Culture medium for inducing adipogenic differentiation of muscle derived stem cells of skeletal muscles, and application and adipogenic differentiation method thereof

An adipogenic differentiation and muscle-derived technology, applied in the medical field, can solve the problem of low adipogenic differentiation rate and achieve the effect of increasing adipogenic differentiation rate

Active Publication Date: 2017-05-31
GUANGZHOU SALIAI STEMCELL SCI & TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, there is no related content specifically aimed at the adipogenic differentiation of skeletal muscle myogenic stem cells. Chinese patent CN104830757A discloses a mesenchymal stem cell adipogenic differentiation medium, which uses DMEM / F12 as the basal medium, which contains FBS, Glutamine, antibiotics, indomethacin, insulin, dexamethasone, 3-isobutyl-1-methylxanthine (IBMX) and fasudil hydrochloride; Chinese patent CN105462917A discloses a periodontal ligament stem cell Adipogenic differentiation induction liquid, which also uses DMEM / F12 as a basal medium, which contains FBS, PRP, indomethacin, insulin, dexamethasone and 3-isobutyl-1-methylxanthine (IBMX); Although the induction medium mentioned above can induce the adipogenic differentiation of mesenchymal stem cells, stem cells from different sources need their own suitable induction and differentiation environment due to the difference in microenvironment. The rate is low, and it is not suitable for the adipogenic differentiation of skeletal muscle myogenic stem cells

Method used

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  • Culture medium for inducing adipogenic differentiation of muscle derived stem cells of skeletal muscles, and application and adipogenic differentiation method thereof
  • Culture medium for inducing adipogenic differentiation of muscle derived stem cells of skeletal muscles, and application and adipogenic differentiation method thereof
  • Culture medium for inducing adipogenic differentiation of muscle derived stem cells of skeletal muscles, and application and adipogenic differentiation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040]Example 1: Primary isolation and purification, subculture and identification of MDSCs

[0041] 1. Drawing materials

[0042] Take adult normal temporalis muscle specimens (taken from patients undergoing craniotomy, the patients are informed and voluntary, and the specimen collection is carried out in a regular hospital, and does not affect the patient's health), washed with PBS buffer containing double antibodies and transferred to sterile culture In a dish, cut into 1mm with ophthalmic scissors 3 Fragments of about the size, and then transferred to a 50mL centrifuge tube, blown and washed with PBS buffer 3 times, left to stand for 1 minute, then discarded the supernatant and floating tissues.

[0043] 2. Enzymolysis

[0044] Add 2 times the volume of mixed enzymes to the muscle fragments obtained above, including 2.4u / mL Dispase II, 1% type I collagenase, 2.5mmol / L CaCl 2 , mix well, and place it in a constant temperature shaker at 37°C for about 60 minutes to digest...

Embodiment 2

[0052] Embodiment 2: the method for adipogenic differentiation of the present invention

[0053] 1. Medium

[0054] (1) Contain 0.1mM 3-isobutyl-1-methylxanthine, 5mg / L insulin, 0.1mM indomethacin, 1μM dexamethasone, 5μM pioglitazone, and 10% volume fraction in high-sugar DMEM medium FBS;

[0055] (2) Contain 0.1mM 3-isobutyl-1-methylxanthine, 5mg / L insulin, 0.1mM indomethacin, 1μM dexamethasone, 10μM pioglitazone, and 10% volume fraction in high-sugar DMEM medium FBS;

[0056] (3) Contain 0.1mM 3-isobutyl-1-methylxanthine, 5mg / L insulin, 0.1mM indomethacin, 1μM dexamethasone, 15μM pioglitazone, and 10% volume fraction in high-sugar DMEM medium FBS;

[0057] 2. Differentiation induction method

Embodiment 3

[0059] Embodiment 3: comparative test of adipogenic differentiation

[0060] In order to verify the effect of adipogenic differentiation induction in the present invention, 4 control groups and 3 experimental groups were set up at the same time, and oil red O staining and expression detection of adipogenic genes PPARγ-2 and LPL were performed on the experimental groups and control groups.

[0061] Control group 1 is a negative control group, which is a conventional culture group, and the culture medium used is: high-sugar DMEM culture medium containing 10% FBS by volume.

[0062] Control group 2 is a positive control group, which is a conventional mesenchymal stem cell adipogenic induction formula, and the adipogenic induction liquid used is 0.1mM 3-isobutyl-1-methylxanthine (IBMX), 5mg / L insulin, 0.1mM Indomethacin, 1 μM dexamethasone, volume fraction 10% FBS, and the balance in high-sugar DMEM culture medium.

[0063] Control group 3 is a positive control group, with refere...

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Abstract

The invention relates to the field of medicine, and discloses a culture medium for inducing adipogenic differentiation of muscle derived stem cells of skeletal muscles, and an application and an adipogenic differentiation method thereof. The culture medium disclosed by the invention comprises 3-isobutyl-1-methylxanthine, insulin, indomethacin, dexamethasone, pioglitazone and FBS in a basic culture medium. The components of the culture medium are optimized and are induced to differentiate, and the conventional component in the existing mesenchymal stem cells is replaced by pioglitazone, so that an effect of significantly improving the adipogenic differentiation rate of the muscle derived stem cells of skeletal muscles is realized.

Description

technical field [0001] The invention relates to the medical field, in particular to a medium for inducing adipogenic differentiation of skeletal muscle myogenic stem cells, its application and adipogenic differentiation method. Background technique [0002] Skeletal muscle myogenic stem cells (muscle-derived stem cells, MDSCs) are the unique mesenchymal stem cells in adult animal or human muscle tissue. As a kind of adult mesenchymal stem cells originating from mesoderm, they have the ability of self-renewal and multiple differentiation potentials, and can differentiate into cells of blood cells, bone cells, endothelial cells, nerve cells and other lines in vitro. The functional characteristics of skeletal muscle stem cells have greatly attracted researchers from basic medicine and clinical medicine, and provided basic science researchers with a model for studying developmental biology. The repair, replacement and regeneration of skeletal muscle stem cells Capabilities also...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/077
CPCC12N5/0653C12N2500/30C12N2500/40C12N2501/30C12N2501/33C12N2506/1392
Inventor 陈海佳葛啸虎王一飞戚康艺张维敏
Owner GUANGZHOU SALIAI STEMCELL SCI & TECH CO LTD
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