Final Report Summary - MICROLIVERMATURATION (Gene network-based maturation of embryonic stem cell-derived hepatocytes in a microfabricated array) Embryonic and genetically-induced pluripotent stem cells (hPSCs) have a limitless capacity for self-renewal and a remarkable ability to differentiate, promising a limitless source of cells for cell therapies and tissue engineering. Organ scarcity and lack of proliferative ability make hPSC-derived hepatocytes one of the few potential sources of cells for clinical applications and pharmaceutical drug discovery. However, current techniques for hepatocyte differentiation result in mixed cell populations, minimal metabolic function, and long-term de-differentiation. The aim of this project is to develop and optimize a protocol for the differentiation of hPSCs to hepatocytes following developmental changes in the hepatic transcription factor network. A fundamental discovery of this project was the role of nutrients in modulating key decision points in stem cell differentiation. Our work showed that enhanced glycolysis in hPSC drives the production of Ac-CoA and subsequent histone acetylation. Mitochondrial activation in the first hours of differentiation shuts down this pathway, driving de-acetylation and stem cell commitment (Moussaieff et al. Cell Metabolism 2015). Our second discovery was the role of gut microbiome in the production of key metabolites controlling late stage hepatocyte maturation. We show that the post-partum transition from placental to enteral nutrition exposes the fetal liver to new microbiome-derived metabolites that induce hepatocyte to acquire a mature phenotype. We showed that microbiome-derived lithocholic acid and vitamin K2, act synergistically to induce PXR and activate nascent drug metabolism in hPSC-derived hepatocytes (Avior et al. Hepatology 2015). Our work was the first to demonstrate fully functional hPSC-derived hepatocytes for drug toxicity screening, demonstrate drug toxicity of 12 compounds with accuracy above 95% and R2 of 0.90.1. Y. Avior, G. Levy, M. Zimerman, D. Kitsberg, R. Schwartz, R. Sadeh, A. Moussaieff, M. Cohen, J. Itskovitz-Eldor, Y. Nahmias. Microbial-Derived Lithocholic Acid and Vitamin K2 Drive the Metabolic Maturation of Pluripotent Stem Cells-Derived and Fetal Hepatocytes, Hepatology epub (2015) 2. A. Moussaieff, M. Rouleau, D. Kitsberg, S.S. Shen-Orr, I.Laevsky M. Amit, D. Bomze, T. Scherf, M. Nissim-Rafinia, E. Meshorer, J. Itskovitz-Eldor, D. Aberdam, Y. Nahmias. Glycolysis-mediated Changes in Acetyl-coA and Histone Acetylation Control the Early Differentiation of Embryonic Stem Cells, Cell Metabolism 21(3): 392 - 402 (2015) 3. P. Roelandt, S. Obeid, J. Paeshuyse, J. Vanhove, A. Van Lommel, Y. Nahmias, F. Nevens, J. Neyts, C.M. Verfaillie. Human pluripotent stem cell derived hepatocytes support complete replication of hepatitis C virus. J Hepatol. 57(2):246-51 (2012)4. E. Keinan, E. Ezra, Y. Nahmias. Frame Rate Free Image Velocimetry for Microfluidic Devices, Appl. Phys. Lett. 103, 063507 (2013)
