biostudies-literatureBone HKMedical Research CouncilWellcome TrustEngineering and Physical Sciences Research Council1992-2000https://www.ebi.ac.uk/biostudies/studies/S-EPMC3104033biostudies-literatureUnknown124(Pt 12)The use of small molecules to 'chemically direct' differentiation represents a powerful approach to promote specification of embryonic stem cells (ESCs) towards particular functional cell types for use in regenerative medicine and pharmaceutical applications. Here, we demonstrate a novel route for chemically directed differentiation of human ESCs (hESCs) into definitive endoderm (DE) exploiting a selective small-molecule inhibitor of glycogen synthase kinase 3 (GSK-3). This GSK-3 inhibitor, termed 1m, when used as the only supplement to a chemically defined feeder-free culture system, effectively promoted differentiation of ESC lines towards primitive streak (PS), mesoderm and DE. This contrasts with the role of GSK-3 in murine ESCs, where GSK-3 inhibition promotes pluripotency. Interestingly, 1m-mediated induction of differentiation involved transient NODAL expression and Nodal signalling. Prolonged treatment of hESCs with 1m resulted in the generation of a population of cells displaying hepatoblast characteristics, that is expressing α-fetoprotein and HNF4α. Furthermore, 1m-induced DE had the capacity to mature and generate hepatocyte-like cells capable of producing albumin. These findings describe, for the first time, the utility of GSK-3 inhibition, in a chemically directed approach, to a method of DE generation that is robust, potentially scalable and applicable to different hESC lines.Journal of cell scienceA novel chemically directed route for the generation of definitive endoderm from human embryonic stem cells based on inhibition of GSK-3.PMC3104033GR/S99419/01MC_G1000732Nelson ASGoldring CETosh DWelham MJBone HKfalseA novel chemically directed route for the generation of definitive endoderm from human embryonic stem cells based on inhibition of GSK-3.The use of small molecules to 'chemically direct' differentiation represents a powerful approach to promote specification of embryonic stem cells (ESCs) towards particular functional cell types for use in regenerative medicine and pharmaceutical applications. Here, we demonstrate a novel route for chemically directed differentiation of human ESCs (hESCs) into definitive endoderm (DE) exploiting a selective small-molecule inhibitor of glycogen synthase kinase 3 (GSK-3). This GSK-3 inhibitor, termed 1m, when used as the only supplement to a chemically defined feeder-free culture system, effectively promoted differentiation of ESC lines towards primitive streak (PS), mesoderm and DE. This contrasts with the role of GSK-3 in murine ESCs, where GSK-3 inhibition promotes pluripotency. Interestingly, 1m-mediated induction of differentiation involved transient NODAL expression and Nodal signalling. Prolonged treatment of hESCs with 1m resulted in the generation of a population of cells displaying hepatoblast characteristics, that is expressing α-fetoprotein and HNF4α. Furthermore, 1m-induced DE had the capacity to mature and generate hepatocyte-like cells capable of producing albumin. These findings describe, for the first time, the utility of GSK-3 inhibition, in a chemically directed approach, to a method of DE generation that is robust, potentially scalable and applicable to different hESC lines.2011-01-01T00:00:00Z2011 Jun2025-04-22T08:48:10.902Z2019-03-27T00:42:06ZS-EPMC31040332161009910.1242/jcs.081679