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Granzyme-mediated regulation of host defense in the liver in experimental Leishmania donovani infection.

ABSTRACT: In the livers of susceptible C57BL/6 (B6) mice infected with Leishmania donovani, CD8(+) T cell mechanisms are required for granuloma assembly, macrophage activation, intracellular parasite killing, and self-cure. Since gene expression of perforin and granzymes A and B (GzmA and GzmB), cytolytic proteins linked to CD8(+) cell effector function, was enhanced in infected liver tissue, B6 mice deficient in these granular proteins were used to gauge host defense roles. Neither perforin nor GzmA was required; however, mice deficient in GzmB (GzmB(-/-), GzmB cluster(-/-), and GzmA×B cluster double knockout [DKO] mice) showed both delayed granuloma assembly and initially impaired control of parasite replication. Since these two defects in B6 mice were limited to early-stage infection, innately resistant 129/Sv mice were also tested. In this genetic setting, expression of both innate and subsequent T (Th1) cell-dependent acquired resistance, including the self-cure phenotype, was entirely derailed in GzmA×B cluster DKO mice. These results, in susceptible B6 mice for GzmB and in resistant 129/Sv mice for GzmA and/or the GzmB cluster, point to granzyme-mediated host defense regulation in the liver in experimental visceral leishmaniasis.

SUBMITTER: Murray HW

PROVIDER: S-EPMC4294235 | biostudies-literature | 2015 Feb

REPOSITORIES: biostudies-literature

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Granzyme-mediated regulation of host defense in the liver in experimental Leishmania donovani infection.

Murray Henry W HW Mitchell-Flack Marisa M Zheng Hua H Ma Xiaojing X

Infection and immunity 20141201 2

In the livers of susceptible C57BL/6 (B6) mice infected with Leishmania donovani, CD8(+) T cell mechanisms are required for granuloma assembly, macrophage activation, intracellular parasite killing, and self-cure. Since gene expression of perforin and granzymes A and B (GzmA and GzmB), cytolytic proteins linked to CD8(+) cell effector function, was enhanced in infected liver tissue, B6 mice deficient in these granular proteins were used to gauge host defense roles. Neither perforin nor GzmA was ...[more]

PMID: 25452549

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Project description:Little is known about the protective role of inflammatory processes in modulating lipid metabolism in infection. Here we report an intimate link between the innate immune response to infection and regulation of the sterol metabolic network characterized by down-regulation of sterol biosynthesis by an interferon regulatory loop mechanism. In time-series experiments profiling genome-wide lipid-associated gene expression of macrophages, we show a selective and coordinated negative regulation of the complete sterol pathway upon viral infection or cytokine treatment with IFNγ or β but not TNF, IL1β, or IL6. Quantitative analysis at the protein level of selected sterol metabolic enzymes upon infection shows a similar level of suppression. Experimental testing of sterol metabolite levels using lipidomic-based measurements shows a reduction in metabolic output. On the basis of pharmacologic and RNAi inhibition of the sterol pathway we show augmented protection against viral infection, and in combination with metabolite rescue experiments, we identify the requirement of the mevalonate-isoprenoid branch of the sterol metabolic network in the protective response upon statin or IFNβ treatment. Conditioned media experiments from infected cells support an involvement of secreted type 1 interferon(s) to be sufficient for reducing the sterol pathway upon infection. Moreover, we show that infection of primary macrophages containing a genetic knockout of the major type I interferon, IFNβ, leads to only a partial suppression of the sterol pathway, while genetic knockout of the receptor for all type I interferon family members, ifnar1, or associated signaling component, tyk2, completely abolishes the reduction of the sterol biosynthetic activity upon infection. Levels of the proteolytically cleaved nuclear forms of SREBP2, a key transcriptional regulator of sterol biosynthesis, are reduced upon infection and IFNβ treatment at both the protein and de novo transcription level. The reduction in srebf2 gene transcription upon infection and IFN treatment is also found to be strictly dependent on ifnar1. Altogether these results show that type 1 IFN signaling is both necessary and sufficient for reducing the sterol metabolic network activity upon infection, thereby linking the regulation of the sterol pathway with interferon anti-viral defense responses. These findings bring a new link between sterol metabolism and interferon antiviral response and support the idea of using host metabolic modifiers of innate immunity as a potential antiviral strategy.

Project description:IntroductionSpatial analysis of lipids in inflammatory microenvironments is key to understand the pathogenesis of infectious disease. Granulomatous inflammation is a hallmark of leishmaniasis and changes in host and parasite lipid metabolism have been observed at the bulk tissue level in various infection models. Here, mass spectrometry imaging (MSI) is applied to spatially map hepatic lipid composition following infection with Leishmania donovani, an experimental mouse model of visceral leishmaniasis.MethodsLivers from naïve and L. donovani-infected C57BL/6 mice were harvested at 14- and 20-days post-infection (n=5 per time point). 12 µm transverse sections were cut and covered with norhamane, prior to lipid analysis using MALDI-MSI. MALDI-MSI was performed in negative mode on a Rapiflex (Bruker Daltonics) at 5 and 50 µm spatial resolution and data-dependent analysis (DDA) on an Orbitrap-Elite (Thermo-Scientific) at 50 µm spatial resolution for structural identification analysis of lipids.ResultsAberrant lipid abundances were observed in a heterogeneous distribution across infected mouse livers compared to naïve mouse liver. Distinctive localized correlated lipid masses were found in granulomas and surrounding parenchymal tissue. Structural identification revealed 40 different lipids common to naïve and d14/d20 infected mouse livers, whereas 15 identified lipids were only detected in infected mouse livers. For pathology-guided MSI imaging, we deduced lipids from manually annotated granulomatous and parenchyma regions of interests (ROIs), identifying 34 lipids that showed significantly different intensities between parenchyma and granulomas across all infected livers.DiscussionOur results identify specific lipids that spatially correlate to the major histopathological feature of Leishmania donovani infection in the liver, viz. hepatic granulomas. In addition, we identified a three-fold increase in the number of unique phosphatidylglycerols (PGs) in infected liver tissue and provide direct evidence that arachidonic acid-containing phospholipids are localized with hepatic granulomas. These phospholipids may serve as important precursors for downstream oxylipin generation with consequences for the regulation of the inflammatory cascade. This study provides the first description of the use of MSI to define spatial-temporal lipid changes at local sites of infection induced by Leishmania donovani in mice.

Dataset Information

Granzyme-mediated regulation of host defense in the liver in experimental Leishmania donovani infection.

Publications

Granzyme-mediated regulation of host defense in the liver in experimental Leishmania donovani infection.

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