Periodontal membrane stem cell proliferation culture medium and proliferation culture method thereof

A technology of periodontal ligament stem cells and proliferation medium, applied in the field of stem cell culture, can solve the problems of limited proliferation of periodontal ligament stem cells, achieve strong osteogenic induction ability, and promote rapid proliferation

Active Publication Date: 2019-04-12
GUANGZHOU SALIAI STEMCELL SCI & TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the existing medium has limited effect on the proliferation of periodontal ligament stem cells

Method used

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  • Periodontal membrane stem cell proliferation culture medium and proliferation culture method thereof
  • Periodontal membrane stem cell proliferation culture medium and proliferation culture method thereof
  • Periodontal membrane stem cell proliferation culture medium and proliferation culture method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] Embodiment 1, mixture proportioning

[0049] The concentration of each component of the mixture:

[0050] (1) The working concentration of tranexamic acid is 500mg / L and 10000mg / L;

[0051] (2) The working concentration of trehalose is 1000mg / L, 20000mg / L;

[0052] (3) The working concentration of EGF is 10ng / ml.

[0053] Experimental group: named group A, which is DMEM / F12+ mixture;

[0054] Control group: named group B, DMEM / F12+10%FBS.

[0055] Table 1 Concentration ratio of each component in Group A

[0056]

Embodiment 2

[0057] Embodiment 2, adopt the gradient concentration culture medium of embodiment 2 to carry out proliferation experiment

[0058] Take a 12-well culture plate, inoculate the revived periodontal ligament stem cells in each well at a density of 10,000 cells / well, add 1 mL of relevant culture medium (medium medium in Example 1) to each well, place at 37°C, 5%CO 2 Cultured in an incubator, the medium was changed every three days. From day 2 (the newly recovered cells need about 24 hours to adapt to the environment), take out the plate every 24 hours, randomly select 3 wells, aspirate the old culture medium and wash it with PBS, add trypsin to digest the cells, stop the digestion, and prepare single cells Take the suspension, blow it evenly, take 10 μL of the cell suspension and mix with 10 μL of 0.4% trypan blue, add the sample to the blood cell counting board, count the number of cells in the large squares at the four corners of the counting board under a 10X microscope, and t...

Embodiment 3

[0063] Embodiment 3, identification of osteogenic ability

[0064] Take the P5 cells (cell number P16QD00198) obtained by A3 ratio culture, and use 5×10 4 Cells / well were inoculated in 6-well plates, divided into osteogenic induction group and control group, and cultured in complete medium. The osteogenic induction medium (containing 500ng / mL BMP-2, 10% FBS, 100nmol / L dexamethasone, 20mmol / Lβ-sodium glycerophosphate, 10mg / L vitamin C) was replaced after the adherent cells in the osteogenic induction group grew and fused. L-DMEM), the control group continued to culture with complete medium and conventional induction medium (L-DMEM of 10% FBS, 100nmol / L dexamethasone, 20mmol / L β-sodium glycerophosphate, 10mg / L vitamin C). The culture medium was changed every 3 days, and Alizarin red staining was performed at 4 weeks for identification. Osteogenic effect see figure 2 .

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Abstract

The invention relates to the technical field of stem cell culture, in particular to a periodontal membrane stem cell proliferation culture medium and a proliferation culture method thereof. The periodontal membrane stem cell proliferation culture medium comprises the components with concentration of tranexamic acid of which the concentration is 500-15000mg / L, trehalose of which the concentration is 500-2000mg / L, EGF of which the concentration is 5-15ng / mL, and the balance a basic culture medium. After the tranexamic acid, the trehalose and the EGF are added to the basic culture medium, quick proliferation of periodontal membrane stem cells can be promoted, and the effects of the periodontal membrane stem cell proliferation culture medium are better than those of a conventional culture medium, and are better than those of a culture medium with a single factor. Besides, the culture medium can effectively maintain stem cells of the periodontal membrane stem cells, and the cells after proliferation culture have higher ossification inducibility.

Description

technical field [0001] The invention relates to the technical field of stem cell culture, in particular to a periodontal ligament stem cell proliferation medium and a proliferation culture method thereof. Background technique [0002] Periodontitis is mainly a chronic inflammation of periodontal supporting tissues caused by local factors. The age of onset is more common after the age of 35. If gingivitis is not treated in time, the inflammation can spread from the gums to the deep layer to the periodontal ligament, alveolar bone and cementum and develop into periodontitis. Because there are no obvious symptoms in the early stage, it is easy to be ignored. When the symptoms appear, they are already serious, and even the teeth cannot be preserved. Periodontal ligament stem cells are adult stem cells derived from periodontal ligament. They have the ability to self-replicate, can produce different types of mature cells with specific phenotypes and functions, can maintain the st...

Claims

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Application Information

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IPC IPC(8): C12N5/0775
CPCC12N5/0668C12N2500/34C12N2501/11C12N2501/999
Inventor 葛啸虎陈海佳黄幸王小燕李学家
Owner GUANGZHOU SALIAI STEMCELL SCI & TECH CO LTD
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