Selina Möbus1, Dakai Yang1, Qinggong Yuan2, Timo H-W Lüdtke3, Asha Balakrishnan2, Malte Sgodda4, Bhavna Rani1, Andreas Kispert3, Marcos J Araúzo-Bravo5, Arndt Vogel6, Michael P Manns6, Michael Ott2, Tobias Cantz7, Amar Deep Sharma8. 1. Junior Research Group MicroRNA in Liver Regeneration, Cluster of Excellence REBIRTH, Hannover Medical School, Hannover, Germany; Translational Hepatology and Stem Cell Biology, Cluster of Excellence REBIRTH, Hannover Medical School, Hannover, Germany. 2. Department of Gastroenterology, Hepatology and Endocrinology, Hannover Medical School, Hannover, Germany; TWINCORE, Centre for Experimental and Clinical Infection Research, Hannover, Germany. 3. Institute for Molecular Biology, Hannover Medical School, Germany. 4. Translational Hepatology and Stem Cell Biology, Cluster of Excellence REBIRTH, Hannover Medical School, Hannover, Germany. 5. Computational Biology and Bioinformatics Group, Max Planck Institute for Molecular Biomedicine, Münster, Germany; Group of Computational Biology and Systems Biomedicine, Biodonostia Health Research Institute, 20014 San Sebastián, Spain; IKERBASQUE, Basque Foundation for Science, 48011 Bilbao, Spain. 6. Department of Gastroenterology, Hepatology and Endocrinology, Hannover Medical School, Hannover, Germany. 7. Translational Hepatology and Stem Cell Biology, Cluster of Excellence REBIRTH, Hannover Medical School, Hannover, Germany; Department of Gastroenterology, Hepatology and Endocrinology, Hannover Medical School, Hannover, Germany; Computational Biology and Bioinformatics Group, Max Planck Institute for Molecular Biomedicine, Münster, Germany. Electronic address: cantz.tobias@mh-hannover.de. 8. Junior Research Group MicroRNA in Liver Regeneration, Cluster of Excellence REBIRTH, Hannover Medical School, Hannover, Germany; Translational Hepatology and Stem Cell Biology, Cluster of Excellence REBIRTH, Hannover Medical School, Hannover, Germany; Department of Gastroenterology, Hepatology and Endocrinology, Hannover Medical School, Hannover, Germany. Electronic address: sharma.amar@mh-hannover.de.
Abstract
BACKGROUND & AIMS: Current hepatic differentiation protocols for human embryonic stem cells (ESCs) require substantial improvements. MicroRNAs (miRNAs) have been reported to regulate hepatocyte cell fate during liver development, but their utility to improve hepatocyte differentiation from ESCs remains to be investigated. Therefore, our aim was to identify and to analyse hepatogenic miRNAs for their potential to improve hepatocyte differentiation from ESCs. METHODS: By miRNA profiling and in vitro screening, we identified miR-199a-5p among several potential hepatogenic miRNAs. Transplantation studies of miR-199a-5p-inhibited hepatocyte-like cells (HLCs) in the liver of immunodeficient fumarylacetoacetate hydrolase knockout mice (Fah(-/-)/Rag2(-/-)/Il2rg(-/-)) were performed to assess their in vivo liver repopulation potential. For target determination, western blot and luciferase reporter assay were carried out. RESULTS: miRNA profiling revealed 20 conserved candidate hepatogenic miRNAs. By miRNA screening, only miR-199a-5p inhibition in HLCs was found to be able to enhance the in vitro hepatic differentiation of mouse as well as human ESCs. miR-199a-5p inhibition in human ESCs-derived HLCs enhanced their engraftment and repopulation capacity in the liver of Fah(-/-)/Rag2(-/-)/Il2rg(-/-) mice. Furthermore, we identified SMARCA4 and MST1 as novel targets of miR-199a-5p that may contribute to the improved hepatocyte generation and in vivo liver repopulation. CONCLUSIONS: Our findings demonstrate that miR-199a-5p inhibition in ES-derived HLCs leads to improved hepatocyte differentiation. Upon transplantation, HLCs were able to engraft and repopulate the liver of Fah(-/-)/Rag2(-/-)/Il2rg(-/-) mice. Thus, our findings suggest that miRNA modulation may serve as a promising approach to generate more mature HLCs from stem cell sources for the treatment of liver diseases.
BACKGROUND & AIMS: Current hepatic differentiation protocols for human embryonic stem cells (ESCs) require substantial improvements. MicroRNAs (miRNAs) have been reported to regulate hepatocyte cell fate during liver development, but their utility to improve hepatocyte differentiation from ESCs remains to be investigated. Therefore, our aim was to identify and to analyse hepatogenic miRNAs for their potential to improve hepatocyte differentiation from ESCs. METHODS: By miRNA profiling and in vitro screening, we identified miR-199a-5p among several potential hepatogenic miRNAs. Transplantation studies of miR-199a-5p-inhibited hepatocyte-like cells (HLCs) in the liver of immunodeficient fumarylacetoacetate hydrolase knockout mice (Fah(-/-)/Rag2(-/-)/Il2rg(-/-)) were performed to assess their in vivo liver repopulation potential. For target determination, western blot and luciferase reporter assay were carried out. RESULTS: miRNA profiling revealed 20 conserved candidate hepatogenic miRNAs. By miRNA screening, only miR-199a-5p inhibition in HLCs was found to be able to enhance the in vitro hepatic differentiation of mouse as well as human ESCs. miR-199a-5p inhibition in human ESCs-derived HLCs enhanced their engraftment and repopulation capacity in the liver of Fah(-/-)/Rag2(-/-)/Il2rg(-/-) mice. Furthermore, we identified SMARCA4 and MST1 as novel targets of miR-199a-5p that may contribute to the improved hepatocyte generation and in vivo liver repopulation. CONCLUSIONS: Our findings demonstrate that miR-199a-5p inhibition in ES-derived HLCs leads to improved hepatocyte differentiation. Upon transplantation, HLCs were able to engraft and repopulate the liver of Fah(-/-)/Rag2(-/-)/Il2rg(-/-) mice. Thus, our findings suggest that miRNA modulation may serve as a promising approach to generate more mature HLCs from stem cell sources for the treatment of liver diseases.
Authors: Kendall R Van Keuren-Jensen; Ivana Malenica; Amanda L Courtright; Layla T Ghaffari; Alex P Starr; Raghu P Metpally; Taylor A Beecroft; Elizabeth W J Carlson; Jeffrey A Kiefer; Paul J Pockros; Jorge Rakela Journal: Liver Int Date: 2015-09-09 Impact factor: 5.828
Authors: James A Heslop; Cliff Rowe; Joanne Walsh; Rowena Sison-Young; Roz Jenkins; Laleh Kamalian; Richard Kia; David Hay; Robert P Jones; Hassan Z Malik; Stephen Fenwick; Amy E Chadwick; John Mills; Neil R Kitteringham; Chris E P Goldring; B Kevin Park Journal: Arch Toxicol Date: 2016-04-02 Impact factor: 5.153