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Mucous cell metaplasia (MCM) and neutrophil-predominant airway inflammation are pathological features of chronic inflammatory airway diseases. A signature feature of MCM is increased expression of a major respiratory tract mucin, MUC5AC. Neutrophil elastase (NE) upregulates MUC5AC in primary airway epithelial cells by generating reactive oxygen species, and this response is due in part to upregulation of NADPH quinone oxidoreductase 1 (NQO1) activity. Delivery of NE directly to the airway triggers inflammation and MCM and increases synthesis and secretion of MUC5AC protein from airway epithelial cells. We hypothesized that NE-induced MCM is mediated in vivo by NQO1. Male wild-type and Nqo1-null mice (C57BL/6 background) were exposed to human NE (50 μg) or vehicle via oropharyngeal aspiration on days 1, 4, and 7. On days 8 and 11, lung tissues and bronchoalveolar lavage (BAL) samples were obtained and evaluated for MCM, inflammation, and oxidative stress. MCM, inflammation, and production of specific cytokines, granulocyte-macrophage colony-stimulating factor, macrophage inflammatory protein-2, interleukin-4, and interleukin-5 were diminished in NE-treated Nqo1-null mice compared with NE-treated wild-type mice. However, in contrast to the role of NQO1 in vitro, we demonstrate that NE-treated Nqo1-null mice had greater levels of BAL and lung tissue lipid carbonyls and greater BAL iron on day 11, all consistent with increased oxidative stress. NQO1 is required for NE-induced inflammation and MCM. This model system demonstrates that NE-induced MCM directly correlates with inflammation, but not with oxidative stress.

Patients with chronic obstructive pulmonary disease (COPD) exhibit dominant features of chronic bronchitis, emphysema, and/or asthma, with a common phenotype of airflow obstruction. COPD pulmonary physiology reflects the sum of pathological changes in COPD, which can occur in large central airways, small peripheral airways, and the lung parenchyma. Quantitative or high-resolution computed tomography is used as a surrogate measure for assessment of disease progression. Different biological or molecular markers have been reported that reflect the mechanistic or pathogenic triad of inflammation, proteases, and oxidants and correspond to the different aspects of COPD histopathology. Similar to the pathogenic triad markers, genetic variations or polymorphisms have also been linked to COPD-associated inflammation, protease-antiprotease imbalance, and oxidative stress. Furthermore, in recent years, there have been reports identifying aging-associated mechanistic markers as downstream consequences of the pathogenic triad in the lungs from COPD patients. For this review, the authors have limited their discussion to a review of mechanistic markers and genetic variations and their association with COPD histopathology and disease status.

BACKGROUND:Although autoimmunity and hyperinflammation secondary to recombination activating gene (RAG) deficiency have been associated with delayed diagnosis and even death, our current understanding is limited primarily to small case series. OBJECTIVE:Understand the frequency, severity, and treatment responsiveness of autoimmunity and hyperinflammation in RAG deficiency. METHODS:In reviewing the literature and our own database, we identified 85 patients with RAG deficiency, reported between 2001 and 2016, and compiled the largest case series to date of 63 patients with prominent autoimmune and/or hyperinflammatory pathology. RESULTS:Diagnosis of RAG deficiency was delayed a median of 5 years from the first clinical signs of immune dysregulation. Most patients (55.6%) presented with more than 1 autoimmune or hyperinflammatory complication, with the most common etiologies being cytopenias (84.1%), granulomas (23.8%), and inflammatory skin disorders (19.0%). Infections, including live viral vaccinations, closely preceded the onset of autoimmunity in 28.6% of cases. Autoimmune cytopenias had early onset (median, 1.9, 2.1, and 2.6 years for autoimmune hemolytic anemia, immune thrombocytopenia, and autoimmune neutropenia, respectively) and were refractory to intravenous immunoglobulin, steroids, and rituximab in most cases (64.7%, 73.7%, and 71.4% for autoimmune hemolytic anemia, immune thrombocytopenia, and autoimmune neutropenia, respectively). Evans syndrome specifically was associated with lack of response to first-line therapy. Treatment-refractory autoimmunity/hyperinflammation prompted hematopoietic stem cell transplantation in 20 patients. CONCLUSIONS:Autoimmunity/hyperinflammation can be a presenting sign of RAG deficiency and should prompt further evaluation. Multilineage cytopenias are often refractory to immunosuppressive treatment and may require hematopoietic cell transplantation for definitive management.

In a mouse model of neutrophil elastase-induced bronchitis that exhibits goblet cell metaplasia and inflammation, we investigated the effects of intratracheal instillation of the MANS peptide, a peptide identical to the NH2 terminus of the myristoylated alanine-rich C kinase substrate (MARCKS) on mucin protein airway secretion, inflammation, and airway reactivity. To induce mucus cell metaplasia in the airways, male BALB/c mice were treated repetitively with the serine protease, neutrophil elastase, on days 1, 4, and 7. On day 11, when goblet cell metaplasia was fully developed and profiles of proinflammatory cytokines were maximal, the animals were exposed to aerosolized methacholine after intratracheal instillation of MANS or a missense control peptide (RNS). MANS, but not RNS, attenuated the methacholine-stimulated secretion of the major respiratory mucin protein, Muc5ac (50% reduction). Concurrently, elastase-induced proinflammatory cytokines typically recovered in bronchoalveolar lavage (BAL), including KC, IL-1β, IL-6, MCP-1, and TNFα, were reduced by the MANS peptide (mean levels decreased 50-60%). Secondary to the effects of MANS on mucin secretion and inflammation, mechanical lung function by forced oscillation technique was characterized with respect to airway reactivity in response to cumulative aerosol stimulation with serotonin. The MANS peptide was also found to effectively attenuate airway hyperresponsiveness to serotonin in this airway hypersecretory model. Collectively, these findings support the concept that even in airway epithelia remodeled with goblet cell metaplasia and in a state of mucin hypersecretion, exogenous attenuation of function of MARCKS protein via the MANS peptide decreases airway mucin secretion, inflammation, and hyperreactivity. [ABSTRACT FROM AUTHOR]

Alpha-1-antitrypsin (A1AT) deficiency is characterized by increased neutrophil elastase (NE) activity and oxidative stress in the lung. We hypothesized that NE exposure generates reactive oxygen species by increasing lung nonheme iron. To test this hypothesis, we measured bronchoalveolar lavage (BAL) iron and ferritin levels, using inductively coupled plasma (ICP) optical emission spectroscopy and an ELISA, respectively, in A1AT-deficient patients and healthy subjects. To confirm the role of NE in regulating lung iron homeostasis, we administered intratracheally NE or control buffer to rats and measured BAL and lung iron and ferritin. Our results demonstrated that A1AT-deficient patients and rats postelastase exposure have elevated levels of iron and ferritin in the BAL. To investigate the mechanism of NE-induced increased iron levels, we exposed normal human airway epithelial cells to either NE or control vehicle in the presence or absence of ferritin, and quantified intracellular iron uptake using calcein fluorescence and ICP mass spectroscopy. We also tested whether NE degraded ferritin in vitro using ELISA and western analysis. We demonstrated in vitro that NE increased intracellular nonheme iron levels and degraded ferritin. Our results suggest that NE digests ferritin increasing the extracellular iron pool available for cellular uptake. [ABSTRACT FROM AUTHOR]

Migration of airway epithelial cells (AEC) is a necessary component of airway mucosal repair after injury. The cytokine IL-1β, present in airway inflammation, has protean effects on constituent cells within the mucosa, but its effects on epithelial repair are not known. We examined migration in differentiated primary human AEC grown in air-liquid interface culture for up to 3 wk and in the 16HBE14o- cell line. Wounds were created by mechanical abrasion and followed to closure using digital microscopy. Concurrent treatment with IL-11.3 (≤10 ng/ml) significantly accelerated migration in primary differentiated cells and in the I6HBE14o- cell line but did not accelerate migration in primary differentiated AEC collected from asthmatic donors. IL-1β treatment did not augment phosphorylation of stress- activated protein kinases normally activated by mechanical injury, such as heat shock protein 27, ERK1/2, and JNK, and did not elicit phosphorylation of signal transducer and activator of transcription-3. However, introduction of a silencing RNA to block expression of the p65 component of NF-κB blocked IL-1β-accelerated migration substantially. Our data demonstrate that IL-1β accelerates migration of normal, but not asthmatic, differentiated AEC by a mechanism that requires activation of the NF-κB signaling complex and suggests a trophic role for this cytokine in airway epithelial repair after injury. [ABSTRACT FROM AUTHOR]

Abstract: Cystic fibrosis is the most common, inherited fatal disease in Caucasians. The major cause of morbidity and mortality is chronic lung disease due to infection and inflammation in the airways leading to bronchiectasis and respiratory failure. The signature pathologic features of CF lung disease including abnormal mucus obstructing airways, chronic infection with Staphylococcus aureus, Pseudomonas aeruginosa and other gram negative bacteria, and a robust neutrophil-dominant airway inflammation, are exacerbated by unopposed proteases present at high concentrations in the ASL. There is strong evidence that proteases, particularly neutrophil elastase, contribute to the pathology of CF by impairing mucociliary clearance, interfering with innate immune functions, and perpetuating neutrophilic inflammation. The mechanisms employed by proteases to impact airway function in CF will be reviewed. [Copyright &y& Elsevier]