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Galectin-1 exerts inhibitory effects during DENV-1 infection.

Toledo KA, Fermino ML, Andrade Cdel C, Riul TB, Alves RT, Muller VD, Russo RR, Stowell SR, Cummings RD, Aquino VH, Dias-Baruffi M - PLoS ONE (2014)

Bottom Line: We found that DENV-1 infection of cells in vitro exhibited caused decreased expression of Gal-1 in several different human cell lines, suggesting that loss of Gal-1 is associated with virus production.Importantly, the inhibition was specific for hrGal-1, since no effect was observed using recombinant human galectin-3.These results provide novel insights into the functions of Gal-1 in resistance to DENV infection and suggest that Gal-1 should be explored as a potential antiviral compound.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Universidade Estadual Paulista - UNESP (FCL-Assis), Assis, Brazil.

ABSTRACT
Dengue virus (DENV) is an enveloped RNA virus that is mosquito-transmitted and can infect a variety of immune and non-immune cells. Response to infection ranges from asymptomatic disease to a severe disorder known as dengue hemorrhagic fever. Despite efforts to control the disease, there are no effective treatments or vaccines. In our search for new antiviral compounds to combat infection by dengue virus type 1 (DENV-1), we investigated the role of galectin-1, a widely-expressed mammalian lectin with functions in cell-pathogen interactions and immunoregulatory properties. We found that DENV-1 infection of cells in vitro exhibited caused decreased expression of Gal-1 in several different human cell lines, suggesting that loss of Gal-1 is associated with virus production. In test of this hypothesis we found that exogenous addition of human recombinant Gal-1 (hrGal-1) inhibits the virus production in the three different cell types. This inhibitory effect was dependent on hrGal-1 dimerization and required its carbohydrate recognition domain. Importantly, the inhibition was specific for hrGal-1, since no effect was observed using recombinant human galectin-3. Interestingly, we found that hrGal-1 directly binds to dengue virus and acts, at least in part, during the early stages of DENV-1 infection, by inhibiting viral adsorption and its internalization to target cells. To test the in vivo role of Gal-1 in DENV infection, Gal-1-deficient-mice were used to demonstrate that the expression of endogenous Galectin-1 contributes to resistance of macrophages to in vitro-infection with DENV-1 and it is also important to physiological susceptibility of mice to in vivo infection with DENV-1. These results provide novel insights into the functions of Gal-1 in resistance to DENV infection and suggest that Gal-1 should be explored as a potential antiviral compound.

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Gal-1 is physiologically relevant to in vivo infection with DENV-1.(A) Newborn WT and Lgals1–/– mice were intracerebraly infected with 106 PFU/ml of DENV-1 mouse-brain adapted or mock (supernatant from mouse brain not infected with DENV-1) and their mortality was monitored for 10 days. Results are shown as the percentage survival from five independent assays performed with 6–8 mice per group. (B) Total mRNA isolated from resident peritoneal macrophages of Gal-1-deficient (Lgals1–/–) and wild-type (WT) mice was converted into cDNA and the Gal-1 expression was analyzed using conventional PCR. The β-actin gene was used as an endogenous control. Alternatively, total protein from Lgals1–/– and WT macrophages were isolated and Gal-1 expression was quantified by western blot assay and normalized to β-actin expression. (C) Peritoneal resident macrophages from Lgals1–/– or WT mice were cultivated (5x105/well) in 24-well plates and inoculated with DENV-1 at a MOI of 0.5. After 72 hours at 37°C, cell-free supernatants were recovered and the viral loads were determined by Real-Time PCR. Results are showed as Viral RNA amounts equivalent to PFU/ml±SD from three experiments assessed in triplicates. ***p<0.0001.
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pone-0112474-g005: Gal-1 is physiologically relevant to in vivo infection with DENV-1.(A) Newborn WT and Lgals1–/– mice were intracerebraly infected with 106 PFU/ml of DENV-1 mouse-brain adapted or mock (supernatant from mouse brain not infected with DENV-1) and their mortality was monitored for 10 days. Results are shown as the percentage survival from five independent assays performed with 6–8 mice per group. (B) Total mRNA isolated from resident peritoneal macrophages of Gal-1-deficient (Lgals1–/–) and wild-type (WT) mice was converted into cDNA and the Gal-1 expression was analyzed using conventional PCR. The β-actin gene was used as an endogenous control. Alternatively, total protein from Lgals1–/– and WT macrophages were isolated and Gal-1 expression was quantified by western blot assay and normalized to β-actin expression. (C) Peritoneal resident macrophages from Lgals1–/– or WT mice were cultivated (5x105/well) in 24-well plates and inoculated with DENV-1 at a MOI of 0.5. After 72 hours at 37°C, cell-free supernatants were recovered and the viral loads were determined by Real-Time PCR. Results are showed as Viral RNA amounts equivalent to PFU/ml±SD from three experiments assessed in triplicates. ***p<0.0001.

Mentions: Based on the results presented so far and to better demonstrate the role of Gal-1 in controlling DENV-1 infection, we next infected newborn Gal-1-deficient (Lgals1–/–) and wild-type mice with mouse-brain adapted DENV-1 virus and analyzed host survival for 10 days (Figure 5A). Although mice from both lineages displayed similar overall mortality rate, Lgals1–/– mice began to succumb earlier than WT mice: within the first 4 days post-infection, almost 60% of Lgals1–/– mice died, while 90% of WT mice survived up to 5 days past challenge (Figure 5A).


Galectin-1 exerts inhibitory effects during DENV-1 infection.

Toledo KA, Fermino ML, Andrade Cdel C, Riul TB, Alves RT, Muller VD, Russo RR, Stowell SR, Cummings RD, Aquino VH, Dias-Baruffi M - PLoS ONE (2014)

Gal-1 is physiologically relevant to in vivo infection with DENV-1.(A) Newborn WT and Lgals1–/– mice were intracerebraly infected with 106 PFU/ml of DENV-1 mouse-brain adapted or mock (supernatant from mouse brain not infected with DENV-1) and their mortality was monitored for 10 days. Results are shown as the percentage survival from five independent assays performed with 6–8 mice per group. (B) Total mRNA isolated from resident peritoneal macrophages of Gal-1-deficient (Lgals1–/–) and wild-type (WT) mice was converted into cDNA and the Gal-1 expression was analyzed using conventional PCR. The β-actin gene was used as an endogenous control. Alternatively, total protein from Lgals1–/– and WT macrophages were isolated and Gal-1 expression was quantified by western blot assay and normalized to β-actin expression. (C) Peritoneal resident macrophages from Lgals1–/– or WT mice were cultivated (5x105/well) in 24-well plates and inoculated with DENV-1 at a MOI of 0.5. After 72 hours at 37°C, cell-free supernatants were recovered and the viral loads were determined by Real-Time PCR. Results are showed as Viral RNA amounts equivalent to PFU/ml±SD from three experiments assessed in triplicates. ***p<0.0001.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4231055&req=5

pone-0112474-g005: Gal-1 is physiologically relevant to in vivo infection with DENV-1.(A) Newborn WT and Lgals1–/– mice were intracerebraly infected with 106 PFU/ml of DENV-1 mouse-brain adapted or mock (supernatant from mouse brain not infected with DENV-1) and their mortality was monitored for 10 days. Results are shown as the percentage survival from five independent assays performed with 6–8 mice per group. (B) Total mRNA isolated from resident peritoneal macrophages of Gal-1-deficient (Lgals1–/–) and wild-type (WT) mice was converted into cDNA and the Gal-1 expression was analyzed using conventional PCR. The β-actin gene was used as an endogenous control. Alternatively, total protein from Lgals1–/– and WT macrophages were isolated and Gal-1 expression was quantified by western blot assay and normalized to β-actin expression. (C) Peritoneal resident macrophages from Lgals1–/– or WT mice were cultivated (5x105/well) in 24-well plates and inoculated with DENV-1 at a MOI of 0.5. After 72 hours at 37°C, cell-free supernatants were recovered and the viral loads were determined by Real-Time PCR. Results are showed as Viral RNA amounts equivalent to PFU/ml±SD from three experiments assessed in triplicates. ***p<0.0001.
Mentions: Based on the results presented so far and to better demonstrate the role of Gal-1 in controlling DENV-1 infection, we next infected newborn Gal-1-deficient (Lgals1–/–) and wild-type mice with mouse-brain adapted DENV-1 virus and analyzed host survival for 10 days (Figure 5A). Although mice from both lineages displayed similar overall mortality rate, Lgals1–/– mice began to succumb earlier than WT mice: within the first 4 days post-infection, almost 60% of Lgals1–/– mice died, while 90% of WT mice survived up to 5 days past challenge (Figure 5A).

Bottom Line: We found that DENV-1 infection of cells in vitro exhibited caused decreased expression of Gal-1 in several different human cell lines, suggesting that loss of Gal-1 is associated with virus production.Importantly, the inhibition was specific for hrGal-1, since no effect was observed using recombinant human galectin-3.These results provide novel insights into the functions of Gal-1 in resistance to DENV infection and suggest that Gal-1 should be explored as a potential antiviral compound.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Universidade Estadual Paulista - UNESP (FCL-Assis), Assis, Brazil.

ABSTRACT
Dengue virus (DENV) is an enveloped RNA virus that is mosquito-transmitted and can infect a variety of immune and non-immune cells. Response to infection ranges from asymptomatic disease to a severe disorder known as dengue hemorrhagic fever. Despite efforts to control the disease, there are no effective treatments or vaccines. In our search for new antiviral compounds to combat infection by dengue virus type 1 (DENV-1), we investigated the role of galectin-1, a widely-expressed mammalian lectin with functions in cell-pathogen interactions and immunoregulatory properties. We found that DENV-1 infection of cells in vitro exhibited caused decreased expression of Gal-1 in several different human cell lines, suggesting that loss of Gal-1 is associated with virus production. In test of this hypothesis we found that exogenous addition of human recombinant Gal-1 (hrGal-1) inhibits the virus production in the three different cell types. This inhibitory effect was dependent on hrGal-1 dimerization and required its carbohydrate recognition domain. Importantly, the inhibition was specific for hrGal-1, since no effect was observed using recombinant human galectin-3. Interestingly, we found that hrGal-1 directly binds to dengue virus and acts, at least in part, during the early stages of DENV-1 infection, by inhibiting viral adsorption and its internalization to target cells. To test the in vivo role of Gal-1 in DENV infection, Gal-1-deficient-mice were used to demonstrate that the expression of endogenous Galectin-1 contributes to resistance of macrophages to in vitro-infection with DENV-1 and it is also important to physiological susceptibility of mice to in vivo infection with DENV-1. These results provide novel insights into the functions of Gal-1 in resistance to DENV infection and suggest that Gal-1 should be explored as a potential antiviral compound.

Show MeSH
Related in: MedlinePlus