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 . 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 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 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 and 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 , 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.