Target regulation system for dedifferentiation of somatic cells, kit and application of kit

A technology for regulating systems and somatic cells, applied in the field of cell biology, can solve the problems of difficult repetition of experiments, unclear components of the transdifferentiation system, and low efficiency.

Active Publication Date: 2018-05-22
YUNNAN JICI INSITUTE FOR REGENERATIVE MEDICINE CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Currently, Schwann cells (THOMA EC, et al, 2014), nerve cells (HU W, et al, 2015), pancreatic islet cells (Sheng The results of Ding, et al, 2015) have been reported successively, but there are still some problems in the above-mentioned studies, such as difficulty in repeating the experiment, low efficiency, unclear composition of the transdifferentiation system, and unstable effect.

Method used

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  • Target regulation system for dedifferentiation of somatic cells, kit and application of kit
  • Target regulation system for dedifferentiation of somatic cells, kit and application of kit
  • Target regulation system for dedifferentiation of somatic cells, kit and application of kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0071] Example 1 Target Regulatory System Used to Prepare Osteoblasts

[0072] 1. Isolation of skin fibroblasts

[0073] 1.1 Obtain a skin tissue block with a diameter of about 1 cm from the donor, separate the skin fibroblasts by the adherent method, and culture the separated cells in the basal culture medium: 10% fetal bovine serum (Hyclone) + 100U / ml penicillin (Sigma) + 100 μg / ml streptomycin (Sigma) + high glucose DMDM.

[0074] 1.2 Subsequent passage of cells, a large number of expansion, the number of cell generation is between the 6th passage and the 12th passage, which is used to induce the transdifferentiation into osteoblasts. The day before initiation of differentiation (Day-1), inoculate cell density at 1-2.5×10 4 / cm 2 Cultured at 37°C, 5% CO 2 in the incubator.

[0075] 2. Reprogramming of skin fibroblasts: the first stage

[0076] After the treatment in the first step above, completely replace it with the induction medium of the first stage for cell cult...

Embodiment 2

[0081] Example 2 Target Regulatory System Used to Prepare Osteoblasts

[0082] 1. Isolation of skin fibroblasts

[0083] 1.1 Obtain a skin tissue block with a diameter of about 1 cm from the donor, separate the skin fibroblasts by the wall-adhesive method, and culture the isolated cells in the basal culture medium: 10% fetal bovine serum (Hyclone) + 100U / ml penicillin (Sigma) + 100 μg / ml streptomycin (Sigma) + high glucose DMDM.

[0084]1.2 Subsequent passage of cells, a large number of expansion, the number of cell generation is between the 6th passage and the 12th passage, which is used to induce the transdifferentiation into osteoblasts. The day before initiation of differentiation (Day-1), inoculate cell density at 1-2.5×10 4 / cm 2 Cultured at 37°C, 5% CO 2 in the incubator.

[0085] 2. Activation of skin fibroblasts

[0086] 2.1 When starting transdifferentiation (Day0), completely replace the basal culture medium with the first-stage culture medium, and culture th...

Embodiment 3

[0092] Example 3 Target Regulatory System Used to Prepare Chondrocytes

[0093] 1. Isolation of skin fibroblasts

[0094] 1.1 Obtain a skin tissue block with a diameter of about 1 cm from the donor, separate the skin fibroblasts by the adherent method, and culture the separated cells in the basal culture medium: 10% fetal bovine serum (Hyclone) + 100U / ml penicillin (Sigma) + 100 μg / ml streptomycin (Sigma) + high glucose DMDM.

[0095] 1.2 Subsequent passage of cells to a large number of expansion, the number of cell generations between the 6th and 12th passages is used for the induction of transdifferentiation into chondrocytes. The day before initiation of differentiation (Day-1), inoculate cell density at 1-2.5×10 4 / cm 2 Cultured at 37°C, 5% CO 2 in the incubator.

[0096] 2. Reprogramming of skin fibroblasts

[0097] After the treatment in the second step above, completely replace it with the second-stage culture medium for cell culture. The culture time is 6-12 day...

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Abstract

The invention discloses a target regulation system for dedifferentiation of somatic cells, a kit and an application of the kit. The target regulation system regulates the somatic cells: inhibition ofPKC activity, inhibition of DNMT activity and/or inhibition of HMT activity and/or inhibition of the activity of histone demethylase; corresponding regulation key targets include PKC, DNMT and/or HMTand/or histone demethylase; the somatic cells are differentiated cells. The target regulation system provided by the invention can promote the efficiency of adopting small molecules to induce the somatic cells to transdifferentiate into different types of functional target cells, and has the advantages of clear targets, clear ingredients, maturity and stability, and easy quality control and large-scale production.

Description

technical field [0001] The invention relates to the fields of cell biology, tissue engineering and regenerative medicine, in particular to a target regulation system, kit and application for somatic cell dedifferentiation. Background technique [0002] Stem cells are the source of the human body and its various tissue cells. They are of great significance in the research of cell therapy of human diseases, regenerative medicine, tissue engineering, drug development, anti-aging, beauty and many other fields. However, due to the ethical issues and potential safety hazards of embryonic stem cells; the scarcity of various types of adult stem cells, limited sources, difficulty in enrichment, complicated acquisition process, restrictions on the health status of donors, easy aging of cells, and the inability to expand a large number of generations, etc., Therefore, they have not been widely used. A variety of differentiated cells such as skin fibroblasts have the advantages of abun...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/077
CPCC12N2501/01C12N2501/415C12N2501/15C12N2501/72C12N5/0656C12N2501/727
Inventor 胡敏李燕皎
Owner YUNNAN JICI INSITUTE FOR REGENERATIVE MEDICINE CO LTD
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