Method and application for promoting induced human multipotent stem cells to differentiate into hepatocyte-like cells
A technology for human pluripotent stem cells and pluripotent stem cells, which is applied in the field of biomedicine, can solve the problems of large differences in differentiation efficiency between different batches and low maturity of liver-like cells, achieves small differences in culture, promotes synchronous differentiation, and matures high effect
Active Publication Date: 2018-02-23
HANGZHOU GRAND BIOTECH CO LTD
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Problems solved by technology
[0004] The use of induced pluripotent stem cells to obtain liver-like cells is a technology with great clinical application prospects. However, the existing methods of obtaining liver-like cells through induced pluripotent stem cells generally have the maturity of hepatic-like cells after directed differentiation. Problems such as not high, large differences in differentiation efficiency between different batches, etc.
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Embodiment 1
[0020] Human fibroblasts were induced into human pluripotent stem cells according to the method of Yamanaka (Okita, K., Ichisaka, T. & Yamanaka, S. Nature 448, 313-317 (2007).
[0021] The obtained induced human pluripotent stem cells were divided into 2.5×10 6 Cells / mL were cultured in the first induction medium containing activin A and / or P13K inhibitors for 96 hours to obtain endoderm cells; the first induction medium was based on RPMI-1640, and added With 1.0 wt % B27 supplement, 100 ng / mL Activin A and 4-6 μM P13K inhibitor;
[0022] The endoderm cells were cultured for 120 hours in a second induction medium containing bone morphogenetic protein 4 and / or fibroblast growth factor 2, and the induced human pluripotent stem cells inoculated in the second induction medium were Carrying out culture and giving electrostatic field stimulation to obtain liver progenitor cells, wherein, during the culture process, giving an electrostatic field stimulation every 3 hours, the field ...
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The invention relates to a method for promoting induced human multipotent stem cells to differentiate into hepatocyte-like cells. The method is characterized by comprising the following steps of 1, culturing the multipotent stem cells in a first induction culture medium with an activin A and / or a P13K inhibitor to obtain endoderm cells; 2, culturing the endoderm cells in a second induction culture medium, culturing the induced human multipotent stem cells inoculated in the second induction culture medium, and exerting electrostatic field stimulation to obtain liver progenitor cells; 3, culturing the liver progenitor cells in a third induction culture medium with hepatocyte growth factors to obtain immature liver cells; 4, culturing the immature liver cells in a fourth induction culture medium to obtain the liver cells. Through the method and application for promoting the induced human multipotent stem cells to differentiate into the hepatocyte-like cells, directional synchronous differentiation from the induced multipotent stem cells to the liver cells can be achieved, and the obtained hepatocyte-like cells after differentiation is small in differentiation batch difference and isrelatively high in maturity.
Description
technical field [0001] The present disclosure relates to the field of biomedicine, in particular to a method and application for promoting the differentiation of induced human pluripotent stem cells into liver-like cells. Background technique [0002] For the treatment of liver diseases, orthotopic liver transplantation is currently the only method with definite efficacy in the treatment of liver failure. However, the current demand for liver donors has far exceeded the actual supply, and there are problems such as immune rejection of allografts, which severely limit the development of orthotopic liver transplantation. Hepatic parenchymal cells from allogeneic sources have certain development prospects in the treatment of liver diseases, but there are also problems such as few sources, low value-added rate, and difficulty in expanding culture, which limit their clinical development and application. [0003] Induced pluripotent stem cells (iPSCs) are sent into somatic cells ...
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IPC IPC(8): C12N5/071C12N13/00A61K35/407A61P1/16
CPCA61K35/407C12N5/067C12N13/00C12N2501/065C12N2501/115C12N2501/12C12N2501/155C12N2501/16C12N2501/237C12N2501/39C12N2501/405C12N2506/45
Inventor 李程茅永智
Owner HANGZHOU GRAND BIOTECH CO LTD