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A2220 - Anti-Fibrotic Effects of Azithromycin in Human Lung Fibroblasts from Idiopathic Pulmonary Fibrosis Patients Compared to Control Fibroblasts
Author Block: M. Funke1, K. Krempaska2, R. Schliep3, L. Knudsen3, T. K. Geiser1; 1Pulmonary Medicine, University of Bern, Bern University Hospital, Bern, Switzerland, 2Department for Biomedical Research (DBMR), University of Bern, Bern, Switzerland, 3Institute of Functional and Applied Anatomy, Hannover Medical School, Hannover, Germany.
RATIONALE: Idiopathic pulmonary fibrosis (IPF) is a devastating progressive lung disease with reduced survival. No cure for IPF is available, but anti-fibrotic treatments are approved, which slow down disease progression. Azithromycin (AZT) is an antibiotic agent that belongs to the group of macrolides. Macrolides possess anti-inflammatory and immunomodulatory actions besides their antibacterial activity and recently anti-fibrotic effects have been described. The effect of AZT on IPF fibroblasts in direct comparison to controls is unknown. METHODS: Primary control human lung fibroblasts and IPF fibroblasts were exposed to TGF-β (5ng/ml), AZT (50μM) alone or combined prior to expression analysis of pro-fibrotic mediators by real-time qPCR. Collagen ELISA assay was used to determine collagen secretion. Cytotoxicity of AZT was assessed by LDH release assay. Microarray analysis was used to determine involved pathways. RESULTS: AZT significantly reduced TGF-β induced gene expression of collagen (Col1α1) in normal human lung fibroblasts (p-value ≤ 0.01). In IPF fibroblasts, AZT significantly reduced TGF-β induced αSMA and Col1α1 gene expression (p-value ≤ 0.001). AZT and TGF-β treatment significantly reduced collagen secretion as measured by ELISA in IPF fibroblasts but not in control lung fibroblasts (p-value ≤ 0.01). In most IPF fibroblasts, AZT induced higher LDH levels compared to control fibroblasts. AZT induced vacuolar structures in both control and IPF fibroblasts. EM revealed that the vacuolar structures are possibly late endosomes or lysosomes. Mitochondria were unaffected. Microarray analysis revealed increased ATP6V1B2 after AZT treatment and suggests involvement of this lysosomal ATPase. CONCLUSIONS: AZT shows enhanced anti-fibrotic effects in TGF-β stimulated IPF fibroblasts compared to controls. Interestingly, most IPF fibroblasts treated with AZT showed cytotoxic effects, without any cytotoxicity in control lung fibroblasts. Altered gene expression of ATP6V1B2 suggests involvement of lysosome acidification. Further studies are required to determine the cause of increased susceptibility of IPF fibroblasts towards AZT treatment.