Chair of Proteomics and Bioanalytics, Technical University of Munich, 85354 Freising, Germany.
Laboratory of Cancer Cell Systems, National Cancer Center Research Institute, Tokyo 104-0045, Japan; Department of Regenerative Medicine, Yokohama City University Graduate School of Medicine, Yokohama, Kanagawa 236-004, Japan.
Cell Rep. 2022 Mar 29;38(13):110604. doi: 10.1016/j.celrep.2022.110604.
Primary human hepatocytes are widely used to evaluate liver toxicity of drugs, but they are scarce and demanding to culture. Stem cell-derived hepatocytes are increasingly discussed as alternatives. To obtain a better appreciation of the molecular processes during the differentiation of induced pluripotent stem cells into hepatocytes, we employ a quantitative proteomic approach to follow the expression of 9,000 proteins, 12,000 phosphorylation sites, and 800 acetylation sites over time. The analysis reveals stage-specific markers, a major molecular switch between hepatic endoderm versus immature hepatocyte-like cells impacting, e.g., metabolism, the cell cycle, kinase activity, and the expression of drug transporters. Comparing the proteomes of two- (2D) and three-dimensional (3D)-derived hepatocytes with fetal and adult liver indicates a fetal-like status of the in vitro models and lower expression of important ADME/Tox proteins. The collective data enable constructing a molecular roadmap of hepatocyte development that serves as a valuable resource for future research.
原代人肝细胞广泛用于评估药物的肝毒性，但它们数量稀少，培养要求高。干细胞来源的肝细胞越来越多地被认为是替代物。为了更好地了解诱导多能干细胞分化为肝细胞过程中的分子过程，我们采用定量蛋白质组学方法来随时间跟踪 9000 种蛋白质、12000 个磷酸化位点和 800 个乙酰化位点的表达。分析揭示了特定于阶段的标志物，这是肝内胚层与未成熟肝细胞样细胞之间的主要分子开关，影响代谢、细胞周期、激酶活性和药物转运体的表达。将二维 (2D) 和三维 (3D) 衍生的肝细胞与胎肝和成人肝的蛋白质组进行比较表明，体外模型具有胎肝样状态，并且重要的 ADME/Tox 蛋白表达水平较低。这些数据使我们能够构建一个肝细胞发育的分子路线图，为未来的研究提供有价值的资源。
