Establishment method of inflammation model of mice primary colonic epithelial cells induced by dextran sodium sulfate combined with lipopolysaccharides

A technology of dextran sodium sulfate and establishment method, which is applied in the field of establishment of mouse primary colonic mucosal epithelial cell inflammation model, can solve the problem of low cost, and achieve the effect of low cost and easy operation

Active Publication Date: 2019-10-18
SOUTH CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The method is easy to operate and low cost

Method used

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  • Establishment method of inflammation model of mice primary colonic epithelial cells induced by dextran sodium sulfate combined with lipopolysaccharides
  • Establishment method of inflammation model of mice primary colonic epithelial cells induced by dextran sodium sulfate combined with lipopolysaccharides
  • Establishment method of inflammation model of mice primary colonic epithelial cells induced by dextran sodium sulfate combined with lipopolysaccharides

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Effects of different concentrations of dextran sodium sulfate treatment on cell viability:

[0042] (1) Experimental grouping and processing

[0043] The cell control group and the dextran sodium sulfate group with different concentration gradients (0.31%, 0.62%, 1.25%, 2.5%, 5%, 10%, 12.5%, 15% w / v) were respectively set up, with 2% (w / v) FBS iCell primary epithelial cell culture medium (product number: PriMed-iCELL-001) to adjust the concentration of mouse primary colonic mucosal epithelial cells to 1×10 5 cells / mL, and seeded in 96-well plate, 100 μL per well, 37°C, 5% CO 2 Cultivate overnight in the incubator; add 100 μL of different concentrations of dextran sodium sulfate diluted with serum-free iCell primary epithelial cell culture medium to each well of the experimental group, and add the same amount of serum-free iCell primary epithelial cell culture medium to the blank control group Cell culture medium, 37°C, 5% CO 2 Continue to grow in the incubator.

[...

Embodiment 2

[0047] Effects of treatment with different concentrations of lipopolysaccharide combined with 10% dextran sodium sulfate on the levels of TNF-α and IL-1β in cell culture supernatant

[0048] (1) Experimental grouping and processing

[0049] Cell control group and lipopolysaccharide group with different concentration gradients (1 μg / mL, 10 μg / mL, 100 μg / mL) were respectively set up, adjusted with iCell primary epithelial cell culture medium (product number: PriMed-iCELL-001) containing 2% FBS. The concentration of mouse primary colonic mucosal epithelial cells was 1×10 5 cells / mL, seeded in 24-well plate, 1000 μL per well, 37°C, 5% CO 2 Cultivate overnight in the incubator; add 1000 μL of 10% dextran sodium sulfate diluted with serum-free iCell primary epithelial cell culture medium to the experimental group, at 37°C, 5% CO 2 After being stimulated in the incubator for 3 hours, suck out the supernatant, add 100 μL PBS to each well and soak for 10 seconds, gently suck out and ...

Embodiment 3

[0053] 1. The experimental data were analyzed by one-way analysis of variance (One-Way ANOVA) using SSP20.0 statistical software, and compared between Duncan groups.

[0054] 2. Experimental results

[0055] (1) Effects of different concentrations of dextran sodium sulfate on the inhibitory rate of primary colonic mucosal epithelial cells in mice

[0056] The result is as figure 1 As shown, as the concentration of dextran sodium sulfate increased, the inhibition rate gradually increased; and as the action time increased, the cell inhibition rate gradually increased. Among them, 10% dextran sodium sulfate treated primary colonic epithelial cells of mice for 2 hours and 3 hours can make the inhibition rate of the cells reach 50% (P<0.05).

[0057] (2) The effect of 10% dextran sodium sulfate combined with lipid treatment on the TNF-α content of primary colonic mucosal epithelial cells of mice

[0058] The result is as figure 2 As shown, in the three stimulation times of 3h,...

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Abstract

The invention discloses an establishment method of an inflammation model of mice primary colonic epithelial cells induced by dextran sodium sulfate combined with lipopolysaccharides, and belongs to the technical field of cellular inflammatory models. According to the establishment method of the inflammation model of the mice primary colonic epithelial cells induced by the dextran sodium sulfate combined with the lipopolysaccharides, dextran sodium sulfate (DSS) and lipopolysaccharides (LPS) are combined as a stimulant for inducing and treating mice primary colonic epithelial cells so as to establish a cellular inflammatory model for the first time; and the colonic epithelial cells are used as model establishing cells of the inflammation model induced by the DSS combined with the LPS for the first time. Moreover, the optimal concentration of the dextran sodium sulfate and the lipopolysaccharides are determined by measuring cell inhibition rate as well as TNF-alpha and IL-1-beta levels of the liquid supernatant of the cells; and thus, the establishment method is easy to operate, relatively low in cost, and capable of helping researchers, who carries out mice primary colonic epithelial cell culture and cellular inflammatory model establishment for the first time, to relatively smoothly culture the mice primary colonic epithelial cells and establish the cellular inflammatory model.

Description

technical field [0001] The invention belongs to the technical field of cell inflammation models, and more specifically relates to a method for establishing a mouse primary colonic mucosal epithelial cell inflammation model induced by sodium dextran sulfate combined with lipopolysaccharide. Background technique [0002] At present, intestinal epithelial cells are considered to be an important role in the regulation of natural and acquired immune systems on the mucosal surface. In the face of bacterial invasion, colonic epithelial cells up-regulate a series of pro-inflammatory cytokines TNF-α, IL-1β, IL-6, IL- 8 and the expression and production of chemokines and prostaglandins. The early signals provided by the intestinal epithelial cells are very important for the initial regulation of the inflammatory response of the bacterial invasion host. From this perspective, the epithelial cells can be considered as the earliest sensors on the mucosal surface for bacterial invasion, a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071
CPCC12N5/0625C12N5/0679C12N2501/90
Inventor 郭世宁黎增权晁利民陈文谦李岳飞吕伟杰
Owner SOUTH CHINA AGRI UNIV
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