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Neutrophils are resistant to Yersinia YopJ/P-induced apoptosis and are protected from ROS-mediated cell death by the type III secretion system.

Spinner JL, Seo KS, O'Loughlin JL, Cundiff JA, Minnich SA, Bohach GA, Kobayashi SD - PLoS ONE (2010)

Bottom Line: In contrast, interaction of human PMNs with the virulence plasmid-deficient Y. pestis strain KIM6 resulted in increased surface exposure of phosphatidylserine (PS) and cell death.PMN reactive oxygen species (ROS) production was inhibited in a virulence plasmid-dependent but YopJ/YopP-independent manner.Following phagocytic interaction with Y. pestis strain KIM6, inhibition of PMN ROS production with diphenyleneiodonium chloride resulted in a reduction of PMN cell death similar to that induced by the virulence plasmid-containing strain Y. pestis KIM5.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Molecular Biology and Biochemistry, University of Idaho, Moscow, Idaho, United States of America.

ABSTRACT

Background: The human innate immune system relies on the coordinated activity of macrophages and polymorphonuclear leukocytes (neutrophils or PMNs) for defense against bacterial pathogens. Yersinia spp. subvert the innate immune response to cause disease in humans. In particular, the Yersinia outer protein YopJ (Y. pestis and Y. pseudotuberculosis) and YopP (Y. enterocolitica) rapidly induce apoptosis in murine macrophages and dendritic cells. However, the effects of Yersinia Yop J/P on neutrophil fate are not clearly defined.

Methodology/principal findings: In this study, we utilized wild-type and mutant strains of Yersinia to test the contribution of YopJ and YopP on induction of apoptosis in human monocyte-derived macrophages (HMDM) and neutrophils. Whereas YopJ and YopP similarly induced apoptosis in HMDMs, interaction of human neutrophils with virulence plasmid-containing Yersinia did not result in PMN caspase activation, release of LDH, or loss of membrane integrity greater than PMN controls. In contrast, interaction of human PMNs with the virulence plasmid-deficient Y. pestis strain KIM6 resulted in increased surface exposure of phosphatidylserine (PS) and cell death. PMN reactive oxygen species (ROS) production was inhibited in a virulence plasmid-dependent but YopJ/YopP-independent manner. Following phagocytic interaction with Y. pestis strain KIM6, inhibition of PMN ROS production with diphenyleneiodonium chloride resulted in a reduction of PMN cell death similar to that induced by the virulence plasmid-containing strain Y. pestis KIM5.

Conclusions: Our findings showed that Yersinia YopJ and/or YopP did not induce pronounced apoptosis in human neutrophils. Furthermore, robust PMN ROS production in response to virulence plasmid-deficient Yersinia was associated with increased PMN cell death, suggesting that Yersinia inhibition of PMN ROS production plays a role in evasion of the human innate immune response in part by limiting PMN apoptosis.

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Caspase-3, -8, -9, and -2 activity in PMNs following incubation with Yersinia.PMNs were incubated alone or with Y. pestis KIM5, KIM5 YopJ-YopP, KIM5ΔyopJ, KIM6 and Y. enterocolitica 8081v grown at 37°C. Caspase-3, -8, -9, and -2 activity was measured after 3 h and 6 h of incubation as described in Materials and Methods and is expressed in relative fluorescence units. Results are expressed as the mean ± SEM of three experiments. *, represents difference from PMN controls (P<0.05).
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pone-0009279-g005: Caspase-3, -8, -9, and -2 activity in PMNs following incubation with Yersinia.PMNs were incubated alone or with Y. pestis KIM5, KIM5 YopJ-YopP, KIM5ΔyopJ, KIM6 and Y. enterocolitica 8081v grown at 37°C. Caspase-3, -8, -9, and -2 activity was measured after 3 h and 6 h of incubation as described in Materials and Methods and is expressed in relative fluorescence units. Results are expressed as the mean ± SEM of three experiments. *, represents difference from PMN controls (P<0.05).

Mentions: To determine if PMN caspase activation occurs similar to macrophages following interaction with Yersinia, we measured PMN caspase-3, -8, -9, and -2 activity after 3 and 6 h of incubation with Yersinia. PMNs incubated with anti-Fas antibody were included as a positive control [36]. At 3 h, there was no difference (P>0.05) among any strain tested and the PMN control (Fig. 5). At 6 h, there was a 2–3 fold increase in caspase activity due to anti-Fas antibody (Fig. 5). After 6 h, caspase activity in PMNs incubated with any of the Yersinia strains was less than or equal to the PMN control (P>0.05) (Fig. 5).


Neutrophils are resistant to Yersinia YopJ/P-induced apoptosis and are protected from ROS-mediated cell death by the type III secretion system.

Spinner JL, Seo KS, O'Loughlin JL, Cundiff JA, Minnich SA, Bohach GA, Kobayashi SD - PLoS ONE (2010)

Caspase-3, -8, -9, and -2 activity in PMNs following incubation with Yersinia.PMNs were incubated alone or with Y. pestis KIM5, KIM5 YopJ-YopP, KIM5ΔyopJ, KIM6 and Y. enterocolitica 8081v grown at 37°C. Caspase-3, -8, -9, and -2 activity was measured after 3 h and 6 h of incubation as described in Materials and Methods and is expressed in relative fluorescence units. Results are expressed as the mean ± SEM of three experiments. *, represents difference from PMN controls (P<0.05).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2823771&req=5

pone-0009279-g005: Caspase-3, -8, -9, and -2 activity in PMNs following incubation with Yersinia.PMNs were incubated alone or with Y. pestis KIM5, KIM5 YopJ-YopP, KIM5ΔyopJ, KIM6 and Y. enterocolitica 8081v grown at 37°C. Caspase-3, -8, -9, and -2 activity was measured after 3 h and 6 h of incubation as described in Materials and Methods and is expressed in relative fluorescence units. Results are expressed as the mean ± SEM of three experiments. *, represents difference from PMN controls (P<0.05).
Mentions: To determine if PMN caspase activation occurs similar to macrophages following interaction with Yersinia, we measured PMN caspase-3, -8, -9, and -2 activity after 3 and 6 h of incubation with Yersinia. PMNs incubated with anti-Fas antibody were included as a positive control [36]. At 3 h, there was no difference (P>0.05) among any strain tested and the PMN control (Fig. 5). At 6 h, there was a 2–3 fold increase in caspase activity due to anti-Fas antibody (Fig. 5). After 6 h, caspase activity in PMNs incubated with any of the Yersinia strains was less than or equal to the PMN control (P>0.05) (Fig. 5).

Bottom Line: In contrast, interaction of human PMNs with the virulence plasmid-deficient Y. pestis strain KIM6 resulted in increased surface exposure of phosphatidylserine (PS) and cell death.PMN reactive oxygen species (ROS) production was inhibited in a virulence plasmid-dependent but YopJ/YopP-independent manner.Following phagocytic interaction with Y. pestis strain KIM6, inhibition of PMN ROS production with diphenyleneiodonium chloride resulted in a reduction of PMN cell death similar to that induced by the virulence plasmid-containing strain Y. pestis KIM5.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Molecular Biology and Biochemistry, University of Idaho, Moscow, Idaho, United States of America.

ABSTRACT

Background: The human innate immune system relies on the coordinated activity of macrophages and polymorphonuclear leukocytes (neutrophils or PMNs) for defense against bacterial pathogens. Yersinia spp. subvert the innate immune response to cause disease in humans. In particular, the Yersinia outer protein YopJ (Y. pestis and Y. pseudotuberculosis) and YopP (Y. enterocolitica) rapidly induce apoptosis in murine macrophages and dendritic cells. However, the effects of Yersinia Yop J/P on neutrophil fate are not clearly defined.

Methodology/principal findings: In this study, we utilized wild-type and mutant strains of Yersinia to test the contribution of YopJ and YopP on induction of apoptosis in human monocyte-derived macrophages (HMDM) and neutrophils. Whereas YopJ and YopP similarly induced apoptosis in HMDMs, interaction of human neutrophils with virulence plasmid-containing Yersinia did not result in PMN caspase activation, release of LDH, or loss of membrane integrity greater than PMN controls. In contrast, interaction of human PMNs with the virulence plasmid-deficient Y. pestis strain KIM6 resulted in increased surface exposure of phosphatidylserine (PS) and cell death. PMN reactive oxygen species (ROS) production was inhibited in a virulence plasmid-dependent but YopJ/YopP-independent manner. Following phagocytic interaction with Y. pestis strain KIM6, inhibition of PMN ROS production with diphenyleneiodonium chloride resulted in a reduction of PMN cell death similar to that induced by the virulence plasmid-containing strain Y. pestis KIM5.

Conclusions: Our findings showed that Yersinia YopJ and/or YopP did not induce pronounced apoptosis in human neutrophils. Furthermore, robust PMN ROS production in response to virulence plasmid-deficient Yersinia was associated with increased PMN cell death, suggesting that Yersinia inhibition of PMN ROS production plays a role in evasion of the human innate immune response in part by limiting PMN apoptosis.

Show MeSH
Related in: MedlinePlus