Mucous Metaplasia Research Articles

The receptor for advanced glycation end products is a critical mediator of type 2 cytokine signaling in the lungs

Asthma is estimated to effect more than 300 million persons worldwide, leading to nearly 250,000 deaths annually. The majority of patients with mild-to-severe asthma have what is deemed "type-2 high" asthma, which is driven by the prototypical type 2 cytokines IL-4, IL-5, and IL-13. Studies have indicated that the receptor for advanced glycation end products (RAGE) is a critical molecule in the pathogenesis of experimental asthma/allergic airway inflammation. More specifically, RAGE expressed on stromal cells, rather than hematopoietic cells, is critical to induction of asthma/allergic airway inflammation by driving type 2 inflammatory responses. However, the role of RAGE in directly mediating type 2 cytokine signaling has never been investigated. The goal of this study was to test the hypothesis that RAGE mediates type 2 cytokine-induced signal transduction, airway inflammation, and mucus metaplasia in the lungs. Wild-type (WT) and RAGE knockout (RAGE-/-) mice, were intranasally administered rIL-5/rIL-13 or rIL-4 alone, and signal transducer and activator of transcription 6 (STAT6) signaling, airway inflammation, and mucus metaplasia were assessed. ARAGE small-molecule antagonist was used to determine the effects of pharmacologically inhibiting RAGE on type 2 cytokine-induced effects. Administration of type 2 cytokines induced pronounced airway inflammation and mucus metaplasia in WT mice, which was nearly completely abrogated in RAGE-/- mice. In addition, treatment with a RAGE-specific antagonist diminished the effects of type 2 cytokines in WT mice and in primary human bronchial epithelial cell cultures. Genetic ablation or pharmacologic inhibition of RAGE blocks the effects of IL-13 and IL-4 by inhibiting sustained STAT6 activation and downstream target gene expression in mice and in human bronchial epithelial cells. This study is the first to indicate that RAGE is a critical component of type 2 cytokine signal transduction mechanisms, which is a driving force behind type 2-high asthma.

Apolipoprotein E is a concentration-dependent pulmonary danger signal that activates the NLRP3 inflammasome and IL-1β secretion by bronchoalveolar fluid macrophages from asthmatic subjects

House dust mite (HDM)-challenged Apoe-/- mice display enhanced airway hyperreactivity and mucous cell metaplasia. We sought to characterize the pathways that induce apolipoprotein E (APOE) expression by bronchoalveolar lavage fluid (BALF) macrophages from asthmatic subjects and identify how APOE regulates IL-1β secretion. Macrophages were isolated from asthmatic BALF and derived from THP-1cells and human monocytes. HDM-derived cysteine and serine proteases induced APOE secretion from BALF macrophages through protease-activated receptor 2. APOE at concentrations of less than 2.5nmol/L, which are similar to levels found in epithelial lining fluid from healthy adults, did not induce IL-1β release from BALF macrophages. In contrast, APOE at concentrations of 25nmol/L or greater induced nucleotide-binding oligomerization domain, leucine-rich repeat-containing protein (NLRP) 3 and pro-IL-1β expression by BALF macrophages, as well as the caspase-1-mediated generation of mature IL-1β secreted from cells. HDM acted synergistically with APOE to both prime and activate the NLRP3 inflammasome. In a murine model of neutrophilic airway inflammation induced by HDM and polyinosinic-polycytidylic acid, APOE reached a concentration of 32nmol/L in epithelial lining fluid, with associated increases in BALF IL-1β levels. APOE-dependent NLRP3 inflammasome activation in macrophages was primarily mediated through a potassium efflux-dependent mechanism. APOE can function as an endogenous, concentration-dependent pulmonary danger signal that primes and activates the NLPR3 inflammasome in BALF macrophages from asthmatic subjects to secrete IL-1β. This might represent a mechanism through which APOE amplifies pulmonary inflammatory responses when concentrations in the lung are increased to greater than normal levels, which can occur during viral exacerbations of HDM-induced asthma characterized by neutrophilic airway inflammation.

Loss of the adhesion G-protein coupled receptor ADGRF5 in mice induces airway inflammation and the expression of CCL2 in lung endothelial cells

BackgroundAdhesion G-protein coupled receptor F5 (ADGRF5) was recently identified as an essential regulator of pulmonary surfactant homeostasis in alveolar type II cells. We previously showed that in addition to abnormal surfactant accumulation, Adgrf5-deficient (Adgrf5−/−) mice exhibit emphysema-like signs, suggesting a possible role for ADGRF5 in immune regulation. Here, we extended the phenotypic analysis of Adgrf5−/− mice to help understand its biological role in the lung, and especially in immune regulation.MethodsHistological features of lungs were evaluated by Alcian blue and Masson’s trichrome staining. Quantitative real-time PCR (qPCR) and western blot analyses were performed to analyze the differential expression of genes/proteins related to airway inflammation in lungs between wildtype and Adgrf5−/− mice. Acid–base status was assessed by performing blood gas tests and urine pH measurements. Inflammatory cell counting was performed using Giemsa-stained bronchoalveolar lavage cells. Serum IgE concentrations were determined by enzyme-linked immunosorbent assay. The expression of Ccl2, S100a8, S100a9, and Saa3 in primary lung endothelial cells (ECs) was determined by qPCR and/or western blotting. Finally, the effect of administrating RS504393 to 2-week-old Adgrf5−/− mice on gene expression in the lungs was analyzed by qPCR.ResultsAdgrf5−/− mice exhibited several features of chronic airway inflammation (mucous cell metaplasia, mucus hyperproduction, subepithelial fibrosis, respiratory acidosis, high serum IgE, mast cell accumulation, and neutrophilia) in parallel with elevated expression of genes involved in mucous cell metaplasia (Muc5ac, Muc5b, Slc26a4, and Clca1), fibrosis (Tgfb1, Col1a1, Fn1, and Tnc), and type 2 immune response (Il4, Il5, Il13, IL-25, and IL-33) at 12 and/or 30 weeks of age. In contrast, mRNA expression of Ccl2, S100a8, and S100a9 was upregulated in embryonic or neonatal Adgrf5−/− lungs as well as in lung ECs of Adgrf5−/− mice at 1 week of age. RS504393 treatment suppressed the upregulation of S100a8, S100a9, Slc26a4, and Il5 in Adgrf5−/− lungs.ConclusionsTargeted disruption of ADGRF5 results in the development of airway inflammation, which is likely mediated by the type 2 immune response and possibly CCL2-mediated inflammation. ADGRF5 also has a potential role in the regulation of genes encoding CCL2 in lung ECs, thereby maintaining immune homeostasis.

Radiation-Induced Rhinitis: Cytological and Olfactory Changes

Oral mucositis is a well-known adverse event of radiotherapy (RT) for head and neck cancer (HNC). Its nasal counterpart, the radiation-induced rhinitis, is poorly studied and considered in clinical practice. The aim of this observational study was to evaluate acute cytological and olfactory alterations during RT and their correlation with RT doses. Ten patients who underwent RT for HNC, excluding tumors of the nasal cavities, were evaluated with nasal scraping for cytological examination, Sniffin' Sticks test for olfactory assessment, and Nasal Obstruction Symptom Evaluation scale. The examinations were performed before (T0), at mid-course (T1), and at the end (T2) of RT. They were repeated 1 and 3 months after RT (T3 and T4). Mean dose (Dmean) and near maximum dose (D2%) to nasal cavities and inferior turbinates were used for correlation analyses. Radiation-induced rhinitis was present in 70% of patients at T2, and it was still observed in 40% of cases after 3 months. Although olfactory function remained within the normal range at the evaluated times, a significant decrease in odor threshold and discrimination was observed during RT, which returned to baseline levels after RT. Nasal cytology showed a radiation-induced rhinitis with neutrophils and sometimes bacteria. Mucous and squamous cell metaplasia appeared in 10% of patients. Dmean and D2% to inferior turbinates were associated to neutrophilic rhinitis at T2, and D2% to inferior turbinates was correlated to mucous cell metaplasia at T2. RT for HNC induces acute rhinitis that may persist after the completion of treatment and can affect patient's quality of life. Nasal cytology can help to choose the best treatment on an individual basis.

Genes and Pathways Regulating Decline in Lung Function and Airway Remodeling in Asthma.

Asthma is a common disorder of the airways characterized by airway inflammation and by decline in lung function and airway remodeling in a subset of asthmatics. Airway remodeling is characterized by structural changes which include airway smooth muscle hypertrophy/hyperplasia, subepithelial fibrosis due to thickening of the reticular basement membrane, mucus metaplasia of the epithelium, and angiogenesis. Epidemiologic studies suggest that both genetic and environmental factors may contribute to decline in lung function and airway remodeling in a subset of asthmatics. Environmental factors include respiratory viral infection-triggered asthma exacerbations, and tobacco smoke. There is also evidence that several asthma candidate genes may contribute to decline in lung function, including ADAM33, PLAUR, VEGF, IL13, CHI3L1, TSLP, GSDMB, TGFB1, POSTN, ESR1 and ARG2. In addition, mediators or cytokines, including cysteinyl leukotrienes, matrix metallopeptidase-9, interleukin-33 and eosinophil expression of transforming growth factor-β, may contribute to airway remodeling in asthma. Although increased airway smooth muscle is associated with reduced lung function (i.e. forced expiratory volume in 1 second) in asthma, there have been few long-term studies to determine how individual pathologic features of airway remodeling contribute to decline in lung function in asthma. Clinical studies with inhibitors of individual gene products, cytokines or mediators are needed in asthmatic patients to identify their individual role in decline in lung function and/or airway remodeling.

RAGE-dependent VCAM-1 expression in the lung endothelium mediates IL-33-induced allergic airway inflammation.

The receptor for advanced glycation endproducts (RAGE) has been implicated as a critical molecule in the pathogenesis of experimental asthma/allergic airway inflammation (AAI). It has been previously shown that RAGE acts both upstream of interleukin-33 (IL-33) release and downstream of IL-33 release via RAGE-dependent IL-33-induced accumulation of type 2 innate lymphoid cells (ILC2s) in the lungs, which perpetuate type 2 inflammation and mucus metaplasia. However, the mechanism by which RAGE mediates downstream IL-33-induced type 2 inflammatory responses is unknown. This study tested the hypothesis that ILC2s are recruited to the lungs via RAGE-dependent vascular cell adhesion molecule 1 (VCAM-1) expression on lung endothelial cells. House dust mite extract, Alternaria alternata extract, or rIL-33 was used to induce AAI/VCAM-1 expression in wild-type (WT) and RAGE-knockout (RAGE-KO) mice. Intravenous (i.v.) anti-VCAM-1 or intraperitoneal (i.p.) β7 blocking antibody administration was used to determine the role of VCAM-1 in IL-33-induced AAI. Enhanced VCAM-1 expression in the lungs by HDM, AA, or rIL-33 exposure was found to be RAGE-dependent. In addition, stimulation of primary mouse lung endothelial cells with IL-33 induced VCAM-1 expression in WT, but not RAGE-KO cells. Administration of VCAM-1 and β7-integrin blocking antibodies reduced IL-33-induced eosinophilic inflammation, mucus metaplasia, and type 2 inflammatory responses. This study demonstrates that allergen- and cytokine-induced VCAM-1 expression is RAGE-dependent and contributes to lung ILC2 accumulation and downstream eosinophilic inflammation, mucus metaplasia, and type 2 inflammatory responses.

MyD88 controls airway epithelial Muc5ac expression during TLR activation conditions from agricultural organic dust exposure.

Inflammation from airborne microbes can overwhelm compensatory mucociliary clearance mechanisms, leading to mucous cell metaplasia. Toll-like receptor (TLR) activation via myeloid differentiation factor 88 (MyD88) signaling is central to pathogen responses. We have previously shown that agricultural organic dust extract (ODE), with abundant microbial component diversity, activates TLR-induced airway inflammation. With the use of an established model, C57BL/6J wild-type (WT) and global MyD88 knockout (KO) mice were treated with intranasal inhalation of ODE or saline, daily for 1 wk. ODE primarily increased mucin (Muc)5ac levels relative to Muc5b. Compared with ODE-challenged WT mice, ODE-challenged, MyD88-deficient mice demonstrated significantly increased Muc5ac immunostaining, protein levels by immunoblot, and expression by quantitative PCR. The enhanced Muc5ac levels in MyD88-deficient mice were not explained by differences in the differentiation program of airway secretory cells in naïve mice. Increased Muc5ac levels in MyD88-deficient mice were also not explained by augmented inflammation, IL-17A, or neutrophil elastase levels. Furthermore, the enhanced airway mucins in the MyD88-deficient mice were not due to defective secretion, as the mucin secretory capacity of MyD88-KO mice remained intact. Finally, ODE-induced Muc5ac levels were enhanced in MyD88-deficient airway epithelial cells in vitro. In conclusion, MyD88 deficiency enhances airway mucous cell metaplasia under environments with high TLR activation.

Inflammation-induced upregulation of P2X4 expression augments mucin secretion in airway epithelia.

Mucus clearance provides an essential innate defense mechanism to keep the airways and lungs free of particles and pathogens. Baseline and stimulated mucin secretion from secretory airway epithelial cells need to be tightly regulated to prevent mucus hypersecretion and mucus plugging of the airways. It is well established that extracellular ATP is a potent stimulus for regulated mucus secretion. Previous studies revealed that ATP acts via metabotropic P2Y2 purinoreceptors on goblet cells. Extracellular ATP, however, is also a potent agonist for ionotropic P2X purinoreceptors. Expression of several P2X isoforms has been reported in airways, but cell type-specific expression and the function thereof remained elusive. With this study, we now provide evidence that P2X4 is the predominant P2X isoform expressed in secretory airway epithelial cells. After IL-13 treatment of either human primary tracheal epithelial cells or mice, P2X4 expression is upregulated in vitro and in vivo under conditions of chronic inflammation, mucous metaplasia, and hyperplasia. Upregulation of P2X4 is strongest in MUC5AC-positive goblet cells. Moreover, activation of P2X4 by extracellular ATP augments intracellular Ca2+ signals and mucin secretion, whereas Ca2+ signals and mucin secretion are dampened by inhibition of P2X4 receptors. These data provide new insights into the purinergic regulation of mucin secretion and add to the emerging picture that P2X receptors modulate exocytosis of large secretory organelles and secretion of macromolecular vesicle cargo.

Abstract 5731: Synergistic effect of cigarette smoke and bacterial-induced chronic obstructive pulmonary disease type airway inflammation on promotion of K-ras mutant lung cancer

Abstract Lung cancer, particularly K-ras mutant lung cancer, is the leading cause of cancer death worldwide, and cigarette smoking (CS) is its main cause. Epidemiologic studies had consistently revealed a strong association (3 to 9 fold increase) between lung cancer and COPD (chronic obstructive pulmonary disease), after controlling for CS exposure. COPD is an inflammatory disease of the airways with smoking being the main cause of it. Importantly, lung inflammation persists and lung function continues to deteriorate as does the increased risk of lung cancer even after cessation of cigarette smoking among former smokers with COPD. These facts suggest a strong link between COPD-related airway inflammation and lung cancer promotion independent of smoking. We have previously shown that weekly exposure to an aerosolized bacterial lysate of nontypeable Haemophilus influenzae (NTHi), induces lung inflammation with a profile of mediators and inflammatory cells similar to that observed in COPD patients excluding mucous metaplasia. We further showed that NTHi-induced COPD type airway inflammation promotes lung cancer 3.2-fold in a K-ras mutant mouse model remarkably similar to that in the epidemiologic literature. NTHi is the most common colonizing bacteria in the lower respiratory tract of patients with COPD and could be a potential cause of perpetuating and promoting persistent airway inflammation after CS exposure in these patients. Therefore, we further studied the effect of combined CS and NTHi exposure in the induction of COPD phenotype and promotion of lung cancer. Briefly, 6-week old K-ras mutant mice were exposed to NTHi lysate once a week for 8 weeks and to cigarette smoke (CS) daily for 2 hours/day, 5 days/week for 8 weeks and studied at the age of 14 weeks. The CS exposure was conducted by burning 3R4F reference cigarettes (University of Kentucky, Tobacco Research Institute), using an InExpose System (SCIREQ Scientific Respiratory Equipment Inc). We found that CS exposure alone caused a mild macrophage dominant airway inflammation, induced airway epithelial mucous metaplasia, and led to a 2.3 fold increase in lung tumor burden. Notably, combined NTHi and CS exposure resulted in a robust neutrophilic lung inflammation and mucous metaplasia and promoted K-ras mutant lung cancer by 4.3 folds (2 times more than CS alone). Our experimental results suggest that CS exposure and colonization of smoke-injured airways with NTHi induce an inflammatory and structural COPD phenotype fully recapitulating human COPD and provide a microenvironment that has a significant promoting effect on K-ras mutant lung cancer. Citation Format: Marco A. Ramos, Misha Umer, Susana Castro, Berenice Adriana Gutierrez, Nassim Khosravi, Seyed Javad Moghaddam. Synergistic effect of cigarette smoke and bacterial-induced chronic obstructive pulmonary disease type airway inflammation on promotion of K-ras mutant lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5731.

A Disintegrin and A Metalloproteinase-9 (ADAM9): A Novel Proteinase Culprit with Multifarious Contributions to COPD.

Proteinases with a disintegrin and a metalloproteinase domain (ADAMs) have not been well studied in COPD. We investigated whether ADAM9 is linked to COPD in humans and mice. ADAM9 blood and lung levels were measured in COPD patients versus controls, and air- versus cigarette smoke (CS)-exposed wild-type (WT) mice. WT and Adam9-/- mice were exposed to air or CS for 1-6 months, and COPD-like lung pathologies were measured. ADAM9 staining was increased in lung epithelial cells and macrophages in smokers and even more so in COPD patients and correlated directly with pack-year smoking history and inversely with airflow obstruction and/or FEV1 % predicted. Bronchial epithelial cell ADAM9 mRNA levels were higher in COPD patients than controls and correlated directly with pack-year smoking history. Plasma, BALF and sputum ADAM9 levels were similar in COPD patients and controls. CS exposure increased Adam9 levels in WT murine lungs. Adam9-/- mice were protected from emphysema development, small airway fibrosis, and airway mucus metaplasia. CS-exposed Adam9-/- mice had reduced lung macrophage counts, alveolar septal cell apoptosis, lung elastin degradation, and shedding of VEGFR2 and EGFR in BALF samples. Recombinant ADAM9 sheds EGF and VEGF receptors from epithelial cells to reduce activation of the Akt pro-survival pathway and increase cellular apoptosis. ADAM9 levels are increased in COPD lungs and linked to key clinical variables. Adam9 promotes emphysema development, and large and small airway disease in mice. Inhibition of ADAM9 could be a therapeutic approach for multiple COPD phenotypes.

Helicobacter pylori-infected C57BL/6 mice with different gastrointestinal microbiota have contrasting gastric pathology, microbial and host immune responses

C57BL/6 (B6) mice from Taconic Sciences (Tac) and the Jackson Laboratory (Jax) were infected with H. pylori PMSS1 (Hp) for 16 week; there was no significant difference in the gastric histologic activity index between Hp infected Tac and Jax B6. However, the degree of gastric mucous metaplasia and Th1-associated IgG2c levels in response to Hp infection were increased in Tac mice over Jax mice, whereas the colonization levels of gastric Hp were higher by 8-fold in Jax B6 compared with Tac B6. Additionally, mRNA expression of gastric Il-1β, Il-17A and RegIIIγ were significantly lower in the infected Tac compared to the infected Jax mice. There were significant differences in the microbial community structures in stomach, colon, and feces between Jax and Tac B6 females. Differences in gastric microbial communities between Jax and Tac B6 females are predicted to affect the metagenome. Moreover, Hp infection perturbed the microbial community structures in the stomach, colon and feces of Jax mice, but only altered the colonic microbial composition of Tac mice. Our data indicate that the GI microbiome of Tac B6 mice is compositionally distinct from Jax B6 mice, which likely resulted in different pathological, immunological, and microbial responses to Hp infection.

Toll-like receptor signalling protects against IL-33 led virus-induced asthmatic response in a Type I Interferon independent manner

Abstract Respiratory viruses are frequent causes of asthma development and exacerbations. However, there are gaps in understanding how viral infections cause immuno-pathology in the host leading to asthma progression. We have previously shown that mice treated with a combination of Toll-like receptor (TLR) 2/6 and 9 agonists (Pam2-ODN) are protected against broad range of respiratory infections induced by viruses, fungi and bacteria. Pam2-ODN treatment one day prior to virus infection leads to enhanced survival with reduced lung viral burden in mice. Further, Pam2-ODN pretreatment also inhibits acute lung pathology caused by CD8+ T cells and prevents chronic asthmatic phenotype of eosinophilia, airway hyperreactivity and mucus metaplasia. This protective effect was accompanied by reduced virus-driven lung IL-33, which is necessary to induce chronic asthmatic phenotype. Interestingly, Pam2-ODN does not induce increased production of Type I or III Interferons (IFN), nor does the protective response require intact interferon signaling. Transcriptome profiling of the mouse lungs in the chronic phase of infection led to identification of genes involved in airway remodelling, epithelial cell differentiation and chemokine signalling. More importantly, we identified genes from the Pam2-ODN treatment group that are involved in reversing the viral infection effect on day 49. Taken together, our findings reveal novel type I IFN independent mechanisms to prevent virus-induced asthma, and may provide an opportunity to protect vulnerable populations.

Ablation of adaptive immune responses facilitates increased bacterial clearance in murine model of defective mucociliary clearance

Abstract Defective mucociliary clearance (MCC) associated with muco-obstructive lung diseases may compromise effective clearance of aspirated bacteria. Mucus hypersecretion is often associated with mucous cell metaplasia (MCM) of airway epithelium, which is a Type 2 immune response. Whether ablation of Type 2 response abolishes MCM/muco-obstruction, and thus, enhances bacterial clearance remains untested. Accordingly, we hypothesized that the ablation of adaptive immune responses will promote bacterial clearance in murine model of defective MCC, i.e., Scnn1b [Sodium channel, non-voltage-gated 1, beta] transgenic (Tg+) mice. The Tg+ model recapitulates hallmarks of muco-obstructive airways, including MCM and bacterial infections. The Type 2 response deficient (Il2rg−/−, Rag1−/− and Il2rg −/−\Rag1−/−) mice strains on Tg+ background were observed at neonatal age (6–7 days) for bacterial clearance response. Our data suggest that while Il2rg−/− mice [lacking innate lymphoid cells (ILC)] had significantly impaired bacterial clearance in control (Tg−) as well as Tg+, the Rag1−/−mice [lacking adaptive immune response] did not exhibit impaired bacterial clearance in Tg− as well as Tg+ neonates. The defective bacterial clearance in Il2rg−/− was not due to reduced phagocyte recruitment. Interestingly, Il2rg −/−\Rag1−/− mice had significantly improved bacterial clearance, an observation that remains a focus of our ongoing investigation. Our analyses revealed absence of IL-10 in Rag1−/−mice was associated with morphological activation of macrophages. Taken together, our current data suggest that while ILCs are essential for bacterial clearance, the adaptive immune responses suppress macrophage’s antibacterial function.

Pre-existing atopy protects mice from paramyxoviral respiratory infection weight loss

Abstract Severe paramyxoviral infections early in life have been associated with development of asthma and allergies; respiratory viral infections are known to exacerbate asthma. We have a mouse model of a severe paramyxviral respiratory infection using murine parainfluenza virus 1, Sendai virus (SeV). C57BL6 mice inoculated with SeV develop a severe acute infection demonstrated by significant weight loss over 8–10 days. The mice recover and are left with long-lasting airway hyper-reactivity and mucous cell metaplasia. Exposure to a non-viral antigen during the acute infection leads to IgE production against the non-viral antigen (allergic sensitization). While this model provides an explanation for development of allergies and asthma, it is not known whether being atopic before viral infection alters the disease state. To address this, we intranasally sensitized and challenged C57BL6 mice with house dust mite extract (HDM) or PBS, and one week later inoculated the mice with 2×105 pfu SeV or ultraviolet light inactivated SeV (UV-SeV). All UV-SeV groups lost no weight. SeV infection in PBS sensitized/challenged (PBS/PBS/SeV) mice led to a 10.8+/−2.5% weight loss from baseline (mean+/−SEM, n=4); surprisingly, HDM sensitized and challenged mice lost no weight with SeV infection (−1.7+/−0.6% weight loss, n=4, p=0.0022 versus PBS/PBS/SeV). Mice HDM sensitized but not challenged loss 10.8+/−1.6% weight with SeV infection (n=4, p>0.99 versus PBS/PBS/SeV). Whether the weight loss protection from HDM sensitization/challenge alters the antiviral immune response and/or protects against post-viral atopic disease is being investigated. These data suggest atopy may protect against symptoms of a severe paramyxoviral respiratory infection.

Targeting MARCKS phosphorylation site domain shows therapeutic potential for allergic asthma

Abstract Prevalence of asthma is increased all over the world, especially in Taiwan, study analyzed data of 997,729 enrolees from the National Health Insurance register from 2000 to 2007, the prevalence of asthma increase from 2.9% to 11.9%. Various signaling pathways and mechanisms have been identified and targeted for therapeutic application. Among them, one treatment, based on the use of MANS peptide, which is corresponding to the myristoylated N-terminal 24-amino acid portion of the MARCKS (myristoylated alanine-rich C kinase substrates) protein, to compete MARCKS’ binding to membrane has produced useful results. There is a lack of information regarding to the level/activity of phosphorylated MARCKS (p-MARCKS) in allergic asthma. It is also unclear if this activity can be directly suppressed for the allergic asthma treatment application. In this study, we have found elevated p-MARCKS in most of asthmatic tissues from human as well as from the animal model. This elevation can be directly suppressed by a peptide, MPS, targeting MARCKS phosphorylation site domain, prior to allergen sensitization. In addition, MPS pre-treatment also suppressed all of the symptoms associated with allergic airway asthma, which includes a reduction of inflammatory cells influx and TH2 cytokine presence in the lumen, and also a reduction of airway mucous cell metaplasia and airway hyperreactivity. In addition to inhibition of TH2 responses, lung dendritic cells migrate to mediastinal lymph nodes also decreased by MPS treatment. These results suggest that MARCKS phosphorylation site domain is a potential therapeutic target for the treatment of allergic asthma.

Pyloric metaplasia, pseudopyloric metaplasia, ulcer-associated cell lineage and spasmolytic polypeptide-expressing metaplasia: reparative lineages in the gastrointestinal mucosa.

The gastrointestinal mucosae provide a critical barrier between the external and internal milieu. Thus, damage to the mucosa requires an immediate response to provide appropriate wound closure and healing. Metaplastic lineages with phenotypes similar to the mucous glands of the distal stomach or Brunner's glands have been associated with various injurious scenarios in the stomach, small bowel, and colon. These lineages have been assigned various names including pyloric metaplasia, pseudopyloric metaplasia, ulcer-associated cell lineage (UACL), and spasmolytic polypeptide-expressing metaplasia (SPEM). A re-examination of the literature on these various forms of mucous cell metaplasia suggests that pyloric-type mucosal gland lineages may provide a ubiquitous response to mucosal injury throughout the gastrointestinal tract as well as in the pancreas, esophagus, and other mucosal surfaces. While the cellular origin of these putative reparative lineages likely varies in different regions of the gut, their final phenotypes may converge on a pyloric-type gland dedicated to mucous secretion. In addition to their healing properties in the setting of acute injury, these pyloric-type lineages may also represent precursors to neoplastic transitions in the face of chronic inflammatory influences. Further investigations are needed to determine how discrete molecular profiles relate to the origin and function of pyloric-type metaplasias previously described by histological characteristics in multiple epithelial mucosal systems in the setting of acute and chronic damage. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

A Novel Orally Available Asthma Drug Candidate That Reduces Smooth Muscle Constriction and Inflammation by Targeting GABAA Receptors in the Lung.

We describe lead compound MIDD0301 for the oral treatment of asthma based on previously developed positive allosteric α5β3γ2 selective GABAA receptor (GABAAR) ligands. MIDD0301 relaxed airway smooth muscle at single micromolar concentrations as demonstrated with ex vivo guinea pig tracheal rings. MIDD0301 also attenuated airway hyperresponsiveness (AHR) in an ovalbumin murine model of asthma by oral administration. Reduced numbers of eosinophils and macrophages were observed in mouse bronchoalveolar lavage fluid without changing mucous metaplasia. Importantly, lung cytokine expression of IL-17A, IL-4, and TNF-α were reduced for MIDD0301-treated mice without changing antiinflammatory cytokine IL-10 levels. Automated patch clamp confirmed amplification of GABA induced current mediated by α1-3,5β3γ2 GABAARs in the presence of MIDD0301. Pharmacodynamically, transmembrane currents of ex vivo CD4+ T cells from asthmatic mice were potentiated by MIDD0301 in the presence of GABA. The number of CD4+ T cells observed in the lung of MIDD0301-treated mice were reduced by an oral treatment of 20 mg/kg b.i.d. for 5 days. A half-life of almost 14 h was demonstrated by pharmacokinetic studies (PK) with no adverse CNS effects when treated mice were subjected to sensorimotor studies using the rotarod. PK studies also confirmed very low brain distribution. In conclusion, MIDD0301 represents a safe and improved oral asthma drug candidate that relaxes airway smooth muscle and attenuates inflammation in the lung leading to a reduction of AHR at a dosage lower than earlier reported GABAAR ligands.

Injury, repair, inflammation and metaplasia in the stomach

The development of intestinal-type gastric cancer is preceded by the emergence of metaplastic cell lineages in the gastric mucosa. In particular, intestinal metaplasia and spasmolytic polypeptide-expressing metaplasia (SPEM) have been associated with the pathological progression to intestinal-type gastric cancer. The development of SPEM represents a physiological response to damage that recruits reparative cells to sites of mucosal injury. Metaplastic cell lineages are characterized by mucus secretion, adding a protective barrier to the epithelium. Increasing evidence indicates that the influence of alarmins and cytokines is required to initiate the process of metaplasia development. In particular, IL-33 derived from epithelial cells stimulates IL-13 production by specialized innate immune cells to induce chief cell transdifferentiation into SPEM following the loss of parietal cells from the corpus of the stomach. While SPEM represents a physiological healing response to acute injury, persistent injury and chronic inflammation can perpetuate a recurring pattern of reprogramming and metaplasia that is a risk factor for gastric cancer development. The transdifferentiation of zymogen secreting cells into mucous cell metaplasia may represent both a general repair mechanism in response to mucosal injury in many epithelia as well as a common pre-neoplastic pathway associated with chronic injury and inflammation.

Acellular filtrate of a microbial-based cleaning product potentiates house dust mite allergic lung inflammation

Microbial-based cleaning products (MBCPs) contain bacteria and chemical constituents. They are used in consumer applications such as odor reduction, unclogging drains, and surface cleaning. To determine the capacity of a model MBCP to contribute to acute allergic lung inflammation, a two-week repeated exposure regimen was used. Mice were exposed by endotracheal instillation to saline alone, MBCP alone, house dust mites (HDM) alone, or sequentially (i.e., MBCP followed by HDM, HDM followed by MBCP, or HDM + MBCP followed by HDM). Both whole MBCP and acellular MBCP filtrate were investigated, and showed minimal differences in the endpoints examined. HDM exposure caused pulmonary perivascular inflammation, bronchiolar mucous cell metaplasia, elevated bronchoalveolar lavage fluid (BALF) eosinophils, and HDM-specific IgG1. For MBCP, notable changes were associated with sequential exposures. MBCP/HDM caused elevated TH2 cytokines in BALF, and elevated neutrophils, eosinophils and IL-5 in peripheral blood. Co-administration of MBCP and HDM followed by HDM resulted in elevated blood and BALF eosinophils and HDM-specific IgE and IgG1. These results demonstrated that acellular MBCP filtrate, and not bacteria within MBCPs, potentiated the acute allergic inflammation to HDM. This methodology could be extended to investigate chronic allergic inflammation and inflammatory potential of other MBCPs and biotechnology products with complex compositions.

Noxa/HSP27 complex delays degradation of ubiquitylated IkBα in airway epithelial cells to reduce pulmonary inflammation.

IFN-γ is known as a pro-inflammatory cytokine, but can also block inflammation in certain chronic diseases although the underlying mechanisms are poorly understood. We found that IFN-γ rapidly induced Noxa expression and that extent of inflammation by repeated house dust mite exposure was enhanced in noxa-/- compared with noxa+/+ mice. Noxa expression blocked TNF-α-induced nuclear translocation of NF-κB and the production of pro-inflammatory cytokines. Noxa did not affect TNF-α-induced IκBα phosphorylation but the degradation of 48-chain-ubiquitylated IκBα. The Cys25 of Noxa was cross-linked with Cys137 of phospho-HSP27 and both proteins were required for blocking the degradation of ubiquitylated IκBα. Because, phospho-HSP27 is present in airway epithelial cells and not in fibroblasts or thymocytes, we generated transgenic mice that inducibly expressed Noxa in airway epithelia. These mice showed protection from allergen-induced inflammation and mucous cell metaplasia by blocking nuclear translocation of NF-κB. Further, we identified a Noxa-derived peptide that prolonged degradation of 48-chain-ubiquitylated IκBα blocked nuclear translocation of NF-κB, and reduced allergen-induced inflammation in mice. These results suggest that the anti-inflammatory role of the Noxa protein may be restricted to airway epithelial cells and the use of Noxa for therapy of chronic lung diseases may be associated with reduced side effects.