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Erica L Martin,
Danielle G Souza,
Caio T Fagundes,
Flavio A Amaral,
Barbara Assenzio,
Valeria Puntorieri,
Lorenzo Del Sorbo,
Vito Fanelli,
Martino Bosco,
Luisa Delsedime,
Jose F Pinho,
Virginia S Lemos,
Fabricio O Souto,
Jose C Alves-Filho,
Fernando Q Cunha,
Arthur S Slutsky, Thomas Ruckle,
Emilio Hirsch,
Mauro M Teixeira,
V Marco Ranieri
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ABSTRACT: Sepsis is a leading cause of death in the intensive care unit, characterized by a systemic inflammatory response (SIRS) and bacterial infection, which can often induce multiorgan damage and failure. Leukocyte recruitment, required to limit bacterial spread, depends on phosphoinositide-3 kinase γ (PI3Kγ) signaling in vitro; however, the role of this enzyme in polymicrobial sepsis has remained unclear.
This study aimed to determine the specific role of the kinase activity of PI3Kγ in the pathogenesis of sepsis and multiorgan damage.
PI3Kγ wild-type, knockout, and kinase-dead mice were exposed to cecal ligation and perforation-induced sepsis and assessed for survival; pulmonary, hepatic, and cardiovascular damage; coagulation derangements; systemic inflammation; bacterial spread; and neutrophil recruitment. Additionally, wild-type mice were treated either before or after the onset of sepsis with a PI3Kγ inhibitor and assessed for survival, neutrophil recruitment, and bacterial spread.
Both genetic and pharmaceutical PI3Kγ kinase inhibition significantly improved survival, reduced multiorgan damage, and limited bacterial decompartmentalization, while modestly affecting SIRS. Protection resulted from both neutrophil-independent mechanisms, involving improved cardiovascular function, and neutrophil-dependent mechanisms, through reduced susceptibility to neutrophil migration failure during severe sepsis by maintaining neutrophil surface expression of the chemokine receptor, CXCR2. Furthermore, PI3Kγ pharmacological inhibition significantly decreased mortality and improved neutrophil migration and bacterial control, even when administered during established septic shock.
This study establishes PI3Kγ as a key molecule in the pathogenesis of septic infection and the transition from SIRS to organ damage and identifies it as a novel possible therapeutic target.
American Journal of Respiratory and Critical Care Medicine 09/2010; 182(6):762-73. · 11.08 Impact Factor
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Vanessa Pinho,
Remo Castro Russo,
Remo de Castro Russo,
Flávio A Amaral,
Lirlândia P de Sousa,
Michele M Barsante,
Danielle G de Souza,
José C Alves-Filho,
Denise C Cara,
Joel S Hayflick,
Christian Rommel, Thomas Ruckle,
Adriano G Rossi,
Mauro M Teixeira
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ABSTRACT: PI3K plays a fundamental role in regulating neutrophil recruitment into sites of inflammation but the role of the different isoforms of PI3K remains unclear. In this study, we evaluated the role of PI3Kgamma and PI3Kdelta for neutrophil influx induced by the exogenous administration or the endogenous generation of the chemokine CXCL1. Administration of CXCL1 in PI3Kgamma(-/-) or wild-type (WT) mice induced similar increases in leukocyte rolling, adhesion, and emigration in the cremaster muscle when examined by intravital microscopy. The induction of neutrophil recruitment into the pleural cavity or the tibia-femoral joint induced by the injection of CXCL1 was not significantly different in PI3Kgamma(-/-) or WT mice. Neutrophil influx was not altered by treatment of WT mice with a specific PI3Kdelta inhibitor, IC87114, or a specific PI3Kgamma inhibitor, AS605240. The administration of IC87114 prevented CXCL1-induced neutrophil recruitment only in presence of the PI3Kgamma inhibitor or in PI3Kgamma(-/-) mice. Ag challenge of immunized mice induced CXCR2-dependent neutrophil recruitment that was inhibited by wortmannin or by blockade of and PI3Kdelta in PI3Kgamma(-/-) mice. Neutrophil recruitment to bronchoalveolar lavage induced by exogenously added or endogenous production of CXCL1 was prevented in PI3Kgamma(-/-) mice. The accumulation of the neutrophils in lung tissues was significantly inhibited only in PI3Kgamma(-/-) mice treated with IC87114. Neutrophil recruitment induced by exogenous administration of C5a or fMLP appeared to rely solely on PI3Kgamma. Altogether, our data demonstrate that there is a tissue- and stimulus-dependent role of PI3Kgamma and PI3Kdelta for neutrophil recruitment induced by different chemoattractants in vivo.
The Journal of Immunology 01/2008; 179(11):7891-8. · 5.79 Impact Factor
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ABSTRACT: The signal transduction pathways that lead activated natural killer (NK) cells to produce cytokines, releases cytotoxic granules, or do both, are not clearly dissected. For example, phosphoinositide 3-kinases (PI3Ks) are key players in the execution of both functions, but the relative contribution of each isoform is unknown. We show here that the catalytic isoform p110delta, not p110gamma, was required for interferon-gamma (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha), and granulocyte macrophage colony-stimulating factor (GM-CSF) secretion, whereas neither was necessary for cytotoxicity. Yet, when both p110delta and p110gamma isoforms were inactivated by a combination of genetic and biochemical approaches, cytotoxicity was decreased. NK-cell numbers were also affected by the lack of p110delta but not p110gamma and more severely so in mice lacking both subunits. These results provide genetic evidence that p110delta is the dominant PI3K isoform for cytokine secretion by NK cells and suggest that PI3Ks cooperate during NK-cell development and cytotoxicity.
Blood 12/2007; 110(9):3202-8. · 9.90 Impact Factor
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ABSTRACT: Mouse gene-targeting studies have documented a central role of the p110delta isoform of phosphoinositide 3-kinase (PI3K) in B-cell development and function. A defect in B-cell antigen receptor (BCR) signaling is key to this B-cell phenotype. Here we further characterize this signaling defect and report that a p110delta-selective small molecule inhibitor mirrors the effect of genetic inactivation of p110delta in BCR signaling. p110delta activity is indispensable for BCR-induced DNA synthesis and phosphorylation of Akt/protein kinase B (PKB), forkhead transcription factor/forkhead box O3a (FOXO3a), and p70 S6 kinase (p70 S6K), with modest effects on the phosphorylation of glycogen synthase kinase 3 alpha/beta (GSK3alpha/beta) and extracellular signal-regulated kinase (Erk). The PI3K-dependent component of intracellular calcium mobilization also completely relies on p110delta catalytic activity. Resting B cells with inactive p110delta fail to enter the cell cycle, correlating with an incapacity to up-regulate the expression of cyclins D2, A, and E, and to phosphorylate the retinoblastoma protein (Rb). p110delta is also critical for interleukin 4 (IL-4)-induced phosphorylation of Akt/PKB and FOXO3a, and protection from apoptosis. Taken together, these data show that defects observed in p110delta mutant mice are not merely a consequence of altered B-cell differentiation, and emphasize the potential utility of p110delta as a drug target in autoimmune diseases in which B cells play a crucial role.
Blood 02/2006; 107(2):642-50. · 9.90 Impact Factor
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Alison M Condliffe,
Keith Davidson,
Karen E Anderson,
Chris D Ellson,
Tom Crabbe,
Klaus Okkenhaug,
Bart Vanhaesebroeck,
Martin Turner,
Louise Webb,
Matthias P Wymann,
Emilio Hirsch, Thomas Ruckle,
Montserrat Camps,
Christian Rommel,
Shaun P Jackson,
Edwin R Chilvers,
Len R Stephens,
Phillip T Hawkins
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ABSTRACT: It is well established that preexposure of human neutrophils to proinflammatory cytokines markedly augments the production of reactive oxygen species (ROS) to subsequent stimuli. This priming event is thought to be critical for localizing ROS to the vicinity of the inflammation, maximizing their role in the resolution of the inflammation, and minimizing the damage to surrounding tissue. We have used a new generation of isoform-selective phosphoinositide 3-kinase (PI3K) inhibitors to show that ROS production under these circumstances is regulated by temporal control of class I PI3K activity. Stimulation of tumor necrosis factor-alpha (TNF-alpha)-primed human neutrophils with N-formyl-methionyl-leucyl-phenylalanine (fMLP) results in biphasic activation of PI3K; the first phase is largely dependent on PI3Kgamma, and the second phase is largely dependent on PI3Kdelta. The second phase of PI3K activation requires the first phase; it is this second phase that is augmented by TNF-alpha priming and that regulates parallel activation of ROS production. Surprisingly, although TNF-alpha-primed mouse bone marrow-derived neutrophils exhibit superficially similar patterns of PI3K activation and ROS production in response to fMLP, these responses are substantially lower and largely dependent on PI3Kgamma alone. These results start to define which PI3K isoforms are responsible for modulating neutrophil responsiveness to infection and inflammation.
Blood 09/2005; 106(4):1432-40. · 9.90 Impact Factor
