The Use of Livers With Metabolic Disorders for Transplantation - Current Status: The Domino Liver World Transplant Register (Dltr) Viewpoint

Abstract

Introduction: Liver ischemia-reperfusion (I/R) injury is an unavoidable and no cure process in clinical liver transplantation. We previously reported to prevent liver I/R through gene silencing using siRNA. Although this strategy is promising in animal models, however, several concerns such as global gene silencing have been raised when the siRNA is administrated systemically. The aim of this study is to develop a hepatocyte-specific delivery system of siRNA for treatment of liver I/R injury. Methods: shRNA expression vectors were constructed for specifically targeting TLR4 gene. shRNA is encapsulated with the liposomes that were ligated with galactose (Gal-lipo-shRNA). BALB/c mice were treated with (Gal-lipo-shRNA) by i.v. injection. Liver I/R injury was induced by interrupting blood supply to the left lateral and median lobes of the liver for 60 min followed by reperfusion 6 hrs. I/R injury was evaluated using liver histopathology, as well as levels of serum alanine transferase (ALT) and aspartate transaminase (AST). Neutrophil accumulation was determined by a myeloperoxidase (MPO) assay and immunohiscochemical staining. Lipid peroxidation was assessed by malondialdehyde (MDA) levels. Quantitative PCR was used to test gene silencing efficacy in vitro and in vivo. Results: The galactose-ligated liposomes was capable of encapsulating >96% shRNA. Heptocyte-specific targeting was confirmed by in vivo delivery experiment, in which most Gal-lipo-shRNA were accumulated in the liver as soon as 6 hrs after administration. In vivo gene silencing was significant in the liver after treatment of Gal-lipo-shRNA. To test the therapeutic effects, we treated the mice with Gal-lipo-shRNA in liver I/R mice. In comparison with control mice, the serum levels of ALT and AST, were significantly reduced in mice treated with Gal-lipo-shRNA. Moreover, histopathology displayed remarkable improvement in the level treated with Gal-lipo-shRNA. Conclusions: This study is the first demonstration to treat liver I/R injury using targeted RNA interference. We successfully developed a new system to deliver siRNA specifically to hepatocytes using galactose-ligated liposomes. Treatment with Gal-lipo-shRNA can prevent liver I/R through inducing liver-specific gene silencing but avoiding adverse effects caused by systemic administration of shRNA.