Role of autophagy in hydrogen-induced reduction of lung injury in septic mice

Objective To evaluate the role of autophagy in hydrogen-induced reduction of lung injury in septic mice. Methods Sixty pathogen-free healthy male ICR mice, aged 6 weeks, weighing 20-25 g, were divided into 5 groups(n=12 each)using a random number table: sham operation group(group Sh), sepsis group(group Sep), sepsis plus hydrogen group(group Sep+ H2), sepsis plus autophagy inhibitor 3-methyladenine(3-MA)group(group Sep+ 3-MA)and sepsis plus 3-MA plus hydrogen group(group Sep+ 3-MA+ H2). Sepsis was produced by cecal ligation and puncture.At 1 h before operation, 3-MA 10 mg/kg was intraperitoneally injected.The mice inhaled 2% H2 for 1 h starting from 1 and 6 h after operation.Blood samples were collected from the common carotid artery at 24 h after operation for measurement of arterial oxygen partial pressure, and the oxygenation index(OI)was calculated.Pulmonary specimens were obtained for examination of the pathological changes which were scored.Pulmonary mitochondria were isolated for determination of mitochondrial membrane potential(MMP)and ATP content using fluorescence spectrophotometry and a bioluminescence assay, respectively, and the respiratory control rate(RCR)was calculated.The expression of autophagy-related protein microtubule-associated protein 1 light chain 3(LC3)was determined by Western blot, and the ratio of LC3-Ⅱ to LC3-Ⅰ expression (LC3-Ⅱ/LC3-Ⅰ ratio)was calculated. Results Compared with group Sh, the pathological scores were significantly increased, the OI and contents of mitochondrial RCR, MMP and ATP were decreased, and the LC3-Ⅱ/LC3-Ⅰratio was increased in Sep and Sep+ H2 groups(P 0.05). Compared with group Sep+ H2, the pathological scores were significantly increased, the OI and contents of mitochondrial RCR, MMP and ATP were decreased, and the LC3-Ⅱ/LC3-Ⅰratio was decreased in group Sep+ 3-MA+ H2(P<0.05). Conclusion The mechanism by which hydrogen ameliorates lung injury is related to enhanced level of autophagy in septic mice. Key words: Hydrogen; Sepsis; Respiratory distress syndrome, adult; Autophagy