Bruce K. Rubin

Mucins, Mucus, and Sputum*

Normal airway mucus lines the epithelial surface and provides an important innate immune function by detoxifying noxious molecules and by trapping and removing pathogens and particulates from the airway via mucociliary clearance. The major macromolecular constituents of normal mucus, the mucin glycoproteins, are large, heavily glycosylated proteins with a defining feature of tandemly repeating sequences of amino acids rich in serine and threonine, the linkage sites for large carbohydrate structures. The mucins are composed of two major families: secreted mucins and membrane-associated mucins. Membrane-associated mucins have been reported to function as cell surface receptors for pathogens and to activate intracellular signaling pathways. The biochemical and cellular functions for secreted mucin glycoproteins have not been definitively assigned. In contrast to normal mucus, sputum production is the hallmark of chronic inflammatory airway diseases such as asthma, chronic bronchitis, and cystic fibrosis (CF). Sputum has altered macromolecular composition and biophysical properties which vary with disease, but unifying features are failure of mucociliary clearance, resulting in airway obstruction, and failure of innate immune properties. Mucin glycoprotein overproduction and hypersecretion are common features of chronic inflammatory airway disease, and this has been the underlying rationale to investigate the mechanisms of mucin gene regulation and mucin secretion. However, in some pathologic conditions such as CF, airway sputum contains little intact mucin and has increased content of several macromolecules including DNA, filamentous actin, lipids, and proteoglycans. This review will highlight the most recent insights on mucus biology in health and disease.

Mechanisms of Action and Clinical Application of Macrolides as Immunomodulatory Medications

SUMMARY Macrolides have diverse biological activities and an ability to modulate inflammation and immunity in eukaryotes without affecting homeostatic immunity. These properties have led to their long-term use in treating neutrophil-dominated inflammation in diffuse panbronchiolitis, bronchiectasis, rhinosinusitis, and cystic fibrosis. These immunomodulatory activities appear to be polymodal, but evidence suggests that many of these effects are due to inhibition of extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation and nuclear factor kappa B (NF-κB) activation. Macrolides accumulate within cells, suggesting that they may associate with receptors or carriers responsible for the regulation of cell cycle and immunity. A concern is that long-term use of macrolides increases the emergence of antimicrobial resistance. Nonantimicrobial macrolides are now in development as potential immunomodulatory therapies.

Macrolide antibiotics as immunomodulatory medications: proposed mechanisms of action.

Macrolide antibiotics administered in sub-antimicrobial doses improve pulmonary function and decrease exacerbation frequency for persons with diffuse panbronchiolitis or cystic fibrosis. Data also suggest a beneficial effect of macrolide antibiotics in the treatment of steroid dependent asthma. Many potential immunomodulatory effects of macrolide antibiotics have been reported including the ability to down-regulate prolonged inflammation, decreasing airway mucus secretion, inhibiting bacterial biofilm, decreasing the production of reactive oxygen species, inhibiting neutrophil activation and mobilization, accelerating neutrophil apoptosis, and blocking the activation of nuclear transcription factors. Macrolides initially decrease, then increase, and have finally a sustained suppression of cytokine secretions from normal human bronchial epithelial cells through inhibition and activation of extracellular signal-regulated kinases (ERK) and then reversibly retard cell proliferation probably through ERK. Consistent with this, macrolide antibiotics possibly reduce mucin production as well as neutrophil migration by interfering with ERK signal transduction.

Familial pulmonary alveolar proteinosis caused by mutations in CSF2RA

Primary pulmonary alveolar proteinosis (PAP) is a rare syndrome characterized by accumulation of surfactant in the lungs that is presumed to be mediated by disruption of granulocyte/macrophage colony-stimulating factor (GM-CSF) signaling based on studies in genetically modified mice. The effects of GM-CSF are mediated by heterologous receptors composed of GM-CSF binding (GM-CSF-Rα) and nonbinding affinity-enhancing (GM-CSF-Rβ) subunits. We describe PAP, failure to thrive, and increased GM-CSF levels in two sisters aged 6 and 8 yr with abnormalities of both GM-CSF-Rα–encoding alleles (CSF2RA). One was a 1.6-Mb deletion in the pseudoautosomal region of one maternal X chromosome encompassing CSF2RA. The other, a point mutation in the paternal X chromosome allele encoding a G174R substitution, altered an N-linked glycosylation site within the cytokine binding domain and glycosylation of GM-CSF-Rα, severely reducing GM-CSF binding, receptor signaling, and GM-CSF–dependent functions in primary myeloid cells. Transfection of cloned cDNAs faithfully reproduced the signaling defect at physiological GM-CSF concentrations. Interestingly, at high GM-CSF concentrations similar to those observed in the index patient, signaling was partially rescued, thereby providing a molecular explanation for the slow progression of disease in these children. These results establish that GM-CSF signaling is critical for surfactant homeostasis in humans and demonstrate that mutations in CSF2RA cause familial PAP.

Inhaled mannitol for the treatment of mucociliary dysfunction in patients with bronchiectasis: Effect on lung function, health status and sputum

Objective:  Inhaled mannitol increases mucus clearance in patients with bronchiectasis by an unclear mechanism. The effect of mannitol on lung function, health status and sputum properties was investigated.

Alternative mechanisms for tiotropium

Tiotropium is commonly used in the treatment of chronic obstructive pulmonary disease. Although largely considered to be a long-acting bronchodilator, its demonstrated efficacy in reducing the frequency of exacerbations and preliminary evidence from early studies indicating that it might slow the rate of decline in lung function suggested mechanisms of action in addition to simple bronchodilation. This hypothesis was examined in the recently published UPLIFT study and, although spirometric and other clinical benefits of tiotropium treatment extended to four years, the rate of decline in lung function did not appear to be reduced by the addition of tiotropium in this study. This article summarizes data from a variety of investigations that provide insights into possible mechanisms to account for the effects of tiotropium. The report summarizes the discussion on basic and clinical research in this field.

Hereditary Pulmonary Alveolar Proteinosis: Pathogenesis, Presentation, Diagnosis, and Therapy

RATIONALE We identified a 6-year-old girl with pulmonary alveolar proteinosis (PAP), impaired granulocyte-macrophage colony-stimulating factor (GM-CSF) receptor function, and increased GM-CSF. OBJECTIVES Increased serum GM-CSF may be useful to identify individuals with PAP caused by GM-CSF receptor dysfunction. METHODS We screened 187 patients referred to us for measurement of GM-CSF autoantibodies to diagnose autoimmune PAP. Five were children with PAP and increased serum GM-CSF but without GM-CSF autoantibodies or any disease causing secondary PAP; all were studied with family members, subsequently identified patients, and controls. MEASUREMENT AND MAIN RESULTS Eight children (seven female, one male) were identified with PAP caused by recessive CSF2RA mutations. Six presented with progressive dyspnea of insidious onset at 4.8 ± 1.6 years and two were asymptomatic at ages 5 and 8 years. Radiologic and histopathologic manifestations were similar to those of autoimmune PAP. Molecular analysis demonstrated that GM-CSF signaling was absent in six and severely reduced in two patients. The GM-CSF receptor β chain was detected in all patients, whereas the α chain was absent in six and abnormal in two, paralleling the GM-CSF signaling defects. Genetic analysis revealed multiple distinct CSF2RA abnormalities, including missense, duplication, frameshift, and nonsense mutations; exon and gene deletion; and cryptic alternative splicing. All symptomatic patients responded well to whole-lung lavage therapy. CONCLUSIONS CSF2RA mutations cause a genetic form of PAP presenting as insidious, progressive dyspnea in children that can be diagnosed by a combination of characteristic radiologic findings and blood tests and treated successfully by whole-lung lavage.

Macrolides as immunomodulatory medications for the therapy of chronic lung diseases

Macrolide antibiotics have potent immunomodulatory activity. The spectrum of action of these antibiotics extends to regulation of leukocyte function and production of inflammatory mediators, control of mucus hypersecretion, resolution of inflammation, and modulation of host defense mechanisms. Macrolides are now being used or investigated to treat chronic lung inflammatory diseases, including diffuse panbronchiolitis (DPB), cystic fibrosis (CF), chronic obstructive pulmonary disease (COPD) and asthma. Intense research is ongoing to further elucidate the targets and mechanism/s of action of macrolides in eukaryotic cells. In this paper, we review recent findings on novel effects of macrolides on epithelial barrier function and resolution of inflammation, which may shed light on the mechanisms underlying the beneficial effects of macrolides in the clinic.

Exogenous surfactant enhances mucociliary clearance in the anaesthetized dog

Therapy with exogenous surfactants is currently used for the treatment of respiratory distress syndrome of the newborn (RDS) and is under investigation for treatments related to adult RDS. However, the possible use of exogenous surfactant as a means of enhancing mucus clearance in other respiratory diseases has not been addressed. We therefore studied the effects of an artificial surfactant (Curosurf) on in vivo tracheal mucus velocity in intubated pentobarbital-anaesthetized dogs. Five dogs were randomly administered, on separate occasions, either vehicle (saline) or 10 mg Curosurf by means of local instillation via a catheter into the right lung. Tracheal mucus was collected by inserting a soft-bristled cytology brush to the level of the carina, and analysed for viscoelasticity by microrheometry. Mucociliary clearability in vivo, tracheal mucus velocity (TMV) in mm.min-1, was determined by bronchoscopic observation of charcoal marker particle transit times. The initial placement of charcoal was at the same level of the lower trachea that mucus was collected from. The effect of ciliary beat frequency was assessed on the frog palate assay by a videoscopic technique. In the dog, TMV was significantly increased after administration of surfactant. The values of TMV in the vehicle- and surfactant-treated dogs were 6.3 +/- 4.0 vs 25.6 +/- 6.5 mm.min-1 (SD), respectively. There were no discernible differences between prevehicle and postvehicle TMV values, and no significant differences in any mucus viscoelastic parameter, as determined by magnetic rheometry.(ABSTRACT TRUNCATED AT 250 WORDS)

Physiotherapy and bronchial mucus transport

Cough and expectoration of mucus are the best-known symptoms in patients with pulmonary disease. The most applied intervention for these symptoms is the use of chest physiotherapy to increase bronchial mucus transport and reduce retention of mucus in the airways. Chest physiotherapy interventions can be evaluated using different outcome variables, such as bronchial mucus transport measurement, measurement of the amount of expectorated mucus, pulmonary function, medication use, frequency of exacerbation and quality of life. Measurement of the transport rate of mucus in the airways using a radioactive tracer appears to be an appropriate outcome variable for short-term studies. Evaluation of chest physiotherapy only with pulmonary function tests appears to be inadequate in short-term studies. The popularity of using pulmonary function tests is probably based more on the availability of the instruments than on a theoretical basis related to the question of chest physiotherapy improving mucus transport. Quality of life and progression of the disease are not often used as outcome variables, but it may be worthwhile to use these in the future.