Naked mole rat microglial cell culture method

A technology of microglia and culture method, which is applied in the field of isolation, purification and culture of naked mole rat microglia, can solve the problems of inapplicability to naked mole rats, and achieve the goal of ensuring proliferation and vitality, high reproducibility, and operation The effect of simple method

Active Publication Date: 2016-06-22
SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] However, in the prior art, the methods for isolating, purifying and culturing microglia in mice and rats are not suitable for naked mole rats. to report

Method used

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  • Naked mole rat microglial cell culture method
  • Naked mole rat microglial cell culture method
  • Naked mole rat microglial cell culture method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Example 1: Isolation, purification and culture of naked mole rat microglia

[0038] 1. Experimental materials

[0039] Clean-grade naked mole rats born 1-7 years old were provided by the Experimental Animal Center of the Second Military Medical University of the Chinese People's Liberation Army. Place the newborn naked mole rats on an ice plate at -20°C for anesthesia, soak them in 75% ethanol for 10 minutes, then decapitate them, and wipe the surface of the naked mole rats' heads with a mixture of penicillin 100 U / ml and 0.1 mg / ml streptomycin , then cut the head skin and skull, peel off the brain and place it in pre-cooled dissecting fluid.

[0040] DNase I was purchased from Solarbio Biotechnology Co., Ltd., trypsin, penicillin-streptomycin mixture, etc. were purchased from Sigma Company, DMEM, low-sugar DMEM, fetal bovine serum from Australia, Neurobasal medium, B27 serum-free additive factors, etc. were purchased from ThermoFisherScientific Company, Petri dishes,...

Embodiment 2

[0046] Example 2: Identification of naked mole rat microglia

[0047] Identification of naked mole rat microglial cells obtained in Example 1: 4% paraformaldehyde was used to fix the microglial cells in the proliferation medium, and combined with morphological identification, immunochemical refinement and other methods for identification .

[0048] 1. Cell morphology identification:

[0049] The identification method was as described in the literature (ChungWS, ClarkeLE, WangGX, StaffordBK, SherA, ChakrabortyC, JoungJ, FooLC, ThompsonA, ChenC, SmithSJ, BarresBA. Astrocytes mediates synapsee elimination through MEGF10 and MERTK pathways. Nature, 2013, 504(7480): 394-400.).

[0050] The result is as figure 2 As shown, under the light microscope, the cell bodies of microglial cells are round or oval, and there are no protrusions around the cell bodies when they are not activated.

[0051] 2. Immunocytochemical identification:

[0052] The identification method was as describ...

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Abstract

The invention relates to the technical field of cell biology and in particular relates to a separation, purification and culture method of naked mole rat microglial cells. The microglial cells are separated from cerebral cortices of newly-born naked mole rats and are purified and cultured by comprehensively utilizing a plurality of types of culture mediums, and a reasonable culture method applicable to poikilothermal rodent mammal naked mole rat microglial cells is explored. The method provided by the invention can be used for simply, conveniently, efficiently and economically obtaining a lot of naked mole rat microglial cells with normal functional activity, and cells can still keep biological characteristics under an in-vivo state in an in-vitro environment through culture under a low-oxygen condition, so that special physiological functions of the naked mole rat microglial cells can be conveniently and directly researched in a pure in-vitro cell culture model, and furthermore, an important theoretical basis is provided for exploring a biological mechanism and applying the biological mechanism to clinically relative fields.

Description

Technical field: [0001] The invention relates to the technical field of cell biology, in particular to a method for separating, purifying and culturing cells, in particular to a method for separating, purifying and culturing naked mole rat microglial cells. Background technique: [0002] Microglia is an important component of the immune function of the central nervous system and plays an important role in the regulation of central nervous system homeostasis. Microglia closely monitor their microenvironment and activate when stimulated. Studies have found that the activation of microglia can be observed in central nervous system diseases such as Alzheimer's disease, multiple sclerosis, and amyotrophic lateral sclerosis. The activation of microglia plays an important role in the injury of the central nervous system. It is possible to regulate the repair process of the central nervous system by inhibiting or promoting the activation of microglia. Regulation of plasma cell fun...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/079
CPCC12N5/0622
Inventor 崔淑芳杨文静汤球孙伟
Owner SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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