The purpose of the present invention is to provide a process or method that can be utilized when deriving a three-dimensional structure of a kidney from pluripotent stem cells such as ES cells or iPS cells. The differentiation inducing method is characterized by culturing pluripotent cells with the following three steps, in order: (a) a step of culturing an embryoid body induced from the pluripotent stem cells in medium containing Bmp4 and a high-concentration (concentration A) Wnt agonist; (b) a step of culturing the embryoid body in medium which contains activin, Bmp4, retinoic acid and a middle-concentration (concentration B) Wnt agonist; and (c) a step of culturing the embryoid body in medium containing Fgf9 and a low-concentration (concentration C) Wnt agonist (herein, the Wnt agonist concentrations is concentration A>concentration B>concentration C).

This disclosure provides an improved system for culturing human pluripotent stem cells. Traditionally, pluripotent stem cells are cultured on a layer of feeder cells (such as mouse embryonic fibroblasts) to prevent them from differentiating. In the system described here, the role of feeder cells is replaced by components added to the culture environment that support rapid proliferation without differentiation. Effective features are a suitable support structure for the cells, and an effective medium that can be added fresh to the culture without being preconditioned by another cell type. Culturing human embryonic stem cells in fresh medium according to this invention causes the cells to expand surprisingly rapidly, while retaining the ability to differentiate into cells representing all three embryonic germ layers. This new culture system allows for bulk proliferation of pPS cells for commercial production of important products for use in drug screening and human therapy.

Methods are provided for producing differentiated cells from stem cells, including producing hepatocytes. Compositions thereof are also provided, as are methods of treating a liver disorder.

This invention relates to the efficient generation of cholangiocyte progenitor (CP) cells. Foregut stem cells (FSCs) are cultured in a hepatic induction medium comprising bone morphogenetic protein (BMP) and a TGF signalling inhibitor to produce a population of hepatoblasts. The hepatoblasts are then cultured in a biliary induction medium comprising fibroblast growth factor (FGF), retinoic acid and a TGF ligand to produce a population of cholangiocyte progenitors (CPs). The cholangiocyte progenitors (CPs) may be matured into cholangiocyte-like cells (CLCs) that display functional properties of Common Bile Duct (CBD) cholangiocytes. Methods, kits, cell populations and uses of these cell populations are provided.

The invention described herein relates to methods of isolating non-embryonic stem cell, e.g., adult stem cell, from a non-embryonic tissue, e.g., an adult tissue or organ. Non-embryonic stem cells (e.g., adult stem cells) thus isolated from the various tissues or organs can self-renew or propagate indefinitely in vitro, are multipotent and can differentiate into the various differentiated cell types normally found within the tissue or organ from which the stem cells are isolated. In addition, the isolated stem cells can be propagated through clonal expansion of a single isolated stem cell, to produce a clone of which at least about 40%, 70%, or 90% or more cells within the clone can be further passaged as single cell originated clones.

Subpopulations of spore-like cells expressing specific cell surface and gene expression markers are provided. In one embodiment, the cells express at least one cell surface or gene expression marker selected from the group consisting of Oct4, nanog, Zfp296, cripto, Gdf3, UtF1, Ecat1, Esg1, Sox2, Pax6, nestin, SCA-1, CD29, CD34, CD90, B1 integrin, cKit, SP-C, CC10, SF1, DAX1, and SCG10. Also provided are methods for purifying a subpopulation of spore-like cells of interest from a population of spore-like cells, and methods for inducing differentiation of the isolated spore-like cells into cells of endodermal, mesodermal or ectodermal origin. The spore-like cells can be used to generate cells originating from all three germ layers and can be used to treat a patient who has a deficiency of functional cells in any of a wide variety of tissues, including the retina, intestine, bladder, kidney, liver, lung, nervous system, or endocrine system.

This invention relates to the differentiation of pluripotent cells (PSCs) into foregut stem cells (FSCs) using a definitive endoderm induction medium comprising a TGFfi ligand, fibroblast growth factor (FGF), bone morphogenetic protein (BMP) and a PI3K inhibitor to differentiate the pluripotent cells into definitive endoderm cells and a foregut induction medium comprising a TGF ligand to differentiate the definitive endoderm cells into foregut stem cells (FSCs). Methods of differentiation, populations of foregut stem cells, culture media and kits are provided.

The present invention relates to a method for improving drug metabolism function of human stem cell-derived hepatocytes. More precisely, the human stem cell-derived hepatocytes are similar in cell morphology to human hepatocytes but display reduced expressions of drug or alcohol metabolism associated enzymes and antioxidant enzymes. So, the inventors co-cultured the hepatocytes differentiated from human stem cells with mouse hepatic stellate cells. As a result, it was confirmed that alcohol mediated toxicity was reduced so that liver cell damage or apoptosis level was reduced. In the meantime, the expressions of drug and alcohol metabolism associated enzymes and antioxidant enzymes were increased.

The present invention pertains to hepatocytes, liver progenitor cells, cholangiocytes, liver sinusoidal endothelial progenitor cells, liver sinusoidal endothelial cells, hepatic stellate progenitor cells, hepatic stellate cells, and liver cellular tissue models, as well as to methods for preparing these cells. The present invention also pertains to a cell fraction comprising liver progenitor cells, liver sinusoidal endothelial progenitor cells, or hepatic stellate progenitor cells. The present invention also pertains to a pharmaceutical composition or kit comprising the above-mentioned cells, a liver cellular tissue model, or a cell fraction. The present invention also pertains to: a method for screening liver disease treatment agents; a method for evaluating the hepatotoxicity of drugs, hepatocytes for infectious disease models, and a method for preparing the same; infectious disease model tissues and a method for preparing the same; as well as a method for screening infectious liver disease treatment agents.

A novel method of inducing or producing hepatocytes from human induced pluripotent stem cells at an unprecedented efficiency and functionality. The core of the invention is the use of experimentally discovered mRNAs at multiple critical differentiation decision points along a pluripotent to mesendoderm to endoderm to hepatocytes pathway in a previously unknown manner.