Highly efficient and expedited hepatic differentiation from human pluripotent stem cells by pure small-molecule cocktails

Abstract

BackgroundThe advent of human-induced pluripotent stem cells holds great promise for producing ample individualized hepatocytes. Although previous efforts have succeeded in generating hepatocytes from human pluripotent stem cells in vitro by viral-based expression of transcription factors and/or addition of growth factors during the differentiation process, the safety issue of viral transduction and high cost of cytokines would hinder the downstream applications. Recently, the use of small molecules has emerged as a powerful tool to induce cell fate transition for their superior stability, safety, cell permeability, and cost-effectiveness.MethodsIn the present study, we established a novel efficient hepatocyte differentiation strategy of human pluripotent stem cells with pure small-molecule cocktails. This method induced hepatocyte differentiation in a stepwise manner, including definitive endoderm differentiation, hepatic specification, and hepatocyte maturation within only 13 days.ResultsThe differentiated hepatic-like cells were morphologically similar to hepatocytes derived from growth factor-based methods and primary hepatocytes. These cells not only expressed specific hepatic markers at the transcriptional and protein levels, but also possessed main liver functions such as albumin production, glycogen storage, cytochrome P450 activity, and indocyanine green uptake and release.ConclusionsHighly efficient and expedited hepatic differentiation from human pluripotent stem cells could be achieved by our present novel, pure, small-molecule cocktails strategy, which provides a cost-effective platform for in vitro studies of the molecular mechanisms of human liver development and holds significant potential for future clinical applications.Electronic supplementary materialThe online version of this article (10.1186/s13287-018-0794-4) contains supplementary material, which is available to authorized users.

Regulated differentiation of stem cells into an artificial 3D liver as a transplantable source

“…Over the past decade, several studies have introduced protocols to induce hepatocyte-differentiation in stem cells; most methods use chemical and conditioned medium stimulations or coculture with other cell types to expedite hepatic differentiation from human bone marrow mesenchymal stem cells, human umbilical cord mesenchymal stem cells, and human iPSCs. [20][21][22][23] Induced hepatocyte-like cells (i-Hep) are shown to possess most of the normal hepatocyte functions (including the secretion of aminotransferases, albumin, and α-fetoprotein), functional biotransformation systems, and the ability to engraft, integrate, and proliferate in an injured liver. Nie et al 24 used human iPSC to generate a functional liver organoid, which exhibited stronger hepatic functions than human iPSC-derived hepatocytelike cells (or i-Hep).…”

Current progress in hepatic tissue regeneration by tissue engineering

“…More recently, different research groups showed that glycogen synthase kinase 3 inhibition with CHIR99021 efficiently directs hPSCs toward the definitive endodermal lineage and primary hepatocytes, via a combination of small-molecule treatments with dimethyl sulfoxide. They stated that the small-molecule based approach is a simple, inexpensive and reproducible platform for the derivation of hepatocytes from hPSCs [182,195]. Itaba et al screened 23 newly synthesized derivatives of small molecule combinations produced from several known Wnt/βcatenin signal inhibitors.…”

The Use of Human Pluripotent Stem Cells for Modeling Liver Development and Disease

“…Overview of protocols used to derive and differentiate PSCs to hepatocyte‐like cells. FH1 ( N,N′ ‐[methylenedi‐4,1‐phenylene]bis‐acetamide) and FPH1 (2‐ N ‐[5‐chloro‐2‐methylphenyl][methylsulfonamido]‐ N ‐[2,6‐difluorophenyl]‐acetamide) are small molecules discovered to enhance hepatic phenotype . Abbreviations: DMSO, dimethyl sulfoxide; GSK3, glycogen synthase kinase 3; HGF, hepatocyte growth factor; OSKM, OCT4, SOX2, KLF4, c‐MYC reprogramming factors; OSM, OCT4, SOX2, c‐MYC reprogramming factors; PI3K, phosphoinositide 3‐kinase; TGF‐β, transforming growth factor beta.s…”

“…Recent studies have also reported the use of small molecules to generate iPSC‐derived hepatocytes, without the need for growth factors (Fig. ) . Small molecule–based protocols have the advantage of reduced cost and should also have enhanced reproducibility and usability when compared to growth factor–based protocols.…”