Establishment of human ectopic liver tissue in immunocompromised mice as a model for Hepatitis C Virus studies

Abstract

Infection with hepatitis C virus (HCV) leads to an inflammatory condition which may progress to liver cirrhosis and in some cases, hepato-cellular carcinoma. Though, new drugs are being added to the list of HCV therapeutics, there is need for newer and safer drugs for the management of the disease. There is a paucity of animal models because HCV infects only humans and chimpanzees. Towards establishing a small animal model that can be used for testing the efficacy of anti-HCV drugs, it was proposed to establish an ectopic human liver in an immunocompromised strain of mouse, namely, nude mouse.

A copolymer scaffold of polyethylene glycol (PEG), gelatin and alginate was synthesized through cryo-gelation technique, on which three different liver cell lines of human origin were cultured and the 3D culture conditions were established and optimized. To assess hepatocyte functionality, hepato-specific marker molecules like urea and human serum albumin (HSA) were measured. The transcript levels of phase I and phase II enzyme genes as also HSA and transthyretin were also measured. Significant up-regulation of these was observed in the 3D cultured cells as compared to the cells grown as 2 D cells. Overall it could be concluded that the scaffold could indeed support continued hepatocyte proliferation and a robust mass of viable cells could be achieved, with the hepato-functions intact. The cells surface receptors required for viral entry in hepatocytes were shown to be expressed as assessed by immunofluorescence microscopy. Of importance was the observation that the human hepatocytes grown on the scaffold were amenable to infection with HCV as tested by measuring the negative strand of the viral RNA.

Having established the 3D culture of hepatocytes successfully, in vivo animal studies were initiated. First the biocompatibility of the scaffold was established in mice involving histopathological testing. Next, the hepatocyte laden scaffolds were transplanted in the peritoneal cavity of nude mice. The health of the mice was monitored closely and the circulatory levels of HSA was measured every second day, over a period of 30 days. The detection of significant levels of HSA in the mice even after 30 days after implantation points to the success in establishment of a small animal model which can be exploited for HCV infection studies.