The present invention discloses a method for establishing a colorectal cancer HK2 reporter gene cell line, specifically including: firstly, designing a site-specific sgRNA sequence of an HK2 gene, and cloning same into a PX459 plasmid; integrating a homologous recombination sequence of an HK2 gene and a green fluorescent protein DNA fragment (EGFP), and transforming the plasmid and the integrated fragment together into a colorectal cancer cell line HCT116 by electroporation; and performing signal cell screening through a flow cytometer to obtain EGFP-expressing cells, and amplifying a monoclonal cell line; and identifying a positive HK2 reporter gene cell line through PCR identification and Western blot, among screened EGFP-expressing cell lines. The colorectal cancer cell line HK2 gene and EGFP are co-expressed, and the expression level of the EGFP is highly consistent with that of the HK2 gene. Therefore, the expression level of the HK2 gene can be accurately determined by detecting a change in the expression level of the EGFP. The method for establishing a cell line in the present invention are simple, easy to implement and efficient, and a gene site can be precisely positioned.

3D native tissue-derived scaffolding materials are made in various formats, including but not limited to hydrogel, sponge, fibers, microspheres, and films, all of which function to better preserve natural extracellular matrix molecules and to recapitulate the natural tissue environment, thereby effectively guiding tissue regeneration. Tissue-derived scaffolds are prepared by incorporating a homogenized tissue-derived suspension into a polymeric solution of synthetic, natural, or hybrid polymers. Such tissue-derived scaffolds and scaffolding materials have a variety of utilities, including: the creation of 3D tissue models such as skin, bone, liver, pancreas, lung, and so on; facilitation of studies on cell-matrix interactions; and the fabrication of implantable scaffolding materials for guided tissue formation in vivo. The tissue-derived scaffolds and scaffolding materials also provide the opportunity to correlate the functions of extracellular matrix with tissue regeneration and cancer metastasis, for example.

Methods and compositions for the production of cardiomyocytes and cardiac organoids from the differentiation of pluripotent stem cells in three-dimensional (3D) culture are provided. Rock inhibitor, which has been ubiquitously used as a medium supplement to prevent apoptosis during handling and culture of pluripotent stem cells, compromises the capacity of cardiac differentiation of pluripotent stem cells in 3D culture at the most commonly used concentrations.

Human cardiac fibroblasts obtained from induced pluripotent stem cells (iPS cells) are provided for use in analysis, screening programs, and the like.

Provided are a culture medium and cultivation method of rapidly expanding primary cells of esophageal squamous carcinoma in vitro, and the use thereof in screening drugs. The culture medium comprises an initial culture medium selected from DMEM/F12, DMEM, F12, or RPMI-1640, a Rho protease inhibitor, an antibiotic, insulin, an N2 additive, insulin-like growth factor 1, a non-essential amino acid, and optionally, hydrocortisone, optionally, glutamine, and optionally, bovine pituitary extract.

The subject matter of the present invention is a method for differentiating epithelial stem cells, comprising culturing one or more epithelial stem cells in contact with an extracellular matrix in the presence of an expansion medium, a bovine pituitary extract, a receptor tyrosine kinase ligand, a supernatant of primary fibroblasts and optionally, a Rho kinase inhibitor.

The invention is directed to an isolated population of mammal cells comprising about 75% or higher B-1a regula e PBS-treated tory cells expressing the cell surface inhibitory receptors lympho-cyte-activation gene 3 (LAG-3), programmed cell death protein 1 (PD-1), and C-X-C chemokine receptor type 4 (CXCR4), and secreting interleukin-27 (IL-27). The invention is also directed to methods of preparing and using the cell population to suppress the immune system and/or to treat or prevent diseases.

Human raphe nuclei organoids or spheroids (hRNS) are generated in vitro, which may be generated at least in part from human pluripotent stem (hPS) cells. Such spheroids model the human raphe nuclei and comprise specific sets of cells, e.g. serotonergic neurons, that are associated with the raphe nuclei of a human, and can be assembled with cortical spheroids (hCS) to generate functional human neuromodulatory circuits.

This disclosure relates to compositions and methods for recruiting brown adipocytes in vitro and in vivo from brown adipocyte progenitor cells found in human skeletal muscle. Methods for treating metabolic disease are also provided. Additionally, methods for treating hypothermia are provided. In some embodiments, the brown adipocyte recruiter is a human protein or peptide. In other embodiments the brown adipocyte recruiter may be a non-human protein or peptide. In still other embodiments, the brown adipocyte recruiter is a small molecule or natural product.

An object of the present invention is to provide a method of producing an intestinal epithelial cell, which has a large number of cells per area and a high accuracy of kinetic prediction for a CYP3A4 substrate drug such as midazolam, by inducing the differentiation of a pluripotent stem cell, as well as the intestinal epithelial cell, a cell sheet, an evaluation method for a test substance, a screening kit for a test substance, and a cell preparation. According to the present invention, there is provided a production method for an intestinal epithelial cell, including a first differentiation step of differentiating a pluripotent stem cell into an intestinal stem cell, a proliferation step of proliferating the intestinal stem cell obtained in the differentiation step, and a second differentiation step of differentiating the intestinal stem cell obtained in the proliferation step into an intestinal epithelial cell, in which the proliferation step is a step of bringing the intestinal stem cell into a specific state.