Complement-mediated regulation of the IL-17A axis is a central genetic determinant of the severity of experimental allergic asthma.

Severe asthma is associated with the production of interleukin 17A (IL-17A). The exact role of IL-17A in severe asthma and the factors that drive its production are unknown. Here we demonstrate that IL-17A mediated severe airway hyperresponsiveness (AHR) in susceptible strains of mice by enhancing IL-13-driven responses. Mechanistically, we demonstrate that IL-17A and AHR were regulated by allergen-driven production of anaphylatoxins, as mouse strains deficient in complement factor 5 (C5) or the complement receptor C5aR mounted robust IL-17A responses, whereas mice deficient in C3aR had fewer IL-17-producing helper T cells (T(H)17 cells) and less AHR after allergen challenge. The opposing effects of C3a and C5a were mediated through their reciprocal regulation of IL-23 production. These data demonstrate a critical role for complement-mediated regulation of the IL-23-T(H)17 axis in severe asthma.

Differential control of CD4(+) T-cell subsets by the PD-1/PD-L1 axis in a mouse model of allergic asthma.

Studies examining the role of PD-1 family members in allergic asthma have yielded conflicting results. Using a mouse model of allergic asthma, we demonstrate that blockade of PD-1/PD-L1 has distinct influences on different CD4(+) T-cell subsets. PD-1/PD-L1 blockade enhances airway hyperreactivity (AHR), not by altering the magnitude of the underlying Th2-type immune response, but by allowing the development of a concomitant Th17-type immune response. Supporting differential CD4(+) T-cell responsiveness to PD-1-mediated inhibition, naïve PD-1(-/-) mice displayed elevated Th1 and Th17 levels, but diminished Th2 cytokine levels, and ligation of PD-1 in WT cells limited cytokine production by in vitro polarized Th1 and Th17 cells, but slightly enhanced cytokine production by in vitro polarized Th2 cells. Furthermore, PD-1 ligation enhanced Th2 cytokine production by naïve T cells cultured under nonpolarizing conditions. These data demonstrate that different CD4(+) T-cell subsets respond differentially to PD-1 ligation and may explain some of the variable results observed in control of allergic asthma by the PD-1 family members. As the PD-1/PD-L1 axis limits asthma severity by constraining Th17 cell activity, this suggests that severe allergic asthma may be associated with a defective PD-1/PD-L1 regulatory axis in some individuals.

CD4+CD25+ T cells protect against experimentally induced asthma and alter pulmonary dendritic cell phenotype and function.

The role of natural CD4+CD25+ regulatory T (T reg) cells in the control of allergic asthma remains poorly understood. We explore the impact of T reg cell depletion on the allergic response in mice susceptible (A/J) or comparatively resistant (C3H) to the development of allergen-induced airway hyperresponsiveness (AHR). In C3H mice, anti-CD25-mediated T reg cell depletion before house dust mite treatment increased several features of the allergic diathesis (AHR, eosinophilia, and IgE), which was concomitant with elevated T helper type 2 (Th2) cytokine production. In similarly T reg cell-depleted A/J mice, we observed a moderate increase in airway eosinophilia but no effects on AHR, IgE levels, or Th2 cytokine synthesis. As our experiments suggested that T reg cell depletion in C3H mice before sensitization was sufficient to enhance the allergic phenotype, we characterized dendritic cells (DCs) in T reg cell-depleted C3H mice. T reg cell-depleted mice had increased numbers of pulmonary myeloid DCs with elevated expression of major histocompatibility complex class II, CD80, and CD86. Moreover, DCs from T reg cell-depleted mice demonstrated an increased capacity to stimulate T cell proliferation and Th2 cytokine production, which was concomitant with reduced IL-12 expression. These data suggest that resistance to allergen-driven AHR is mediated in part by CD4+CD25+ T reg cell suppression of DC activation and that the absence of this regulatory pathway contributes to susceptibility.

Protease-activated receptor 2 activation of myeloid dendritic cells regulates allergic airway inflammation.

BACKGROUND: A common characteristic of allergens is that they contain proteases that can activate protease-activated receptor (PAR-2); however the mechanism by which PAR-2 regulates allergic airway inflammation is unclear. METHODS: Mice (wild type and PAR-2-deficient) were sensitized using German cockroach (GC) feces (frass), the isolated protease from GC frass, or through adoptive transfer of GC frass-treated bone marrow-derived dendritic cells (BMDC) and measurements of airway inflammation (cellular infiltration, cytokine expression, and mucin production), serum IgE levels and airway hyperresponsiveness (AHR) were assessed. BMDC were cultured, treated with GC frass and assessed for cytokine production. PAR-2 expression on pulmonary mDCs was determined by flow cytometry. RESULTS: Exposure to GC frass induced AHR and airway inflammation in wild type mice; however PAR-2-deficient mice had significantly attenuated responses. To directly investigate the role of the protease, we isolated the protease from GC frass and administered the endotoxin-free protease into the airways of mice in the presence of OVA. GC frass proteases were sufficient to promote the development of AHR, serum IgE, and Th2 cytokine production. PAR-2 expression on mDC was upregulated following GC frass exposure, but the presence of a functional PAR-2 did not alter antigen uptake. To determine if PAR-2 activation led to differential cytokine production, we cultured BMDC in the presence of GM-CSF and treated these cells ex vivo with GC frass. PAR-2-deficient BMDC released significantly less IL-6, IL-23 and TNFα compared to BMDC from wild type mice, suggesting PAR-2 activation was important in Th2/Th17 skewing cytokine production. To determine the role for PAR-2 on mDCs on the initiation of allergic airway inflammation, BMDCs from wild type and PAR-2-deficient mice were treated in the presence or absence of GC frass and then adoptively transferred into the airway of wild type mice. Importantly, GC frass-stimulated wild type BMDCs were sufficient to induce AHR and allergic airway inflammation, while GC frass-stimulated PAR-2-deficient BMDC had attenuated responses. CONCLUSIONS: Together these data suggest an important role for allergen activation of PAR-2 on mDCs in mediating Th2/Th17 cytokine production and allergic airway responses.

Mucosal sensitization to German cockroach involves protease-activated receptor-2.

BACKGROUND: Allergic asthma is on the rise in developed countries. A common characteristic of allergens is that they contain intrinsic protease activity, and many have been shown to activate protease-activated receptor (PAR)-2 in vitro. The role for PAR-2 in mediating allergic airway inflammation has not been assessed using a real world allergen. METHODS: Mice (wild type or PAR-2-deficient) were sensitized to German cockroach (GC) feces (frass) or protease-depleted GC frass by either mucosal exposure or intraperitoneal injection and measurements of airway inflammation (IL-5, IL-13, IL-17A, and IFNgamma levels in the lung, serum IgE levels, cellular infiltration, mucin production) and airway hyperresponsiveness were performed. RESULTS: Following systemic sensitization, GC frass increased airway hyperresponsiveness, Th2 cytokine release, serum IgE levels, cellular infiltration and mucin production in wild type mice. Interestingly, PAR-2-deficient mice had similar responses as wild type mice. Since these data were in direct contrast to our finding that mucosal sensitization with GC frass proteases regulated airway hyperresponsiveness and mucin production in BALB/c mice (Page et. al. 2007 Resp Res 8:91), we backcrossed the PAR-2-deficient mice into the BALB/c strain. Sensitization to GC frass could now occur via the more physiologically relevant method of intratracheal inhalation. PAR-2-deficient mice had significantly reduced airway hyperresponsiveness, Th2 and Th17 cytokine release, serum IgE levels, and cellular infiltration compared to wild type mice when sensitization to GC frass occurred through the mucosa. To confirm the importance of mucosal exposure, mice were systemically sensitized to GC frass or protease-depleted GC frass via intraperitoneal injection. We found that removal of proteases from GC frass had no effect on airway inflammation when administered systemically. CONCLUSIONS: We showed for the first time that allergen-derived proteases in GC frass elicit allergic airway inflammation via PAR-2, but only when allergen was administered through the mucosa. Importantly, our data suggest the importance of resident airway cells in the initiation of allergic airway disease, and could make allergen-derived proteases attractive therapeutic targets.

Unique and overlapping gene expression patterns driven by IL-4 and IL-13 in the mouse lung.

BACKGROUND: Allergic asthma results from inappropriate T(H)2-mediated inflammation. Both IL-4 and IL-13 contribute to asthma pathogenesis, but IL-4 predominantly drives T(H)2 induction, whereas IL-13 is necessary and sufficient for allergen-induced airway hyperresponsiveness and goblet cell hyperplasia. Although these 2 cytokines share signaling components, the molecular mechanisms by which they mediate different phases of the allergic asthmatic response remain elusive. OBJECTIVE: We sought to clarify the role or roles of IL-4 and IL-13 in asthma-pathogenesis. METHODS: We used DNA Affymetrix microarrays to profile pulmonary gene expression in BALB/c mice inoculated intratracheally with ragweed pollen, house dust mite, IL-4, IL-13, or both cytokines. IL-13 dependence was confirmed by comparing pulmonary gene expression in house dust mite-inoculated wild-type and IL-13 knockout mice. RESULTS: A signature gene expression profile consisting of 23 genes was commonly induced by means of inoculation with house dust mite, ragweed pollen, or IL-4 plus IL-13. Although rIL-4 and rIL-13 treatment induced an overlapping set of genes, IL-4 uniquely induced 21 genes, half of which were interferon response genes and half of which were genes important in immunoregulation. IL-13 uniquely induced 8 genes, most of which encode proteins produced by epithelial cells. CONCLUSIONS: IL-4 and IL-13 together account for most allergen-induced pulmonary genes. Selective IL-4 induction of IFN-gamma response genes and other genes that might negatively regulate allergic inflammation could partially explain the greater importance of IL-13 in the effector phase of allergic airway disease.

Elevated cytokine levels in children with autism spectrum disorder.

UNLABELLED: This study compared production of IL-2, IFN-gamma, IL-4, IL-13, IL-5 and IL-10 in peripheral blood mononuclear cells from 20 children with autism spectrum disorder to those from matched controls. Levels of all Th2 cytokines were significantly higher in cases after incubation in media alone, but the IFN-gamma/IL-13 ratio was not significantly different between cases and controls. Cases had significantly higher IL-13/IL-10 and IFN-gamma/IL-10 than controls. CONCLUSION: Children with ASD had increased activation of both Th2 and Th1 arms of the adaptive immune response, with a Th2 predominance, and without the compensatory increase in the regulatory cytokine IL-10.

Trefoil factor 2 rapidly induces interleukin 33 to promote type 2 immunity during allergic asthma and hookworm infection.

The molecular mechanisms that drive mucosal T helper type 2 (T(H)2) responses against parasitic helminths and allergens remain unclear. In this study, we demonstrate in mice that TFF2 (trefoil factor 2), an epithelial cell-derived repair molecule, is needed for the control of lung injury caused by the hookworm parasite Nippostrongylus brasiliensis and for type 2 immunity after infection. TFF2 is also necessary for the rapid production of IL-33, a T(H)2-promoting cytokine, by lung epithelia, alveolar macrophages, and inflammatory dendritic cells in infected mice. TFF2 also increases the severity of allergic lung disease caused by house dust mite antigens or IL-13. Moreover, TFF2 messenger RNA expression is significantly increased in nasal mucosal brushings during asthma exacerbations in children. These experiments extend the biological functions of TFF2 from tissue repair to the initiation and maintenance of mucosal T(H)2 responses.

Early immunological response to German cockroach frass exposure induces a Th2/Th17 environment.

Cockroach exposure is a major risk factor for the development of asthma; however, the early immune events induced by cockroach leading to the Th2 response are not fully understood. Exposure of naïve mice to German cockroach (GC) feces (frass) was sufficient to induce dendritic cell (DC) recruiting and activating chemokines C-C motif ligand 20, granulocyte macrophage colony-stimulating factor, granulocyte colony-stimulating factor and macrophage inflammatory protein-1α into the airways. This corresponded with an increase in myeloid DCs (mDCs) in the airways as well as increased expression of CD80 and CD86 on the mDCs. Plasmacytoid DCs in the lung were unchanged. Levels of IL-5, IL-17A and IL-6 cytokines in whole lung cultures were significantly increased 18 h following GC frass exposure demonstrating the early development of a mixed Th2/Th17 response. In addition, GC frass stimulated the production of IL-23, IL-6 and IL-12p70 from bone marrow-derived mDCs. Adoptive transfer of GC frass-pulsed mDCs induced airway reactivity, airway inflammation as well as eosinophilia and induced a strong Th2/Th17 response in the lung. MyD88-deficient bone marrow-derived mDCs did not respond to GC frass treatment, suggesting a functional Toll-like receptor pathway was important to induce the Th2/Th17 response. Together, our data show that GC frass activated the innate immune response to augment DC recruitment and activation of mDCs which promoted robust T cell-skewing cytokines and ultimately drive the development of airway inflammation.