1082. Delivery of Anti-Oxidative Enzyme Genes Protects Against Ischemia/Reperfusion-Induced Liver Injury in Mice

Abstract

BACKGROUND: Hepatic ischemic/reperfusion (I/R) injury is characterized by the generation of reactive oxygen species (ROS), such as superoxide anions and hydrogen peroxide. The aim of this study is to investigate whether anti-oxidative gene delivery by our polycationic liposomes will be an effective approach for prevention of the injury. METHODS: Polycationic liposome-mediated extracellular superoxide dismutase (EC-SOD) or/and catalase gene delivery by portal vein injection was undertaken one day prior to warm I/R injury in mice. The effects of the gene delivery were determined 6 hours after starting reperfusion. RESULTS: Polycationic liposome-mediated anti-oxidative gene delivery led to marked hepatic transgene expression. There was a 56-fold increase in human EC-SOD gene expression in the liver 30 hours after injection of EC-SOD lipoplexes as shown by real time RT-PCR; Western blot analysis demonstrated a 10-fold increase in catalase protein levels in the liver as compared to controls. Liver SOD and catalase activity increased 16.6-fold and 9-fold, respectively. The overexpression of these two anti-oxidative genes significantly suppressed the subsequent I/R-induced acute liver injury as reflected by markedreduction of serum ALT levels (520±55 and 643±24 vs. 1217±193 units/ml, p<0.01). The I/R procedure significantly increased liver superoxide anions (699±83 vs. 20±13 RLU/sec/mg, p<0.01) as detected by a chemoluminescent method with a superoxide tracer, MCLA. These levels were markedly reduced to a nearly normal level (24±7 RLU/sec/mg) by the prior delivery of the EC-SOD gene. In addition, there were reduced liver malondialdehyde levels, and restored glutathione levels, in mice receiving either EC-SOD or catalase gene delivery. Moreover, the enhanced activation of nuclear factor-κB and AP-1 in hepatic I/R injury was minimized by the anti-oxidative gene delivery. The control lipoplexes did not affect the extent of liver injury and oxidant stress levels in this I/R model. The combination of EC-SOD and catalase lipoplex injection (in half amount of each plasmid) resulted in a slight improvement in hepatic I/R injury when compared to either alone. CONCLUSIONS: The findings demonstrate that polycationic liposome-mediated EC-SOD or catalase gene delivery led to high levels of the transgene activity in the liver, and markedly attenuated liver I/R injury. The protection is associated with enhanced anti-oxidative effects in the liver. To our knowledge, this is the first successful demonstration that polycationic liposome-mediated EC-SOD and/or catalase gene delivery ameliorates warm I/R-associated liver injury in mice. This approach may become a potential novel therapy to improve graft function and survival after liver transplantation.