ADAM9 promotes type I interferon-mediated innate immunity during encephalomyocarditis virus infection.

Viral myocarditis, an inflammatory disease of the heart, causes significant morbidity and mortality. Type I interferon (IFN)-mediated antiviral responses protect against myocarditis, but the mechanisms are poorly understood. We previously identified A Disintegrin And Metalloproteinase domain 9 (ADAM9) as an important factor in viral pathogenesis. ADAM9 is implicated in a range of human diseases, including inflammatory diseases; however, its role in viral infection is unknown. Here, we demonstrate that mice lacking ADAM9 are more susceptible to encephalomyocarditis virus (EMCV)-induced death and fail to mount a characteristic type I IFN response. This defect in type I IFN induction is specific to positive-sense, single-stranded RNA (+ ssRNA) viruses and involves melanoma differentiation-associated protein 5 (MDA5)-a key receptor for +ssRNA viruses. Mechanistically, ADAM9 binds to MDA5 and promotes its oligomerization and thereby downstream mitochondrial antiviral-signaling protein (MAVS) activation in response to EMCV RNA stimulation. Our findings identify a role for ADAM9 in the innate antiviral response, specifically MDA5-mediated IFN production, which protects against virus-induced cardiac damage, and provide a potential therapeutic target for treatment of viral myocarditis.

Mechanistic study of broadly neutralizing human monoclonal antibodies against dengue virus that target the fusion loop.

There are no available vaccines for dengue, the most important mosquito-transmitted viral disease. Mechanistic studies with anti-dengue virus (DENV) human monoclonal antibodies (hMAbs) provide a rational approach to identify and characterize neutralizing epitopes on DENV structural proteins that can serve to inform vaccine strategies. Here, we report a class of hMAbs that is likely to be an important determinant in the human humoral response to DENV infection. In this study, we identified and characterized three broadly neutralizing anti-DENV hMAbs: 4.8A, D11C, and 1.6D. These antibodies were isolated from three different convalescent patients with distinct histories of DENV infection yet demonstrated remarkable similarities. All three hMAbs recognized the E glycoprotein with high affinity, neutralized all four serotypes of DENV, and mediated antibody-dependent enhancement of infection in Fc receptor-bearing cells at subneutralizing concentrations. The neutralization activities of these hMAbs correlated with a strong inhibition of virus-liposome and intracellular fusion, not virus-cell binding. We mapped epitopes of these antibodies to the highly conserved fusion loop region of E domain II. Mutations at fusion loop residues W101, L107, and/or G109 significantly reduced the binding of the hMAbs to E protein. The results show that hMAbs directed against the highly conserved E protein fusion loop block viral entry downstream of virus-cell binding by inhibiting E protein-mediated fusion. Characterization of hMAbs targeting this region may provide new insights into DENV vaccine and therapeutic strategies.

Genetic control of severe egg-induced immunopathology and IL-17 production in murine schistosomiasis.

Infection with the trematode parasite Schistosoma mansoni results in a distinct heterogeneity of disease severity, both in humans and in an experimental mouse model. Severe disease is characterized by pronounced hepatic egg-induced granulomatous inflammation in a proinflammatory cytokine environment, whereas mild disease corresponds with reduced hepatic inflammation in a Th2 skewed cytokine environment. This marked heterogeneity indicates that genetic differences play a significant role in disease development, yet little is known about the genetic basis of dissimilar immunopathology. To investigate the role of genetic susceptibility in murine schistosomiasis, quantitative trait loci analysis was performed on F(2) progeny derived from SJL/J and C57BL/6 mice, which develop severe and mild pathology, respectively. In this study, we show that severe liver pathology in F(2) mice 7 wk after infection significantly correlated with an increase in the production of the proinflammatory cytokines IL-17, IFN-gamma, and TNF-alpha by schistosome egg Ag-stimulated mesenteric lymph node cells. Quantitative trait loci analysis identified several genetic intervals controlling immunopathology as well as IL-17 and IFN-gamma production. Egg granuloma size exhibited significant linkage to two loci, D4Mit203 and D17Mit82, both of which were inherited in a BL/6 dominant manner. Furthermore, a significant reduction of hepatic granulomatous inflammation and IL-17 production in interval-specific congenic mice demonstrated that the two identified genetic loci have a decisive effect on the development of immunopathology in murine schistosomiasis.

Coinfection with the intestinal nematode Heligmosomoides polygyrus markedly reduces hepatic egg-induced immunopathology and proinflammatory cytokines in mouse models of severe schistosomiasis.

Infection with the trematode helminth Schistosoma mansoni results in a parasite egg-induced, CD4 T-cell-mediated, hepatointestinal granulomatous and fibrosing inflammation that varies greatly in severity, with a higher frequency of milder forms typically occurring in regions where the disease is endemic. One possible explanation for this is that in these regions the degree of inflammation is lessened by widespread concurrent infection with gastrointestinal nematodes. We tested this hypothesis by establishing a murine coinfection model in which mice were infected with the intestinal nematode parasite Heligmosomoides polygyrus prior to infection with S. mansoni. In CBA mice that naturally display a severe form of schistosomiasis, preinfection with H. polygyrus resulted in a marked reduction in schistosome egg-induced hepatic immunopathology, which was associated with significant decreases in the levels of interleukin-17 (IL-17), gamma interferon, tumor necrosis factor alpha, IL-23, IL-6, and IL-1beta and with increases in the levels of IL-4, IL-5, IL-10, and transforming growth factor beta in mesenteric lymph node cells, purified CD4 T cells, and isolated liver granuloma cells. There also were increases in liver Ym1 and forkhead box P3 transcription factor expression. In another model of high-pathology schistosomiasis induced in C57BL/6 mice by immunization with schistosome egg antigens in complete Freund's adjuvant, coinfection with the nematodes also resulted in a marked inhibition of hepatic immunopathology accompanied by similar shifts in cytokine production. These findings demonstrate that intestinal nematodes prevent Th1- and Th17-cell-mediated inflammation by promoting a strong Th2-polarized environment associated with increases in the levels of alternatively activated macrophages and T regulatory cells, which result in significant amelioration of schistosome-induced immunopathology.

Disruption of interleukin-27 signaling results in impaired gamma interferon production but does not significantly affect immunopathology in murine schistosome infection.

In schistosomiasis mansoni, parasite eggs cause hepatointestinal granulomatous inflammation and fibrosis mediated by CD4 T cells specific for egg antigens. The severity of disease varies extensively in humans and among mouse strains. Marked disease exacerbation induced in typically low-pathology C57BL/6 mice by immunization with schistosome egg antigens (SEA) in complete Freund's adjuvant (SEA/CFA) correlates with elevated production of the proinflammatory cytokines gamma interferon (IFN-gamma) and interleukin-17 (IL-17), which are regulated by IL-12 and IL-23, respectively. Here we examined the effect on the schistosome infection of a third member of the IL-12 family of heterodimeric cytokines, IL-27, using SEA/CFA-immunized and unimmunized mice deficient in the IL-27 receptor chain WSX-1 (WSX-1(-/-)). SEA-stimulated bulk mesenteric lymph node cells or CD4 T cells from 7-week-infected WSX-1(-/-) mice produced significantly less IFN-gamma than did those from C57BL/6 mice, even though there was no difference between these mice in exacerbated hepatic egg-induced granulomatous inflammation or in the levels of IL-17 induced by immunization with SEA/CFA. A fraction of the cells in the granulomas stained positive for IL-27, but there were no significant differences between WSX-1(-/-) and BL/6 mice, nor were there differences in the number of CD4 T cells and eosinophils. A 24-week chronic infection resulted in markedly reduced levels of proinflammatory cytokines, including IFN-gamma, in WSX-1(-/-) mice, but again the magnitude of immunopathology was not significantly different between the two groups. These findings indicate that despite the impaired IFN-gamma production, IL-27 signaling has no significant effect on either the magnitude of egg-induced immunopathology or on its closest in vitro correlate, IL-17.