IFN-γ mediates Paneth cell death via suppression of mTOR.

Paneth cells constitutively produce antimicrobial peptides and growth factors that allow for intestinal homeostasis, host protection, and intestinal stem cell replication. Paneth cells rely heavily on the glycolytic metabolic program, which is in part controlled by the kinase complex Mechanistic target of rapamycin (mTORC1). Yet, little is known about mTOR importance in Paneth cell integrity under steady-state and inflammatory conditions. Our results demonstrate that IFN-γ, a crucial mediator of the intestinal inflammation, acts directly on murine Paneth cells to alter their mitochondrial integrity and membrane potential, resulting in an TORC1-dependent cell death mechanism distinct from canonical cell death pathways including apoptosis, necroptosis, and pyroptosis. These results were established with the purified cytokine and a physiologically relevant common Th1-inducing human parasite Toxoplasma gondii. Given the crucial role for IFN-γ, which is a cytokine frequently associated with the development of inflammatory bowel disease and compromised Paneth cell functions, the identified mechanisms underlying mTORC1-dependent Paneth cell death downstream of IFN-γ may provide promising novel approaches for treating intestinal inflammation.

An Assessment of Siderophore Production, Mucoviscosity, and Mouse Infection Models for Defining the Virulence Spectrum of Hypervirulent Klebsiella pneumoniae.

Hypervirulent Klebsiella pneumoniae (hvKp) bacteria are more virulent than classical K. pneumoniae (cKp) with resultant differences in clinical manifestations and management. It is unclear whether all hvKp isolates share a similar pathogenic potential. This report assessed the utility of siderophore production, mucoviscosity, and murine infection for defining the virulence spectrum of hvKp. Three strain cohorts were identified and defined based on the CD1 mouse subcutaneous (SQ) challenge model: (i) fully virulent hvKp strains (fvhvKp), lethal at a challenge inoculum (CI) of ≤103 CFU; (ii) partially virulent hvKp strains (pvhvKp), lethal at a CI of >103 to 107 CFU; (iii) classical K. pneumoniae, not lethal at a CI of 107 CFU. Quantitative siderophore and mucoviscosity assays differentiated fvhvKp and pvhvKp strains from cKp strains but were unable to differentiate between the fvhvKP and pvhvKP strain cohorts. However, SQ challenge of CD1 mice and intraperitoneal (IP) challenge of CD1 and BALB/c mice, but not C57BL/6 mice, were able to discriminate between an fvhvKp and a pvhvKp strain; SQ challenge of CD1 mice may have the greatest sensitivity. cKp was differentiated from hvKp both by SQ challenge of CD1 mice and IP challenge of all three mouse strains. These data identify a means to define the relative virulence of hvKP strains. It remains unclear whether the observed differences of hvKp virulence in mice translates to human infection. However, these data can be used to sort random collections of K. pneumoniae strains into fvhvKp and pvhvKp strain cohorts and assess for differences in clinical manifestations and outcomes.IMPORTANCE The pathogenic potential of hvKp strains is primarily mediated by a large virulence plasmid. The minimal set of genes required for the full expression of the hypervirulent phenotype is undefined. A number of reports describe hvKp strains possessing only a portion of the virulence plasmid; the clinical consequences of this are unclear. Therefore, the goal of this report was to determine whether virulence among hvKp strains varied and, if so, how to best identify the relative virulence of hvKp isolates. Data demonstrate hvKp pathogenic potential varies in CD1 and BALB/c murine infection models. In contrast, measurements of siderophore production and mucoviscosity were unable to discriminate the differences in hvKp isolate virulence observed in mice. This information can be used in future studies to determine the mechanisms responsible for differences between fully virulent hvKp and partially virulent hvKp and whether the differences observed in mice translate to disease in humans.

T-bet-dependent ILC1- and NK cell-derived IFN-γ mediates cDC1-dependent host resistance against Toxoplasma gondii.

Host resistance against intracellular pathogens requires a rapid IFN-γ mediated immune response. We reveal that T-bet-dependent production of IFN-γ is essential for the maintenance of inflammatory DCs at the site of infection with a common protozoan parasite, Toxoplasma gondii. A detailed analysis of the cellular sources for T-bet-dependent IFN-γ identified that ILC1s and to a lesser degree NK, but not TH1 cells, were involved in the regulation of inflammatory DCs via IFN-γ. Mechanistically, we established that T-bet dependent innate IFN-γ is critical for the induction of IRF8, an essential transcription factor for cDC1s. Failure to upregulate IRF8 in DCs resulted in acute susceptibility to T. gondii infection. Our data identifies that T-bet dependent production of IFN-γ by ILC1 and NK cells is indispensable for host resistance against intracellular infection via maintaining IRF8+ inflammatory DCs at the site of infection.

TLR11-independent inflammasome activation is critical for CD4+ T cell-derived IFN-γ production and host resistance to Toxoplasma gondii.

Innate recognition of invading intracellular pathogens is essential for regulating robust and rapid CD4+ T cell effector function, which is critical for host-mediated immunity. The intracellular apicomplexan parasite, Toxoplasma gondii, is capable of infecting almost any nucleated cell of warm-blooded animals, including humans, and establishing tissue cysts that persist throughout the lifetime of the host. Recognition of T. gondii by TLRs is essential for robust IL-12 and IFN-γ production, two major cytokines involved in host resistance to the parasite. In the murine model of infection, robust IL-12 and IFN-γ production have been largely attributed to T. gondii profilin recognition by the TLR11 and TLR12 heterodimer complex, resulting in Myd88-dependent IL-12 production. However, TLR11 or TLR12 deficiency failed to recapitulate the acute susceptibility to T. gondii infection seen in Myd88-/- mice. T. gondii triggers inflammasome activation in a caspase-1-dependent manner resulting in cytokine release; however, it remains undetermined if parasite-mediated inflammasome activation impacts IFN-γ production and host resistance to the parasite. Using mice which lack different inflammasome components, we observed that the inflammasome played a limited role in host resistance when TLR11 remained functional. Strikingly, in the absence of TLR11, caspase-1 and -11 played a significant role for robust CD4+ TH1-derived IFN-γ responses and host survival. Moreover, we demonstrated that in the absence of TLR11, production of the caspase-1-dependent cytokine IL-18 was sufficient and necessary for CD4+ T cell-derived IFN-γ responses. Mechanistically, we established that T. gondii-mediated activation of the inflammasome and IL-18 were critical to maintain robust CD4+ TH1 IFN-γ responses during parasite infection in the absence of TLR11.

Alarmin S100A11 initiates a chemokine response to the human pathogen Toxoplasma gondii.

Toxoplasma gondii is a common protozoan parasite that infects up to one third of the world's population. Notably, very little is known about innate immune sensing mechanisms for this obligate intracellular parasite by human cells. Here, by applying an unbiased biochemical screening approach, we show that human monocytes recognized the presence of T. gondii infection by detecting the alarmin S100A11 protein, which is released from parasite-infected cells via caspase-1-dependent mechanisms. S100A11 induced a potent chemokine response to T. gondii by engaging its receptor RAGE, and regulated monocyte recruitment in vivo by inducing expression of the chemokine CCL2. Our experiments reveal a sensing system for T. gondii by human cells that is based on the detection of infection-mediated release of S100A11 and RAGE-dependent induction of CCL2, a crucial chemokine required for host resistance to the parasite.