Role of autophagy in reduction of apoptosis in liver cells by hydrogen-rich saline in rats undergoing liver transplantation

Objective To investigate the role of autophagy in reduction of apoptosis in liver cells by hydrogen-rich saline in the rats undergoing liver transplantation. Methods Fifty-six healthy adult male Sprague-Dawley rats, weighing 200-250 g, were randomly divided into 4 groups using a random number table: sham operation group (group S, n=8); orthotopic liver transplantation (OLT) group (n=16); hydrogen-rich saline group (group HS, n=16); autophagy inhibitor 3-methyladenine group (group M, n=16). The number of donors was 8, and recipients 8 in OLT, HS and M groups.3-methyladenine 2.5 mg/kg was injected intraperitoneally at 1 h before surgery in group M. The model of OLT was established according to the modified Kamada′s two-cuff technique.Immediately after opening of infra-hepatic vena cava, 4 ℃ hydrogen-rich saline 6 ml/kg was injected through the infra-hepatic vena cava in HS and M groups, and 4 ℃ normal saline 6 ml/kg was given in group OLT.At 6 h after portal vein opening in OLT, HS and M groups, or at 6 h after the end of surgery in group S, blood samples were collected from the infra-hepatic vena cava for determination of serum alanine transaminase (ALT) and aspartate transaminase (AST) concentrations.After blood sampling, livers were removed for microscopic examination of the pathological changes which were scored and ultrastructure, and for determination of the malondialdehyde (MDA) content, superoxide dismutase (SOD) activity, apoptotic cells, and expression of total p53 in cytoplasm, phosphorylated p53 (p-p53), microtubule-associated proteins light chain 3Ⅱ (LC3 Ⅱ)and Beclin-1, caspase-3 and cytochrome c (Cyt c) in liver tissues (by Western blot). The ratio of p-p53 to total p53 was used to reflect the phosphorylation of p53. Results Compared with group S, the serum ALT and AST concentrations, pathological score, MDA content and apoptosis index were significantly increased, the SOD activity was decreased, and the expression of caspase-3 and Cyt c was up-regulated in OLT, HS and M groups, the expression of LC3Ⅱ and Beclin-1 was up-regulated in OLT and HS groups, and the ratio of p-p53 to total p53 was increased in HS and M groups (P 0.05), and autophagosomes were reduced in group M. Conclusion The mechanism by which hydrogen-rich saline mitigates apoptosis in liver cells is related to enhancement of p53 activity in cytoplasm and inhibition of autophagy in the rats undergoing liver transplantation. Key words: Hydrogen; Liver transplantation; Apoptosis; Autophagy