A common polymorphism in the Intelectin-1 gene influences mucus plugging in severe asthma.

By incompletely understood mechanisms, type 2 (T2) inflammation present in the airways of severe asthmatics drives the formation of pathologic mucus which leads to airway mucus plugging. Here we investigate the molecular role and clinical significance of intelectin-1 (ITLN-1) in the development of pathologic airway mucus in asthma. Through analyses of human airway epithelial cells we find that ITLN1 gene expression is highly induced by interleukin-13 (IL-13) in a subset of metaplastic MUC5AC+ mucus secretory cells, and that ITLN-1 protein is a secreted component of IL-13-induced mucus. Additionally, we find ITLN-1 protein binds the C-terminus of the MUC5AC mucin and that its deletion in airway epithelial cells partially reverses IL-13-induced mucostasis. Through analysis of nasal airway epithelial brushings, we find that ITLN1 is highly expressed in T2-high asthmatics, when compared to T2-low children. Furthermore, we demonstrate that both ITLN-1 gene expression and protein levels are significantly reduced by a common genetic variant that is associated with protection from the formation of mucus plugs in T2-high asthma. This work identifies an important biomarker and targetable pathways for the treatment of mucus obstruction in asthma.

A Novel DNase Assay Reveals Low DNase Activity in Severe Asthma.

Secreted deoxyribonucleases (DNases), such as DNase-1 and DNase-IL3, degrade extracellular DNA, and endogenous DNases have roles in resolving airway inflammation and guarding against autoimmune responses to nucleotides. Subsets of patients with asthma have high airway DNA levels, but information about DNase activity in health and in asthma is lacking. To characterize DNase activity in health and in asthma, we developed a novel kinetic assay using a Taqman probe sequence that is quickly cleaved by DNase-I to produce a large product signal. We used this kinetic assay to measure DNase activity in sputum from participants in the Severe Asthma Research Program (SARP)-3 (n=439) and from healthy controls (n=89). We found that DNase activity was lower than normal in asthma (78.7 RFU/min vs 120.4 RFU/min, p<0.0001). Compared to asthma patients with sputum DNase activity levels in the upper tertile activity levels, those in the lower tertile of sputum DNase activity were characterized clinically by more severe disease and pathologically by airway eosinophilia and airway mucus plugging. Carbamylation of DNase-I, a post translational modification that can be mediated by eosinophil peroxidase, inactivated DNase-I. In summary, a Taqman probe-based DNase activity assay uncovers low DNase activity in the asthma airway which is associated with more severe disease and airway mucus plugging and may be caused, at least in part, by eosinophil-mediated carbamylation.

Lung Interstitial Macrophages Can Present Soluble Antigens and Induce Foxp3+ Regulatory T Cells.

Lung macrophages constitute a sophisticated surveillance and defense system that contributes to tissue homeostasis, host defense, and allows the host to cope with the myriad of insults and antigens to which the lung mucosa is exposed. As opposed to alveolar macrophages, lung interstitial macrophages express high levels of type 2 major histocompatibility complex (MHC-II), a hallmark of antigen-presenting cells. Here, we showed that lung IMs, like dendritic cells (DCs), possess the machinery to present soluble antigens in an MHC-II-restricted way. Using ex vivo ovalbumin (OVA)-specific T cell proliferation assays, we found that OVA-pulsed IMs could trigger OVA-specific CD4+ T cell proliferation and Foxp3 expression via MHC-II-, IL-10 and Tgfß-dependent mechanisms. Moreover, we showed that IMs efficiently captured locally instilled antigens in vivo, did not migrate to the draining lymph nodes and enhanced local interactions with CD4+ T cells in a model of OVA-induced allergic asthma. These results support that IMs can present antigens to CD4+ T cells and trigger regulatory T cells, which might attenuate lung immune responses and have functional consequences for lung immunity and T-cell-mediated disorders.

Airway Macrophages Encompass Transcriptionally and Functionally Distinct Subsets Altered by Smoking.

Alveolar macrophages (AMs) are functionally important innate cells involved in lung homeostasis and immunity and whose diversity in health and disease is a subject of intense investigations. Yet, it remains unclear to what extent conditions like smoking or chronic obstructive pulmonary disease (COPD) trigger changes in the AM compartment. Here, we aimed to explore heterogeneity of human AMs isolated from healthy nonsmokers, smokers without COPD, and smokers with COPD by analyzing BAL fluid cells by flow cytometry and bulk and single-cell RNA sequencing. We found that subpopulations of BAL fluid CD206+ macrophages could be distinguished based on their degree of autofluorescence in each subject analyzed. CD206+ autofluorescenthigh AMs were identified as classical, self-proliferative AM, whereas autofluorescentlow AMs were expressing both monocyte and classical AM-related genes, supportive of a monocytic origin. Of note, monocyte-derived autofluorescentlow AMs exhibited a functionally distinct immunoregulatory profile, including the ability to secrete the immunosuppressive cytokine IL-10. Interestingly, single-cell RNA-sequencing analyses showed that transcriptionally distinct clusters of classical and monocyte-derived AM were uniquely enriched in smokers with and without COPD as compared with healthy nonsmokers. Of note, such smoking-associated clusters exhibited gene signatures enriched in detoxification, oxidative stress, and proinflammatory responses. Our study independently confirms previous reports supporting that monocyte-derived macrophages coexist with classical AM in the airways of healthy subjects and patients with COPD and identifies smoking-associated changes in the AM compartment that may favor COPD initiation or progression.

Myoferlin targeting triggers mitophagy and primes ferroptosis in pancreatic cancer cells.

Myoferlin, an emerging oncoprotein, has been associated with a low survival in several cancer types including pancreas ductal adenocarcinoma where it controls mitochondria structure and respiratory functions. Owing to the high susceptibility of KRAS-mutated cancer cells to iron-dependent cell death, ferroptosis, and to the high iron content in mitochondria, we investigated the relation existing between mitochondrial integrity and iron-dependent cell death. We discovered that myoferlin targeting with WJ460 pharmacological compound triggered mitophagy and ROS accumulation culminating with lipid peroxidation and apoptosis-independent cell death. WJ460 caused a reduction of the abundance of ferroptosis core regulators xc- cystine/glutamate transporter and GPX-4. Mitophagy inhibitor Mdivi1 and iron chelators inhibited the myoferlin-related ROS production and restored cell growth. Additionally, we reported a synergic effect between ferroptosis inducers, erastin and RSL3, and WJ460.

The interstitial macrophage: A long-neglected piece in the puzzle of lung immunity.

Lung macrophages have mostly been studied considering only their most accessible and well-defined representative, the alveolar macrophage (AM). In contrast, the identity and putative immune functions of their tissue counterpart, the interstitial macrophage (IM), have long remained much more elusive. Yet, recent evidence supports the notion that IMs perform important immune functions in the lung, notably in terms of innate immunoregulation. Here, we review current knowledge on the phenotype, ontogeny and function of IMs and propose strategies for the unambiguous identification and study of this important and dynamic lung innate immune cell population.