oral , Animales , Anticuerpos Antibacterianos/análisis , Células Cultivadas , Infecciones por Chlamydia/fisiopatología , Modelos Animales de Enfermedad , Femenino , Enfermedades de los Genitales Femeninos/fisiopatología , Células HeLa , Humanos , Inmunidad Mucosa , Inmunoglobulina A Secretora/análisis , Inmunoglobulina G/análisis , Inyecciones Intravenosas , Inyecciones Subcutáneas , Ratones , Ratones Endogámicos BALB C , Linfocitos T/metabolismo , Linfocitos T/microbiología , Vagina/inmunologíaRESUMEN
Type 1 CD4+-T-cell-mediated immunity is crucial for the resolution of chlamydial infection of the murine female genital tract. Previous studies demonstrating a correlation between CD4+-T-cell-mediated inhibition of chlamydial growth and gamma interferon (IFN-gamma)-mediated induction of nitric oxide synthase suggested a potential role for the nitric oxide (NO) effector pathway in the clearance of Chlamydia from genital epithelial cells by the immune system. To clarify the role of this pathway, the growth levels of Chlamydia trachomatis organisms in normal (iNOS+/+) mice and in genetically engineered mice lacking the inducible nitric oxide synthase (iNOS) gene (iNOS-/- mice) were compared. There was no significant difference in the course of genital chlamydial infections in iNOS+/+ and iNOS-/- mice as determined by recovery of Chlamydia organisms shed from genital epithelial cells. Dissemination of Chlamydia to the spleen and lungs occurred to a greater extent in iNOS-/- than in iNOS+/+ mice, which correlated with a marginal increase in the susceptibility of macrophages from iNOS-/- mice to chlamydial infection in vitro. However, infections were rapidly cleared from all affected tissues, with no clinical signs of disease. The finding of minimal dissemination in iNOS-/- mice suggested that activation of the iNOS effector pathway was not the primary target of IFN-gamma during CD4+-T-cell-mediated control of chlamydial growth in macrophages because previous reports demonstrated extensive and often fatal dissemination of Chlamydia in mice lacking IFN-gamma. In summary, these results indicate that the iNOS effector pathway is not required for elimination of Chlamydia from epithelial cells lining the female genital tract of mice although it may contribute to the control of dissemination of C. trachomatis by infected macrophages.
Asunto(s)
Infecciones por Chlamydia/inmunología , Chlamydia trachomatis , Enfermedades de los Genitales Femeninos/inmunología , Óxido Nítrico Sintasa/fisiología , Animales , Femenino , Interferón gamma/fisiología , Macrófagos Peritoneales/microbiología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Óxido Nítrico/biosíntesis , Óxido Nítrico Sintasa/genética , Óxido Nítrico Sintasa de Tipo IIRESUMEN
It was previously shown that murine T cell clones could inhibit the intracellular growth of the mouse strain of Chlamydia trachomatis by cytokine-mediated induction of the inducible nitric oxide synthase (iNOS) system in epithelial cells, an effect enhanced by direct epithelial-T cell interaction via specific adhesion molecules. These findings and other recent reports showing that human mucosal epithelial cells secrete nitric oxide (NO) via iNOS expression would suggest that mucosal epithelial-derived NO may be involved in mucosal defense against Chlamydia and other pathogens that infect epithelial cells. As an initial approach to investigating whether NO contributes to chlamydial control in humans, the present studies evaluated the susceptibility of human isolates of C. trachomatis to NO delivered by chemical donors or via induction of the epithelial iNOS system by a cytokine-secreting T cell clone. It was found that a chlamydial-specific, cytokine-secreting, murine T lymphocyte clone (clone 2.14-0) could inhibit the intraepithelial growth of human strains of Chlamydia trachomatis (serovar E and H, and Lymphogranuloma venerum type L2) via the iNOS pathway when the clone was co-cultured with chlamydial-infected epithelial cells. Furthermore, treatment of infected epithelial cells with 50 microM of the NO donor, S-nitroso-L-glutathione, resulted in significant inhibition (approximately 70%) of chlamydial multiplication, while the NO scavenger, myoglobin plus ascorbate, could reverse the effect, demonstrating that NO could directly inhibit human strains of Chlamydia. The results are consistent with the hypothesis that the IFN-gamma-inducible iNOS pathway can contribute to chlamydial control in humans.
Asunto(s)
Chlamydia trachomatis/efectos de los fármacos , Chlamydia trachomatis/inmunología , Óxido Nítrico/farmacología , Animales , Antibacterianos/farmacología , Línea Celular , Infecciones por Chlamydia/inmunología , Infecciones por Chlamydia/metabolismo , Infecciones por Chlamydia/prevención & control , Chlamydia trachomatis/crecimiento & desarrollo , Epitelio/inmunología , Epitelio/metabolismo , Femenino , Humanos , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Óxido Nítrico/metabolismo , Óxido Nítrico Sintasa/metabolismo , Linfocitos T/inmunología , Linfocitos T/metabolismoRESUMEN
T cell-mediated immunity against Chlamydia in mice is mediated at least in part by T cell-derived interferon-gamma (IFN-gamma) induction of the nitric oxide synthase (iNOS) system in infected epithelial cells. Although IFN-gamma alone could stimulate nitric oxide (NO) production from epithelial cells and inhibit the intracellular growth of Chlamydia, the effectiveness was less than when infected epithelial cells were co-cultured with IFN-gamma-producing T cell clones. In co-cultures containing T cells and infected epithelial cells, additional NO produced by activated T cells could augment chlamydial killing; however, T cell-derived NO was insufficient to account for the total NO present in the co-culture and therefore could not explain the dramatic increase in chlamydial inhibition under those conditions. To determine whether direct cell-to-cell interaction involving adhesion molecules was involved in increased NO induction, the ability of neutralizing monoclonal antibodies directed against intercellular adhesion molecule type 1 (ICAM-1) and leukocyte function antigen-1 (LFA-1) to suppress NO production and lower intracellular chlamydial inhibition was investigated. It was found that monoclonal antibodies against ICAM-1/LFA-1 could significantly reduce the capacity of a protective CD4+ type 1 (Thl) clone (clone 2.14-0) to inhibit the intracellular growth of the C. trachomatis agent of mouse pneumonitis (MoPn). The suppression of the anti-chlamydial action of the clone by antibodies correlated with approximately 50% decrease in NO production. Also, paraformaldehyde-fixed clone 2.14-0 could enhance NO induction and chlamydial inhibition mediated by IFN-gamma, and this effect could be reversed by anti-ICAM-1/LFA-1 antibodies. The results indicated that epithelial-T cell interaction via adhesion molecules enhances NO production and increased chlamydial inhibition by IFN-gamma-secreting T cells.