Asthma Lung Research Articles

Interaction of Interleukin-17A with a Th2 Response in a Mouse Model of Allergic Airway Inflammation.

A total of 262 million people worldwide suffer from asthma and 461000 people died from it in 2019. Asthma is a disease with different endotypes defined by the granulocytes found in the asthmatic lung. In allergic asthma, the eosinophilic endotype is present, driven by a TH2 response. A TH17 immune response leads to the neutrophil endotype. This often causes uncontrolled asthma and is triggered by pollutants, microbes, and oxidative stress. It has been described that a significant number of patients with eosinophilic asthma develop mixed granulocytic asthma over time. The severity of asthma in the mixed endotype is related to the proportion of neutrophils in the lungs. In this report, we address the question of how a TH2 response interacts with IL-17A in allergic asthma. To this end, we used a mouse model to induce allergic asthma followed by an aerosol challenge with ovalbumin. To investigate the role of IL-17A, we administered IL-17A intranasally during the challenge phase. IL-17A alone did not elicit an immune response, whereas in combination with allergic asthma, it resulted in a shift of the asthmatic endotype from eosinophilic to neutrophilic. TGFβ1 was increased in these lungs compared to asthmatic lungs without IL-17A, as was the expression of the IL-17A receptor subunits IL-17RA and IL-17RC. In cultures with human cells, we also found that IL-17A increased the expression of its receptors only in combination with IL-13. We also found this effect for IL-8, which attracts neutrophils in humans. The TH2 response increased the sensitivity to IL-17A in a mouse asthma model as well as in human cell lines.

Formulation and Evaluation of the Anti-inflammatory, Anti-oxidative, and Anti-remodelling Effects of the Niosomal Myrtenol on the Lungs of Asthmatic Rats.

Asthma is a common chronic allergic disease that affects a significant percentage of the world's population. Niosomes are nanoparticles consisting of non-ionic surfactants that can be used for drug delivery. This research was designed to investigate the impacts of inhalation of simple and niosomal forms of myrtenol against adverse consequences of asthma in rats. Asthma induction was performed via injection of ovalbumin, followed by its inhalation. Niosomes were created by a heating protocol, and their physicochemical features were evaluated. Forty-nine male Wistar rats were allotted into 7 groups (n=7 each): Control (CTL), vacant niosome (VN), Asthma, Asthma+VN, Asthma+SM (simple myrtenol), Asthma+NM (niosomal myrtenol), and Asthma+B (budesonide). Lung remodeling, serum immunoglobulin E (IgE), inflammatory and cytokines, and antioxidant factors in the lung tissue and bronchoalveolar fluid (BALF), as well as), were evaluated. The results showed that myrtenol-loaded niosomes had appropriate encapsulation efficiency, kinetic release, size, and zeta potential. The thickness of the epithelial cell layer in the lungs, as well as cell infiltration, fibrosis, IgE, reactive oxygen species, interleukin (IL)-6, and tumor nuclear factor alpha (TNF-α) levels, decreased significantly. In contrast, superoxide dismutase and glutathione peroxide activity increased significantly in the serum and BALF of the treated groups. The niosomal form of myrtenol revealed a higher efficacy than simple myrtenol and was similar to budesonide in ameliorating asthma indices. Inhalation of simple and niosomal forms of myrtenol improved the detrimental changes in the asthmatic lung. The niosomal form induced more prominent anti-asthmatic effects comparable to those of budesonide.

The emerging role of extracellular vesicles as communicators between adipose tissue and pathologic lungs with a special focus on asthma.

Extracellular vesicles (EVs) gain increasing attention due to their (patho-)physiological role in intercellular signaling, specifically in the communication between distant organs. Recent studies highlight a connection between the adipose tissue (AT) and the lung via (immuno-)modulatory EVs in disorders such as obesity-associated asthma and lung cancer-associated cachexia. Although lung cancer-derived EVs induce lipolysis and myotube atrophy in vivo, pathogenic effects were also reported in the opposite direction with the involvement of AT-derived EVs in cancer-promoting responses and potentially in asthma development. In contrast, the majority of studies on AT-derived EVs demonstrate their protective influence on the asthmatic lung. Beneficial effects, such as induction of anti-inflammatory pathways in vitro and in ovalbumin (OVA)-induced asthma mouse models, were particularly conveyed by EVs enriched from AT-derived mesenchymal stem/stromal cells (AT-MSCs), which therefore pose an interesting subject in possible future therapeutic applications. Likewise, AT-MSC-derived EVs exerted beneficial effects in several other pulmonary abnormalities, such as different types of lung injury or pathological changes related to chronic obstructive pulmonary disease. These contradictory findings highlight the need for extensive research to widen the understanding of the role of EVs in the development of diseases and interconnectivity between organs.

Targeting Multiple Costimulatory Molecules Prevents Persistence of Lung Tissue-resident Memory T cells and Leads to Airway Tolerance

Abstract Pools of tissue-resident memory T (Trm) cells are thought to maintain and contribute to exacerbations of inflammatory or autoimmune disease. We hypothesized that several costimulatory molecules would control the expansion and maintenance of these populations when responding to antigen, and tested the potential of targeting OX40L, CD30L, and ICOSL, using a house dust mite (HDM) allergen-based mouse model of asthmatic lung inflammation. We found that allergen induced persisting CD4+CD44hiCD62Llo CD69+ and CD69− lung-localized Trm cells and RNA-seq revealed the heterogeneity of these memory pools, containing Th2, Tfh, Th17, and Th1 clusters. Showing their reliance on costimulation for recall activity to allergen, blocking CD30L and OX40L together, or blocking ICOSL alone, resulted in ~50% reduction in the expansion of both CD69+ and CD69− tissue-resident effector T cells during a secondary allergen exposure or a tertiary allergen exposure. Blocking all three molecules at once further inhibited T cell accumulation by ~90%, corresponding to a similar reduction in lung tissue inflammation, including eosinophilia. Moreover, triple costimulation blockade resulted in a diminished persisting lung tissue-resident memory T cell pool, which failed to expand to normal levels and drive further exacerbations of lung inflammation upon subsequent exposure to allergen, resulting in a state of airway tolerance. Overall, our data suggest that the concurrent inhibition of OX40, CD30, and ICOS signaling, can impair the expansion and persistence of lung-localized Trm cells and could engender long-lasting tolerance in asthma and similar autoimmune or inflammatory diseases that are maintained by tissue-resident memory T cells.

Multi-Resolution 3D Optical Mapping of Immune Cell Infiltrates in Mouse Asthmatic Lung.

Asthma is a chronic inflammatory airway disease driven by various infiltrating immune cell types into the lung. Optical microscopy has been used to study immune infiltrates in the asthmatic lungs. Confocal laser scanning microscopy (CLSM) identifies the phenotypes and locations of individual immune cells in lung tissue sections by employing high-magnification objectives and multiplex immunofluorescence staining. In contrast, light-sheet fluorescence microscopy (LSFM) can visualize the macroscopic and mesoscopic architecture of whole-mount lung tissues in three dimensions (3D) by adopting an optical tissue clearing method. Despite each microscopy method producing image data with unique resolution from a tissue sample, CLSM and LSFM have not been applied together due to the difference in the tissue preparation procedures. Here we introduce a new approach combining LSFM and CLSM into a sequential imaging pipeline. We built a new optical tissue clearing workflow in which the immersion clearing agent can be switched from an organic solvent to an aqueous sugar solution for sequential 3D LSFM and CLSM of mouse lungs. This sequential combination microscopy offered quantitative 3D spatial analyses of the distribution of immune infiltrates in the same mouse asthmatic lung tissue at the organ, tissue, and cell level. These results show that our method facilitates multi-resolution 3D fluorescence microscopy as a new imaging approach providing comprehensive spatial information for a better understanding of inflammatory lung diseases. This article is open access and distributed under the terms of the Creative Commons Attribution Non-Commercial No Derivatives License 4.0 (http://creativecommons.org/licenses/by-nc-nd/4.0/).

Lymphotoxin beta receptor signaling directly controls airway smooth muscle deregulation and asthmatic lung dysfunction

Dysregulation of airway smooth muscle cells (ASM) is central to the severity of asthma. Which molecules dominantly control ASM in asthma is unclear. High levels of the cytokine LIGHT (aka TNFSF14) have been linked to asthma severity and lower baseline predicted FEV1 percentage, implying that signals through its receptors might directly control ASM dysfunction. Our study sought to determine whether signaling via lymphotoxin beta receptor (LTβR) or herpesvirus entry mediator from LIGHT dominantly drives ASM hyperreactivity induced by allergen. Conditional knockout mice deficient for LTβR or herpesvirus entry mediator in smooth muscle cells were used to determine their role in ASM deregulation and airway hyperresponsiveness (AHR) invivo. Human ASM were used to study signals induced by LTβR. LTβR was strongly expressed in ASM from normal and asthmatic subjects compared to several other receptors implicated in smooth muscle deregulation. Correspondingly, conditional deletion of LTβR only in smooth muscle cells in smMHCCreLTβRfl/fl mice minimized changes in their numbers and mass as well as AHR induced by house dust mite allergen in a model of severe asthma. Intratracheal LIGHT administration independently induced ASM hypertrophy and AHR invivo dependent on direct LTβR signals to ASM. LIGHT promoted contractility, hypertrophy, and hyperplasia of human ASM invitro. Distinguishing LTβR from the receptors for IL-13, TNF, and IL-17, which have also been implicated in smooth muscle dysregulation, LIGHT promoted NF-κB-inducing kinase-dependent noncanonical nuclear factor kappa-light-chain enhancer of activated B cells in ASM invitro, leading to sustained accumulation of F-actin, phosphorylation of myosin light chain kinase, and contractile activity. LTβR signals directly and dominantly drive airway smooth muscle hyperresponsiveness relevant for pathogenesis of airway remodeling in severe asthma.

DOCK2 Promotes Asthma Development by Eliciting Airway Epithelial-Mesenchymal Transition.

Epithelial-mesenchymal transition (EMT) contributes to airway remodeling, a predominant feature of asthma. DOCK2 (dedicator of cytokinesis 2) is an innate immune signaling molecule involved in vascular remodeling. However, it is unknown if DOCK2 plays a role in airway remodeling during asthma development. In this study, we found that DOCK2 is highly induced in both normal human bronchial epithelial cells treated with house dust mite (HDM) extract and human asthmatic airway epithelium. DOCK2 is also upregulated by TGF-β1 (transforming growth factor β1) during EMT of human bronchial epithelial cells. Importantly, knockdown of DOCK2 inhibits, and overexpression of DOCK2 promotes, TGF-β1-induced EMT. Consistently, DOCK2 deficiency suppresses the EMT of airway epithelium, attenuates the subepithelial fibrosis, and improves pulmonary function in HDM-induced asthmatic lungs. These data suggest that DOCK2 plays an important role in EMT and asthma development. Mechanistically, DOCK2 interacts with transcription factor FoxM1 (forkhead box M1), which enhances FoxM1 binding to mesenchymal marker gene promoters and further promotes mesenchymal marker gene transcription and expression, leading to EMT. Taken together, our study identifies DOCK2 as a novel regulator for airway EMT in an HDM-induced asthma model, thus providing a potential therapeutic target for treatment of asthma.

Airway Epithelium: A Neglected but Crucial Cell Type in Asthma Pathobiology.

The features of allergic asthma are believed to be mediated mostly through the Th2 immune response. In this Th2-dominant concept, the airway epithelium is presented as the helpless victim of Th2 cytokines. However, this Th2-dominant concept is inadequate to fill some of the vital knowledge gaps in asthma pathogenesis, like the poor correlation between airway inflammation and airway remodeling and severe asthma endotypes, including Th2-low asthma, therapy resistance, etc. Since the discovery of type 2 innate lymphoid cells in 2010, asthma researchers started believing in that the airway epithelium played a crucial role, as alarmins, which are the inducers of ILC2, are almost exclusively secreted by the airway epithelium. This underscores the eminence of airway epithelium in asthma pathogenesis. However, the airway epithelium has a bipartite functionality in sustaining healthy lung homeostasis and asthmatic lungs. On the one hand, the airway epithelium maintains lung homeostasis against environmental irritants/pollutants with the aid of its various armamentaria, including its chemosensory apparatus and detoxification system. Alternatively, it induces an ILC2-mediated type 2 immune response through alarmins to amplify the inflammatory response. However, the available evidence indicates that restoring epithelial health may attenuate asthmatic features. Thus, we conjecture that an epithelium-driven concept in asthma pathogenesis could fill most of the gaps in current asthma knowledge, and the incorporation of epithelial-protective agents to enhance the robustness of the epithelial barrier and the combative capacity of the airway epithelium against exogenous irritants/allergens may mitigate asthma incidence and severity, resulting in better asthma control.

Targeted GATA3 knockdown in activated T cells via pulmonary siRNA delivery as novel therapy for allergic asthma.

GATA3 gene silencing in activated T cells displays a promising option to early-on undermine pathological pathways in the disease formation of allergic asthma. The central transcription factor of T helper 2 (Th2) cell cytokines IL-4, IL-5, and IL-13 plays a major role in immune and inflammatory cascades underlying asthmatic processes in the airways. Pulmonary delivery of small interfering RNAs (siRNA) to induce GATA3 knockdown within disease related T cells of asthmatic lungs via RNA interference (RNAi) presents an auspicious base to realize this strategy, however, still faces some major hurdles. Main obstacles for successful siRNA delivery in general comprise stability and targeting issues, while in addition the transfection of T cells presents a particularly challenging task itself. In previous studies, we have developed and advanced an eligible siRNA delivery system composed of polyethylenimine (PEI) as polycationic carrier, transferrin (Tf) as targeting ligand and melittin (Mel) as endosomolytic agent. Resulting Tf-Mel-PEI polyplexes exhibited ideal characteristics for targeted siRNA delivery to activated T cells and achieved efficient and sequence-specific gene knockdown in vitro. In this work, the therapeutic potential of this carrier system was evaluated in an optimized cellular model displaying the activated status of asthmatic T cells. Moreover, a suitable siRNA sequence combination was found for effective gene silencing of GATA3. To confirm the translatability of our findings, Tf-Mel-PEI polyplexes were additionally tested ex vivo in activated human precision-cut lung slices (PCLS). Here, the formulation showed a safe profile as well as successful delivery to the lung epithelium with 88% GATA3 silencing in lung explants. These findings support the feasibility of Tf-Mel-PEI as siRNA delivery system for targeted gene knockdown in activated T cells as a potential novel therapy for allergic asthma.

Asthmatic lung fibroblasts promote type 2 immune responses via endoplasmic reticulum stress response dependent thymic stromal lymphopoietin secretion.

Lung fibroblasts contribute to asthma pathology partly through modulation of the immune environment in the airway. Tumor necrosis factor-α (TNFα) expression is upregulated in asthmatic lungs. How asthmatic lung fibroblasts respond to TNFα stimulation and subsequently regulate immune responses is not well understood. Endoplasmic reticulum (ER) stress and unfolded protein responses (UPR) play important roles in asthma, but their functional roles are still under investigation. In this study, we investigated TNFα-induced cytokine production in primary lung fibroblasts from asthmatic vs. non-asthmatic human subjects, and downstream effects on type 2 immune responses. TNFα significantly upregulated IL-6, IL-8, C-C motif chemokine ligand 5 (CCL5), and thymic stromal lymphopoietin (TSLP) mRNA expression and protein secretion by lung fibroblasts. Asthmatic lung fibroblasts secreted higher levels of TSLP which promoted IL-33-induced IL-5 and IL-13 production by peripheral blood mononuclear cells. TNFα exposure enhanced expression of ER stress/UPR pathways in both asthmatic and non-asthmatic lung fibroblasts, especially inositol-requiring protein 1α in asthmatics. ER stress/UPR inhibitors decreased IL-6, CCL5, and TSLP protein secretion by asthmatic lung fibroblasts. Our data suggest that TNFα and lung fibroblasts form an important axis in asthmatic lungs to promote asthmatic inflammation that can be attenuated by inhibiting ER stress/UPR pathway.

Evaluation Anti-inflammatory Effect of Conjugated Gold Nanoparticles with Cortistatin Peptide as Drug Delivery to Asthmatic Lung Tissue

Evaluation Anti-inflammatory Effect of Conjugated Gold Nanoparticles with Cortistatin Peptide as Drug Delivery to Asthmatic Lung Tissue

PLZF promotes the development of asthma tolerance via affecting memory phenotypes of immune cells.

Clarifying the pathogenesis of asthma and/or identifying the specific pathway underlying oral asthma tolerance (OT) would be of great significance. In our previous study, promyelocytic leukemia zinc finger (PLZF), which reportedly regulates memory phenotypes, was found to promote ovalbumin (OVA)-induced OT. Therefore, this study aimed to explore the regulatory effects of PLZF on memory phenotypes in asthma and OT mouse models. We found that Zbtb16 (encoding PLZF) and PLZF+ cells were highly increased in OT lungs compared with asthmatic lungs. PLZF was co-expressed with GATA3, and IL-4+PLZF+ cells were significantly lower in OT lungs than in asthmatic lungs. Notably, memory cells were decreased in OT mice, and these mice had PLZF+ cells that expressed lower levels of CD44 than those of asthmatic mice. When Zbtb16 was overexpressed in splenic lymphocytes, the number of CD44+ cells decreased. There were increased memory cells in splenic lymphocytes after treatment with the supernatant of OVA-treated airway epithelial cells; however, this was reversed by Zbtb16 overexpression. Early respiratory syncytial virus infection increased memory cells and reduced PLZF+ cells in the OT mice. Collectively, these results indicate that PLZF may reduce the proportion of memory cells, thereby, promoting the establishment of OT.

Ozone impairs endogenous compensatory responses in allergic asthma

Asthma is a chronic inflammatory airway disease characterized by acute exacerbations triggered by inhaled allergens, respiratory infections, or air pollution. Ozone (O3), a major component of air pollution, can damage the lung epithelium in healthy individuals. Despite this association, little is known about the effects of O3 and its impact on chronic lung disease. Epidemiological data have demonstrated that elevations in ambient O3 are associated with increased asthma exacerbations. To identify mechanisms by which O3 exposure leads to asthma exacerbations, we developed a two-hit mouse model where mice were sensitized and challenged with three common allergens (dust mite, ragweed and Aspergillus fumigates, DRA) to induce allergic inflammation prior to exposure to O3 (DRAO3). Changes in lung physiology, inflammatory cells, and inflammation were measured. Exposure to O3 following DRA significantly increased airway hyperreactivity (AHR), which was independent of TLR4. DRA exposure resulted in increased BAL eosinophilia while O3 exposure resulted in neutrophilia. Additionally, O3 exposure following DRA blunted anti-inflammatory and antioxidant responses. Finally, there were significantly less monocytes and innate lymphoid type 2 cells (ILC2s) in the dual challenged DRA-O3 group suggesting that the lack of these immune cells may influence O3-induced AHR in the setting of allergic inflammation. In summary, we developed a mouse model that mirrors some aspects of the clinical course of asthma exacerbations due to air pollution and identified that O3 exposure in the asthmatic lung leads to impaired endogenous anti-inflammatory and antioxidant responses and alterations inflammatory cell populations.

"Liquid biopsy" - extracellular vesicles as potential novel players towards precision medicine in asthma.

Extracellular vesicles (EVs) have emerged as vital mediators in intracellular communication in the lung microenvironment. Environmental exposure to various triggers (e.g., viruses, allergens) stimulates the EV-mediated cascade of pro-inflammatory responses that play a key role in the asthma pathomechanism. This complex EV-mediated crosstalk in the asthmatic lung microenvironment occurs between different cell types, including airway epithelial cells and immune cells. The cargo composition of EVs mirrors hereby the type and activation status of the parent cell. Therefore, EVs collected in a noninvasive way (e.g., in nasal lavage, serum) could inform on the disease status as a "liquid biopsy", which is particularly important in the pediatric population. As a heterogeneous disease, asthma with its distinct endotypes and phenotypes requires more investigation to develop novel diagnostics and personalized case management. Filling these knowledge gaps may be facilitated by further EV research. Here, we summarize the contribution of EVs in the lung microenvironment as potential novel players towards precision medicine in the development of asthma. Although rapidly evolving, the EV field is still in its infancy. However, it is expected that a better understanding of the role of EVs in the asthma pathomechanism will open up new horizons for precision medicine diagnostic and therapeutic solutions.

Thrombin induces IL-8/CXCL8 expression by DCLK1-dependent RhoA and YAP activation in human lung epithelial cells

BackgroundDoublecortin-like kinase 1 (DCLK1) has been recognized as a marker of cancer stem cell in several malignancies. Thrombin is crucial in asthma severity as it can promote IL-8/CXCL8 production in lung epithelial cells, which is a potent chemoattractant for neutrophils. However, the pathologic role of DCLK1 in asthma and its involvement in thrombin-stimulated IL-8/CXCL8 expression remain unknown.MethodsIL-8/CXCL8, thrombin, and DCLK1 expression were observed in the lung tissues of severe asthma patients and ovalbumin (OVA)-induced asthmatic mice model. A549 and BEAS-2B cells were either pretreated with inhibitors or small interfering RNAs (siRNAs) before being treated with thrombin. IL-8/CXCL8 expression and the molecules involved in signaling pathway were performed using ELISA, luciferase activity assay, Western blot, or ChIP assay.ResultsIL-8/CXCL8, thrombin, and DCLK1 were overexpressed in the lung tissues of severe asthma patients and ovalbumin (OVA)-induced asthmatic mice model. Our in vitro study found that DCLK siRNA or LRKK2-IN-1 (DCLK1 inhibitor) attenuated IL-8/CXCL8 release after thrombin induction in A549 and BEAS-2B cells. Thrombin activated DCLK1, RhoA, and YAP in a time-dependent manner, in which DCLK1 siRNA inhibited RhoA and YAP activation. YAP was dephosphorylated on the Ser127 site after thrombin stimulation, resulting in YAP translocation to the nucleus from the cytosol. DCLK1, RhoA and YAP activation following thrombin stimulation were inhibited by U0126 (ERK inhibitor). Moreover, DCLK1 and YAP siRNA inhibited κB-luciferase activity. Thrombin stimulated the recruitment of YAP and p65 to the NF-κB site of the IL-8/CXCL8 promoter and was inhibited by DCLK1 siRNA.ConclusionsThrombin activates the DCLK1/RhoA signaling pathway, which promotes YAP activation and translocation to the nucleus from the cytosol, resulting in YAP/p65 formation, and binding to the NF-κB site, which enhances IL-8/CXCL8 expression. DCLK1 might be essential in thrombin-stimulated IL-8/CXCL8 expression in asthmatic lungs and indicates a potential therapeutic strategy for severe asthma treatment.

Single-cell transcriptomic characterization reveals the landscape of airway remodeling and inflammation in a cynomolgus monkey model of asthma.

Monkey disease models, which are comparable to humans in terms of genetic, anatomical, and physiological characteristics, are important for understanding disease mechanisms and evaluating the efficiency of biological treatments. Here, we established an A.suum-induced model of asthma in cynomolgus monkeys to profile airway inflammation and remodeling in the lungs by single-cell RNA sequencing (scRNA-seq). The asthma model results in airway hyperresponsiveness and remodeling, demonstrated by pulmonary function test and histological characterization. scRNA-seq reveals that the model elevates the numbers of stromal, epithelial and mesenchymal cells (MCs). Particularly, the model increases the numbers of endothelial cells (ECs), fibroblasts (Fibs) and smooth muscle cells (SMCs) in the lungs, with upregulated gene expression associated with cell functions enriched in cell migration and angiogenesis in ECs and Fibs, and VEGF-driven cell proliferation, apoptotic process and complement activation in SMCs. Interestingly, we discover a novel Fib subtype that mediates type I inflammation in the asthmatic lungs. Moreover, MCs in the asthmatic lungs are found to regulate airway remodeling and immunological responses, with elevated gene expression enriched in cell migration, proliferation, angiogenesis and innate immunological responses. Not only the numbers of epithelial cells in the asthmatic lungs change at the time of lung tissue collection, but also their gene expressions are significantly altered, with an enrichment in the biological processes of IL-17 signaling pathway and apoptosis in the majority of subtypes of epithelial cells. Moreover, the ubiquitin process and DNA repair are more prevalent in ciliated epithelial cells. Last, cell-to-cell interaction analysis reveals a complex network among stromal cells, MCs and macrophages that contribute to the development of asthma and airway remodeling. Our findings provide a critical resource for understanding the principle underlying airway remodeling and inflammation in a monkey model of asthma, as well as valuable hints for the future treatment of asthma, especially the airway remodeling-characterized refractory asthma.

Leptin-producing monocytes in the airway submucosa may contribute to asthma pathogenesis

BackgroundObesity leads to an increase in the incidence and severity of asthma. Adipokines, such as leptin, secreted by adipocytes induce systemic inflammation, causing airway inflammation. We previously reported that leptin activates both inflammatory and structural cells, including lung fibroblasts. However, little is known about the differential leptin expression and responsiveness to leptin in asthmatic individuals and healthy controls (HC). In this study, we investigated the expression and origin of leptin in asthmatic airways. We also compared the effect of leptin on asthmatic and HC fibroblasts. MethodsLung specimens from asthmatic and non-asthmatic patients were analyzed by immunohistochemical staining using anti-leptin and anti-CD163 antibodies. Leptin mRNA and protein levels in human monocytes were detected by real-time PCR and western blotting and ELISA, respectively. We used flow cytometry to analyze asthmatic and HC lung fibroblasts for leptin receptor (Ob-R) expression. Further, we determined cytokine levels using cytometric bead array and ELISA and intracellular phosphorylation of specific signaling molecules using western blotting. ResultsAsthma specimens displayed accumulation of leptin-positive inflammatory cells, which were also positive for CD163, a high-affinity scavenger receptor expressed by monocytes and macrophages. Leptin expression was observed at both transcript and protein levels in human blood-derived monocytes. No significant differences were observed between asthmatic and HC lung fibroblasts in Ob-R expression, cytokine production, and intracellular phosphorylation of p38 mitogen-activated protein kinase. ConclusionsOur findings reveal similar responsiveness of control and asthmatic fibroblasts to leptin. However, the accumulation of inflammatory leptin-producing monocytes in the airway may contribute to the pathogenesis of asthma.

Role of FK506-sensitive signals in asthmatic lung inflammation.

Asthma is airway inflammatory diseases caused by the activation of group 2 innate lymphoid cells (ILC2s) and type 2 helper T (TH2) cells. Cysteine proteases allergen cause tissue damage to airway epithelial cells and activate ILC2-mediated type 2 airway inflammation. FK506 is an immunosuppressive agent against calcium-dependent NFAT activation that is also effective against asthmatic inflammation. However, the effects of FK506 on cysteine protease allergen-mediated airway inflammation remain unclear. In this study, we investigated the suppressive effects of FK506 on airway inflammation. FK506 had a partial inhibitory effect on ILC2-dependent eosinophil inflammation and a robust inhibitory effect on T cell-dependent eosinophil inflammation in a cysteine protease-induced mouse asthma model. The infiltration of T1/ST2+ CD4 T cells in the lungs contributed to the persistence of eosinophil infiltration in the airway; FK506 completely inhibited the infiltration of T1/ST2+ CD4 T cells. In the initial phase, FK506 treatment targeted lung ILC2 activation induced by leukotriene B4 (LTB4)-mediated calcium signaling, but not IL-33 signaling. FK506 also inhibited the IL-13-dependent accumulation of T1/ST2+ CD4 T cells in the lungs of the later responses. These results indicated that FK506 potently suppressed airway inflammation by targeting ILC2 activation and T1/ST2+ CD4 T cell accumulation.

Intravenous Mesenchymal Stem Cell Administration Modulates Monocytes/Macrophages and Ameliorates Asthmatic Airway Inflammation in a Murine Asthma Model.

Although asthma is a common chronic airway disease that responds well to anti-inflammatory agents, some patients with asthma are unresponsive to conventional treatment. Mesenchymal stem cells (MSCs) have therapeutic potential for the treatment of inflammatory diseases owing to their immunomodulatory properties. However, the target cells of MSCs are not yet clearly known. This study aimed to determine the effect of human umbilical cord-derived MSCs (hUC-MSCs) on asthmatic lungs by modulating innate immune cells and effector T cells using a murine asthmatic model. Intravenously administered hUC-MSCs reduced airway resistance, mucus production, and inflammation in the murine asthma model. hUC-MSCs attenuated not only T helper (Th) 2 cells and Th17 cells but also augmented regulatory T cells (Tregs). As for innate lymphoid cells (ILC), hUC-MSCs effectively suppressed ILC2s by downregulating master regulators of ILC2s, such as Gata3 and Tcf7. Finally, regarding lung macrophages, hUC-MSCs reduced the total number of macrophages, particularly the proportion of the enhanced monocyte-derived macrophage population. In a closer examination of monocyte-derived macrophages, hUC-MSCs reduced the M2a and M2c populations. In conclusion, hUC-MSCs can be considered as a potential anti- asthmatic treatment given their therapeutic effect on the asthmatic airway inflammation in a murine asthma model by modulating innate immune cells, such as ILC2s, M2a, and M2c macrophages, as well as affecting Tregs and effector T cells.

Oral antibiotics relieve allergic asthma in post-weaning mice via reducing iNKT cells and function of ADRB2

The role of normal gut microbiota in asthma or ovalbumin (OVA)-induced asthma tolerance (OT) remains unclear. Here, we established mouse models of asthma and OT followed by 2 weeks of antibiotic treatment, to clear the gut microbiota. Antibiotic treatment was found to alleviate allergic asthma accompanied with a reduction of invariant natural killer (iNKT) cells. By RNA-seq analysis, we found that β-adrenergic receptor (ADRB) genes, including Adrb1, Adrb2, and Adrb3, were downregulated in asthmatic lungs, but these changes were reversed in OT lungs. Moreover, Adrb2 and Adrb3 were significantly upregulated in asthmatic lungs after antibiotic treatment. Surprisingly, blocking ADRB with propranolol relieved allergic asthma while reducing T helper 2 (Th2) and Treg cell numbers. Further analyses using flow cytometry and immunofluorescence showed that the protein expression level of ADRB2 was higher in asthmatic lungs than that in the control and OT lungs. Notably, dendritic cells (DCs), especially the ADRB2+ DCs, were increased in asthmatic lungs compared to that in the control and OT lungs. In addition, ADRB2+ DCs were significantly reduced following the administration of the ADRB2-specific antagonist ICI118551. Our findings suggest that antibiotic treatment can alleviate OVA-induced allergic asthma via reducing the frequency of iNKT cells and function of ADRB2.