Small-molecular compound combination and method for inducing differentiated cells to prepare vascular smooth muscle cells by using small-molecular compound combination

A small molecular compound and small molecule technology, applied in the field of cell biology, can solve the problems of induced cell transdifferentiation and the low feasibility of similar cell transdifferentiation

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

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Problems solved by technology

[0005] Since there are about 25% genetic differences between humans and mice, it is not feasible to apply the above-mentioned successful patent application technical scheme in mouse cell experiments to human cells of the same kind; on the other hand, due to the induction of similar cells The specific theoretical basis and technical means of transdifferentiation to obtain different target cells are not the same. The applicant used the technical solutions reported above to repeat the experiments, but failed to successfully apply the transdifferentiation technical solutions applied to mouse cells to similar human cells. Transdifferentiation, also failed to induce transdifferentiation of human differentiated cells into vascular smooth muscle cells

Method used

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  • Small-molecular compound combination and method for inducing differentiated cells to prepare vascular smooth muscle cells by using small-molecular compound combination
  • Small-molecular compound combination and method for inducing differentiated cells to prepare vascular smooth muscle cells by using small-molecular compound combination
  • Small-molecular compound combination and method for inducing differentiated cells to prepare vascular smooth muscle cells by using small-molecular compound combination

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0146] 1. Isolation of skin fibroblasts

[0147] 1.1 Obtain a skin tissue block with a diameter of about 1 cm from any position on the donor body, and isolate skin fibroblasts by the adherent method. The isolated cells are cultured in a basal culture medium. The basal culture medium refers to: 10% fetal bovine serum (Hyclone )+100 U / ml penicillin (Sigma)+100 μg / ml streptomycin (Sigma)+high sugar DMDM.

[0148] 1.2 Subsequent expansion of cells, the number of passages between the 6th and 12th passages is used to induce transdifferentiation into vascular smooth muscle cells. The day before activation (Day-1), inoculate cell density 4~6×10 3 / cm 2 Cultured at 37°C, 5% CO 2 in the incubator.

[0149] 2. Activation of skin fibroblasts

[0150] 2.1 When starting the transactivation (Day0), completely replace the basal culture medium with the first-stage culture medium A, and culture the cells for 2 to 10 days. The first-stage culture medium A refers to: 10% fetal bovine serum (...

Embodiment 2

[0159] 1. Isolation of skin fibroblasts

[0160] 1.1 Obtain a skin tissue block with a diameter of about 1 cm from any position on the donor body, and isolate skin fibroblasts by the adherent method. The isolated cells are cultured in a basal culture medium. The basal culture medium refers to: 10% fetal bovine serum (Hyclone )+100 U / ml penicillin (Sigma)+100 μg / ml streptomycin (Sigma)+high sugar DMDM.

[0161] 1.2 Subsequent expansion of cells, the number of passages between the 6th and 12th passages is used to induce transdifferentiation into vascular smooth muscle cells. The day before activation (Day-1), inoculate cell density 4~6×10 3 / cm 2 Cultured at 37°C, 5% CO 2 in the incubator.

[0162] 2. Activation of skin fibroblasts

[0163]2.1 When starting the transactivation (Day0), completely replace the basal culture medium with the first-stage culture medium, and culture the cells for 2 to 10 days. The first-stage culture medium A refers to: 10% fetal bovine serum (Hyc...

Embodiment 3

[0171] 1. Isolation of skin fibroblasts

[0172] 1.1 Obtain a skin tissue block with a diameter of about 1 cm from any position on the donor body, and isolate skin fibroblasts by the adherent method. The isolated cells are cultured in a basal culture medium. The basal culture medium refers to: 10% fetal bovine serum (Hyclone )+100 U / ml penicillin (Sigma)+100 μg / ml streptomycin (Sigma)+high sugar DMDM.

[0173] 1.2 Subsequent expansion of cells, the number of passages between the 6th and 12th passages is used to induce transdifferentiation into vascular smooth muscle cells. The day before activation (Day-1), inoculate cell density 4~6×10 3 / cm 2 Cultured at 37°C, 5% CO 2 in the incubator.

[0174] 2. Activation of skin fibroblasts

[0175] 2.1 When starting the transactivation (Day0), completely replace the basal culture medium with the first-stage culture medium, and culture the cells for 2 to 10 days. The first-stage culture medium A refers to: 10% fetal bovine serum (Hy...

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Abstract

The invention discloses a small-molecular compound combination and a method for inducing differentiated cells to prepare vascular smooth muscle cells by using the small-molecular compound combination.The small-molecular compound combination includes a first-stage small-molecular compound and a second-stage small-molecular compound used separately according to time sequence, wherein the first-stage small-molecular compound includes at least one of DNMT inhibitors and lysine deacetylases inhibitors; the second-stage small-molecular compound includes WNT / beta-catenin agonists, cAMP agonists andTG F-beta receptor inhibitors. Through staged induction of the small-molecular compounds, the differentiated cells can be differentiated into the vascular smooth muscle cells, precision quality control can be achieved in each step, and standardization operation and large-scale production are facilitated. The method provided by the invention has wide donor sources, a patient himself can be used asa donor, and the vascular smooth muscle cells needed for basic research, clinical treatment, or tissue engineering production can be obtained in a relatively short period of time.

Description

technical field [0001] The invention relates to the fields of cell biology, tissue engineering and regenerative medicine, in particular to a combination of small molecular compounds and a method for preparing vascular smooth muscle cells from cells induced by the combination of small molecular compounds. Background technique [0002] Due to the large number of patients with trauma, cardiovascular disease, organ transplantation, and hemodialysis, and the irreplaceability of autologous blood vessels, the limited source of allogeneic blood vessels, the risk of disease transmission, and immune rejection, the clinical demand for vascular substitutes huge. Vascular substitutes can be divided into two categories: polymers and biomaterials according to the stent material. Vascular substitutes should have good strength, toughness, compliance, non-bleeding, tissue compatibility, and other biological functions of normal blood vessels. Acellular matrix artificial vascular substitutes ...

Claims

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Application Information

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IPC IPC(8): C12N5/077
CPCC12N5/0661C12N2501/01C12N2501/15C12N2501/405C12N2501/415C12N2501/72C12N2501/73
Inventor 胡敏李燕皎
Owner YUNNAN JICI INSITUTE FOR REGENERATIVE MEDICINE CO LTD
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