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Method for obtaining mesenchymal stem cells from mouse gingival tissue

A technology of mesenchymal stem cells and gums, applied in the field of obtaining mesenchymal stem cells

Pending Publication Date: 2022-03-15
SUN YAT SEN MEMORIAL HOSPITAL SUN YAT SEN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The extraction method of mouse GMSCs mentioned in the previous literature uses a variety of enzymes and complex medium conditions

Method used

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  • Method for obtaining mesenchymal stem cells from mouse gingival tissue
  • Method for obtaining mesenchymal stem cells from mouse gingival tissue
  • Method for obtaining mesenchymal stem cells from mouse gingival tissue

Examples

Experimental program
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Embodiment 1

[0024] The isolation and culture of embodiment 1 mouse GMSCs

[0025] The C57 BL / 6 mice were killed, the mandible was removed, excess skin and muscle tissue were removed, and the mandible was washed several times with PBS containing 5% double antibody. Using a syringe needle, gently separate the gingiva in the molar area. Collect the tissue in a culture dish, add 1 mg / L type IV collagenase, digest at 37°C for 30 minutes, add α-MEM containing FBS to stop the digestion, centrifuge, and discard the supernatant. Spread the tissue in a six-well plate, add 1ml of α-MEM containing 20% ​​FBS to each well, and place at 37°C, 5% CO 2 cultured in an incubator. After 2 to 3 days, the cells climbed out, and the remaining tissue pieces were washed out by changing the medium. Add 2ml of 20% FBS α-MEM to each well, and change the medium every 3 days. The cells were subcultured after covering 80% of the bottom of the plate, and then replaced with complete medium containing 10% FBS to contin...

Embodiment 2

[0026] Example 2 Detection of proliferation ability of mouse GMSCs

[0027] According to the instructions of the CCK8 cell proliferation and cytotoxicity detection kit, the absorbance at 450nm was measured with a microplate reader at the same time for 8 consecutive days, and the growth curve of GMSCs was drawn according to the results.

Embodiment 3

[0028] Example 3 Surface Marker Flow Cytometry Detection of Mouse GMSCs

[0029] The 2nd generation GMSCs (about 1×10 6 Cell suspension) was washed twice with PBS, resuspended in 100 μl PBS, added flow cytometry antibodies CD29, CD34, CD45, CD44, CD73, CD90, CD105 respectively, incubated at 4°C for 20 min in the dark, washed three times with PBS, and flowed Cytometry was used to detect the expression of the above surface markers.

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Abstract

The invention relates to the technical field of biomedicine, in particular to a method for obtaining mesenchymal stem cells from mouse gingival tissue. The method comprises the following steps: collecting gingival tissues of a mouse, and digesting the gingival tissues of the mouse by adopting collagenase to obtain the mesenchymal stem cells. The mouse GMSCs obtained by the invention highly express CD29, CD44, CD73, CD90 and CD105, and hardly express CD34 and CD45. After the mouse GMSCs are cultured for 21 days by using the adipogenesis induction liquid, the mouse GMSCs are differentiated into adipocytes and secrete lipid droplets; after the GMSCs are cultured by an osteogenesis induction solution, the cells gradually deform from a long spindle to a polygon and grow in a multi-layer covering manner, small calcification appears successively, and a new research method is provided for autotransplantation of the GMSCs in animal models of inflammation, autoimmune diseases, tissue repair and the like.

Description

technical field [0001] The invention relates to the technical field of biomedicine, in particular to a method for obtaining mesenchymal stem cells from mouse gingival tissue. Background technique [0002] Mesenchymal stem cells (MSCs) are a kind of pluripotent stem cells, which were first discovered in the bone marrow. They have all the common characteristics of stem cells, namely self-renewal, multi-directional differentiation and immune regulation. The immunoregulatory properties of MSCs are reflected in many aspects: when immune inflammatory injury occurs in the body and inflammatory signals (such as IL-6, TNF-α, IFN-γ, etc.) are released, the activation of MSCs’ immune regulation can continuously inhibit abnormal immune B cells, T cell, NK cell proliferation and cytokine production. It has been reported that the application of umbilical cord blood-derived mesenchymal stem cells (UC-MSCs) or bone marrow-derived mesenchymal stem cells (BM-MSCs) has achieved curative effec...

Claims

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Application Information

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IPC IPC(8): C12N5/0775C12N5/077G01N15/14
CPCC12N5/0668C12N5/0653G01N15/14C12N2509/00
Inventor 徐振健徐安平
Owner SUN YAT SEN MEMORIAL HOSPITAL SUN YAT SEN UNIV
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