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Host defense against viral infection involves interferon mediated down-regulation of sterol biosynthesis.

Blanc M, Hsieh WY, Robertson KA, Watterson S, Shui G, Lacaze P, Khondoker M, Dickinson P, Sing G, Rodríguez-Martín S, Phelan P, Forster T, Strobl B, Müller M, Riemersma R, Osborne T, Wenk MR, Angulo A, Ghazal P - PLoS Biol. (2011)

Bottom Line: Experimental testing of sterol metabolite levels using lipidomic-based measurements shows a reduction in metabolic output.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.

View Article: PubMed Central - PubMed

Affiliation: Division of Pathway Medicine and Centre for Infectious Diseases, University of Edinburgh, Edinburgh, United Kingdom.

ABSTRACT
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.

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Effect of statins on mCMV growth in vitro and in vivo.(A) NIH/3T3 cells were infected with mCMV-GFP (MOI of 0.2) and subsequently treated with varying concentrations of Simvastatin or Gancyclovir immediately after infection. GFP expression was measured to monitor the level of infection (Materials and Methods). Graph represents the percentage of viral inhibition as a function of drug treatment. Data points represent mean ± SD of two independent experiments with six biological replicates for each experiment. (B) Mice were fed with simvastatin (50 mg/kg/mice) daily for 5 d by gavages and at day 1 post-treatment, were challenged with 2×106 PFU of mCMV by intraperitoneal injection, and sacrificed; at 4 dpi, viral titers in different organs were measured by plaque assay and are expressed per gram of tissue. Data points represent mean ± SD of two independent experiments with five mice per group for each experiment. *p<0.05, **p<0.01, ***p<0.001, determined with a Mann-Whitney U test.
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pbio-1000598-g004: Effect of statins on mCMV growth in vitro and in vivo.(A) NIH/3T3 cells were infected with mCMV-GFP (MOI of 0.2) and subsequently treated with varying concentrations of Simvastatin or Gancyclovir immediately after infection. GFP expression was measured to monitor the level of infection (Materials and Methods). Graph represents the percentage of viral inhibition as a function of drug treatment. Data points represent mean ± SD of two independent experiments with six biological replicates for each experiment. (B) Mice were fed with simvastatin (50 mg/kg/mice) daily for 5 d by gavages and at day 1 post-treatment, were challenged with 2×106 PFU of mCMV by intraperitoneal injection, and sacrificed; at 4 dpi, viral titers in different organs were measured by plaque assay and are expressed per gram of tissue. Data points represent mean ± SD of two independent experiments with five mice per group for each experiment. *p<0.05, **p<0.01, ***p<0.001, determined with a Mann-Whitney U test.

Mentions: To assess whether the sterol biosynthesis pathway plays a pro- or anti-viral role in regulating mCMV replication, we exploited the pharmacologic compound “simvastatin,” a potent and selective inhibitor of HMGCR [48]. Inhibition of HMGCR is known to result in a reduction of the metabolic intermediate mevalonate (Figure 3) and an accompanying drop in cholesterol synthesis by the cell [49]. The treatment of cells with simvastatin resulted in a dose-dependent inhibition of mCMV plaque formation (unpublished data) and in live cell replication assays (Figure 4A) with an IC50 of 2 µM that is comparable to the “gold standard” anti-viral Gancyclovir (Figure 4A) in the murine model system. Notably, the observed inhibitory effect of simvastatin occurred below a level at which non-specific toxic effects to cells were observed (15 µM) (Figure S7). These experiments pointed to a potential protective anti-viral role via a targeted disruption of the sterol pathway and raised the question of whether pharmacologic treatment in vivo also develops an inhibitory effect. To investigate whether simvastatin could play an anti-infective role in vivo, mice were administered with an established pre-clinical pharmacologic dose of simvastatin or vehicle alone and infected by intra-peritoneal inoculation with mCMV. Viral titres were then determined in a variety of organs at day 4 post-inoculation. Markedly, viral titres are reduced by over one order of magnitude in multiple organs following treatment with simvastatin (Figure 4B).


Host defense against viral infection involves interferon mediated down-regulation of sterol biosynthesis.

Blanc M, Hsieh WY, Robertson KA, Watterson S, Shui G, Lacaze P, Khondoker M, Dickinson P, Sing G, Rodríguez-Martín S, Phelan P, Forster T, Strobl B, Müller M, Riemersma R, Osborne T, Wenk MR, Angulo A, Ghazal P - PLoS Biol. (2011)

Effect of statins on mCMV growth in vitro and in vivo.(A) NIH/3T3 cells were infected with mCMV-GFP (MOI of 0.2) and subsequently treated with varying concentrations of Simvastatin or Gancyclovir immediately after infection. GFP expression was measured to monitor the level of infection (Materials and Methods). Graph represents the percentage of viral inhibition as a function of drug treatment. Data points represent mean ± SD of two independent experiments with six biological replicates for each experiment. (B) Mice were fed with simvastatin (50 mg/kg/mice) daily for 5 d by gavages and at day 1 post-treatment, were challenged with 2×106 PFU of mCMV by intraperitoneal injection, and sacrificed; at 4 dpi, viral titers in different organs were measured by plaque assay and are expressed per gram of tissue. Data points represent mean ± SD of two independent experiments with five mice per group for each experiment. *p<0.05, **p<0.01, ***p<0.001, determined with a Mann-Whitney U test.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3050939&req=5

pbio-1000598-g004: Effect of statins on mCMV growth in vitro and in vivo.(A) NIH/3T3 cells were infected with mCMV-GFP (MOI of 0.2) and subsequently treated with varying concentrations of Simvastatin or Gancyclovir immediately after infection. GFP expression was measured to monitor the level of infection (Materials and Methods). Graph represents the percentage of viral inhibition as a function of drug treatment. Data points represent mean ± SD of two independent experiments with six biological replicates for each experiment. (B) Mice were fed with simvastatin (50 mg/kg/mice) daily for 5 d by gavages and at day 1 post-treatment, were challenged with 2×106 PFU of mCMV by intraperitoneal injection, and sacrificed; at 4 dpi, viral titers in different organs were measured by plaque assay and are expressed per gram of tissue. Data points represent mean ± SD of two independent experiments with five mice per group for each experiment. *p<0.05, **p<0.01, ***p<0.001, determined with a Mann-Whitney U test.
Mentions: To assess whether the sterol biosynthesis pathway plays a pro- or anti-viral role in regulating mCMV replication, we exploited the pharmacologic compound “simvastatin,” a potent and selective inhibitor of HMGCR [48]. Inhibition of HMGCR is known to result in a reduction of the metabolic intermediate mevalonate (Figure 3) and an accompanying drop in cholesterol synthesis by the cell [49]. The treatment of cells with simvastatin resulted in a dose-dependent inhibition of mCMV plaque formation (unpublished data) and in live cell replication assays (Figure 4A) with an IC50 of 2 µM that is comparable to the “gold standard” anti-viral Gancyclovir (Figure 4A) in the murine model system. Notably, the observed inhibitory effect of simvastatin occurred below a level at which non-specific toxic effects to cells were observed (15 µM) (Figure S7). These experiments pointed to a potential protective anti-viral role via a targeted disruption of the sterol pathway and raised the question of whether pharmacologic treatment in vivo also develops an inhibitory effect. To investigate whether simvastatin could play an anti-infective role in vivo, mice were administered with an established pre-clinical pharmacologic dose of simvastatin or vehicle alone and infected by intra-peritoneal inoculation with mCMV. Viral titres were then determined in a variety of organs at day 4 post-inoculation. Markedly, viral titres are reduced by over one order of magnitude in multiple organs following treatment with simvastatin (Figure 4B).

Bottom Line: Experimental testing of sterol metabolite levels using lipidomic-based measurements shows a reduction in metabolic output.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.

View Article: PubMed Central - PubMed

Affiliation: Division of Pathway Medicine and Centre for Infectious Diseases, University of Edinburgh, Edinburgh, United Kingdom.

ABSTRACT
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.

Show MeSH
Related in: MedlinePlus