David Dombrowicz

Activation of the Prostaglandin D2 Receptor DP2/CRTH2 Increases Allergic Inflammation in Mouse

Allergic pathologies are often associated with IgE production, mast cell activation, and eosinophilia. PGD2 is the major eicosanoid, among several inflammatory mediators, released by mast cells. PGD2 binds to two membrane receptors, D prostanoid receptor (DP)1 and DP2, endowed with antagonistic properties. In humans, DP2 is preferentially expressed on type 2 lymphocytes, eosinophils, and basophils and mediates chemotaxis in vitro. Although not yet supported by in vivo studies, DP2 is thought to be important in the promotion of Th2-related inflammation. Herein, we demonstrate that mouse eosinophils express both DP1 and DP2 and that PGD2 exerts in vitro chemotactic effects on eosinophils through DP2 activation. Furthermore, 13,14-dihydro-15-keto-PGD2, a specific DP2 agonist not only increases eosinophil recruitment at inflammatory sites but also the pathology in two in vivo models of allergic inflammation: atopic dermatitis and allergic asthma. By contrast, DP1 activation tends to ameliorate the pathology in asthma. Taken together, these results support the hypothesis that DP2 might play a critical role in allergic diseases and underline the interest of DP2 antagonists in human therapy.

Eosinophils, allergy and parasites.

Recently, known eosinophil functions have been extended considerably: previously the cells were thought to have an exclusive role in the release of cytotoxic mediators; now they are known to have roles in antigen presentation and immunoregulation through the release of cytokines. Although questionable, animal models indicate a rather beneficial role of eosinophils in parasitic infections but a detrimental one, together with other cells, in allergy.

Receptors and Signaling Mechanisms Required for Prostaglandin E2-Mediated Regulation of Mast Cell Degranulation and IL-6 Production

Mast cells are implicated in the pathogenesis of a broad spectrum of immunological disorders. These cells release inflammatory mediators in response to a number of stimuli, including IgE-Ag complexes. The degranulation of mast cells is modified by PGs. To begin to delineate the pathway(s) used by PGs to regulate mast cell function, we examined bone marrow-derived mast cells (BMMC) cultured from mice deficient in the EP1, EP2, EP3, and EP4 receptors for PGE2. Although BMMCs express all four of these PGE2 receptors, potentiation of Ag-stimulated degranulation and IL-6 cytokine production by PGE2 is dependent on the EP3 receptor. Consistent with the coupling of this receptor to Gαi, PGE2 activation of the EP3 receptor leads to both inhibition of adenylate cyclase and increased intracellular Ca2+. The magnitude of increase in intracellular Ca2+ induced by EP3 activation is similar to that observed after activation of cells with IgE and Ag. Although PGE alone is not sufficient to initiate BMMC degranulation, stimulation of cells with PGE along with PMA induces degranulation. These actions are mediated by the EP3 receptor through signals involving Ca2+ mobilization and/or decreased cAMP levels. Accordingly, these studies identify PGE2/EP3 as a proinflammatory signaling pathway that promotes mast cell activation.

Human Eosinophils and Human High Affinity IgE Receptor Transgenic Mouse Eosinophils Express Low Levels of High Affinity IgE Receptor, but Release IL-10 upon Receptor Activation

FcεRI expressed by human eosinophils is involved in IgE-mediated cytotoxicity reactions toward the parasite Schistosoma mansoni in vitro. However, because receptor expression is low on these cells, its functional role is still controversial. In this study, we have measured surface and intracellular expression of FcεRI by blood eosinophils from hypereosinophilic patients and normal donors. The number of unoccupied receptors corresponded to ∼4,500 Ab binding sites per cell, whereas 50,000 Ab binding sites per cell were detected intracellularly. Eosinophils from patients displayed significantly more unoccupied receptors than cells from normal donors. This number correlated to both serum IgE concentrations and to membrane-bound IgE. The lack of FcεRI expression by mouse eosinophils has hampered further studies. To overcome this fact and experimentally confirm our findings on human eosinophils, we engineered IL-5 × hFcεRIα double-transgenic mice, whose bone marrow, blood, spleen, and peritoneal eosinophils expressed FcεRI levels similar to levels of human eosinophils, after 4 days culture with IgE in the presence of IL-5. Both human and mouse eosinophils were able to secrete IL-10 upon FcεRI engagement. Thus, comparative analysis of cells from patients and from a relevant animal model allowed us to clearly demonstrate that FcεRI-mediated eosinophil activation leads to IL-10 secretion. Through FcεRI expression, these cells are able to contribute to both the regulation of the immune response and to its effector mechanisms.

Heterogeneity of Expression of IgA Receptors by Human, Mouse, and Rat Eosinophils

IgA is the most abundant class of Abs at mucosal surfaces where eosinophils carry out many of their effector functions. Most of the known IgA-mediated functions require interactions with IgA receptors, six of which have been identified in humans. These include the IgA FcR FcαRI/CD89 and the receptor for the secretory component, already identified on human eosinophils, the polymeric IgR, the Fcα/μR, asialoglycoprotein (ASGP)-R, and transferrin (Tf)R/CD71. In rodents, the existence of IgA receptors on mouse and rat eosinophils remains unclear. We have compared the expression and function of IgA receptors by human, rat, and mouse eosinophils. Our results show that human eosinophils express functional polymeric IgR, ASGP-R, and TfR, in addition to CD89 and the receptor for the secretory component, and that IgA receptors are expressed by rodent eosinophils. Indeed, mouse eosinophils expressed only TfR, whereas rat eosinophils expressed ASGP-R and CD89 mRNA. These results provide a molecular basis for the differences observed between human, rat, and mouse regarding IgA-mediated immunity.

Beneficial metabolic effects of rapamycin are associated with enhanced regulatory cells in diet-induced obese mice.

The “mechanistic target of rapamycin” (mTOR) is a central controller of growth, proliferation and/or motility of various cell-types ranging from adipocytes to immune cells, thereby linking metabolism and immunity. mTOR signaling is overactivated in obesity, promoting inflammation and insulin resistance. Therefore, great interest exists in the development of mTOR inhibitors as therapeutic drugs for obesity or diabetes. However, despite a plethora of studies characterizing the metabolic consequences of mTOR inhibition in rodent models, its impact on immune changes associated with the obese condition has never been questioned so far. To address this, we used a mouse model of high-fat diet (HFD)-fed mice with and without pharmacologic mTOR inhibition by rapamycin. Rapamycin was weekly administrated to HFD-fed C57BL/6 mice for 22 weeks. Metabolic effects were determined by glucose and insulin tolerance tests and by indirect calorimetry measures of energy expenditure. Inflammatory response and immune cell populations were characterized in blood, adipose tissue and liver. In parallel, the activities of both mTOR complexes (e. g. mTORC1 and mTORC2) were determined in adipose tissue, muscle and liver. We show that rapamycin-treated mice are leaner, have enhanced energy expenditure and are protected against insulin resistance. These beneficial metabolic effects of rapamycin were associated to significant changes of the inflammatory profiles of both adipose tissue and liver. Importantly, immune cells with regulatory functions such as regulatory T-cells (Tregs) and myeloid-derived suppressor cells (MDSCs) were increased in adipose tissue. These rapamycin-triggered metabolic and immune effects resulted from mTORC1 inhibition whilst mTORC2 activity was intact. Taken together, our results reinforce the notion that controlling immune regulatory cells in metabolic tissues is crucial to maintain a proper metabolic status and, more generally, comfort the need to search for novel pharmacological inhibitors of the mTOR signaling pathway to prevent and/or treat metabolic diseases.

Eosinophils and urticaria

Previously believed to have an exclusive role in the release of cytotoxic mediators in the defense against helminthic infections, eosinophils are now considered key players in inflammatory reaction and regulation of immune response. Through activation of a large variety of membrane receptors and production of various pharmacologically active mediators, eosinophils may exert a detrimental role in tissues in which they have been recruited and may contribute to the perennization of inflammatory processes. The crucial role of eosinophils has been documented in several eosinophilic skin diseases, such as hypereosinophilic syndrome and bullous pemphigoid, and the literature provides strong evidence for their role in urticaria. The aim of this article is to discuss the mechanisms of specific tissue recruitment of eosinophils, the factors of eosinophil activation, and the contribution of these cells to inflammation and immunoregulation in urticaria. Recent advances in the knowledge of eosinophils will certainly help toward developing new strategies for the management of antihistamines resistant to urticaria.

CX3CL1 (fractalkine) and its receptor CX3CR1 regulate atopic dermatitis by controlling effector T cell retention in inflamed skin

Fractalkine interactions with its receptor, CX3CR1, regulate CD4+ T cell retention in atopic dermatitis and offer a potential therapeutic target in allergic disease.

Anti-folate receptor-α IgE but not IgG recruits macrophages to attack tumors via TNFa/MCP-1 signaling

IgE antibodies are key mediators of antiparasitic immune responses, but their potential for cancer treatment via antibody-dependent cell-mediated cytotoxicity (ADCC) has been little studied. Recently, tumor antigen-specific IgEs were reported to restrict cancer cell growth by engaging high-affinity Fc receptors on monocytes and macrophages; however, the underlying therapeutic mechanisms were undefined and in vivo proof of concept was limited. Here, an immunocompetent rat model was designed to recapitulate the human IgE-Fcε receptor system for cancer studies. We also generated rat IgE and IgG mAbs specific for the folate receptor (FRα), which is expressed widely on human ovarian tumors, along with a syngeneic rat tumor model expressing human FRα. Compared with IgG, anti-FRα IgE reduced lung metastases. This effect was associated with increased intratumoral infiltration by TNFα+ and CD80+ macrophages plus elevated TNFα and the macrophage chemoattractant MCP-1 in lung bronchoalveolar lavage fluid. Increased levels of TNFα and MCP-1 correlated with IgE-mediated tumor cytotoxicity by human monocytes and with longer patient survival in clinical specimens of ovarian cancer. Monocytes responded to IgE but not IgG exposure by upregulating TNFα, which in turn induced MCP-1 production by monocytes and tumor cells to promote a monocyte chemotactic response. Conversely, blocking TNFα receptor signaling abrogated induction of MCP-1, implicating it in the antitumor effects of IgE. Overall, these findings show how antitumor IgE reprograms monocytes and macrophages in the tumor microenvironment, encouraging the clinical use of IgE antibody technology to attack cancer beyond the present exclusive reliance on IgG. Cancer Res; 77(5); 1127-41. ©2017 AACR.

ILC2 and ILC3 contribute to house dust mite (HDM)-induced asthma in obese mice

Many epidemiological and clinical evidences suggest that obesity is an important risk factor for asthma, but the underlying mechanisms remain poorly understood. Besides adaptive immunity and adipokines, innate mechanisms have been involved in both diseases. Given the recent implication of group 2 innate lymphoid cells (ILC2) in asthma and adipose tissue homeostasis, and of ILC3 in obesity-associated airway hyperresponsiveness (AHR), the purpose of this study was to characterize the contribution of adaptive T helper cells versus ILC in a model of house dust mite (HDM)-induced asthma exacerbation by diet-induced obesity. Obesity was induced by a 60% high fat diet (HFD), and lean mice were fed with a low fat diet (LFD). Twelve weeks later, asthma was induced by intranasal administration of HDM. The cardinal features of experimental asthma were evaluated in combination with a flow cytometric analysis of the different adaptive or innate lymphoid population proportions and cytokine production. In lean mice, HDM induced the classical parameters of allergic asthma including increased airway hyperresponsiveness, IgE production, cell recruitment in the bronchoalveolar lavage with eosinophil, lymphocyte and neutrophil infiltrations. Histologic staining confirmed increased mucus accumulation in lungs of asthmatic mice. Feeding mice with HFD resulted in the exacerbation of some features of asthma including serum concentrations of IgE, recruitment of eosinophils in the bronchoalveolar lavage and airway hyperreactivity. Evaluation of the T cell cytokine profile in the lung protein extracts showed an exacerbation of the Th2 and Th17 profiles in obese mice compared to lean mice after HDM challenge. Moreover, the number of ILC2 and ILC3 NCR− cells was increased in HDM-challenged obese mice compared to HDM-challenged lean mice, with an increased intracellular expression of IL-13, IL-17A and IL-22. At the mRNA level, IL-33 mRNA was strongly increased in HDM-challenged obese mice as compared with HDM-challenged lean mice, suggesting that IL-33 might contribute to the induction of ILC2 in obese asthmatic mice. These results support the concept that obesity may exacerbate allergen-induced asthma by a mechanism involving in part ILC2 and ILC3.