Downregulation of miR122 by grainyhead like-2 restricts the hepatocytic differentiation potential of adult liver progenitor cells.

肝臓の組織幹細胞は，肝細胞と胆管上皮細胞に分化可能な細胞である．我々は，転写因子 grainyhead like-2 が，microRNA122 の発現を抑制することによって，肝幹細胞の分化能を制御していることを明らかにした

Self-renewal capability of hepatocytic parental progenitor cells derived from adult rat liver is maintained long term when cultured on laminin 111 in serum-free medium

ラット小型肝細胞の前駆細胞は，Laminin(LN)111上で自己複製能と肝細胞としての基本機能を維持しながら継代培養可能であることがわかった．また，継代培養した前駆細胞はMatrigel積層培養で成熟化する

Transplantation of Thy1+ cells accelerates liver regeneration by enhancing the growth of small hepatocyte-like progenitor cells via IL17RB Signaling

障害肝から単離したThy1陽性細胞を，肝細胞の増殖を抑制した後に肝臓の2/3を部分切除したラット肝臓に移植すると，ドナー細胞が細胞外小胞を分泌し，レシピエント肝臓の内在性細胞に作用してサイトカインを分泌させることで，レシピエントに元々存在する肝前駆細胞の増殖を促進させ，肝再生を促進する，というメカニズムを解明しました

Hepatocytic parental progenitor cells of rat small hepatocytes maintain self-renewal capability after long-term culture

ラット小型肝細胞の前駆細胞は，長期培養後も自己複製能を維持し，肝細胞としての基本機能を維持していることがわかった．また，肝前駆細胞は生体内において肝細胞と胆管細胞への二分化能を有していることも示された

Use of Biliary Organoids in Cholestasis Research.

Cholangiocytes play a crucial role in the pathophysiology of cholestasis. However, research on human cholangiocytes has been restricted by challenges in long-term propagation and large-scale expansion of primary biliary epithelium. The advent of organoid technology has overcome this limitation allowing long-term culture of a variety of epithelia from multiple organs. Here, we describe two methods for growing human cholangiocytes in organoid format. The first applies to the generation of intrahepatic bile ducts using human induced pluripotent stem cells using a protocol of differentiation that recapitulates physiological bile duct development. The second method allows the propagation of primary biliary epithelium from the extrahepatic ducts or gallbladder. Both protocols result in large numbers of cholangiocyte organoids expressing biliary markers and maintaining key cholangiocyte functions

Generation of Glucose-Responsive Functional Islets with a Three-Dimensional Structure from Mouse Fetal Pancreatic Cells and iPS Cells In Vitro

Islets of Langerhans are a pancreatic endocrine compartment consisting of insulin-producing β cells together with several other hormone-producing cells. While some insulin-producing cells or immature pancreatic cells have been generated in vitro from ES and iPS cells, islets with proper functions and a three-dimensional (3D) structure have never been successfully produced. To test whether islets can be formed in vitro, we first examined the potential of mouse fetal pancreatic cells. We found that E16.5 pancreatic cells, just before forming islets, were able to develop cell aggregates consisting of β cells surrounded by glucagon-producing α cells, a structure similar to murine adult islets. Moreover, the transplantation of these cells improved blood glucose levels in hyperglycemic mice. These results indicate that functional islets are formed in vitro from fetal pancreatic cells at a specific developmental stage. By adopting these culture conditions to the differentiation of mouse iPS cells, we developed a two-step system to generate islets, i.e. immature pancreatic cells were first produced from iPS cells, and then transferred to culture conditions that allowed the formation of islets from fetal pancreatic cells. The islets exhibited distinct 3D structural features similar to adult pancreatic islets and secreted insulin in response to glucose concentrations. Transplantation of the islets improved blood glucose levels in hyperglycemic mice. In conclusion, the two-step culture system allows the generation of functional islets with a 3D structure from iPS cells

Notch Signaling Regulates Bile Duct Morphogenesis in Mice

BACKGROUND: Alagille syndrome is a developmental disorder caused predominantly by mutations in the Jagged1 (JAG1) gene, which encodes a ligand for Notch family receptors. A characteristic feature of Alagille syndrome is intrahepatic bile duct paucity. We described previously that mice doubly heterozygous for Jag1 and Notch2 mutations are an excellent model for Alagille syndrome. However, our previous study did not establish whether bile duct paucity in Jag1/Notch2 double heterozygous mice resulted from impaired differentiation of bile duct precursor cells, or from defects in bile duct morphogenesis. METHODOLOGY/PRINCIPAL FINDINGS: Here we characterize embryonic biliary tract formation in our previously described Jag1/Notch2 double heterozygous Alagille syndrome model, and describe another mouse model of bile duct paucity resulting from liver-specific deletion of the Notch2 gene. CONCLUSIONS/SIGNIFICANCE: Our data support a model in which bile duct paucity in Notch pathway loss of function mutant mice results from defects in bile duct morphogenesis rather than cell fate specification

High efficient differentiation of functional hepatocytes from porcine induced pluripotent stem cells

Hepatocyte transplantation is considered to be a promising therapy for patients with liver diseases. Induced pluripotent stem cells (iPSCs) provide an unlimited source for the generation of functional hepatocytes. In this study, we generated iPSCs from porcine ear fibroblasts (PEFs) by overexpressing Sox2, Klf4, Oct4, and c-Myc (SKOM), and developed a novel strategy for the efficient differentiation of hepatocyte-like cells from porcine iPSCs by following the processes of early liver development. The differentiated cells displayed the phenotypes of hepatocytes, exhibited classic hepatocyte-associated bio-functions, such as LDL uptake, glycogen storage and urea secretion, as well as possessed the metabolic activities of cytochrome P-450 (CYP) 3A and 2C. Furthermore, we compared the hepatocyte differentiation efficacy of our protocol with another published method, and the results demonstrated that our differentiation strategy could significantly improve the generation of morphological and functional hepatocyte-like cells from porcine iPSCs. In conclusion, this study establishes an efficient method for in vitro generation of functional hepatocytes from porcine iPSCs, which could represent a promising cell source for preclinical testing of cell-based therapeutics for liver failure and for pharmacological applications. © 2014 Ao et al

Cholangiocytes derived from human induced pluripotent stem cells for disease modeling and drug validation.

The study of biliary disease has been constrained by a lack of primary human cholangiocytes. Here we present an efficient, serum-free protocol for directed differentiation of human induced pluripotent stem cells into cholangiocyte-like cells (CLCs). CLCs show functional characteristics of cholangiocytes, including bile acids transfer, alkaline phosphatase activity, γ-glutamyl-transpeptidase activity and physiological responses to secretin, somatostatin and vascular endothelial growth factor. We use CLCs to model in vitro key features of Alagille syndrome, polycystic liver disease and cystic fibrosis (CF)-associated cholangiopathy. Furthermore, we use CLCs generated from healthy individuals and patients with polycystic liver disease to reproduce the effects of the drugs verapamil and octreotide, and we show that the experimental CF drug VX809 rescues the disease phenotype of CF cholangiopathy in vitro. Our differentiation protocol will facilitate the study of biological mechanisms controlling biliary development, as well as disease modeling and drug screening.This work was funded by ERC starting grant Relieve IMDs (L.V., N.H.), the Cambridge Hospitals National Institute for Health Research Biomedical Research Center (L.V., N.H., F.S.), the Evelyn trust (N.H.) and the EU Fp7 grant TissuGEN (M.CDB.). FS has been supported by an Addenbrooke’s Charitable Trust Clinical Research Training Fellowship and a joint MRC-Sparks Clinical Research Training Fellowship.This is the author accepted manuscript. The final version is available from Nature Publishing Group via http://dx.doi.org/10.1038/nbt.327