Azithromycin Augments Bacterial Uptake and Anti-Inflammatory Macrophage Polarization in Cystic Fibrosis.

BACKGROUND: Azithromycin (AZM) is widely being used for treating patients with cystic fibrosis (pwCF) following clinical trials demonstrating improved lung function and fewer incidents of pulmonary exacerba-tions. While the precise mechanisms remain elusive, immunomodulatory actions are thought to be involved. We previously reported impaired phagocytosis and defective anti-inflammatory M2 macrophage polarization in CF. This study systematically analyzed the effect of AZM on the functions of unpolarized and M1/M2 polarized macrophages in CF. METHODS: Monocytes, isolated from the venous blood of patients with CF (pwCF) and healthy controls (HCs), were differentiated into monocyte-derived macrophages (MDMs) and subsequently infected with P. aeruginosa. P. aeruginosa uptake and killing by MDMs in the presence or absence of AZM was studied. M1 and M2 macrophage polarizations were induced and their functions and cytokine release were analyzed. RESULTS: Following AZM treatment, both HC and CF MDMs exhibited a significant increase in P. aeruginosa uptake and killing, however, lysosomal acidification remained unchanged. AZM treatment led to higher activation of ERK1/2 in both HC and CF MDMs. Pharmacological inhibition of ERK1/2 using U0126 significantly reduced P. aeruginosa uptake in HC MDMs. M1 macrophage polarization remained unaffected; however, AZM treatment led to increased IL-6 and IL-10 release in both HC and CF M1 macrophages. AZM also significantly increased the phagocytic index for both pHrodo E. coli and S. aureus in CF M1 macrophages. In CF, AZM treatment promoted anti-inflammatory M2 macrophage polarization, with an increased percentage of CD209+ M2 macrophages, induction of the M2 gene CCL18, along with its secretion in the culture supernatant. However, AZM d'd not restore endocytosis in CF, another essential feature of M2 macrophages. CONCLUSIONS: This study highlights the cellular functions and molecular targets of AZM which may involve an improved uptake of both Gram-positive and Gram-negative bacteria, restored anti-inflammatory macrophage polarization in CF. This may in turn shape the reduced lung inflammation observed in clinical trials. In addition, we confirmed the role of ERK1/2 activation for bacterial uptake.

Neutrophil respiratory burst activity is not exaggerated in cystic fibrosis.

BACKGROUND: Exaggerated neutrophil-dominated inflammation underlies progressive cystic fibrosis (CF) lung disease. Older studies reported a defective respiratory burst in CF, but more recent studies suggest neutrophil function is normal. METHODS: We measured the amount and rate of reactive oxygen species (ROS) during PMA-stimulated respiratory burst activity in children [70 CF, 13 disease controls, 19 health controls] and adults [31 CF, 14 health controls] in neutrophils harvested from peripheral blood. Blood was collected from participants with CF when clinically stable (60 children, 9 adults) and on hospital admission (38 children, 24 adults) and discharge (18 children, 21 adults) for acute pulmonary exacerbations. RESULTS: When clinically stable, children with CF had lower ROS production [median 318,633, 25% 136,810 - 75% 569,523 RLU] than disease controls [median 599,459, 25% 425,566 - 75% 730,527 RLU] and healthy controls [median 534,073, 25% 334,057 - 75% 738,593 RLU] (p = 0.008). The rate of ROS production was also lower (p = 0.029). In neither children nor adults with CF did ROS production increase on hospital admission for acute pulmonary exacerbation, nor fall prior to discharge. There were no associations between ROS production and high-sensitivity C-reactive protein (indicating systemic inflammation) in either children or adults with CF. CONCLUSIONS: Our data do not support a role for exaggerated respiratory burst activity contributing to the exaggerated neutrophil-dominated inflammation seen with CF lung disease.

Altered skeletal muscle glucose-fatty acid flux in amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis is characterized by the degeneration of upper and lower motor neurons, yet an increasing number of studies in both mouse models and patients with amyotrophic lateral sclerosis suggest that altered metabolic homeostasis is also a feature of disease. Pre-clinical and clinical studies have shown that modulation of energy balance can be beneficial in amyotrophic lateral sclerosis. However, the capacity to target specific metabolic pathways or mechanisms requires detailed understanding of metabolic dysregulation in amyotrophic lateral sclerosis. Here, using the superoxide dismutase 1, glycine to alanine substitution at amino acid 93 (SOD1G93A) mouse model of amyotrophic lateral sclerosis, we demonstrate that an increase in whole-body metabolism occurs at a time when glycolytic muscle exhibits an increased dependence on fatty acid oxidation. Using myotubes derived from muscle of amyotrophic lateral sclerosis patients, we also show that increased dependence on fatty acid oxidation is associated with increased whole-body energy expenditure. In the present study, increased fatty acid oxidation was associated with slower disease progression. However, within the patient cohort, there was considerable heterogeneity in whole-body metabolism and fuel oxidation profiles. Thus, future studies that decipher specific metabolic changes at an individual patient level are essential for the development of treatments that aim to target metabolic pathways in amyotrophic lateral sclerosis.