Reduction of Liver Ischemia Reperfusion Injury by Silencing of TNF-α Gene with shRNA

Background. Tumor necrosis factor-alpha (TNF-alpha) is a central mediator in the hepatic response to ischemia/reperfusion. Short hairpin RNA (shRNA) has been proven to be an effective means of harnessing the RNA interference pathway in mammalian cells. In the current study, we investigated whether silencing TNF-alpha gene with shRNA can prevent liver ischemic reperfusion injury (IRI). Methods. Male BalB/c mice were randomized to TNF-alpha shRNA, scramble shRNA, or sham operation groups. TNF-alpha shRNA and scramble shRNA groups were injected 48 h before inducing IRI. IRI was induced via microaneurysm clamps applied to the left hepatic artery and portal vein. Six hours after reperfusion, IRI injury was examined by serum level of alanine aminotransferase (ALT) and aspartate aminotransferase (AST), liver histopathology, MPO, and MDA level, as well as by relative quantities of TNF-alpha mRNA. Results. TNF-alpha expression induced by ischemia reperfusion in the liver was significantly suppressed after treatment with TNF-alpha shRNA compared with the group treated with scramble shRNA (P < 0.001). Mice treated with TNF-alpha shRNA showed lower peak values of AST and ALT than scramble shRNA treated mice (P < 0.001). On histopathologic slides, mice treated with TNF-alpha shRNA had significantly less ischemia/reperfusion injury based on Suzuki score than the scramble shRNA group, 3.57 +/- 2.30 and 8.83 +/- 0.98 respectively (P < 0.001), while the sham group was not significantly different from the TNF-alpha shRNA group, 0 +/- 0 and 3.57 +/- 2.30, respectively (P = 0.075). Liver tissue MDA levels were significantly lower in mice treated with TNF-alpha shRNA as compared with the group treated with scramble shRNA (P < 0.01). Immunohistochemical staining for MPO was significantly lower in mice treated with TNF-alpha shRNA compared with the group treated with shRNA (compared with treated with scramble shRNA group.) Conclusions. Liver IRI can be minimized through gene silencing of TNF-alpha. This may represent a novel therapy in the setting of transplantation and in other conditions associated with IRI of the liver. (C) 2012 Elsevier Inc. All rights reserved.