Emilie Bourdonnay

Prostaglandin E2 suppresses allergic sensitization and lung inflammation by targeting the E prostanoid 2 receptor on T cells

BACKGROUND Endogenous prostanoids have been suggested to modulate sensitization during experimental allergic asthma, but the specific role of prostaglandin (PG) E₂ or of specific E prostanoid (EP) receptors is not known. OBJECTIVE Here we tested the role of EP2 signaling in allergic asthma. METHODS Wild-type (WT) and EP2(-/-) mice were subjected to ovalbumin sensitization and acute airway challenge. The PGE2 analog misoprostol was administered during sensitization in both genotypes. In vitro culture of splenocytes and flow-sorted dendritic cells and T cells defined the mechanism by which EP2 exerted its protective effect. Adoptive transfer of WT and EP2(-/-) CD4 T cells was used to validate the importance of EP2 expression on T cells. RESULTS Compared with WT mice, EP2(-/-) mice had exaggerated airway inflammation in this model. Splenocytes and lung lymph node cells from sensitized EP2(-/-) mice produced more IL-13 than did WT cells, suggesting increased sensitization. In WT but not EP2(-/-) mice, subcutaneous administration of misoprostol during sensitization inhibited allergic inflammation. PGE₂ decreased cytokine production and inhibited signal transducer and activator of transcription 6 phosphorylation by CD3/CD28-stimulated CD4(+) T cells. Coculture of flow cytometry-sorted splenic CD4(+) T cells and CD11c(+) dendritic cells from WT or EP2(-/-) mice suggested that the increased IL-13 production in EP2(-/-) mice was due to the lack of EP2 specifically on T cells. Adoptive transfer of CD4(+) EP2(-/-) T cells caused greater cytokine production in the lungs of WT mice than did transfer of WT CD4(+) T cells. CONCLUSION We conclude that the PGE2-EP2 axis is an important endogenous brake on allergic airway inflammation and primarily targets T cells and that its agonism represents a potential novel therapeutic approach to asthma.

PTEN Directly Activates the Actin Depolymerization Factor Cofilin-1 During PGE2-Mediated Inhibition of Phagocytosis of Fungi

By promoting actin depolymerization, the protein phosphatase activity of PTEN impairs macrophage phagocytosis of a fungal pathogen. Preventing Phagocytosis The fungus Candida albicans is normally a commensal microbe found on mucosal surfaces, including those in the lung. However, C. albicans can cause systemic infections that are a leading cause of morbidity and mortality in immunocompromised individuals. A key innate immune response to C. albicans is its ingestion (phagocytosis) by macrophages, a process that requires polymerization of the actin cytoskeleton. Another component of the macrophage response to fungus is the production of prostaglandin E2 (PGE2), a lipid mediator whose synthesis is initiated by cyclooxygenase (COX) enzymes. Serezani et al. found that infection of alveolar macrophages with C. albicans triggered the production of PGE2, which prevented polymerization of the actin cytoskeleton and inhibited phagocytosis of C. albicans by alveolar macrophages. The authors defined the signaling pathway involved. These results suggest that COX inhibitors, such as aspirin, which are in widespread clinical use, may stimulate innate immune responses. In addition, immunosuppression is associated with increased production of PGE2, which may help to explain how antifungal responses are attenuated in immunocompromised individuals. Macrophage ingestion of the yeast Candida albicans requires its recognition by multiple receptors and the activation of diverse signaling programs. Synthesis of the lipid mediator prostaglandin E2 (PGE2) and generation of cyclic adenosine monophosphate (cAMP) also accompany this process. Here, we characterized the mechanisms underlying PGE2-mediated inhibition of phagocytosis and filamentous actin (F-actin) polymerization in response to ingestion of C. albicans by alveolar macrophages. PGE2 suppressed phagocytosis and F-actin formation through the PGE2 receptors EP2 and EP4, cAMP, and activation of types I and II protein kinase A. Dephosphorylation and activation of the actin depolymerizing factor cofilin-1 were necessary for these inhibitory effects of PGE2. PGE2-dependent activation of cofilin-1 was mediated by the protein phosphatase activity of PTEN (phosphatase and tensin homolog deleted on chromosome 10), with which it directly associated. Because enhanced production of PGE2 accompanies many immunosuppressed states, the PTEN-dependent pathway described here may contribute to impaired antifungal defenses.

Alveolar Epithelial Cell–Derived Prostaglandin E2 Serves as a Request Signal for Macrophage Secretion of Suppressor of Cytokine Signaling 3 during Innate Inflammation

Preservation of gas exchange mandates that the pulmonary alveolar surface restrain unnecessarily harmful inflammatory responses to the many challenges to which it is exposed. These responses reflect the cross-talk between alveolar epithelial cells (AECs) and resident alveolar macrophages (AMs). We recently determined that AMs can secrete suppressor of cytokine signaling (SOCS) proteins within microparticles. Uptake of these SOCS-containing vesicles by epithelial cells inhibits cytokine-induced STAT activation. However, the ability of epithelial cells to direct AM release of SOCS-containing vesicles in response to inflammatory insults has not been studied. In this study, we report that SOCS3 protein was elevated in bronchoalveolar lavage fluid of both virus- and bacteria-infected mice, as well as in an in vivo LPS model of acute inflammation. In vitro studies revealed that AEC-conditioned medium (AEC-CM) enhanced AM SOCS3 secretion above basal levels. Increased amounts of PGE2 were present in AEC-CM after LPS challenge, and both pharmacologic inhibition of PGE2 synthesis in AECs and neutralization of PGE2 in AEC-CM implicated this prostanoid as the major AEC-derived factor mediating enhanced AM SOCS3 secretion. Moreover, pharmacologic blockade of PGE2 synthesis or genetic deletion of a PGE2 synthase similarly attenuated the increase in bronchoalveolar lavage fluid SOCS3 noted in lungs of mice challenged with LPS in vivo. These results demonstrate a novel tunable form of cross-talk in which AECs use PGE2 as a signal to request SOCS3 from AMs to dampen their endogenous inflammatory responses during infection.

Regulation of alveolar macrophage p40phox: hierarchy of activating kinases and their inhibition by PGE2

PGE2, produced in the lung during infection with microbes such as Klebsiella pneumoniae, inhibits alveolar macrophage (AM) antimicrobial functions by preventing H2O2 production by NADPH oxidase (NADPHox). Activation of the NADPHox complex is poorly understood in AMs, although in neutrophils it is known to be mediated by kinases including PI3K/Akt, protein kinase C (PKC) δ, p21‐activated protein kinase (PAK), casein kinase 2 (CK2), and MAPKs. The p40phox cytosolic subunit of NADPHox has been recently recognized to function as a carrier protein for other subunits and a positive regulator of oxidase activation, a role previously considered unique to another subunit, p47phox. The regulation of p40phox remains poorly understood, and the effect of PGE2 on its activation is completely undefined. We addressed these issues in rat AMs activated with IgG‐opsonized K. pneumoniae. The kinetics of kinase activation and the consequences of kinase inhibition and silencing revealed a critical role for a PKCδ‐PAK‐class I PI3K/Akt1 cascade in the regulation of p40phox activation upon bacterial challenge in AMs; PKCα, ERK, and CK2 were not involved. PGE2 inhibited the activation of p40phox, and its effects were mediated by protein kinase A type II, were independent of interactions with anchoring proteins, and were directed at the distal class I PI3K/Akt1 activation step. Defining the kinases that control AM p40phox activation and that are the targets for inhibition by PGE2 provides new insights into immunoregulation in the infected lung.

Prostaglandin E2 reduces Toll-like receptor 4 expression in alveolar macrophages by inhibition of translation.

Alveolar macrophages (AMs) represent the first line of innate immune defense in the lung. AMs use pattern recognition receptors (PRRs) to sense pathogens. The best studied PRR is Toll-like receptor (TLR)4, which detects LPS from gram-negative bacteria. The lipid mediator prostaglandin (PG)E2 dampens AM immune responses by inhibiting the signaling events downstream of PRRs. We examined the effect of PGE2 on TLR4 expression in rat AMs. Although PGE2 did not reduce the mRNA levels of TLR4, it decreased TLR4 protein levels. The translation inhibitor cycloheximide reduced TLR4 protein levels with similar kinetics as PGE2, and its effects were not additive with those of the prostanoid, suggesting that PGE2 inhibits TLR at the translational level. The action of PGE2 could be mimicked by the direct stimulator of cAMP formation, forskolin, and involved E prostanoid receptor 2 ligation and cAMP-dependent activation of unanchored type I protein kinase A. Cells pretreated with PGE2 for 24 hours exhibited decreased TNF-α mRNA and protein levels in response to LPS stimulation. Knockdown of TLR4 protein by small interfering RNA to the levels achieved by PGE2 treatment likewise decreased TNF-α mRNA and protein in response to LPS, establishing the functional significance of this PGE2 effect. We provide the first evidence of a lipid mediator acting through its cognate G protein-coupled receptor to affect PRR translation. Because PGE2 is produced in abundance at sites of infection, its inhibitory effects on AM TLR4 expression have important implications for host defense in the lung.

EP4 and EP2 receptor activation of protein kinase A by prostaglandin E2 impairs macrophage phagocytosis of Clostridium sordellii.

Clostridium sordellii causes endometrial infections, but little is known regarding host defenses against this pathogen.