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Blood serum/blood plasma substitute material for culturing and amplifying dental pulp stem cells

A dental pulp stem cell and plasma replacement technology, which is used in the field of serum/plasma substitute for culturing and expanding dental pulp stem cells, and can solve the problems of inactivation of various cytokines, loss of concentrated platelets, support for cell proliferation, and transmission of infectious diseases. , to achieve the effect of fast cell proliferation, avoiding the spread of infectious diseases, and clear ingredients

Active Publication Date: 2018-09-11
北京科霖恩生物科技有限公司 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, concentrated platelets are a blood product, and there is still a risk of transmitting infectious diseases without inactivating the virus; and conventional inactivation procedures, such as albumin preparation procedures, incubation at 60°C for 12 hours, often make multiple Inactivation of cytokines, which deprives concentrated platelets of their ability to support cell proliferation

Method used

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  • Blood serum/blood plasma substitute material for culturing and amplifying dental pulp stem cells
  • Blood serum/blood plasma substitute material for culturing and amplifying dental pulp stem cells
  • Blood serum/blood plasma substitute material for culturing and amplifying dental pulp stem cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] 1. Primary culture and expansion of human dental pulp stem cells in different media

[0033] 1. Experimental materials

[0034] Specimens: Deciduous teeth were provided by the Department of Stomatology, Fujian Armed Police General Hospital, and the collection of dental specimens complied with the regulations of the Hospital Ethics and Technical Committee on the use of human specimens.

[0035] Materials: Calf serum was purchased from Sigma (USA). Recombinant human fibronectin (FN) fragments are products of Beijing Kexin Biotechnology Co., Ltd. Recombinant human albumin is a product of Huabei Pharmaceutical in Shijiazhuang City, Hebei Province, and is a pharmaceutical excipient with a dosage of 2g / L. alpha-MEM was a product of Invitrogen Gibco (USA), and insulin-transferrin-selenium solution (ITS, item number I3146) was purchased from Sigma. Both ethanolamine and O-phosphoethanolamine are products of Sigma, with final concentrations of 4 μl / L and 6 mg / L, respectively,...

Embodiment 2

[0090] Example 2. Phenotype analysis of dental pulp stem cells cultured in different systems

[0091] 1. Experimental materials

[0092] Reagents: Recombinant human FN fragments were purchased from Beijing Kexin Biotechnology Co., Ltd., and the components of the serum-free basal medium were as described in Example 1. Calf serum was provided by Sigma. Alpha-MEM is a product of Gibco. FITC-labeled mouse anti-human CD73 and CD105 monoclonal antibodies, PE-labeled mouse anti-human CD31 and CD45 are all products of BD Company in the United States.

[0093] Cells: cells obtained as described in Experiment 1. Subculture in different media.

[0094] 2. Experimental procedure

[0095] 1) Cultivate dental pulp stem cells as described in Experiment 1. When the cell density reaches about 80%, digest with 0.05% trypsin to obtain cells.

[0096] 2) Count the number of cells and suspend cells in different media (with and without serum).

[0097] 3) Follow 10 5 / dish / 10ml was inoculat...

Embodiment 3

[0107] Embodiment 3, the expansion effect of different culture media on human dental pulp stem cells

[0108] 1. Experimental materials

[0109] Specimens: Deciduous teeth were provided by the Department of Stomatology, Fujian Armed Police General Hospital, and the collection of dental specimens complied with the regulations of the Hospital Ethics and Technical Committee on the use of human specimens.

[0110] Materials: Calf serum was purchased from Sigma Company (USA), and used as a 10% solution in alpha-MEM as a positive control for cell culture. The serum-free medium basal medium formulation was as described in Experiment 1. The medium in the experimental group was serum-free basal medium containing 5-HT, PGE-2, citric acid and Vitronectin, the specific dosage was 5-HT 60 μg / L, PGE-2 60 μg / L, citric acid 20 mg / L, Vitronectin 10 mg / L.

[0111] 2. Experimental procedure

[0112] 1) The isolation of human dental pulp stem cells was carried out according to the method des...

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Abstract

The invention discloses a blood serum / blood plasma substitute material for culturing and amplifying dental pulp stem cells. The substitute material is prepared from (1) 5-hydroxytryptamine substances,(2) vitronectin, (3) prostaglandin substances and (4) citric acid or citrate. When the substitute material is used for culturing the dental pulp stem cells, the different property possibility of thecultured mesenchymal stem cells caused by batch differences of the culture system can be avoided. In addition, when the system is used for culturing the dental pulp stem cells, the zoonoses caused byanimal blood serum or blood plasma in the cell treatment process can be avoided in the cell treatment process; the human body immune reaction risk caused by cell treatment can be avoided. The blood serum / blood plasma / blood platelet substitute material is applicable to the human dental pulp stem cell culture amplification, is also applicable to the culture of other tissue sourced mesenchymal stem cells and has wide application prospects.

Description

technical field [0001] The invention belongs to the field of cell proliferation in the field of cell biology, and specifically a serum / plasma / platelet substitute for the cultivation and expansion of human dental pulp stem cells, which is used when culturing human mesenchymal stem cells including dental pulp stem cells Fetal bovine serum / plasma, or human serum / plasma and platelet-rich human plasma substitute, so as to achieve the purpose of rapidly culturing and expanding human mesenchymal stem cells, especially dental pulp stem cells. Background technique [0002] Mesenchymal stem cells (mesenchymal stem cells, MSCs) are a type of adult stem cells with the ability to form osteogenesis, adipogenesis and cartilage. In addition to the ability to differentiate into multiple lines, MSCs cultured in vitro still have the following functions: (1) tissue repair ability based on the mechanism of promoting angiogenesis; (2) immune regulation based on the mechanism of paracrine; Hemato...

Claims

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Application Information

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IPC IPC(8): C12N5/0775
CPCC12N5/0664C12N2500/30C12N2501/30C12N2501/998
Inventor 郭子宽朱笑飞
Owner 北京科霖恩生物科技有限公司
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