Method for repolarizing primary hepatocytes in vitro

A primary hepatocyte, repolarization technology, applied in artificial cell constructs, animal cells, vertebrate cells, etc., can solve the problems of insignificant improvement and late appearance of repolarization, achieving compact arrangement, The effect of enhancing cell function in vitro

Inactive Publication Date: 2015-12-09
NANFANG HOSPITAL OF SOUTHERN MEDICAL UNIV
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Problems solved by technology

[0008] 2. The primary hepatocytes cultured in the sandwich configuration improved the repolarization ability of the physiological primary hepatocytes, and the hepatocytes formed a hepatic plate-like structure, accompanied by the formation of bile canalicular network, and the proteins in the hepatocyte membrane region were specifically distributed, but The degree of improvement is not significant
[0009] 3. Repolarization occurs late

Method used

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  • Method for repolarizing primary hepatocytes in vitro
  • Method for repolarizing primary hepatocytes in vitro
  • Method for repolarizing primary hepatocytes in vitro

Examples

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Embodiment 1

[0041] Embodiment 1, microfluidic perfusion culture method

[0042] This example provides a method for in vitro repolarization of primary hepatocytes, comprising the following steps: 50 μL of 6 The cell / mL primary hepatocyte suspension flowed through at a flow rate of 0.02 mL / h by means of circulatory perfusion and naturally retained in the cell culture pool. At the same time, under an ordinary inverted optical microscope, a simple compact force is applied to the primary hepatocyte population by gently wriggling with ophthalmic tweezers, so that the cell density per unit volume reaches more than 90%. Cells were cultured at 37°C, 5% CO in serum-free medium for hepatocyte phenotype after planting. 2 perfusion culture conditions. In the first hour, hepatocyte phenotype serum-free culture medium was used for perfusion culture at a flow rate of 0.02mL / h, and in the second to 4h, hepatocyte phenotype serum-free culture medium was used for culture at a low flow rate (0.02-0.03mL / h)...

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Abstract

The invention relates to a method for repolarizing primary hepatocytes in vitro. The method is characterized by comprising the following steps of planting a primary hepatocyte suspension into a cell culture insert in a perfusion planting mode; meanwhile, applying a compact force to the primary hepatocytes for enabling cell density in a unit volume to reach over 90 percent; then performing perfusion culture by using a serum-free medium with a hepatocyte phenotype. According to the method disclosed by the invention, a primary mature hepatocyte population is planted in a compact mechanical planting mode, and the repolarization of the hepatocytes in structural and functional mechanics are both formed early at twelfth hour after planting, so that the repolarizing speed of an in vitro hepatocyte population in a three-dimensional environment is improved, and primary mature hepatocytes with an approximate physiological function phenotype are provided for liver tissue engineering.

Description

technical field [0001] The invention relates to the technical field of liver tissue engineering, in particular to a method for in vitro repolarization of primary liver cells. Background technique [0002] Liver tissue engineering has important basic research and clinical application values ​​in the fields of drug screening, in vitro supportive therapy, and liver tissue physiology and disease simulation models. Primary mature hepatocytes with a similar physiological function phenotype are important cellular components for liver tissue engineering. How to obtain engineered liver tissue with the most similar in vivo functions is the core goal of research in this field and one of the key technologies for success. [0003] Mature hepatocytes have complex polarity characteristics different from other epithelial cells in function and structure, and this polarity is the basis for hepatocytes to perform physiological functions. After enzymatic cleavage from the internal environment...

Claims

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Application Information

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IPC IPC(8): C12N5/071
Inventor 汪艳杨金连
Owner NANFANG HOSPITAL OF SOUTHERN MEDICAL UNIV
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