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Osteogenic induction method of periodontal ligament stem cells (PDLSCs)

A technology of periodontal ligament stem cells and inducing differentiation, applied in the fields of medicine and cell biology, can solve problems such as inability to cure periodontitis, and achieve the effects of promoting development, realizing reuse, and expanding sources

Inactive Publication Date: 2014-12-24
丁刚
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, the commonly used clinical treatment methods for periodontitis include: basic periodontal treatment (scaling, scaling, root planing), periodontal flap surgery and periodontal tissue regeneration, etc., but these methods can only relieve the symptoms of patients , To prevent the further development of the disease, it is not possible to completely rebuild the healthy periodontal tissue, that is, it cannot cure periodontitis

Method used

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  • Osteogenic induction method of periodontal ligament stem cells (PDLSCs)
  • Osteogenic induction method of periodontal ligament stem cells (PDLSCs)
  • Osteogenic induction method of periodontal ligament stem cells (PDLSCs)

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Example 1: Isolation and cultivation of human PDLSCs:

[0021] Extracted normal third molars or premolars of healthy patients (provided by the Department of Stomatology, Yidu Central Hospital, Weifang City) were selected, and PDLSCs were isolated and cultured according to the methods reported in the literature. The brief description is as follows: immediately put the extracted teeth into a sterile centrifuge tube filled with pre-cooled PBS, transfer them into the cell chamber, and isolate and culture PDLSCs within 24 hours. Gently peel off the periodontal tissue around the teeth, take the periodontal tissue in the middle, wash it repeatedly with PBS, cut it into pieces, put it in the digestive solution containing type Ⅰ collagenase (3g / L) and neutral protease (4g / L), Digest at 37°C for 1 hour, collect cells through a 70 μm cell sieve, centrifuge at 1000 rpm for 10 minutes, and resuspend with culture medium to form a single cell suspension. Inoculate the single cell sus...

Embodiment 2

[0022] Example 2: Osteotropic differentiation of PDLSCs

[0023] The PDLSCs obtained in Example 1 were inoculated in the osteotropic differentiation medium, and the inoculum size was 2.0×10 4 / cm 2 , and then placed in a 37°C cell culture incubator for induction and differentiation for 2 weeks. –3 mol / L icariin, 10 -6 mol / L emodin, 10 -5 mol / L Puerarin, 10 -6 mol / L berberine, 10 -5 mol / L eucommia alcohol extract, 2mg / L drynaria total flavonoids, 4mg / mL psoralen, 4mg / mL oleanolic acid and 10 -6 mol / L Dipsacus saponin Ⅵ.

[0024] The formulation of the basal medium in the above osteotropic induction differentiation medium is α-MEM.

Embodiment 3

[0025] Example 3: Detection of osteogenic differentiation of PDLSCs

[0026] The following three methods were used to detect the osteotropic induced PDLSCs of Example 2:

[0027] 1. Detection of alkaline phosphatase activity:

[0028] The alkaline phosphatase activity kit was used to detect PDLSCs 1 day, 3 days, and 7 days after osteotropic induction. The specific operation was carried out according to the instructions of the kit.

[0029] Results: The alkaline phosphatase activity of PDLSCs was detected after 1 day, 3 days, and 7 days of osteotropic induction, and it was found that the alkaline phosphatase activity of PDLSCs and uninduced PDLSCs on the 1st day of osteotropic induction was similar, and there was no significant difference between the two, while The alkaline phosphatase activity of PDLSCs on the 3rd and 7th day of osteotropic induction was significantly higher than that of uninduced PDLSCs, indicating that PDLSCs had obvious osteogenic differentiation.

[0030]...

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Abstract

The invention provides an osteogenic induction method of periodontal ligament stem cells (PDLSCs), which comprises the following steps: by using a third molar or premolar of a healthy patient as a raw material, separating and culturing the PDLSCs; and inoculating the PDLSCs in an osteogenic induction differentiation culture medium with the inoculum size of 2.0*10<4> / cm<2>, putting in a 37-DEG C cell culture box, and carrying out induction differentiation for 2 weeks, wherein the osteogenic induction differentiation culture medium is prepared by adding 10<-3> mol / L icariin, 10<-6> mol / L emodin, 10<-5> mol / L puerarin, 10<-6> mol / L berberine, 10<-5> mol / L encommiol extract, 2 mg / L drynaria total flavone, 4 mg / ml psoralen, 4 mg / ml oleanolic acid and 10<-6> mol / L Chinese teasel root saponin VI into a basal culture medium. The method can successfully implement osteogenic induction of the PDLSCs without influencing the stem cell characteristics and immunologic characteristics of the PDLSCs.

Description

technical field [0001] The invention relates to the fields of medicine and cell biology technology, in particular to an osteotropic induction method of periodontal ligament stem cells. Background technique [0002] Periodontitis is a common clinical disease in stomatology. It is an infectious disease characterized by the destruction of periodontal tissue. bit until lost. Periodontitis is common in the population, among which chronic periodontitis accounts for more than 60%, aggressive periodontitis accounts for 5%-15%, and periodontitis accounts for 30%-44% of tooth extraction reasons. Periodontitis is not only the main cause of tooth loss, but also related to the occurrence and development of certain systemic diseases, such as diabetes and cardiovascular diseases. Once the destruction of periodontal attachment and alveolar bone has occurred, the ideal way is to completely rebuild healthy periodontal tissue, form new periodontal attachment and new bone, but the loss of per...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/074
Inventor 丁刚
Owner 丁刚
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