Gamma interferon-independent effects of interleukin-12 on immunity to Salmonella enterica serovar Typhimurium

Interleukin-12 (IL-12) and IL-18 are both central to the induction of gamma interferon (IFN-γ), and various roles for IL-12 and IL-18 in control of intracellular microbial infections have been demonstrated. We used IL-12p40^−/− and IL-18^−/− mice to further investigate the role of IL-12 and IL-18 in control of Salmonella enterica serovar Typhimurium. While C57BL/6 and IL-18^−/− mice were able to resolve attenuated S. enterica serovar Typhimurium infections, the IL-12p40^−/− mice succumbed to a high bacterial burden after 60 days. Using ovalbumin (OVA)-specific T-cell receptor transgenic T cells (OT-II cells), we demonstrated that following oral infection with recombinant S. enterica serovar Typhimurium expressing OVA, the OT-II cells proliferated in the mesenteric lymph nodes of C57BL/6 and IL-18^−/− mice but not in IL-12p40^−/− mice. In addition, we demonstrated by flow cytometry that equivalent or increased numbers of T cells produced IFN-γ in IL-12p40^−/− mice compared with the numbers of T cells that produced IFN-γ in C57BL/6 and IL-18^−/− mice. Finally, we demonstrated that removal of macrophages from S. enterica serovar Typhimurium-infected C57BL/6 and IL-12p40^−/− mice did not affect the bacterial load, suggesting that impaired control of S. enterica serovar Typhimurium infection in the absence of IL-12p40 is not due to reduced macrophage bactericidal activities, while IL-18^−/− mice did rely on the presence of macrophages for control of the infection. Our results suggest that IL-12p40, but not IL-18, is critical to resolution of infections with attenuated S. enterica serovar Typhimurium and that especially the effects of IL-12p40 on proliferative responses of CD4⁺ T cells, but not the ability of these cells to produce IFN-γ, are important in the resolution of infection by this intracellular bacterial pathogen.     

Dendritic Cell and NK Cell Reciprocal Cross Talk Promotes Gamma Interferon-Dependent Immunity to Blood-Stage Plasmodium chabaudi AS Infection in Mice

Dendritic cells (DCs) are important accessory cells for promoting NK cell gamma interferon (IFN-γ) production in vitro in response to Plasmodium falciparum-infected red blood cells (iRBC). We investigated the requirements for reciprocal activation of DCs and NK cells leading to Th1-type innate and adaptive immunity to P. chabaudi AS infection. During the first week of infection, the uptake of iRBC by splenic CD11c⁺ DCs in resistant wild-type (WT) C57BL/6 mice was similar to that in interleukin 15^−/− (IL-15^−/−) and IL-12p40^−/− mice, which differ in the severity of P. chabaudi AS infection. DCs from infected IL-15^−/− mice expressed costimulatory molecules, produced IL-12, and promoted IFN-γ secretion by WT NK cells in vitro as efficiently as WT DCs. In contrast, DCs from infected IL-12p40^−/− mice exhibited alterations in maturation and cytokine production and were unable to induce NK cell IFN-γ production. Coculture of DCs and NK cells demonstrated that DC-mediated NK cell activation required IL-12 and, to a lesser extent, IL-2, as well as cell-cell contact. In turn, NK cells from infected WT mice enhanced DC maturation, IL-12 production, and priming of CD4⁺ T-cell proliferation and IFN-γ secretion. Infected WT mice depleted of NK cells, which exhibit increased parasitemia, had impaired DC maturation and DC-induced CD4⁺ Th1 cell priming. These findings indicate that DC-NK cell reciprocal cross talk is critical for control and rapid resolution of P. chabaudi AS infection and provide in vivo evidence for the importance of this interaction in IFN-γ-dependent immunity to malaria.     

Attenuated Coxiella burnetii phase II causes a febrile response in gamma interferon knockout and Toll-like receptor 2 knockout mice and protects against reinfection

Coxiella burnetii is a highly infectious obligate intracellular bacterium. The phase I form is responsible for Q fever, a febrile illness with flu-like symptoms that often goes undiagnosed. The attenuated C. burnetii phase II (having a truncated “O” chain of its lipopolysaccharide) does not cause disease in immunocompetent animals; however, phase II organisms remain infectious, and we questioned whether disease could be produced in immunodeficient mice. To study C. burnetii phase II infections, febrile responses in gamma interferon knockout (IFN-γ^−/−), BALB/c, Toll-like receptor 2 knockout (TLR2^−/−), and C57BL/6 mice were measured using the Nine Mile phase II (NMII) strain of C. burnetii. Immunocompetent mice showed minimal febrile responses, unlike those obtained with IFN-γ^−/− and TLR2^−/− mice, which showed elevated rectal temperatures that were sustained for ~15 days with transient increases in splenic weights. Reinfection of IFN-γ^−/− and TLR2^−/− mice with C. burnetii NMII 30 days after primary infection protected mice as evident by reduced febrile responses and a lack of splenic inflammation. Although minimal detection of Coxiella in TLR2^−/− mouse spleens was observed, greater colonization was evident in the IFN-γ^−/− mice. Cytokine analysis was performed on infected peritoneal macrophages isolated from these mice, and immunocompetent macrophages showed robust tumor necrosis factor alpha, IFN-γ, and granulocyte-macrophage colony-stimulating factor (GM-CSF) but no interleukin-12 (IL-12) responses. IFN-γ^−/− macrophages produced elevated levels of IL-6, IL-10, and IL-12, while TLR2^−/− macrophages produced GM-CSF, IL-12, and minimal IL-10. To distinguish immunity conferred by innate or adaptive systems, adoptive transfer studies were performed and showed that immune lymphocytes obtained from immunocompetent mice protected against a subsequent challenge with NMII, indicating that adaptive immunity mediates the observed protection. Thus, our data show that NMII is capable of eliciting disease in immunocompromised mice, which may help in evaluation of vaccine candidates as well as the study of host-pathogen interactions.     

Effects of CXCL10 on dendritic cell and CD4+ T-cell functions during Leishmania amazonensis infection

Leishmania amazonensis can cause progressive disease in most inbred strains of mice. We have previously reported that treatment with CXCL10 activates macrophage (MΦ) effector function(s) in parasite killing and significantly delays lesion development in susceptible C57BL/6 mice via enhanced gamma interferon (IFN-γ) and interleukin 12 (IL-12) secretion; however, the mechanism underlying this enhanced immunity against L. amazonensis infection remains largely unresolved. In this study, we utilized stationary promastigotes to infect bone marrow-derived dendritic cells (DCs) of C57BL/6 mice and assessed the activation of DC subsets and the capacity of these DC subsets to prime CD4⁺ T cells in vitro. We found that CXCL10 induced IL-12 p40 production but reduced IL-10 production in uninfected DCs. Yet L. amazonensis-infected DCs produced elevated levels of IL-10 despite CXCL10 treatment. Elimination of endogenous IL-10 led to increased IL-12 p40 production in DCs as well as increased proliferation and IFN-γ production by in vitro-primed CD4⁺ T cells. In addition, CXCL10-treated CD4⁺ T cells became more responsive to IL-12 via increased expression of the IL-12 receptor β₂ chain and produced elevated levels of IFN-γ. This report indicates the utility of CXCL10 in generating a Th1-favored, proinflammatory response, which is a prerequisite for controlling Leishmania infection.     

Gamma Interferon Production by Cytotoxic T Lymphocytes Is Required for Resolution of Chlamydia trachomatis Infection

In this study, we used mice in which the gene for gamma interferon (IFN-γ) has been disrupted (IFN-γ^−/− mice) to study the role of this cytokine in the resolution of Chlamydia trachomatis infection. We show that IFN-γ^−/− mice are impaired in the ability to clear infection with C. trachomatis compared to IFN-γ^+/+ control mice. Activated CD8⁺ cytotoxic T lymphocytes (CTL) secrete IFN-γ in response to intracellular infection, and we have shown previously that a Chlamydia-specific CTL line can reduce C. trachomatis infection when adoptively transferred into infected mice. In the present study, we found that when these IFN-γ^+/+ CTL lines are transferred into Chlamydia-infected IFN-γ^−/− mice, the transferred CTL cannot overcome the immune defect seen in the IFN-γ^−/− mice. We also show that Chlamydia-specific CTL can be cultured from IFN-γ-deficient mice infected with C. trachomatis; however, the adoptive transfer of IFN-γ^−/− CTL into infected IFN-γ^+/+ mice does not reduce the level of infection. These results suggest that IFN-γ production by CTL is not sufficient to overcome the defect that IFN-γ^−/− mice have in the resolution of Chlamydia infection, yet IFN-γ production by CTL is required for the protective effect seen upon adoptive transfer of CTL into IFN-γ^+/+ mice.     

Defective Nitric Oxide Effector Functions Lead to Extreme Susceptibility of Trypanosoma cruzi-Infected Mice Deficient in Gamma Interferon Receptor or Inducible Nitric Oxide Synthase

Trypanosoma cruzi, the causative agent of Chagas’ disease, induces an innate and adaptive host immune response during the acute phase of infection. These responses were analyzed by comparing mouse lines deficient for the gamma interferon (IFN-γ) receptor (IFN-γR^−/−) or deficient for inducible nitric oxide synthase (iNOS^−/−). Both lines were highly susceptible, with similar and dramatically increased parasite burdens and severe histopathology and were incapable of surviving even very low doses, exhibiting similar mortality kinetics. This pathophysiological correlation has a common cause, since both mutant mouse strains were unable to respond to infection by producing nitric oxide (NO) with the consequence that mutant macrophages had impaired trypanocidal activities. These in vivo and subsequent in vitro studies further demonstrated that an IFN-γ-dependent pathway of iNOS induction is crucial for efficient NO production and mandatory for resisting acute infection with T. cruzi. Despite this defect, both mutant mouse strains had a rather normal proinflammatory cytokine response (interleukin-12 [IL-12], IFN-γ, IL-6), with the exception of an impaired tumor necrosis factor alpha and IL-1α response in IFN-γR^−/− mice, demonstrating that only the latter two cytokines are dependent on IFN-γ activation. Moreover, polarization of T cells in type 1 and type 2 T-helper (Th1/Th2) and cytotoxic T (Tc1/Tc2) cells as well as T. cruzi-specific antibody responses were normal in IFN-γR^−/− mice, demonstrating that IFN-γ is not necessary for the promotion of T-cell differentiation and T. cruzi-specific antibody responses.     

Neither Classical nor Alternative Macrophage Activation Is Required for Pneumocystis Clearance during Immune Reconstitution Inflammatory Syndrome

Pneumocystis is a respiratory fungal pathogen that causes pneumonia (Pneumocystis pneumonia [PcP]) in immunocompromised patients. Alveolar macrophages are critical effectors for CD4⁺ T cell-dependent clearance of Pneumocystis, and previous studies found that alternative macrophage activation accelerates fungal clearance during PcP-related immune reconstitution inflammatory syndrome (IRIS). However, the requirement for either classically or alternatively activated macrophages for Pneumocystis clearance has not been determined. Therefore, RAG2^−/− mice lacking either the interferon gamma (IFN-γ) receptor (IFN-γR) or interleukin 4 receptor alpha (IL-4Rα) were infected with Pneumocystis. These mice were then immune reconstituted with wild-type lymphocytes to preserve the normal T helper response while preventing downstream effects of Th1 or Th2 effector cytokines on macrophage polarization. As expected, RAG2^−/− mice developed severe disease but effectively cleared Pneumocystis and resolved IRIS. Neither RAG/IFN-γR^−/− nor RAG/IL-4Rα^−/− mice displayed impaired Pneumocystis clearance. However, RAG/IFN-γR^−/− mice developed a dysregulated immune response, with exacerbated IRIS and greater pulmonary function deficits than those in RAG2 and RAG/IL-4Rα^−/− mice. RAG/IFN-γR^−/− mice had elevated numbers of lung CD4⁺ T cells, neutrophils, eosinophils, and NK cells but severely depressed numbers of lung CD8⁺ T suppressor cells. Impaired lung CD8⁺ T cell responses in RAG/IFN-γR^−/− mice were associated with elevated lung IFN-γ levels, and neutralization of IFN-γ restored the CD8 response. These data demonstrate that restricting the ability of macrophages to polarize in response to Th1 or Th2 cytokines does not impair Pneumocystis clearance. However, a cell type-specific IFN-γ/IFN-γR-dependent mechanism regulates CD8⁺ T suppressor cell recruitment, limits immunopathogenesis, preserves lung function, and enhances the resolution of PcP-related IRIS.     

Interferon-gamma is crucial for surviving a Brucella abortus infection in both resistant C57BL/6 and susceptible BALB/c mice

Brucella abortus is an intracellular bacterial pathogen that causes chronic infections in humans and a number of agriculturally important species of animals. It has been shown that BALB/c mice are more susceptible to infections with virulent strains of Brucella abortus than C57BL/6 or C57BL/10 strains. In experiments described here, gene knock-out mice were utilized to elucidate some of the salient components of resistance. Resistant C57BL/6 mice with gene deletions or disruptions in the interferon-γ (IFN-γ), perforin or β₂-microglobulin genes had decreased abilities to control intracellular infections with B. abortus strain 2308 during the first week after infection. However, only the IFN-γ knock-out mice had a sustained inability to control infections and this resulted in death of the mice at approximately 6 weeks post-infection. These mice had a continual increase in the number of bacterial colony-forming units (CFU) in their spleens until death. When BALB/c mice with the disrupted IFN-γ gene were infected they had more splenic CFU at one week post-infection than control mice but the increase was not statistically significant and by 3 weeks they did not have more CFU than control mice. Moreover, the number of splenic bacteria did not increase in the BALB/c IFN-γ knock-out mice between 6 and 10·5 weeks, although they died at 10·5 weeks, the time by which normal BALB/c mice were clearing the infection. Death in both strains of IFN-γ gene disrupted mice coincided with symptoms of cachexia and macrophages comprised ≥75% of the splenic leucocytes.     

CD11c+ CD103+ cells of Mycobacterium tuberculosis‐infected C57BL/6 but not of BALB/c mice induce a high frequency of interferon‐γ‐ or interleukin‐17‐producing CD4+ cells

The magnitude of the cellular adaptive immune response is critical for the control of Mycobacterium tuberculosis infection in the chronic phase. In addition, the genetic background is equally important for resistance or susceptibility to tuberculosis. In this study, we addressed whether lung populations of dendritic cells, obtained from genetically different hosts, would play a role in the magnitude and function of CD4⁺ populations generated after M. tuberculosis infection. Thirty days post-infection, C57BL/6 mice, which generate a stronger interferon-γ (IFN-γ)-mediated immune response than BALB/c mice, exhibited a higher number and frequency of lung CD11c⁺ CD11b⁻ CD103⁺ cells compared with BALB/c mice, which exhibited a high frequency of lung CD11c⁺ CD11b⁺ CD103⁻ cells. CD11c⁺ CD11b⁻ CD103⁺ cells, purified from lungs of infected C57BL/6 mice, but not from infected BALB/c mice, induced a higher frequency of IFN-γ-producing or interleukin-17 (IL-17)-producing CD4⁺ cells. Moreover, CD4⁺ cells also arrive at the lung of C57BL/6 mice faster than in BALB/c mice. This pattern of immune response seems to be associated with higher gene expression for CCL4, CCL19, CCL20 and CCR5 in the lungs of infected C57BL/6 mice compared with infected BALB/c mice. The results described here show that the magnitude of IFN-γ-producing or IL-17-producing CD4⁺ cells is dependent on CD11c⁺ CD11b⁻ CD103⁺ cells, and this pattern of immune response is directly associated with the host genetic background. Therefore, differences in the genetic background contribute to the identification of immunological biomarkers that can be used to design human assays to predict progression of M. tuberculosis infection.     

Role of Gamma Interferon in Helicobacter pylori-Induced Gastric Inflammatory Responses in a Mouse Model

The immune responses to Helicobacter pylori infection play important roles in gastroduodenal diseases. The contribution of gamma interferon (IFN-γ) to the immune responses, especially to the induction of gastric inflammation and to protection from H. pylori infection, was investigated with IFN-γ gene knockout (IFN-γ^−/−) mice. We first examined the colonizing abilities of eight H. pylori strains with a short-term infection test in order to select H. pylori strains which could colonize the mouse stomach. Only three strains (ATCC 43504, CPY2052, and HPK127) colonized C57BL/6 wild-type mice, although all of the strains except for ATCC 51110 could colonize IFN-γ^−/− mice. The number of H. pylori organisms colonizing the stomach in wild-type mice was lower than that in IFN-γ^−/− mice. Oral immunization with the CPY2052 sonicate and cholera toxin protected against infection with strain CPY2052 in both types of mouse. These findings suggested that IFN-γ may play a protective role in H. pylori infection, although the degree of its protective ability was estimated to be low. In contrast, in a long-term infection test done to examine the contribution of IFN-γ to gastric inflammation, CPY2052-infected wild-type mice developed a severe infiltration of mononuclear cells in the lamina propria and erosions in the gastric epithelium 15 months after infection, whereas CPY2052-infected IFN-γ^−/− mice showed no inflammatory symptoms. This result clearly demonstrated that IFN-γ plays an important role in the induction of gastric inflammation caused by H. pylori infection.     

Expansion of a Novel Pulmonary CD3− CD4+ CD8+ Cell Population in Mice during Chlamydia pneumoniae Infection

A new pulmonary T-cell-like lymphocyte population with the phenotype CD3⁻ CD4⁺ CD8⁺ was discovered in mice. CD4⁺ CD8⁺ but CD3⁺ cells among murine intestinal intraepithelial lymphocytes have previously been described. We describe herein a dramatic expansion of the CD3⁻ CD4⁺ CD8⁺ cell population in response to experimental respiratory infection. After intranasal Chlamydia pneumoniae infection, CD4⁺ CD8⁺ cells became transiently the dominant lymphocyte type (maximum of 87% of all lymphocytes) in the lungs of NIH/S mice but remained virtually undetectable in spleen and blood. The enrichment of these cells was not a C. pneumoniae-specific event, since infection of NIH/S mice with influenza A virus also resulted in an increase in the number of CD4⁺ CD8⁺ cells (maximum of 42% of all lymphocytes). In addition to outbred NIH/S mice, two other mouse strains were studied: BALB/c (H-2^d) and C57BL/6 (H-2^b). C. pneumoniae-infected BALB/c mice responded with an intermediate increase in the number of CD4⁺ CD8⁺ cells in lungs, whereas C57BL/6 mice did not respond. The double-positive CD4⁺ CD8⁺ cells lacked a major part of the T-cell receptor complex, being both CD3⁻ and TCR αβ⁻. However, when they were stimulated in vitro with a T-cell mitogen, they responded by proliferation but did not secrete gamma interferon. The dramatic expansion of this cell population at the infection site suggests an active role for them in respiratory infection, but the specification of this requires further study.     

Regulatory Actions of Toll-Like Receptor 2 (TLR2) and TLR4 in Leishmania donovani Infection in the Liver

In livers of susceptible but self-curing C57BL/6 mice, intracellular Leishmania donovani infection enhanced Toll-like receptor 4 (TLR4) and TLR2 gene expression. In the liver, infected TLR4^−/− mice showed reduced gamma interferon (IFN-γ), tumor necrosis factor (TNF), and inducible nitric oxide synthase (iNOS) mRNA expression, higher-level and slowly resolving infection, delayed granuloma formation, and little response to low-dose chemotherapy; in serum, the ratio of IFN-γ to interleukin 10 (IL-10) activity was decreased by 50%. In contrast, in TLR2^−/− mice, control of liver infection, parasite killing, and granuloma assembly were accelerated and chemotherapy''s efficacy enhanced. In livers of infected TLR2^−/− mice, mRNA expression was not increased for inflammatory cytokines or iNOS or decreased for IL-10; however, the serum IFN-γ/IL-10 ratio was increased 6.5-fold and minimal responses to IL-10 receptor blockade suggested downregulated IL-10. In established infection in wild-type mice, blockading TLR2 induced parasite killing and triggering TLR4 strengthened resistance and promoted chemotherapy''s effect. Thus, in experimental L. donovani infection in the liver, TLR4 signaling upregulates and TLR2 signaling downregulates macrophage antileishmanial activity, making both receptors potential therapeutic targets in visceral leishmaniasis for engagement (TLR4) or blockade (TLR2).     

Interleukin-17A Enhances Host Defense against Cryptococcal Lung Infection through Effects Mediated by Leukocyte Recruitment,Activation, and Gamma Interferon Production

Infection of C57BL/6 mice with the moderately virulent Cryptococcus neoformans strain 52D models the complex adaptive immune response observed in HIV-negative patients with persistent fungal lung infections. In this model, Th1 and Th2 responses evolve over time, yet the contribution of interleukin-17A (IL-17A) to antifungal host defense is unknown. In this study, we show that fungal lung infection promoted an increase in Th17 T cells that persisted to 8 weeks postinfection. Our comparison of fungal lung infection in wild-type mice and IL-17A-deficient mice (IL-17A^−/− mice; C57BL/6 genetic background) demonstrated that late fungal clearance was impaired in the absence of IL-17A. This finding was associated with reduced intracellular containment of the organism within lung macrophages and deficits in the accumulation of total lung leukocytes, including specific reductions in CD11c⁺ CD11b⁺ myeloid cells (dendritic cells and exudate macrophages), B cells, and CD8⁺ T cells, and a nonsignificant trend in the reduction of lung neutrophils. Although IL-17A did not alter the total number of CD4 T cells, decreases in the total number of CD4 T cells and CD8 T cells expressing gamma interferon (IFN-γ) were observed in IL-17A^−/− mice. Lastly, expression of major histocompatibility complex class II (MHC-II) and the costimulatory molecules CD80 and CD86 on CD11c⁺ CD11b⁺ myeloid cells was diminished in IL-17A^−/− mice. Collectively, these data indicate that IL-17A enhances host defenses against a moderately virulent strain of C. neoformans through effects on leukocyte recruitment, IFN-γ production by CD4 and CD8 T cells, and the activation of lung myeloid cells.     

Gamma Interferon (IFN-γ) and IFN-γ-Inducing Cytokines Interleukin-12 (IL-12) and IL-18 Do Not Augment Infection-Stimulated Bone Resorption In Vivo

Periapical granulomas are induced by bacterial infection of the dental pulp and result in destruction of the surrounding alveolar bone. In previous studies we have reported that the bone resorption in this model is primarily mediated by macrophage-expressed interleukin-1 (IL-1). The expression and activity of IL-1 is in turn modulated by a network of Th1 and Th2 regulatory cytokines. In the present study, the functional roles of the Th1 cytokine gamma interferon (IFN-γ) and IFN-γ-inducing cytokines IL-12 and IL-18 were determined in a murine model of periapical bone destruction. IL-12^−/−, IL-18^−/−, and IFN-γ^−/− mice were subjected to surgical pulp exposure and infection with a mixture of four endodontic pathogens, and bone destruction was determined by microcomputed tomography on day 21. The results indicated that all IL-12^−/−, IL-18^−/−, and IFN-γ^−/− mice had similar infection-stimulated bone resorption in vivo as wild-type control mice. Mice infused with recombinant IL-12 also had resorption similar to controls. IFN-γ^−/− mice exhibited significant elevations in IL-6, IL-10, IL-12, and tumor necrosis factor alpha in lesions compared to wild-type mice, but these modulations had no net effect on IL-1α levels. Recombinant IL-12, IL-18, and IFN-γ individually failed to consistently modulate macrophage IL-1α production in vitro. We conclude that, at least individually, endogenous IL-12, IL-18, and IFN-γ do not have a significant effect on the pathogenesis of infection-stimulated bone resorption in vivo, suggesting possible functional redundancy in proinflammatory pathways.     

Reciprocal Regulation of Th1- and Th2-Cytokine-Producing T Cells during Clearance of Parasitemia in Plasmodium falciparum Malaria

Flow cytometry for the intracellular detection of T-cell cytokines was performed for 15 Gabonese patients during acute uncomplicated Plasmodium falciparum malaria. A striking expansion of CD4⁺ and CD8⁺ T cells producing gamma interferon (IFN-γ) was found during drug-induced clearance of parasitemia, paralleled by a decrease of interleukin-2 (IL-2) production. The frequency of IL-4- and IL-13-producing CD4⁺ cells gradually decreased, whereas the frequency of T cells producing IL-2⁺–IFN-γ⁺, IL-4⁻–IL-5⁺, and IL-4⁺–IL-5⁺ cytokines as well as IL-4⁺–IFN-γ⁺ and IL-13⁺–IFN-γ⁺ cytokines was not significantly altered. The capacity for IL-10 production within the CD4⁺ subset increased due to an expansion of both IL-10⁺–IFN-γ⁻ and IL-10⁺–IFN-γ⁺ cytokine-expressing cells. Thus, a more pronounced Th2-driven immune response during acute untreated P. falciparum infection with a shift towards Th1 responsiveness induced by parasite clearance is suggested.     

Interleukin 18 Contributes to Host Resistance and Gamma Interferon Production in Mice Infected with Virulent Salmonella typhimurium

Spleen and peritoneal macrophages obtained from innately resistant A/J mice released low levels of interleukin 18 (IL-18) upon infection with Salmonella typhimurium C5 RP4. Incubating the cells with recombinant gamma interferon (rIFN-γ) enhanced IL-18 production. A/J mice treated in vivo with anti-IL-18 antibodies showed impaired resistance to infection, with increased bacterial loads in the liver and spleen. Administration of rIL-18 could protect A/J mice from challenge with a lethal dose of virulent salmonellae, with a dramatic reduction in bacterial numbers in the tissues. rIL-18 administration did not ameliorate the disease in IFN-γ-R^−/− mice. IL-18 proved to be required for IFN-γ production by mouse splenocytes from conventional, scid, and rag-1^−/− mice; in vivo IL-18 neutralization caused a decrease in circulating IFN-γ levels. Thus, IL-18 is a key factor in early host resistance to Salmonella and probably acts via IFN-γ.     

TLR4 inactivation protects from graft-versus-host disease after allogeneic hematopoietic stem cell transplantation

Graft-versus-host disease (GVHD) is the most common complication after hematopoietic stem cell transplantation. To clarify the role of Toll-like receptor 4 (TLR4), which is a major receptor for bacterial lipopolysaccharides (LPS), in the development of acute GVHD, we used a TLR4-knockout (TLR4^−/−) mouse GVHD model and analyzed the underlying immunological mechanisms. When TLR4^−/− mice were used as bone marrow and splenocyte cell graft donors or recipients, GVHD symptom occurrence and mortality were delayed compared to wild-type (TLR4^+/+) mice. In addition, histopathological analyses revealed that in TLR4^−/−→BALB/c chimeras, liver and small intestine tissue damage was reduced with minimal lymphocytic infiltration. In contrast to TLR4^+/+, TLR4^−/− mice dendritic cells did not express CD80, CD86, CD40, MHC-II or IL-12 during LPS induction and remained in an immature state. Furthermore, the ability of TLR4^−/− mice spleen dendritic cells to promote allogeneic T-cell proliferation and, in particular, T-helper cell 1 (Th1) development was obviously attenuated compared with TLR4^+/+ mice dendritic cells, and the levels of interferon-γ (IFN-γ) and IL-10, Th2-cell specific cytokines, were significantly higher in the serum of TLR4^−/−→BALB/c than in TLR4^+/+→BALB/c chimeric mice. Overall, our data revealed that TLR4 may play a role in the pathogenesis of GVHD and that targeted TLR4 gene therapy might provide a new treatment approach to reduce the risk of GVHD.     

The alternative activation pathway and complement component C3 are critical for a protective immune response against Pseudomonas aeruginosa in a murine model of pneumonia

Pseudomonas aeruginosa is a leading cause of hospital-acquired pneumonia, and approximately 80% of patients with cystic fibrosis are infected with this bacterium. To investigate the overall role of complement and the complement activation pathways in the host defense against P. aeruginosa pulmonary infection, we challenged C3-, C4-, and factor B-deficient mice with P. aeruginosa via intranasal inoculation. In these studies, C3^−/− mice had a higher mortality rate than C3^+/+ mice. Factor B^−/− mice, but not C4^−/− mice, infected with P. aeruginosa had a mortality rate similar to that of C3^−/− mice, indicating that in this model the alternative pathway of complement activation is required for the host defense against Pseudomonas infection. C3^−/− mice had 6- to 7-fold more bacteria in the lungs and 48-fold more bacteria in the blood than did C3^+/+ mice at 24 h postinfection. In vitro, phagocytic cells from C3^+/+ or C3^−/− mice exhibited a decreased ability to bind and/or ingest P. aeruginosa in the presence of C3-deficient serum compared to phagocytic cells in the presence of serum with sufficient C3. C3^−/− mice displayed a significant increase in neutrophils in the lungs and had higher levels of interleukin-1β (IL-1β), IL-6, IL-10, KC, and MIP-2 in the lungs at 24 h postinfection than did C3^+/+ mice. Collectively, these results indicate that complement activation by the alternative pathway is critical for the survival of mice infected with P. aeruginosa and that the protection provided by complement is at least in part due to C3-mediated opsonization and phagocytosis of P. aeruginosa.     

Type 1 interferon-induced IL-7 maintains CD8+ T-cell responses and homeostasis by suppressing PD-1 expression in viral hepatitis

Type 1 interferon (IFN-I) promotes antigen-presenting cell maturation and was recently shown to induce hepatic IL-7 production during infection. Herein, we further explored the underlying mechanisms used by IFN-I to orchestrate antiviral immune responses in the liver. Acute viral hepatitis was induced by i.v. injection of adenovirus (Ad) in IFN-α receptor knockout (IFNAR^−/−) and control mice. To disrupt signaling, monoclonal antibodies (mAbs) against IL-7 receptor alpha (IL-7Rα) or PD-L1 were i.p. injected. We found that CD8⁺ T cells in IFNAR^−/− mice were less effective than those in control mice. The reduced T-cell function was accompanied by increased levels of PD-1 expression, apoptosis and decreased IFN-γ production. The lack of IFN-I signaling also impaired the expression of accessory molecules in both intrahepatic dendritic cell (DCs) and hepatocytes. PD-L1 was comparably and highly expressed on hepatocytes in both IFNAR^−/− and control mice. Injection of PD-L1-specific mAb in IFNAR^−/− mice reversed the compromised immune responses in the liver. Further investigation showed that hepatic IL-7 elevation was less pronounced in IFNAR^−/− mice compared to the controls. A treatment with recombinant IL-7 suppressed PD-1 expression on CD8⁺ T cells in vitro. Accordingly, blocking IL-7R signaling in vivo resulted in increased PD-1 expression on CD8⁺ T cells in Ad-infected mice. Collectively, the results suggest that IFN-I-induced hepatic IL-7 production maintains antiviral CD8⁺ T-cell responses and homeostasis by suppressing PD-1 expression in acute viral hepatitis.