Lung autophagic response following exposure of mice to whole body irradiation, with and without amifostine.

ABSTRACT

PURPOSE: The effect of ionizing irradiation on the autophagic response of normal tissues is largely unexplored. Abnormal autophagic function may interfere the protein quality control leading to cell degeneration and dysfunction. This study investigates its effect on the autophagic machinery of normal mouse lung. METHODS AND MATERIALS Mice were exposed to 6 Gy of whole body γ-radiation and sacrificed at various time points. The expression of MAP1LC3A/LC3A/Atg8, beclin-1, p62/sequestosome-1 and of the Bnip3 proteins was analyzed. RESULTS: Following irradiation, the LC3A-I and LC3A-II protein levels increased significantly at 72 h and 7 days. Strikingly, LC3A-II protein was increased (5.6-fold at 7 days; p<0.001) only in the cytosolic fraction, but remained unchanged in the membrane fraction. The p62 protein, was significantly increased in both supernatant and pellet fraction (p<0.001), suggesting an autophagosome turnover deregulation. These findings contrast the patterns of starvation-induced autophagy up-regulation. Beclin-1 levels remained unchanged. The Bnip3 protein was significantly increased at 8 h, but it sharply decreased at 72 h (p<0.05). Administration of amifostine (200 mg/kg), 30 min before irradiation, reversed all the LC3A and p62 findings on blots, suggesting restoration of the normal autophagic function. The LC3A and Beclin1 mRNA levels significantly declined following irradiation (p<0.01), whereas Bnip3 levels increased. CONCLUSIONS: It is suggested that irradiation induces dysfunction of the autophagic machinery in normal lung, characterized by decreased transcription of the LC3A/Beclin-1 mRNA and accumulation of the LC3A, and p62 proteins. Whether this is due to defective maturation or to aberrant degradation of the autophagosomes requires further investigation.