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Method for constructing in vitro model of interaction of human intervertebral disc nucleus pulposus cell and immune cell

A technology of nucleus pulposus cells and immune cells, which is applied in the field of in vitro model construction of the interaction between human intervertebral disc nucleus pulposus cells and immune cells, and can solve problems such as the co-culture research model of no immune cells and intervertebral disc nucleus pulposus cells

Inactive Publication Date: 2013-07-24
FOURTH MILITARY MEDICAL UNIVERSITY
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  • Abstract
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AI Technical Summary

Problems solved by technology

However, there is no co-culture research model of immune cells and disc nucleus pulposus cells

Method used

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Experimental program
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Embodiment Construction

[0022] 1) Isolation and culture of human nucleus pulposus cells

[0023] The collected nucleus pulposus tissue was carefully separated under sterile conditions, the fibrous annular tissue segmental cartilage plate was discarded, and the remaining nucleus pulposus tissue was cut into 1×1mm 3 Fragments were digested with 0.25% trypsin at room temperature for 40 minutes, then centrifuged at a low speed (1000r / min) for 5 minutes, discarded the supernatant, digested with 0.025% type II collagenase at room temperature for 4 hours, and centrifuged at a low speed (1000r / min) after 5min, the supernatant was discarded, the cells were washed 3 times with PBS, centrifuged at low speed for 5min, and the remaining tissue was filtered out with a nylon filter with a pore size of 45μm, and the cells were counted on a counting plate. The cells were inoculated in culture flasks with DMEM / F12 medium containing 15% fetal bovine serum, and placed at 37°C with 5% CO 2 cultured in an incubator. The...

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Abstract

The invention provides a method for constructing an in vitro model of interaction of human intervertebral disc nucleus pulposus cell and immune cell, and researches mechanism of action of immune privilege of human intervertebral disc through establishment of a model. A co-culture model provided by the invention simulates in vitro interaction of human nucleus pulposus cells and immune cells, and can provide a novel experimental ideal for in vitro intervertebral disc immunity study. The inventor has been in the research of the mechanism of action of FasL in immune privilege of the intervertebral disc by using the co-culture model. According to the research, FasL expression level of nucleus pulposus cells of patients with degeneration of intervertebral disc is upregulated, and nucleus pulposus cells are co-cultured with immune cells. The research shows that upregulation of FasL in nucleus pulposus cells can effectively increase apoptosis rate of immune cells.

Description

technical field [0001] The invention belongs to cell co-cultivation in basic medicine, in particular to an in vitro model construction method for the interaction between human intervertebral disc nucleus pulposus cells and immune cells. Background technique [0002] Nucleus pulposus cells are the main cells in human intervertebral disc tissue, and play an important role in maintaining the normal shape of intervertebral discs. Related studies have found that a large number of nucleus pulposus cells in degenerated intervertebral discs are apoptotic [Jones P, Gardner L, Menage J, Williams GT, Roberts S:Intervertebral disc cells as competent phagocytes in vitro:implications for cell death in disc degeneration.Arthritis research&therapy2008,10(4):R86.];[Tschoeke SK,Hellmuth M,Hostmann A,Robinson Y,Ertel W,Oberholzer A, Heyde CE: Apoptosis of human intervertebral discs after trauma compares to degenerated discs involving both receptor-mediated and mitochondria-dependent pathways. ...

Claims

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Application Information

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IPC IPC(8): C12N5/0786C12N5/0783C12N5/077
Inventor 王海强刘志恒李新奎李昂高扬孙振陈宇飞张威林张泳照万中元
Owner FOURTH MILITARY MEDICAL UNIVERSITY
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