A Simple Method for Isolating Human and Rabbit Polymorphonuclear Neutrophils(PMNs).

Kouoh, Ferdinand; Gressier, Bernard; Luyckx, M.; Brunet, C.; Dine, T.; Ballester, L.; Cazin, M.; Cazin, J. C.

doi:10.1248/bpb.23.1382

Cited by 11 publications

(3 citation statements)

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“…PMNs were isolated from human blood samples obtained by venipuncture of healthy volunteers [75], [76]. 10 mL venous blood were collected into syringes containing 250 U heparin and diluted back 2-fold in 0.9% saline solution.…”

Section: Methodsmentioning

confidence: 99%

Extracellular Superoxide Dismutase Protects Histoplasma Yeast Cells from Host-Derived Oxidative Stress

et al. 2012

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In order to establish infections within the mammalian host, pathogens must protect themselves against toxic reactive oxygen species produced by phagocytes of the immune system. The fungal pathogen Histoplasma capsulatum infects both neutrophils and macrophages but the mechanisms enabling Histoplasma yeasts to survive in these phagocytes have not been fully elucidated. We show that Histoplasma yeasts produce a superoxide dismutase (Sod3) and direct it to the extracellular environment via N-terminal and C-terminal signals which promote its secretion and association with the yeast cell surface. This localization permits Sod3 to protect yeasts specifically from exogenous superoxide whereas amelioration of endogenous reactive oxygen depends on intracellular dismutases such as Sod1. While infection of resting macrophages by Histoplasma does not stimulate the phagocyte oxidative burst, interaction with polymorphonuclear leukocytes (PMNs) and cytokine-activated macrophages triggers production of reactive oxygen species (ROS). Histoplasma yeasts producing Sod3 survive co-incubation with these phagocytes but yeasts lacking Sod3 are rapidly eliminated through oxidative killing similar to the effect of phagocytes on Candida albicans yeasts. The protection provided by Sod3 against host-derived ROS extends in vivo. Without Sod3, Histoplasma yeasts are attenuated in their ability to establish respiratory infections and are rapidly cleared with the onset of adaptive immunity. The virulence of Sod3-deficient yeasts is restored in murine hosts unable to produce superoxide due to loss of the NADPH-oxidase function. These results demonstrate that phagocyte-produced ROS contributes to the immune response to Histoplasma and that Sod3 facilitates Histoplasma pathogenesis by detoxifying host-derived reactive oxygen thereby enabling Histoplasma survival.

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Section: Methodsmentioning

confidence: 99%

Extracellular Superoxide Dismutase Protects Histoplasma Yeast Cells from Host-Derived Oxidative Stress

et al. 2012

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“…PMNs were isolated from the blood of healthy human donors through venipuncture as previously described (40,41). Briefly, 10 ml of blood was collected into syringes containing 250 U heparin and diluted into 10 ml 0.9% saline solution.…”

Section: Methodsmentioning

confidence: 99%

Redundant Catalases Detoxify Phagocyte Reactive Oxygen and Facilitate Histoplasma capsulatum Pathogenesis

et al. 2013

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Histoplasma capsulatum is a respiratory pathogen that infects phagocytic cells. The mechanisms allowing Histoplasma to overcome toxic reactive oxygen molecules produced by the innate immune system are an integral part of Histoplasma's ability to survive during infection. To probe the contribution of Histoplasma catalases in oxidative stress defense, we created and analyzed the virulence defects of mutants lacking CatB and CatP, which are responsible for extracellular and intracellular catalase activities, respectively. Both CatB and CatP protected Histoplasma from peroxide challenge in vitro and from antimicrobial reactive oxygen produced by human neutrophils and activated macrophages. Optimal protection required both catalases, as the survival of a double mutant lacking both CatB and CatP was lower than that of single-catalase-deficient cells. Although CatB contributed to reactive oxygen species defenses in vitro, CatB was dispensable for lung infection and extrapulmonary dissemination in vivo. Loss of CatB from a strain also lacking superoxide dismutase (Sod3) did not further reduce the survival of Histoplasma yeasts. Nevertheless, some catalase function was required for pathogenesis since simultaneous loss of both CatB and CatP attenuated Histoplasma virulence in vivo. These results demonstrate that Histoplasma's dual catalases comprise a system that enables Histoplasma to efficiently overcome the reactive oxygen produced by the innate immune system.

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“…A metodologia para separação dos polimorfonucleares de coelhos foi adaptada dos trabalhos de Boyum (1968) e de Kouoh et al (2000). Um volume de 5 mL de sangue periférico heparinizado (15 U/mL) foi adicionado a uma solução de Dextran a 3% diluído em PBS e submetido a sedimentação por aproximadamente 20 minutos.…”

Section: Fagocitose Via C3b Por Polimormofonucleares De Coelhounclassified

Contribuição ao estudo da ação in vitro do veneno da aranha marrom, Loxosceles gaucho, sobre plaquetas humanas e de coelho. Participação das plaquetas na dermonecrose induzida experimentalmente pelo veneno loxoscélico em coelhos

Tavares¹

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TAVARES, F. L. Contribuição ao estudo da ação in vitro do veneno da aranha marrom, Loxosceles gaucho, sobre plaquetas humanas e de coelho. Participação das plaquetas na dermonecrose induzida experimentalmente em coelhos pelo veneno loxoscélico. [Contribution to the study of the in vitro action induced by the venom of the brown spider, Loxosceles gaucho, on human and rabbit platelets. Platelet participation in the dermonecrotic lesion experimentally induced in rabbits by the loxoscelic venom]. 2007.

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A Simple Method for Isolating Human and Rabbit Polymorphonuclear Neutrophils(PMNs).

Cited by 11 publications

References 0 publications

Extracellular Superoxide Dismutase Protects Histoplasma Yeast Cells from Host-Derived Oxidative Stress

Extracellular Superoxide Dismutase Protects Histoplasma Yeast Cells from Host-Derived Oxidative Stress

Redundant Catalases Detoxify Phagocyte Reactive Oxygen and Facilitate Histoplasma capsulatum Pathogenesis

Contribuição ao estudo da ação in vitro do veneno da aranha marrom, Loxosceles gaucho, sobre plaquetas humanas e de coelho. Participação das plaquetas na dermonecrose induzida experimentalmente pelo veneno loxoscélico em coelhos

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