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Role of sphingomyelinase in infectious diseases caused by Bacillus cereus.

Oda M, Hashimoto M, Takahashi M, Ohmae Y, Seike S, Kato R, Fujita A, Tsuge H, Nagahama M, Ochi S, Sasahara T, Hayashi S, Hirai Y, Sakurai J - PLoS ONE (2012)

Bottom Line: Treatment of mouse macrophages with Bc-SMase resulted in a reduction in the generation of H(2)O(2) and phagocytosis of macrophages induced by peptidoglycan (PGN), but no effect on the release of TNF-α and little release of LDH under our experimental conditions.A photobleaching analysis suggested that the cells treated with Bc-SMase exhibited a reduction in membrane fluidity.The results suggest that Bc-SMase is essential for the hydrolysis of SM in membranes, leading to a reduction in phagocytosis.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Faculty of Pharmaceutical Sciences, Tokushima Bunri University, Yamashiro-cho, Tokushima, Japan.

ABSTRACT
Bacillus cereus (B. cereus) is a pathogen in opportunistic infections. Here we show that Bacillus cereus sphingomyelinase (Bc-SMase) is a virulence factor for septicemia. Clinical isolates produced large amounts of Bc-SMase, grew in vivo, and caused death among mice, but ATCC strains isolated from soil did not. A transformant of the ATCC strain carrying a recombinant plasmid containing the Bc-SMase gene grew in vivo, but that with the gene for E53A, which has little enzymatic activity, did not. Administration of an anti-Bc-SMase antibody and immunization against Bc-SMase prevented death caused by the clinical isolates, showing that Bc-SMase plays an important role in the diseases caused by B. cereus. Treatment of mouse macrophages with Bc-SMase resulted in a reduction in the generation of H(2)O(2) and phagocytosis of macrophages induced by peptidoglycan (PGN), but no effect on the release of TNF-α and little release of LDH under our experimental conditions. Confocal laser microscopy showed that the treatment of mouse macrophages with Bc-SMase resulted in the formation of ceramide-rich domains. A photobleaching analysis suggested that the cells treated with Bc-SMase exhibited a reduction in membrane fluidity. The results suggest that Bc-SMase is essential for the hydrolysis of SM in membranes, leading to a reduction in phagocytosis.

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Effect of Bc-SMase on activation of mouse macrophages.Mouse macrophages were incubated with or without Bc-SMase at 37°C for 60 min (D), and then treated with PGN (5 µg/ml) for 60 min (A, B, C). H2O2 production, phagocytosis, TNF-α release, and LDH release were measured as described in Materials and Methods. Values represent the mean ± SEM; n = 7; * P<0.01 compared with H2O2 production or phagocytosis induced by PGN alone.
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pone-0038054-g006: Effect of Bc-SMase on activation of mouse macrophages.Mouse macrophages were incubated with or without Bc-SMase at 37°C for 60 min (D), and then treated with PGN (5 µg/ml) for 60 min (A, B, C). H2O2 production, phagocytosis, TNF-α release, and LDH release were measured as described in Materials and Methods. Values represent the mean ± SEM; n = 7; * P<0.01 compared with H2O2 production or phagocytosis induced by PGN alone.

Mentions: González Zorn et al. reported that the SMase from Listeria ivanovii mediates bacterial escape from phagocytic cells [15]. The activation of macrophages is known to be related to bactericidal action in vivo. To investigate the effect of Bc-SMase on the activation of macrophages, we assessed the effect of PGN, an activator of macrophages, on macrophages treated with Bc-SMase. Bc-SMase attenuated PGN-activated H2O2 generation and phagocytosis of macrophages in a dose-dependent manner (Fig. 6A and 6B). However, Bc-SMase had no effect on the release of TNF-α induced by PGN from macrophages and induced no release of lactate dehydrogenase (LDH) from the cells (Fig. 6C and 6D). It therefore is likely that Bc-SMase specifically influences H2O2 generation and phagocytosis without impairing membranes of macrophages, suggesting that treatment of macrophages with Bc-SMase results in a change in function of the membranes. It was thought that the frustrated phagocytosis may be dependent on the formation of ceramide in macrophage membranes.


Role of sphingomyelinase in infectious diseases caused by Bacillus cereus.

Oda M, Hashimoto M, Takahashi M, Ohmae Y, Seike S, Kato R, Fujita A, Tsuge H, Nagahama M, Ochi S, Sasahara T, Hayashi S, Hirai Y, Sakurai J - PLoS ONE (2012)

Effect of Bc-SMase on activation of mouse macrophages.Mouse macrophages were incubated with or without Bc-SMase at 37°C for 60 min (D), and then treated with PGN (5 µg/ml) for 60 min (A, B, C). H2O2 production, phagocytosis, TNF-α release, and LDH release were measured as described in Materials and Methods. Values represent the mean ± SEM; n = 7; * P<0.01 compared with H2O2 production or phagocytosis induced by PGN alone.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3368938&req=5

pone-0038054-g006: Effect of Bc-SMase on activation of mouse macrophages.Mouse macrophages were incubated with or without Bc-SMase at 37°C for 60 min (D), and then treated with PGN (5 µg/ml) for 60 min (A, B, C). H2O2 production, phagocytosis, TNF-α release, and LDH release were measured as described in Materials and Methods. Values represent the mean ± SEM; n = 7; * P<0.01 compared with H2O2 production or phagocytosis induced by PGN alone.
Mentions: González Zorn et al. reported that the SMase from Listeria ivanovii mediates bacterial escape from phagocytic cells [15]. The activation of macrophages is known to be related to bactericidal action in vivo. To investigate the effect of Bc-SMase on the activation of macrophages, we assessed the effect of PGN, an activator of macrophages, on macrophages treated with Bc-SMase. Bc-SMase attenuated PGN-activated H2O2 generation and phagocytosis of macrophages in a dose-dependent manner (Fig. 6A and 6B). However, Bc-SMase had no effect on the release of TNF-α induced by PGN from macrophages and induced no release of lactate dehydrogenase (LDH) from the cells (Fig. 6C and 6D). It therefore is likely that Bc-SMase specifically influences H2O2 generation and phagocytosis without impairing membranes of macrophages, suggesting that treatment of macrophages with Bc-SMase results in a change in function of the membranes. It was thought that the frustrated phagocytosis may be dependent on the formation of ceramide in macrophage membranes.

Bottom Line: Treatment of mouse macrophages with Bc-SMase resulted in a reduction in the generation of H(2)O(2) and phagocytosis of macrophages induced by peptidoglycan (PGN), but no effect on the release of TNF-α and little release of LDH under our experimental conditions.A photobleaching analysis suggested that the cells treated with Bc-SMase exhibited a reduction in membrane fluidity.The results suggest that Bc-SMase is essential for the hydrolysis of SM in membranes, leading to a reduction in phagocytosis.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Faculty of Pharmaceutical Sciences, Tokushima Bunri University, Yamashiro-cho, Tokushima, Japan.

ABSTRACT
Bacillus cereus (B. cereus) is a pathogen in opportunistic infections. Here we show that Bacillus cereus sphingomyelinase (Bc-SMase) is a virulence factor for septicemia. Clinical isolates produced large amounts of Bc-SMase, grew in vivo, and caused death among mice, but ATCC strains isolated from soil did not. A transformant of the ATCC strain carrying a recombinant plasmid containing the Bc-SMase gene grew in vivo, but that with the gene for E53A, which has little enzymatic activity, did not. Administration of an anti-Bc-SMase antibody and immunization against Bc-SMase prevented death caused by the clinical isolates, showing that Bc-SMase plays an important role in the diseases caused by B. cereus. Treatment of mouse macrophages with Bc-SMase resulted in a reduction in the generation of H(2)O(2) and phagocytosis of macrophages induced by peptidoglycan (PGN), but no effect on the release of TNF-α and little release of LDH under our experimental conditions. Confocal laser microscopy showed that the treatment of mouse macrophages with Bc-SMase resulted in the formation of ceramide-rich domains. A photobleaching analysis suggested that the cells treated with Bc-SMase exhibited a reduction in membrane fluidity. The results suggest that Bc-SMase is essential for the hydrolysis of SM in membranes, leading to a reduction in phagocytosis.

Show MeSH
Related in: MedlinePlus