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Background Group 2 innate lymphoid cells (ILC2s) expand in the lungs of mice during type 2 inflammation induced by the fungal allergen Alternaria alternata . The increase in ILC2 numbers in the lung has been largely attributed to local proliferation and whether ILC2s migrate from the circulation to the lung after Alternaria exposure is unknown. Objective We examined whether human (lung, lymph node, and blood) and mouse lung ILC2s express β 1 and β 2 integrin adhesion molecules and whether these integrins are required for trafficking of ILC2s into the lungs of mice. Methods Human and mouse ILC2s were assessed for surface expression of β 1 and β 2 integrin adhesion molecules by using flow cytometry. The role of β 1 and β 2 integrins in ILC2 trafficking to the lungs was assessed by in vivo blocking of these integrins before airway exposure to Alternaria in mice. Results Both human and mouse lung ILC2s express high levels of β 1 and β 2 integrin adhesion receptors. Intranasal administration of Alternaria challenge reduced ILC2 numbers in the bone marrow and concurrently increased blood and lung ILC2 numbers. In vivo blocking of β 2 integrins (CD18) significantly reduced ILC2 numbers in the lungs but did not alter ILC2 proliferation, apoptosis, and function. In contrast, in vivo blocking of β 1 integrins or α 4 integrins did not affect lung ILC2 numbers. Conclusion ILC2 numbers increase in the mouse lung not only through local proliferation but also through trafficking from the circulation into the lung using β 2 rather than β 1 or α 4 integrins.

RV infections frequently trigger exacerbations of respiratory diseases, such as asthma, yet treatment and intervention options remain limited. Statin drugs are the treatment of choice for dyslipidemia and can also modulate immune cell function. To determine whether statin drugs modify antiviral responses of human monocytic cells, we obtained blood monocytes from donors with allergies and/or asthma and treated the cells with sim prior to challenge with RV. RV-induced secretion of CXCL10 was attenuated significantly, irrespective of RV type (RV-16, -14, or -1A), which corresponded with decreases in IFN-α secretion and pSTAT1. Sim pretreatment also reduced RV-induced CXCL10 secretion from human alveolar macrophages. The addition of mev and GGPP—two intermediates of the cholesterol biosynthesis pathway—was able to rescue CXCL10 release fully, demonstrating that effects of sim were related to inhibition of cholesterol biosynthesis and not to an off-target effect. In addition, sim pretreatment attenuated IFN-α-induced pSTAT1 and CXCL10 secretion, providing evidence that sim additionally can affect type I IFNR signaling. SOCS1 and 3 mRNA are both induced with RV stimulation, but sim did not elevate SOCS1 or SOCS3 mRNA expression basally or in the presence of RV. Our findings suggest that sim inhibition of the cholesterol biosynthesis pathway leads to decreased RV-induced chemokine secretion in monocytes and macrophages. These findings suggest that statin drugs have the potential to curb the inflammatory response to RV infection.

Electron microscopy demonstrates that mouse lung ILC2 expressing PSGL-1 have platelets attached to their surface and that platelet depletion reduces lung ILC2 proliferation and Th2 cytokines suggesting ILC2 function is influenced by attachment to platelets.

Gasdermin B (GSDMB) on chromosome 17q21 demonstrates a strong genetic linkage to asthma, but its function in asthma is unknown. Here we identified that GSDMB is highly expressed in lung bronchial epithelium in human asthma. Overexpression of GSDMB in primary human bronchial epithelium increased expression of genes important to both airway remodeling [TGF-β1, 5-lipoxygenase (5-LO)] and airway-hyperresponsiveness (AHR) (5-LO). Interestingly, hGSDMBZp3-Cre mice expressing increased levels of the human GSDMB transgene showed a significant spontaneous increase in AHR and a significant spontaneous increase in airway remodeling, with increased smooth muscle mass and increased fibrosis in the absence of airway inflammation. In addition, hGSDMBZp3-Cre mice showed increases in the same remodeling and AHR mediators (TGF-β1, 5-LO) observed in vitro in GSDMB-overexpressing epithelial cells. GSDMB induces TGF-β1 expression via induction of 5-LO, because knockdown of 5-LO in epithelial cells overexpressing GSDMB inhibited TGF-β1 expression. These studies demonstrate that GSDMB, a gene highly linked to asthma but whose function in asthma is previously unknown, regulates AHR and airway remodeling without airway inflammation through a previously unrecognized pathway in which GSDMB induces 5-LO to induce TGF-β1 in bronchial epithelium.

Recent discoveries have led to the identification of a novel group of immune cells, the innate lymphoid cells (ILCs). The members of this group are divided into three subpopulations: ILC1s, ILC2s, and ILC3s. ILC2s produce Th2 cytokines, IL-4, IL-5, and IL-13, upon activation by epithelial cell-derived cytokines, lipid mediators (cysteinyl leukotrienes and prostaglandin D2), and TNF family member TL1A and promote structural and immune cell responses in the airways after antigen exposure. In addition, ILC2 function is also influenced by inducible T cell costimulator (ICOS)/ ICOS-ligand (ICOS-L) interactions via direct contact between immune cells. The most common airway antigens are allergens and viruses which are highly linked to the induction of airway diseases with underlying type 2 inflammation including asthma and allergic rhinitis. Based on recent findings linking ILC2s and airway Th2 responses, there is intensive investigation into the role of ILC2s in human disease with the hope of a better understanding of the pathophysiology and the discovery of novel potential therapeutic targets. This review summarizes the recent advances made in elucidating ILC2 involvement in human Th2 airway disease.

ILC2s were originally found to be activated by epithelial cell derived cytokines to induce the secretion of Th2 cytokines, IL-5 and IL-13. Recent research has shown that lipid mediators play a large role in the activation and inhibition of ILC2 function. Unlike the traditional epithelial cell derived cytokines IL-33 and IL-25, lipid mediators have been shown to promote ILC2 secretion of not only IL-5 and IL-13, but the secretion of IL-4 as well. Prostaglandin D2 has been shown to be a potent chemoattractant of ILC2s as well as a potent activator of ILC2s to release Th2 cytokines. In addition to prostaglandin D2, cysteinyl leukotrienes also activate ILC2s to secrete Th2 cytokines during inflammation. Notably, lipid mediators have been shown to work in concert with epithelial cell derived cytokines to increase IL-5 and IL-13 secretion from ILC2s. On the other hand, lipid meditators prostaglandin I2 and lipoxin A4 are the first identified lipid mediator inhibitors of ILC2 function, and thus limit ILC2 contribution to Th2 inflammation. ILC2s play a potential significant role in Th2 mediated inflammation in a variety of allergic disease states, such as asthma, atopic dermatitis, and chronic rhinosinusitis. The identification of lipid mediators as activators and inhibitors of ILC2 function provides additional therapeutic targets for altering ILC2 function during disease states.

Chronic asthma is associated with airway remodeling and decline in lung function. In this article, we show that follistatin-like 1 (Fstl1), a mediator not previously associated with asthma, is highly expressed by macrophages in the lungs of humans with severe asthma. Chronic allergen-challenged Lys-Cretg /Fstl1Δ/Δ mice in whom Fstl1 is inactivated in macrophages/myeloid cells had significantly reduced airway remodeling and reduced levels of oncostatin M (OSM), a cytokine previously not known to be regulated by Fstl1. The importance of the Fstl1 induction of OSM to airway remodeling was demonstrated in murine studies in which administration of Fstl1 induced airway remodeling and increased OSM, whereas administration of an anti-OSM Ab blocked the effect of Fstl1 on inducing airway remodeling, eosinophilic airway inflammation, and airway hyperresponsiveness, all cardinal features of asthma. Overall, these studies demonstrate that the Fstl1/OSM pathway may be a novel pathway to inhibit airway remodeling in severe human asthma.

Background Aspirin-exacerbated respiratory disease (AERD) is characterized by tissue eosinophilia and mast cell activation, including abundant production of prostaglandin D 2 (PGD 2 ). Group 2 innate lymphoid cells (ILC2s), which promote tissue eosinophilia and mast cell responses, undergo chemotaxis and cytokine production in response to PGD 2 , but it is unknown whether ILC2s are active in patients with AERD. Objective We sought to determine whether ILC2 numbers change in peripheral blood and the nasal mucosa during COX-1 inhibitor–induced reactions in patients with AERD. Methods Blood and nasal scrapings were collected at baseline, during reactions, and after completion of ketorolac/aspirin challenge/desensitization in 12 patients with AERD. ILC2s and eosinophils were quantitated by means of flow cytometry. Urine was also collected, and quantification of PGD 2 metabolite and leukotriene E 4 levels was done by using ELISA. Baseline and nonsteroidal anti-inflammatory drug reaction clinical data were correlated with cell changes. Results ILC2 numbers significantly increased in nasal mucosal samples and decreased in blood at the time of COX-1 inhibitor reactions in 12 patients with AERD. These changes were not observed in 2 patients without AERD. Furthermore, eosinophil numbers decreased in blood concurrently with significant increases in urinary PGD 2 metabolite and leukotriene E 4 levels. The magnitude of increases in nasal mucosal ILC2 numbers positively correlated with maximum symptom scores during challenges. Furthermore, blood ILC2 numbers during the reaction correlated with time for the reaction to resolve, possibly reflecting reaction severity. Conclusions ILC2s are recruited to the nasal mucosa during COX-1 inhibitor–induced reactions in patients with AERD, correlating with enhanced production of prostaglandins and leukotrienes.

Recent studies suggest that both bacteria and rhinoviruses (RVs) contribute to asthma exacerbations. We hypothesized that bacteria might alter antiviral responses early in the course of infection by modifying monocyte-lineage chemokine responses to RV infection. To test this hypothesis, human blood monocytes or bronchoalveolar lavage (BAL) macrophages were treated with RV types A016, B014, A001, and/or A002 in the presence or absence of LPS, and secretion of chemokines (CXCL10, CXCL11, CCL2, and CCL8) and IFN-α was measured by ELISA. Treatment with RV alone induced blood monocytes and BAL macrophages to secrete CXCL10, CXCL11, CCL2, and CCL8. Pretreatment with LPS significantly attenuated RV-induced CXCL10, CXCL11, and CCL8 secretion by 68–99.9% on average (P < 0.0001, P < 0.004, and P < 0.002, respectively), but did not inhibit RV-induced CCL2 from blood monocytes. Similarly, LPS inhibited RV-induced CXCL10 and CXCL11 secretion by over 88% on average from BAL macrophages (P < 0.002 and P < 0.0001, respectively). Furthermore, LPS inhibited RV-induced signal transducer and activator of transcription 1 phosphorylation (P < 0.05), as determined by immunoblotting, yet augmented RV-induced IFN-α secretion (P < 0.05), and did not diminish expression of RV target receptors, as measured by flow cytometry. In summary, major and minor group RVs strongly induce chemokine expression and IFN-α from monocytic cells. The bacterial product, LPS, specifically inhibits monocyte and macrophage secretion of RV-induced CXCL10 and CXCL11, but not other highly induced chemokines or IFN-α. These effects suggest that airway bacteria could modulate the pattern of virus-induced cell recruitment and inflammation in the airways.

Activation of the ionotropic P2RX7 nucleotide receptor by extracellular ATP has been implicated in modulating inflammatory disease progression. Continuous exposure of P2RX7 to ligand can result in apoptosis in many cell types, including monocytic cells, whereas transient activation of P2RX7 is linked to inflammatory mediator production and the promotion of cell growth. Given the rapid hydrolysis of ATP in the circulation and interstitial space, transient activation of P2RX7 appears critically important for its action, yet its effects on gene expression are unclear. The present study demonstrates that short-term stimulation of human and mouse monocytic cells as well as mouse osteoblasts with P2RX7 agonists substantially induces the expression of several activating protein-1 (AP-1) members, particularly FosB. The potent activation of FosB after P2RX7 stimulation is especially noteworthy considering that little is known concerning the role of FosB in immunological regulation. Interestingly, the magnitude of FosB activation induced by P2RX7 stimulation appears greater than that observed with other known inducers of FosB expression. In addition, we have identified a previously unrecognized role for FosB in osteoblasts with respect to nucleotide-induced expression of cyclooxygenase-2 (COX-2), which is the rate-limiting enzyme in prostaglandin biosynthesis from arachidonic acid and is critical for osteoblastic differentiation and immune behavior. The present studies are the first to link P2RX7 action to FosB/AP-1 regulation in multiple cell types, including a role in nucleotide-induced COX-2 expression, and support a role for FosB in the control of immune and osteogenic function by P2RX7.

To the Editor: Allergy and viral infection are the 2 main risk factors for the inception of persistent wheezing and asthma in early childhood and for acute exacerbations of established asthma.1 The underlying mechanisms of how these 2 distinct sources of inflammation contribute to asthma inception and exacerbation are incompletely understood. Macrophages are the most numerous leukocyte (>90%) in the lower airways. Although rhinovirus (RV) does not replicate in macrophages, it directly interacts with macrophages in airway tissues.2 Once activated by the RV, macrophages can secrete various inflammatory chemokines, depending on the differentiation state and phenotype of the cells. For example, macrophages can secrete CXCL10 and CXCL11, which recruit adaptive immune cells that direct viral clearance,3 and/or CCL2 and CCL8, which can recruit myeloid cells such as monocytes and eosinophils.4 In mouse models of RV infection, macrophage responses contribute to airway inflammation and hyperresponsiveness. Furthermore, RV activation of monocytes and macrophages can potentiate antiviral responses of airway epithelial cells.5 These findings provide evidence that macrophages are important immunoregulatory cells during RV infections. A recent study demonstrated reduced antiviral responses to RV stimulation in allergic asthmatic children, corresponding with increased susceptibility to RV-induced exacerbations.6 Therefore, we hypothesized that allergen exposure modifies RV-induced chemokine responses of airway macrophages to impair the antiviral response and promote inflammation. Together, these effects could increase the severity of viral respiratory infections and lead to lower respiratory tract symptoms in patients with asthma. To test this hypothesis, in accordance with institutional review board–approved protocols from the Human Subjects Committee at the University of Wisconsin-Madison, bronchoalveolar lavage (BAL) was performed to obtain airway mononuclear cells (MNCs) from 10 donors with a history of mild atopic asthma (see Table E1 in this article's Online Repository at www.jacionline.org). BAL was performed before (D0) and 48 hours after (D2) segmental bronchoprovocation with allergen as previously described.7 Contaminating granulocytes were removed from D2 cells by Percoll density gradient centrifugation (see this article's Methods section in the Online Repository at www.jacionline.org), and cell populations were comparable between the 2 isolations (see Fig E1 in this article's Online Repository at www.jacionline.org). Macrophages were isolated from D0 cell and D2 MNC populations by adherence to plastic (2 hours). FIG E1 Cell differentials in BAL cell population on D0 and D2. Eos, Eosinophils; Lymph, lymphocytes; Macs, macrophages; PMNs, polymorphonuclear leukocytes. The values are means ± SEM from 9 individual subjects. *P < .05 by 2-way repeated-measures ... TABLE E1 Subject characteristics To determine the effect of allergen challenge on RV-induced macrophage responses, D0 and D2 BAL macrophages were incubated (24 hours) with RV A016, B014, and A002 (see this article's Methods section). The supernatants were then analyzed for CXCL10, CXCL11, CCL2, and CCL8 using ELISA (Fig 1). In the absence of virus, D0 macrophages secreted low levels of CXCL10 and CCL2 (geometric mean [GM], 95 and 3739 pg/mL, respectively), but not CXCL11 and CCL8, and incubation with RVA016, B014, and A002 significantly induced the secretion of all the 4 chemokines tested (P < .001). FIG 1 Allergen challenge alters RV-induced chemokine secretion in primary human BAL macrophages: CXCL10 (A), CXCL11 (B), CCL2 (C), and CCL8 (D). NS, Not significant; *P < .05, **P < .01, and ***P < .001 by 2-way repeated-measures ANOVA ... After allergen challenge, RV stimulation still significantly induced the secretion of CXCL10, CXCL11, CCL2, and CCL8 (P