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Modeling hepatitis B-dependent hepatocellular carcinoma in human embryonic stem cells

Abstract

Hepatocellular carcinoma (HCC) is the third leading cause of cancer-related death worldwide; 80% of the cases are caused by chronic hepatitis B (HBV) infections that result in liver cirrhosis. Current disease models are composed primarily of cell lines transfected with HBx, a HBV protein shown to promote tumorigenesis. These models can be improved upon, as the transfected cell lines are usually cancerous or immortal already. Transfection of primary hepatocyte cell lines is ideal, but studies have demonstrated that the standard plasmid-based method has a very low targeting efficiency of ̃3%, and must be replenished due to the limited life span of the cells. Transfection of human embryonic stem cells (hESCs) using a bacterial artificial chromosome (BAC) recombineering system followed by differentiation of positive clones into hepatocytes can be a feasible alternative. Firstly, the BAC recombineering system has been shown to have a vastly improved targeting efficiency of ̃20%. Secondly, the immortal lifespan of stem cells ensures that transfection and screening only be performed once. Here, we have created a stem-cell model for HBV-dependent HCC by transfecting HUES 8 embryonic stem cell lines with HBx using a BAC recombineering system. Once differentiated into hepatocytes, they can serve as a cell culture model for HCC as well as an in vivo model via human hepatocyte repopulation in immunodeficient mice and subsequent development into cancerous human liver tissue

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