Rat hepatic stellate cell separation method

A technology of hepatic stellate cells and separation method, applied in the field of rat hepatic stellate cell separation, can solve the problems of inability to activate, less stellate cells, and low activity, and achieve high activity, high yield and high purity. Effect

Inactive Publication Date: 2017-11-10
JIANGYIN CHI SCI
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Problems solved by technology

The hepatic stellate cells isolated by the commonly used liver in situ perfusion method are often mixed with Kupffer cells, the amount of stellate cells obtained is small, the activity is low, and the obtained hepatic stellate cells are in a static state and cannot be activated.

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  • Rat hepatic stellate cell separation method
  • Rat hepatic stellate cell separation method
  • Rat hepatic stellate cell separation method

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Embodiment Construction

[0022] In order to present the purpose and advantages of the present invention more clearly, specific embodiments will now be further described. The specific embodiments described here are only for explaining the present invention, and are not intended to limit the present invention.

[0023] In the present invention, male SD rats with a body weight of 400-600 g are selected to be fed for 6-8 months, and the hepatic stellate cells are isolated. The specific operation is as follows:

[0024] 1. Take male SD rats, inject 10% chloral hydrate intraperitoneally to anesthetize the rats, and open the abdominal cavity of the rats in a U-shape to expose the liver and portal vein;

[0025] 2. Inject 0.2 mL of heparin (1500 U / L) into the inferior vena cava near the left kidney and have a lot of fatty tissue, and place a sterilized cotton ball;

[0026] 3. Insert the venous indwelling needle into the portal vein in parallel, pull out the needle core, ligate and fix the indwelling tube n...

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Abstract

The invention provides a rat hepatic stellate cell separation method, which comprises the following steps of (a) injecting 10-percent chloral hydrate into the abdominal cavity for anaesthetizing a rat, and laparotomizing the abdominal cavity of the rat so as to expose the liver and the portal vein; (b) performing heparin intravenous injection, and preheating perfusion liquid for liver perfusion; (d) taking the liver subjected to sufficient perfusion, cutting the liver into pieces, and performing oscillation digestion in preheated digestion liquid; (e) filtering digested liver tissues through a multilayer sieve screen; performing low-speed centrifugation; sucking supernatant; performing centrifugation again to obtain cell precipitates; and performing once cleaning by cleaning liquid; (f) resuspending the obtained cell precipitates, and performing Nycoden gradient centrifugal separation; (g) cleaning the cells for many times, resuspending the cell precipitates by a complete culture medium, and performing inoculated culture; and (h) performing cell immunofluorescent identification on rat hepatic stellate cells. The rat hepatic stellate cell separation method provided by the invention can be used for obtaining rat hepatic stellate cells which have high yield, high purity, high activity and can be activated.

Description

technical field [0001] The invention belongs to the field of cell biology, and in particular relates to a method for isolating rat hepatic stellate cells. Background technique [0002] Hepatic stellate cells (HSCs) are non-parenchymal cells unique to the liver, accounting for about 8%-13% of the total number of liver cells, 33% of the total number of non-parenchymal cells in the liver, and 1.4% of the total volume of the liver. , is a single layer of adherent growth cells, the cells are star-shaped, spindle-shaped, polygonal, spindle-shaped, with pseudopodia, and the cytoplasm contains many lipid droplets, also known as hepatic fat storage cells. Hepatic stellate cells are the main cell group that synthesizes extracellular matrix in the liver. They can not only secrete proteoglycans, glycoproteins and other extracellular matrix components, but also synthesize a certain amount of collagenase to maintain the normal basement membrane structure. Participate in the microcirculat...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071
CPCC12N5/067C12N2509/00
Inventor 张清张亚洲蒋敏齐来俊
Owner JIANGYIN CHI SCI
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