Azithromycin blocks autophagy and may predispose cystic fibrosis patients to mycobacterial infection

Azithromycin is a potent macrolide antibiotic with poorly understood antiinflammatory properties. Long-term use of azithromycin in patients with chronic inflammatory lung diseases, such as cystic fibrosis (CF), results in improved outcomes. Paradoxically, a recent study reported that azithromycin use in patients with CF is associated with increased infection with nontuberculous mycobacteria (NTM). Here, we confirm that long-term azithromycin use by adults with CF is associated with the development of infection with NTM, particularly the multi-drug-resistant species Mycobacterium abscessus, and identify an underlying mechanism. We found that in primary human macrophages, concentrations of azithromycin achieved during therapeutic dosing blocked autophagosome clearance by preventing lysosomal acidification, thereby impairing autophagic and phagosomal degradation. As a consequence, azithromycin treatment inhibited intracellular killing of mycobacteria within macrophages and resulted in chronic infection with NTM in mice. Our findings emphasize the essential role for autophagy in the host response to infection with NTM, reveal why chronic use of azithromycin may predispose to mycobacterial disease, and highlight the dangers of inadvertent pharmacological blockade of autophagy in patients at risk of infection with drug-resistant pathogens.     

Antibiotic therapy for pulmonary exacerbations in adults with cystic fibrosis.

Individuals with CF are living longer but often with chronic lung infections. Effective antibiotic therapy is necessary to treat life-threatening infectious exacerbations. Knowledge of the particular requirements of antibiotic therapy for the patient with CF allows the nurse to anticipate management plans, prepare for inpatient and outpatient care, and assess response to treatment.     

Azithromycin selectively reduces tumor necrosis factor alpha levels in cystic fibrosis airway epithelial cells

Azithromycin (AZM) ameliorates lung function in cystic fibrosis (CF) patients. This macrolide has been suggested to have anti-inflammatory properties as well as other effects potentially relevant for therapy of CF. In this study, we utilized three CF (IB3-1, 16HBE14o- AS3, and 2CFSMEo-) and two isogenic non-CF (C38 and 16HBE14o- S1) airway epithelial cell lines to investigate whether AZM could reduce tumor necrosis factor alpha (TNF-alpha) mRNA and protein levels by real-time quantitative PCR analysis and an enzyme-linked immunosorbent assay (ELISA), respectively. We studied the effects on the DNA binding of NF-kappaB and specificity protein 1 (Sp1) by an ELISA. Non-CF cells express significantly lower TNF-alpha mRNA and protein levels than an isogenic CF cell line. In CF cells, AZM treatment causes a 30% reduction of TNF-alpha mRNA levels (P < 0.05) and a 45% decrease in TNF-alpha secretion (P < 0.05), reaching approximately the levels of the untreated isogenic non-CF cells. In CF cells, NF-kappaB and Sp1 DNA binding activities were also significantly decreased (about 45 and 60%, respectively; P < 0.05) after AZM treatment. Josamycin, a macrolide lacking clinically described anti-inflammatory effects, was ineffective. Finally, AZM did not alter the mRNA expression levels of interleukin-6, a proinflammatory molecule not differentially expressed in CF and isogenic non-CF cells. The results of our study support the anti-inflammatory activities of this macrolide, since we show that AZM reduced the levels of TNF-alpha and propose inhibitions of NF-kappaB and Sp1 DNA binding as possible mechanisms of this effect.     

Structural equations to model relationships between pulmonary function,fatty acids and oxidation in cystic fibrosis

Objective. To illustrate the advantages of structural equation models in biomedical research using the complex example of cystic fibrosis. Material and methods. 595 blood samples from 312 patients were tested. The model studied the effects of age, BMI and clinical condition on seven major latent variables: pulmonary function, lipid oxidation status, vitamins A and E, glutathione, carotenoids, two essential fatty acids and arachidonic acid. Results. The model confirmed previous associations: positive (fatty acids, arachidonic acid, carotenoids and vitamins with pulmonary function and with lipid oxidation) and negative (glutathione with pulmonary function). It also verified the decrease in fatty acids during bronchial exacerbation and the increase in fatty acids and lipid oxidation after antibiotic treatment. Above all, the model revealed new positive associations between lipid oxidation and carotenoid levels and between lipid oxidation and vitamin A and E levels. Conclusions. Structural equations dealt easily with the great number of outcome variables of the example. They deserve a central place in biomedical issues involving too many correlated factors to help physicians and statisticians conceive biological models that best represent reality.     

Effects of alpha-tocopherol on platelet membrane function in cystic fibrosis

Patients with cystic fibrosis and their parents were reported to have abnormal platelet aggregation responses to prostaglandin E1. To determine whether this is a property of the platelets, we studied the adenosine 3':5'-cyclic monophosphate (cAMP) response of washed platelets to prostaglandin E1. The cAMP response to prostaglandin E1 was the same in platelets from obligate heterozygotes for cystic fibrosis and from those of healthy controls. Patients with cystic fibrosis who had deficient vitamin E levels (plasma alpha-tocopherol, less than 500 micrograms/dl) had significantly (p less than 0.01) reduced platelet cAMP response to prostaglandin E1 compared with patients who had sufficient vitamin E, and supplementation with water-miscible vitamin E in these patients resulted in significant increases in plasma alpha-tocopherol levels (p less than 0.01) and in cAMP response to prostaglandin E1 (p less than 0.05). Plasma alpha-tocopherol levels correlated significantly with platelet cAMP response to prostaglandin E1 in patients with cystic fibrosis (r = 0.58, p less than 0.05). However, plasma alpha-tocopherol level was unrelated to the lymphocyte and granulocyte cAMP response to prostaglandin E1 or to the platelet cAMP response to alpha 2-adrenergic stimulation. Our data suggest that patients with cystic fibrosis have no inherited defect in platelet cAMP response to prostaglandin E1. In patients who have sufficient vitamin E, cAMP responses to prostaglandin E1 are normal in all the formed elements of the blood.     

Recovery of airway cystic fibrosis transmembrane conductance regulator function in mice with cystic fibrosis after single-dose lentivirus-mediated gene transfer

The potential for gene therapy to be an effective treatment for cystic fibrosis (CF) airway disease has been limited by inefficient gene transfer vector particle delivery and lack of persistent gene expression. We have developed an airway conditioning process that, when combined with a human immunodeficiency virus (HIV)-derived lentivirus (LV) vector, resulted in persistent in vivo expression of transgenes in airway epithelium. Pretreatment of mouse nasal epithelium with the detergent lysophosphatidylcholine (LPC) prior to instillation of a single dose of an LV vector carrying the LacZ marker gene produced significant LacZ gene expression in nasal airway epithelium for at least 92 days. Transduction of the cystic fibrosis transmembrane conductance regulator (CFTR) gene using the same LV vector system resulted in partial recovery of electrophysiologic function in the nasal airway epithelium of CF mice (cftr(tm1Unc) knockout) for at least 110 days. This first demonstration of LV-mediated in vivo recovery of CFTR function in CF airway epithelium illustrates the potential of combining a preconditioning of the airway surface with a simple and brief LV vector exposure to produce therapeutic gene expression in airway.     

SELDI-TOF biomarker signatures for cystic fibrosis,asthma and chronic obstructive pulmonary disease

Objectives:The aim of this work was to establish protein profiles in serum and nasal epithelial cells of cystic fibrosis individuals in comparison with controls, asthma and chronic obstructive pulmonary disease patients for specific biomarker signatures identification.Design and methods:Protein extracts were analyzed by Surface Enhanced Laser Desorption/Ionization Time-Of-Flight Mass-Spectrometry (SELDI-TOF-MS).Results:The mass spectra revealed a set of peaks with differential expression in serum and nasal cells among the different groups studied, resulting into peak signatures representative/specific of each pathology. Logistic regressions were applied to those peaks; sensitivity, specificity, Youden's indexes and area under the curve (AUC) of the respective receiver operating characteristic (ROC) curves were compared.Discussion:Multivariate analysis demonstrated that combination of peaks has a better predictive value than the individual ones. These protein signatures may serve as diagnostic/prognostic markers for the studied diseases with common clinical features, or as follow-up assessment markers of therapeutic interventions.     

Cell therapy for cystic fibrosis

Currently there is no cure for cystic fibrosis (CF). Treatments are focused on addressing the disease symptoms, with varying degrees of success. Regenerative medicine holds the promise of regenerating dysfunctional or damaged tissues and to enhance the body's own endogenous repair mechanisms. The discovery of endogenous and exogenous stem cells has provided valuable tools for development of novel treatments for CF. The ability of stem cells to differentiate into functional pulmonary cells, modulate inflammatory responses and contribute to pulmonary function has provided researchers with multiple approaches to develop effective treatment strategies. Several approaches show promise to produce viable therapeutic treatments to treat the underlying cause of CF, reduce the symptoms and mitigate long‐term damage, and generate functional replacement organs for end‐stage transplantation. This review provides an overview of the rapidly progressing field of cell therapy for CF, focusing on the various cell types utilized and current strategies that show promise to improve life expectancy and quality of life for CF patients. Copyright © 2013 John Wiley & Sons, Ltd.     

Prenatal screening for cystic fibrosis

This article focuses on essential components related to prenatal screening for cystic fibrosis, including the clinical disease, inheritance, prognosis and treatment, birth prevalence, and ethnic variability. The molecular basis of this disease is presented, including a discussion of the gene, mutations, and genotype/phenotype correlations. The models that have been used for delivering prenatal screening services in pilot trials are described, along with lessons learned, expected screening performance, and relevant ELSI considerations. A realistic view of laboratory issues is considered, including current standards of performance, guidelines and oversight, and quality assurance. Examples of current laboratory technologies for cystic fibrosis testing are displayed.     

Metabolic and inflammatory responses to pulmonary exacerbation in adults with cystic fibrosis

BACKGROUND: We hypothesized that increased resting energy expenditure in adults with cystic fibrosis was related to chronic inflammation secondary to pulmonary infection and could be modified by treatment of the underlying infection. METHOD: To determine the relationship between resting energy expenditure and the inflammatory and metabolic responses, we studied 22 adults with cystic fibrosis and chronic Pseudomonas aeruginosa infection before and after treatment of a respiratory exacerbation. Resting energy expenditure was measured by indirect calorimetry. Spirometry and circulating concentrations of C-reactive protein, neutrophil elastase alpha1-antiproteinase complex, catecholamines, non-esterified fatty acids and glycerol were determined. RESULTS: The mean (95% confidence interval)% predicted FEV1 was 28.5% (20.6, 36.4) and mean body weight 50.7 kg (47.4, 54.1). Following treatment, 1-s forced expiratory volume (FEV1) and weight increased, while C-reactive protein (P<0.0001) and neutrophil elastase alpha1-antiproteinase complex concentrations (P<0.0001) were reduced. Resting energy expenditure decreased from 6.8 (6.3, 7.2) to 6.25 (5.9, 6.6) MJ day-1 by day 15 (P<0.001). Changes in resting energy expenditure and C-reactive protein were related (r = 0.66, P< 0.0001). Weight gain was inversely related to resting energy expenditure (r = 0.43, P = 0.02) and unrelated to energy intake (r = 0.02, P = 0.47). Post-treatment reduction in norepinephrine was related to changes in heart rate (r = 0.57, P<0.01), resting energy expenditure (r = 0.51, P = 0.001) and non-esterified fatty acids (r = 0.42, P< 0.05). CONCLUSIONS: A parallel reduction in the host inflammatory and catabolic responses followed treatment of a respiratory exacerbation and may have contributed to weight gain.