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In-vitro construction method of liver cancer organ model

A construction method and organoid technology, applied in artificial cell constructs, biochemical equipment and methods, liver cells, etc., can solve problems such as ignoring the interaction between various cells or cells-extracellular matrix, and achieve rapid construction and method Simple and operable effects

Active Publication Date: 2019-07-12
AFFILIATED HOSPITAL OF NANTONG UNIV
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Problems solved by technology

Although it has certain reference significance in the mechanism of hepatocarcinogenesis and liver cancer drug screening, this single-cell component model often ignores various intercellular or cell-extracellular matrix interactions

Method used

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  • In-vitro construction method of liver cancer organ model
  • In-vitro construction method of liver cancer organ model
  • In-vitro construction method of liver cancer organ model

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Embodiment Construction

[0025] see figure 1 , figure 1 The process flow diagram for the preparation of liver fibrosis organoids provided by the present invention.

[0026] Step 1: primary sorting of hepatic stellate cells and hepatic sinusoidal endothelial cells by flow cytometry, and cryopreservation in liquid nitrogen; or directly purchase frozen hepatic stellate cells and hepatic sinusoidal endothelial cells. Liver cancer cell lines were routinely cultured in advance (DMEM high-glucose medium supplemented with 1% penicillin / streptomycin, 10% fetal bovine serum; cultured in 25cm 2 Cell culture flasks, placed in a 37°C carbon dioxide incubator). When the confluence is about 80%, the liver cancer cell line is digested with trypsin, centrifuged at 800rps, and resuspended in medium A for counting. At the same time, hepatic stellate cells and hepatic sinusoidal endothelial cells were carefully recovered using an automatic cell recovery system, centrifuged at 800rps, resuspended in medium A, and count...

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Abstract

The invention discloses an in-vitro construction method of a liver cancer organ model. Liver cancer cells, hepatic stellate cells and liver sinusoidal endothelial cells are suspended in a culture medium A in a specific proportion; on the fourth day of culture, half of culture liquid is replaced, and culture is maintained; from the seventh day, the culture medium A is replaced with a culture mediumB, and then culture is continuously conducted for seven days; amplification subculture is conducted on the fourteenth day. On the basis of complex cellularity in a tumor microenvironment, the 3D liver cancer organ in-vitro model which is uniform in size, stable in structure, capable of being used for detecting the effectiveness of anticancer drugs and capable of achieving multiplication culture is quickly constructed. The constructed liver cancer organ model is simple in method, quick to construct, high in operability and suitable for researching a liver cancer generation and development mechanism, high-throughput screening of liver cancer drugs and the like, and has industrialization significance.

Description

technical field [0001] The invention relates to an in vitro construction method of a liver cancer organoid model. Background technique [0002] Liver cancer is a common malignant tumor of the digestive system, with the highest incidence and tumor-related mortality worldwide. Etiology shows that there are many factors leading to liver cancer, such as hepatitis B or C virus infection, non-alcoholic fatty liver, alcoholism, obesity and so on. Liver cancer has heterogeneity, drug resistance, and metastasis and recurrence, which brings great challenges to the treatment of liver cancer. Currently, research models for liver cancer are mainly divided into in vivo research models and in vitro research models. Among them, the in vitro research models include the traditional planar 2D culture and the three-dimensional culture that appeared in recent years. Although it has certain reference significance in the mechanism of liver cancer and the screening of liver cancer drugs, this si...

Claims

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Application Information

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IPC IPC(8): C12N5/071C12N5/09
CPCC12N5/0693C12N5/0671C12N2513/00
Inventor 郑文杰姚登福张捷金云峰倪温恺徐育青
Owner AFFILIATED HOSPITAL OF NANTONG UNIV
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