Isolation and transcriptional profiling of purified hepatic cells derived from human embryonic stem cells.

摘要

The differentiation of human embryonic stem cells (hESCs) into functional hepatocytes provides a powerful in vitro model system for studying the molecular mechanisms governing liver development. Furthermore, a well-characterized renewable supply of hepatocytes differentiated from hESCs could be used for in vitro assays of drug metabolism and toxicology, screening of potential antiviral agents, and cell-based therapies to treat liver disease. In this study, we describe a protocol for the differentiation of hESCs toward hepatic cells with complex cellular morphologies. Putative hepatic cells were identified and isolated using a lentiviral vector, containing the alpha-fetoprotein promoter driving enhanced green fluorescent protein expression (AFP:eGFP). Whole-genome transcriptional profiling was performed on triplicate samples of AFP:eGFP+ and AFP:eGFP- cell populations using the recently released Affymetrix Exon Array ST 1.0 (Santa Clara, CA, http://www.affymetrix.com). Statistical analysis of the transcriptional profiles demonstrated that the AFP:eGFP+ population is highly enriched for genes characteristic of hepatic cells. These data provide a unique insight into the complex process of hepatocyte differentiation, point to signaling pathways that may be manipulated to more efficiently direct the differentiation of hESCs toward mature hepatocytes, and identify molecular markers that may be used for further dissection of hepatic cell differentiation from hESCs. Disclosure of potential conflicts of interest is found at the end of this article.