Sara Hieny

CCR5 provides a signal for microbial induced production of IL-12 by CD8α + dendritic cells

The activation of dendritic cells (DC) to produce interleukin 12 (IL-12) is thought to be a key step in the initiation of cell-mediated immunity to intracellular pathogens. Here we show that ligation of the C–C chemokine receptor (CCR) 5 can provide a major signal for the induction of IL-12 synthesis by the CD8α+ subset of DC and that this pathway is important in establishing interferon γ-dependent resistance to the protozoan parasite Toxoplasma gondii. These findings support the concept that the early induction of chemokines by invading pathogens is a critical step not only for the recruitment of DC but also for the determination of their subsequent immunologic function.

Cutting Edge: Caspase-1 Independent IL-1β Production Is Critical for Host Resistance to Mycobacterium tuberculosis and Does Not Require TLR Signaling In Vivo

To investigate the respective contributions of TLR versus IL-1R mediated signals in MyD88 dependent control of Mycobacterium tuberculosis, we compared the outcome of M. tuberculosis infection in MyD88, TRIF/MyD88, IL-1R1, and IL-1β–deficient mice. All four strains displayed acute mortality with highly increased pulmonary bacterial burden suggesting a major role for IL-1β signaling in determining the MyD88 dependent phenotype. Unexpectedly, the infected MyD88 and TRIF/MyD88-deficient mice, rather than being defective in IL-1β expression, displayed increased cytokine levels relative to wild-type animals. Similarly, infected mice deficient in caspase-1 and ASC, which have critical functions in inflammasome-mediated IL-1β maturation, showed unimpaired IL-1β production and importantly, were considerably less susceptible to infection than IL-1β deficient mice. Together our findings reveal a major role for IL-1β in host resistance to M. tuberculosis and indicate that during this infection the cytokine can be generated by a mechanism that does not require TLR signaling or caspase-1.

In the Absence of IL-12, CD4+ T Cell Responses to Intracellular Pathogens Fail to Default to a Th2 Pattern and Are Host Protective in an IL-10−/− Setting

IL-12-deficient mice exposed to nonlethal infections with intracellular pathogens or repeatedly immunized with a pathogen extract developed lowered but nevertheless substantial numbers of IFN-gamma(+) CD4(+) T cells compared to those observed in wild-type animals. Moreover, the CD4(+) responses in these knockout animals failed to default to a Th2 pattern. The protective efficacy of the Th1 cells developing in an IL-12-deficient setting was found to be limited by IL-10 since mice doubly deficient in IL-10 and IL-12 survived, while animals deficient in IL-12 alone succumbed to pathogen challenge. In contrast to IL-12 knockout mice, MyD88-deficient animals exposed to a Th1 microbial stimulus developed a pure Th2 response, arguing that this signaling element plays a more critical function than IL-12 in determining pathogen-induced CD4 polarization.

Lipoxin-mediated inhibition of IL-12 production by DCs: a mechanism for regulation of microbial immunity

Lipoxins are eicosanoid mediators that show potent inhibitory effects on the acute inflammatory process. We show here that the induction of lipoxin A4 (LXA4) accompanied the in vivo suppression of interleukin 12 (IL-12) responsiveness of murine splenic dendritic cells (DCs) after microbial stimulation with an extract of Toxoplasma gondii. This paralysis of DC function could not be triggered in mice that were deficient in a key lipoxygenase involved in LXA4 biosynthesis. In addition, DCs pre-treated with LXA4 became refractory to microbial stimulation for IL-12 production in vitro and mice injected with a stable LXA4 analog showed reduced splenic DC mobilization and IL-12 responses in vivo. Together, these findings indicate that the induction of lipoxins in response to microbial stimulation can provide a potent mechanism for regulating DC function during the innate response to pathogens.

Molecular mimicry of a CCR5 binding-domain in the microbial activation of dendritic cells

Toxoplasma gondii releases factors that potently stimulate production of interleukin-12 (IL-12) from dendritic cells (DCs). Purification of this activity showed that cyclophilin-18 (C-18) was its principal component, and antibodies generated against recombinant C-18 inhibited tachyzoite extract–induced synthesis of IL-12. Recombinant C-18 showed high affinity for and triggered cell signaling through CCR5, a chemokine receptor important in parasite-induced IL-12 production by DCs. These findings suggest that the unusual potency of T. gondii in inducing IL-12 from DCs results from its synthesis of a unique chemokine mimic that signals through CCR5. The ability to generate this strong protective response may benefit parasite transmission by preventing the protozoan from overwhelming its intermediate hosts.

Innate and adaptive interferons suppress IL-1α and IL-1β production by distinct pulmonary myeloid subsets during Mycobacterium tuberculosis infection.

Interleukin-1 (IL-1) receptor signaling is necessary for control of Mycobacterium tuberculosis (Mtb) infection, yet the role of its two ligands, IL-1α and IL-1β, and their regulation in vivo are poorly understood. Here, we showed that both IL-1α and IL-1β are critically required for host resistance and identified two multifunctional inflammatory monocyte-macrophage and DC populations that coexpressed both IL-1 species at the single-cell level in lungs of Mtb-infected mice. Moreover, we demonstrated that interferons (IFNs) played important roles in regulating IL-1 production by these cells in vivo. Type I interferons inhibited IL-1 production by both subsets whereas CD4(+) T cell-derived IFN-γ selectively suppressed monocyte-macrophages. These data provide a cellular basis for both the anti-inflammatory effects of IFNs and probacterial functions of type I IFNs during Mtb infection and reveal differential regulation of IL-1 production by distinct cell populations as an additional layer of complexity in the activity of IL-1 in vivo.

Paralysis of Dendritic Cell IL-12 Production by Microbial Products Prevents Infection-Induced Immunopathology

Interleukin-12 plays a major role in immunity to intracellular pathogens by governing the development of IFNgamma-dependent host resistance. Nevertheless, unregulated IL-12 synthesis can lead to immunopathology, an outcome prevented by the concurrent expression of interleukin-10. Dendritic cells (DC) are an important source of the initial IL-12 stimulated by microbial agents. Here, we show that, following systemic triggering, DC can no longer be restimulated to produce IL-12 in vivo while continuing to respond in vitro. When infected with Toxoplasma gondii during this refractory state, mice mount impaired acute IFNgamma responses and, in the case of IL-10-deficient animals, are protected from cytokine-induced mortality. These findings demonstrate a previously unrecognized form of immunologic paralysis involving DC that can protect from infection-induced immunopathology.

Mice lacking myeloid differentiation factor 88 display profound defects in host resistance and immune responses to Mycobacterium avium infection not exhibited by Toll-like receptor 2 (TLR2)- and TLR4-deficient animals.

To assess the role of Toll-like receptor (TLR) signaling in host resistance to Mycobacterium avium infection, mice deficient in the TLR adaptor molecule myeloid differentiation factor 88 (MyD88), as well as TLR2−/− and TLR4−/− animals, were infected with a virulent strain of M. avium, and bacterial burdens and immune responses were compared with those in wild-type (WT) animals. MyD88−/− mice failed to control acute and chronic M. avium growth and succumbed 9–14 wk postinfection. Infected TLR2−/− mice also showed increased susceptibility, but displayed longer survival and lower bacterial burdens than MyD88−/− animals, while TLR4−/− mice were indistinguishable from their WT counterparts. Histopathological examination of MyD88−/− mice revealed massive destruction of lung tissue not present in WT, TLR2−/−, or TLR4−/− mice. In addition, MyD88−/− and TLR2−/−, but not TLR4−/−, mice displayed marked reductions in hepatic neutrophil infiltration during the first 2 h of infection. Although both MyD88−/− and TLR2−/− macrophages showed profound defects in IL-6, TNF, and IL-12p40 responses to M. avium stimulation in vitro, in vivo TNF and IL-12p40 mRNA induction was impaired only in infected MyD88−/− mice. Similarly, MyD88−/− mice displayed a profound defect in IFN-γ response that was not evident in TLR2−/− or TLR4−/− mice or in animals deficient in IL-18. These findings indicate that resistance to mycobacterial infection is regulated by multiple MyD88-dependent signals in addition to those previously attributed to TLR2 or TLR4, and that these undefined elements play a major role in determining bacterial induced proinflammatory as well as IFN-γ responses.

MyD88-deficient mice display a profound loss in resistance to Mycobacterium tuberculosis associated with partially impaired Th1 cytokine and nitric oxide synthase 2 expression.

ABSTRACT Mycobacterium tuberculosis possesses agonists for several Toll-like receptors (TLRs), yet mice with single TLR deletions are resistant to acute tuberculosis. MyD88−/− mice were used to examine whether TLRs play any role in protection against aerogenic M. tuberculosis H37Rv infection. MyD88−/− mice failed to control mycobacterial replication and rapidly succumbed. Moreover, expressions of interleukin 12, tumor necrosis factor alpha, gamma interferon, and nitric oxide synthase 2 were markedly decreased in the knockout animals. These results argue that resistance to M. tuberculosis must depend on MyD88-dependent signals mediated by an as-yet-undetermined TLR or a combination of TLRs.

NK Cell-Derived IFN-γ Differentially Regulates Innate Resistance and Neutrophil Response in T Cell-Deficient Hosts Infected with Mycobacterium tuberculosis

Although it is known that IFN-γ-secreting T cells are critical for control of Mycobacterium tuberculosis infection, the contribution of IFN-γ produced by NK cells to host resistance to the pathogen is less well understood. By using T cell-deficient RAG−/− mice, we showed that M. tuberculosis stimulates NK cell-dependent IFN-γ production in naive splenic cultures and in lungs of infected animals. More importantly, common cytokine receptor γ-chain−/−RAG−/− animals deficient in NK cells, p40−/−RAG−/−, or anti-IFN-γ mAb-treated RAG−/− mice displayed significantly increased susceptibility to M. tuberculosis infection compared with untreated NK-sufficient RAG−/− controls. Studies comparing IL-12 p40- and p35-deficient RAG−/− mice indicated that IL-12 plays a more critical role in the induction of IFN-γ-mediated antimycobacterial effector functions than IL-23 or other p40-containing IL-12 family members. The increased susceptibility of IL-12-deficient or anti-IFN-γ mAb-treated RAG−/− mice was associated not only with elevated bacterial loads, but also with the development of granulocyte-enriched foci in lungs. This tissue response correlated with increased expression of the granulocyte chemotactic chemokines KC and MIP-2 in NK as well as other leukocyte populations. Interestingly, depletion of granulocytes further increased bacterial burdens and exacerbated pulmonary pathology in these animals, revealing a compensatory function for neutrophils in the absence of IFN-γ. The above observations indicate that NK cell-derived IFN-γ differentially regulates T-independent resistance and granulocyte function in M. tuberculosis infection and suggest that this response could serve as an important barrier in AIDS patients or other individuals with compromised CD4+ T cell function.