Hong Bai

IL-17/Th17 Promotes Type 1 T Cell Immunity against Pulmonary Intracellular Bacterial Infection through Modulating Dendritic Cell Function

Although their contribution to host defense against extracellular infections has been well defined, IL-17 and Th17 are generally thought to have limited impact on intracellular infections. In this study, we investigated the role and mechanisms of IL-17/Th17 in host defense against Chlamydia muridarum, an obligate intracellular bacterium, lung infection. Our data showed rapid increase in IL-17 production and expansion of Th17 cells following C. muridarum infection and significant detrimental impact of in vivo IL-17 neutralization by anti-IL-17 mAb on disease course, immune response, and dendritic cell (DC) function. Specifically, IL-17-neutralized mice exhibited significantly greater body weight loss, higher organism growth, and much more severe pathological changes in the lung compared with sham-treated control mice. Immunological analysis showed that IL-17 neutralization significantly reduced Chlamydia-specific Th1 responses, but increased Th2 responses. Interestingly, the DC isolated from IL-17-neutralized mice showed lower CD40 and MHC II expression and IL-12 production, but higher IL-10 production compared with those from sham-treated mice. In two DC-T cell coculture systems, DC isolated from IL-17-neutralized mice induced higher IL-4, but lower IFN-γ production by Ag-specific T cells than those from sham-treated mice in cell priming and reaction settings. Adoptive transfer of DC isolated from IL-17-neutralized mice, unlike those from sham-treated mice, failed to protect the recipients against challenge infection. These findings provide in vivo evidence that IL-17/Th17 plays an important role in host defense against intracellular bacterial infection, and suggest that IL-17/Th17 can promote type 1 T cell immunity through modulating DC function.

IL-17A synergizes with IFN-γ to upregulate iNOS and NO production and inhibit chlamydial growth.

IFN-γ-mediated inducible nitric oxide synthase (iNOS) expression is critical for controlling chlamydial infection through microbicidal nitric oxide (NO) production. Interleukin-17A (IL-17A), as a new proinflammatory cytokine, has been shown to play a protective role in host defense against Chlamydia muridarum (Cm) infection. To define the related mechanism, we investigated, in the present study, the effect of IL-17A on IFN-γ induced iNOS expression and NO production during Cm infection in vitro and in vivo. Our data showed that IL-17A significantly enhanced IFN-γ-induced iNOS expression and NO production and inhibited Cm growth in Cm-infected murine lung epithelial (TC-1) cells. The synergistic effect of IL-17A and IFN-γ on Chlamydia clearance from TC-1 cells correlated with iNOS induction. Since one of the main antimicrobial mechanisms of activated macrophages is the release of NO, we also examined the inhibitory effect of IL-17A and IFN-γ on Cm growth in peritoneal macrophages. IL-17A (10 ng/ml) synergizes with IFN-γ (200 U/ml) in macrophages to inhibit Cm growth. This effect was largely reversed by aminoguanidine (AG), an iNOS inhibitor. Finally, neutralization of IL-17A in Cm infected mice resulted in reduced iNOS expression in the lung and higher Cm growth. Taken together, the results indicate that IL-17A and IFN-γ play a synergistic role in inhibiting chlamydial lung infection, at least partially through enhancing iNOS expression and NO production in epithelial cells and macrophages.

Less inhibition of interferon‐γ to organism growth in host cells may contribute to the high susceptibility of C3H mice to Chlamydia trachomatis lung infection

T‐helper‐1‐like cytokine response and cell‐mediated immunity have been shown to be critical in host resistance to Chlamydia trachomatis infection. Using a murine pneumonia model, we compared the susceptibility of C3H/HeN (C3H) and C57BL/6 mice to C. trachomatis mouse pneumonitis (MoPn) infection. C3H mice exhibited significantly higher mortality, greater organism growth and much more severe pathological changes in the lung compared with C57BL/6 mice. However, the pattern of adaptive immune responses including organism‐specific delayed‐type hypersensitivity, antibody responses and cytokine [interferon‐γ (IFN‐γ), interleukin‐12 (IL‐12), IL‐4, IL‐10 and tumour necrosis factor α] production by spleen and local draining lymph node cells in these two strains of mice appeared comparable during the process of infection. Interestingly, MoPn growth in the cultured ex vivo macrophages from C3H mice was found to be significantly less inhibited by the exogenous IFN‐γ present in the culture compared to C57BL/6 mice. The lower inhibition of MoPn growth in C3H mice was associated with significantly lower nitric oxide production by the infected macrophages following IFN‐γ stimulation. The data suggest that the cellular events downstream of cytokine production in chlamydia host cells may be important in determining the different susceptibility of hosts to chlamydial infection.

CD8α+ and CD8α- DC subsets from BCG-infected mice inhibit allergic Th2-cell responses by enhancing Th1-cell and Treg-cell activity respectively.

The hygiene hypothesis has suggested an inhibitory effect of infections on allergic diseases, but the related mechanism remains unclear. We recently reported that DCs played a critical role in Mycobacterium bovis Bacille Calmette–Guérin (BCG)‐mediated inhibition of allergy, which depended on IL‐12 and IL‐10‐related mechanisms. Here, we tested the hypothesis that BCG infection could modulate the function of DC subsets, which might in turn inhibit allergic responses through different mechanisms. We sorted CD8α+ and CD8α− DCs from BCG‐infected mice and tested their ability to modulate Th2‐cell responses to ovalbumin (OVA) using in vitro and in vivo approaches. We found that both DC subsets could inhibit the allergic Th2‐cell response in both a DC:T‐cell co‐culture system and after adoptive transfer. These subsets exhibited different co‐stimulatory marker expression and cytokine production patterns and were different in inducing Th1 and Treg cells. Specifically, we found that CD8α+ DCs produced higher IL‐12, inducing higher Th1 cell response, while CD8α− DCs expressed higher ICOS‐L and produced higher IL‐10, inducing CD4+CD25+FoxP3+Treg cells with IL‐10 production and membrane‐bound TGF‐β expression. The finding suggests that one infection may inhibit allergy by both immune deviation and regulation mechanisms through modulation of DC subsets.

CD8+ DC, but Not CD8−DC, Isolated from BCG-Infected Mice Reduces Pathological Reactions Induced by Mycobacterial Challenge Infection

Background Tuberculosis is a mycobacterial infection causing worldwide public health problems but the available vaccine is far from ideal. Type-1 T cell immunity has been shown to be critical for host defence against tuberculosis infection, but the role of dendritic cell (DC) subsets in pathogenesis of mycobacterial infection remains unclear. Methodology/Principal Findings We examined the effectiveness of dendritic cell (DC) subsets in BCG-infected mice in generating immune responses beneficial for pathogen clearance and reduction of pathological reactions in the tissues following challenge infection. Our data showed that only the adoptive transfer of the subset of CD8α+ DC isolated from infected mice (iCD8+ DC) generated significant protection, demonstrated by less mycobacterial growth and pathological changes in the lung and liver tissues in iCD8+ DC recipients than sham-treated control mice. The adoptive transfer of the CD8α−DC from the infected mice (iCD8− DC) not only failed to reduce bacterial growth, but enhanced inflammation characterized by diffuse heavy cellular infiltration. Notably, iCD8− DC produced significantly higher levels of IL-10 than iCD8+ DC and promoted more Th2 cytokine responses in in vitro DC-T cell co-culture and in vivo adoptive transfer experiments. Conclusions/Significance The data indicate that in vivo BCG-primed CD8+ DC is the dominant DC subset in inducing protective immunity especially for reducing pathological reactions in infected tissues. The finding has implications for the rational improvement of the prophylactic and therapeutic approaches for controlling tuberculosis infection and related diseases.

Differential modulating effect of natural killer (NK) T cells on interferon‐γ production and cytotoxic function of NK cells and its relationship with NK subsets in Chlamydia muridarum infection

Natural killer T (NKT) cells are a newly identified T‐cell population with potential immunomodulatory functions. Several studies have shown modulating effects of NKT cells activated by α‐galactosylceramide, a model antigen, on NK cell function. We here report a differential modulating effect of NKT cells on the interferon‐γ (IFN‐γ) production and cytolytic function of NK cells in a chlamydial infection model, using NKT‐cell‐deficient mice and antibody blocking (anti‐CD1d monoclonal antibody) approaches. Our results showed that both NKT and NK cells became activated and produced IFN‐γ following Chlamydia muridarum infection in vitro and in vivo. The NK cells in NKT‐cell‐deficient mice and CD1d‐blocked mice showed decreased CD69 expression, cellular expansion and IFN‐γ production but surprisingly showed increased cytolytic activity (degranulation) of immature and more mature NK cell subsets, suggesting an inhibitory role of NKT cells on NK cell killing activity. The results suggest that NKT cells preferentially promote IFN‐γ production but are inhibitory for the cytotoxic function of NK cells in this infection model. Furthermore, the differential modulating effect of NKT cells on the IFN‐γ production and cytotoxicity of NK cells was observed in immature and mature NK cell subsets, although it was more dramatic in the relatively mature CD11bhigh CD27high NK cell subset. This finding demonstrates the complexity of innate cell interactions in infection and the possible differential impact of NKT cells on the variable functional aspects of other cell(s) even in one infection setting.

NK cells modulate the lung dendritic cell‐mediated Th1/Th17 immunity during intracellular bacterial infection

The impact of the interaction between NK cells and lung dendritic cells (LDCs) on the outcome of respiratory infections is poorly understood. In this study, we investigated the effect and mechanism of NK cells on the function of LDCs during intracellular bacterial lung infection of Chlamydia muridarum in mice. We found that the naive mice receiving LDCs from C. muridarum‐infected NK‐cell‐depleted mice (NK‐LDCs) showed more serious body weight loss, bacterial burden, and pathology upon chlamydial challenge when compared with the recipients of LDCs from infected sham‐treated mice (NK+LDCs). Cytokine analysis of the local tissues of the former compared with the latter exhibited lower levels of Th1 (IFN‐γ) and Th17 (IL‐17), but higher levels of Th2 (IL‐4), cytokines. Consistently, NK‐LDCs were less efficient in directing C. muridarum‐specific Th1 and Th17 responses than NK+LDCs when cocultured with CD4+ T cells. In NK cell/LDC coculture experiments, the blockade of NKG2D receptor reduced the production of IL‐12p70, IL‐6, and IL‐23 by LDCs. The neutralization of IFN‐γ in the culture decreased the production of IL‐12p70 by LDCs, whereas the blockade of TNF‐α resulted in diminished IL‐6 production. Our findings demonstrate that NK cells modulate LDC function to elicit Th1/Th17 immunity during intracellular bacterial infection.

Type 1 T-cell responses in chlamydial lung infections are associated with local MIP-1α response

Chemokines and their receptors are important mediators of leukocyte trafficking and recruitment and sometimes work as modulators of T-cell responses during infections and inflammation. Modulating the biological activity of chemokines has been found to influence the course of diseases. However, little is known about the role of chemokine responses during chlamydial lung infections. We therefore analyzed the dynamics of multiple chemokines, which are frequently associated with type 1 (Th1) T cell immune responses, and their receptors for their expression in the lungs during Chlamydia muridarum (Cm) infections. We also examined the relationship between chemokine responses and the development of Th1 responses as well as the clearance of infection. Our results showed that in parallel with the high levels of gamma interferon (IFN-γ) and IL-12 production in the lungs and draining lymph nodes, and the expansion of IFN-γ-producing CD4 and CD8+ T cells, the production of the cell-related chemokines RANTES, IFN-γ-inducible protein-10 (IP-10) and macrophage inflammatory protein-1α (MIP-1α) and their receptor CCR1 was elevated in the lung tissues after infection. Interestingly, in a later phase of infection, the expression of RANTES and IP-10 remained elevated but the expression of MIP-1α and CCR1 decreased to a low level, which suggests a closer association with the pattern of Th1 cytokine responses in the process of infection. These results suggest a close association between the MIP-1α response and the Th1-type T-cell responses in chlamydial lung infections.

Respective IL-17A production by γδ T and Th17 cells and its implication in host defense against chlamydial lung infection

The role of IL-17A is important in protection against lung infection with Chlamydiae, an obligate intracellular bacterial pathogen. In this study, we explored the producers of IL-17A in chlamydial lung infection and specifically tested the role of major IL-17A producers in protective immunity. We found that γδT cells and Th17 cells are the major producers of IL-17A at the early and later stages of chlamydial infection, respectively. Depletion of γδT cells in vivo at the early postinfection (p.i.) stage, when most γδT cells produce IL-17A, failed to alter Th1 responses and bacterial clearance. In contrast, the blockade of IL-17A at the time when IL-17A was mainly produced by Th17 (day 7 p.i.) markedly reduced the Th1 response and increased chlamydial growth. The data suggest that the γδ T cell is the highest producer of IL-17A in the very early stages of infection, but the protection conferred by IL-17A is mainly mediated by Th17 cells. In addition, we found that depletion of γδ T cells reduced IL-1α production by dendritic cells, which was associated with a reduced Th17 response. This finding is helpful to understand the variable role of IL-17A in different infections and to develop preventive and therapeutic approaches against infectious diseases by targeting IL-17A.

Inefficiency of C3H/HeN Mice to Control Chlamydial Lung Infection Correlates with Downregulation of Neutrophil Activation during the Late Stage of Infection

We previously reported that massive infiltration of neutrophils in C3H/HeN (C3H) mice could not efficiently control Chlamydia muridarum (Cm) infection and might contribute to the high susceptibility of these mice to lung infection. To further define the nature of neutrophil responses in C3H mice during chlamydial infection, we examine the expression of adhesion molecules and CD11b related to neutrophils infiltration and activation, respectively, following intranasal Cm infection. The results showed that the expression of selectins (E-selectin, P-selectin and L-selectin), and intercellular cell adhesion molecule-1 (ICAM-1) in the lung of C3H mice increased more significantly than in C57BL/6 (B6) mice, the more resistant strain. These results correlated well with the massive neutrophils infiltration in C3H mice. In contrast, CD11b expression on peripheral blood and lung neutrophils in C3H mice exhibited a significant reduction compared with B6 mice during the late phage of infection (day 14). These findings suggest that the high-level expression of adhesion molecules in C3H mice may enhance neutrophils recruitment to the lung, but the decline of CD11b expression on neutrophils may attenuate neutrophil function. Therefore, CD11b down-regulation on neutrophils may contribute to the failure of C3H mice to control chlamydial lung infection.