Pneumologie 2011; 65 - A35
DOI: 10.1055/s-0031-1296126

Role of autophagy in the development of amiodarone induced pulmonary fibrosis

P Mahavadi 1, I Henneke 1, L Knudsen 2, R Chambers 3, M Ochs 2, M Korfei 1, P Markart 1, W Seeger 1, C Ruppert 1, A Guenther 1
  • 1University of Gießen Lung Center, Medical Clinic II, Gießen, Germany
  • 2Institute for Functional and Applied Anatomy, Hannover Medical School, Germany
  • 3University College London, Rayne Institute, UK

Rationale: Amiodarone (AD) is an anti arrhythmic drug, and is well known for its vasodilatory properties. But this drug is limited by its potential side effects. Severe pulmonary toxicity is reported in patients receiving amiodarone even at low doses, with the most common histological finding being chronic interstitial pneumonia. Amiodarone induced pulmonary toxicity was studied in experimental rat model where apoptosis of alveolar epithelial cells type II (AECII) was reported. However, the molecular mechanisms underlying amiodarone induced pulmonary fibrosis remain unknown. In addition, amiodarone is known to induce autophagy in some cell types. A potential role for autophagy in the development of lung fibrosis has not been elucidated so far.

Aims: We aimed to establish a murine model of amiodarone induced pulmonary fibrosis and assess the role of autophagy in the development of the disease.

Methods: AD or vehicle was instilled intra tracheally into C57BL/6 mice, which were sacrificed on days 7, 14, 21 and 28. Right lung was lavaged & snap frozen for biochemical analysis. Left lung was fixed in formalin and embedded in paraffin. Fibrosis development was shown by trichrome staining & hydroxyproline measurement. (MLE)-12 cells and primary AECII were treated with AD/vehicle and harvested after 8, 16 & 24 hours. Surfactant proteins and markers for autophagy, apoptosis and lysosomal stress were studied by western blotting, immunofluorescence and immunohistochemistry.

Results: Development of pulmonary fibrosis was observed in AD treated mice from day 7. Accumulation of surfactant proteins-B & -C was observed, paralleled by an induction of autophagy (LC3B-II), lysosomal stress (Cathepsin D) and apoptosis (cleaved caspase 3) within the AECII.. MLE12 and AECII cells treated with AD showed the presence of LC3 positive vacuolar structures as early as 4 hours after AD treatment. AD treated cells showed an increase in the apoptotic marker cleaved caspase-3.

Conclusion: Mice exposed to amiodarone develop severe pulmonary fibrosis, along with AECII specific induction of autophagy, lysosomal stress and apoptosis. In vitro data support these findings. Thus, the autophagic pathway might be involved in the development of pulmonary fibrosis, but its interplay with apoptosis and lysosomal stress has yet to be revealed in this disease.