Method for large-scale culture of bionic microspheres of iPSC-derived hepatocytes and application

Abstract

The invention relates to a method for large-scale culture of bionic microspheres of iPSC-derived hepatocytes. The method comprises the following steps of: preparing a GelMA cell microcarrier by utilizing a microfluidic technology, coating the surface of the GelMA cell microcarrier with Bcl-2 recombinant protein so as to enhance specific anti-apoptosis activity, observing the morphology of the GelMA cell microcarrier under a microscope, combining the GelMA cell microcarrier coated with the Bcl-2 recombinant protein with the iPSC-derived hepatocytes, screening the optimal size according to cell adhesion conditions of cell microcarriers with different diameters, observing a three-dimensional structure of the cells by virtue of a light microscope and a transmission electron microscope after large-scale culture, and detecting the activity and functional expression of the iPSC-derived hepatocytes on the cell microcarriers. The method disclosed by the invention is a bionic microcarrier culture method based on the iPSC-derived hepatocytes, and the method is capable of meeting the clinical cell number, high in cell activity, excellent in biocompatibility and high in efficiency.

Description

technical field [0001] The invention relates to the technical field of biomedical engineering, in particular to a method and application for large-scale cultivation of bionic microspheres of iPSC-derived hepatic cells. Background technique [0002] The treatment options for liver failure caused by end-stage liver disease are limited, and simple liver cell transplantation or bioartificial liver are relatively promising treatments. Liver support in patients with liver failure is entirely dependent on the specific biological functions of the hepatocytes used. At present, it is believed that at least 10 10 Cells of the above order of magnitude, and under the condition of ensuring cell activity, the more the number of liver cells, the better the support and treatment effect. Therefore, the large-scale culture technology of liver cells in vitro has become the core technology of the development of bioartificial liver technology. [0003] Human hepatocytes are an ideal source of l...

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Problems solved by technology

[0003] Human hepatocytes are an ideal source of liver cells, but there are ethical risks; liver tumor cell lines have low metabolic function and have tumorigenic potential; stem cells are an ideal source of emerging cells, but their differentiation is unstable; therefore, choose suitable and powerful , low-risk functional liver cell lines are also hot spots and difficulties in current research

[0004] Although the large-scale culture technology of hepatocytes in vitro has made many significant progresses, such as co-culture, biomaterial encapsulation, etc., the emergence of microcarriers provides new ideas for the large-scale culture of hepatocytes, but there are differences in the size and scale of microcarriers. etc., unstable charge properties, poor cell adhesion, and hepatocyte-bound microcarriers are still vulnerable to damage from various factors such as insufficient supply of oxygen nutrients and accumulation of metabolites during large-scale culture in vitro, which leads to the activity and loss of hepatocytes. The function declines, and eventually the therapeutic effect is affected by apoptosis; and how to transfer cells from the microcarriers that have grown to new microcarriers without cell production for cell amplification is also a problem that needs to be solved

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