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Preparation for enhancing differentiation capacity of chicken skeletal muscle myoblast and application thereof

A technology of myoblasts and skeletal muscles, applied in the field of bioengineering, can solve the problem of not taking into account the role of the myogenic microenvironment in the body, achieve the promotion of enlarged and functional contractile protein expression, promote muscle cell maturation, use Simple and effective effect

Active Publication Date: 2014-11-19
NORTHEAST AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Most of the existing myogenic culture systems do not take into account the effect of the myogenic microenvironment in vivo, so establishing an in vitro culture system close to the in vivo myogenic model is of great significance for the research on the mechanism of muscle development and regeneration and the application in the fields of medical treatment and animal husbandry. positive meaning

Method used

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  • Preparation for enhancing differentiation capacity of chicken skeletal muscle myoblast and application thereof
  • Preparation for enhancing differentiation capacity of chicken skeletal muscle myoblast and application thereof
  • Preparation for enhancing differentiation capacity of chicken skeletal muscle myoblast and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Embodiment 1 Acquisition and Proliferation of Chicken Skeletal Muscle Myoblasts

[0031] Fresh eggs were hatched in an incubator at 38°C and 63% humidity for 11 days. After the eggshell is sterilized by 70% ethanol, the eggshell is broken and the chicken embryo is taken out and placed in a petri dish. Use tweezers to remove the breast skin of the chicken embryo, separate the pectoralis major muscle, and remove ligaments, connective tissue and blood vessels under a dissecting microscope. Wash the pectoralis tissue 3 times with Hank's solution, shred it thoroughly, add 0.1% collagenase I and incubate at 38°C for 30 minutes, during which time it is blown every 10 minutes. Centrifuge, add Hank's solution to blow off the cells, and filter the cells with a 200-mesh stainless steel filter to obtain a single-cell suspension.

[0032] Prepare 20%, 27.5% and 40% Percoll solutions, add them into 15ml centrifuge tubes according to the concentration from high to low (the action sh...

Embodiment 2

[0034] Example 2 Obtaining of Myogenic Fibroblast Extract

[0035]Fresh eggs were hatched in an incubator at 38°C and 63% humidity for 11 days. After the eggshell is sterilized by 70% ethanol, the eggshell is broken and the chicken embryo is taken out and placed in a petri dish. Use tweezers to remove the breast skin of the chicken embryo, separate the pectoralis major muscle, and remove ligaments, connective tissue and blood vessels under a dissecting microscope. Wash the pectoralis tissue 3 times with Hank's solution, shred it thoroughly, add 0.1% collagenase I and incubate at 38°C for 30 minutes, during which time it is blown every 10 minutes. Centrifuge, add Hank's solution to blow off the cells, and filter the cells with a 200-mesh stainless steel filter to obtain a single-cell suspension. Cells were resuspended in high-sugar DMEM containing 10% FBS, 100U / ml penicillin, and 100ug / ml streptomycin, inoculated on a culture dish, cultured at 39°C for 2 hours, and adherent c...

Embodiment 3

[0037] The differentiation culture of embodiment 3 chicken skeletal muscle myoblasts

[0038] When the chicken skeletal muscle myoblasts cultured in Example 1 reached 70% confluence, the myogenic proliferation medium was removed, washed 3 times with PBS, and the A-F group medium shown in Table 1 was slowly added along the inner wall of the culture dish. Wherein, group A is the control group, only containing the myogenic differentiation medium (the myogenic differentiation medium is high-sugar DMEM containing 2% horse serum, 2mM L-glutamine, 100U / ml penicillin, 100ug / ml streptomycin ); B-F groups are experimental groups, which are myogenic differentiation medium added with at least one preparation component.

[0039] Table 1 The composition and ratio of culture medium in groups A-F

[0040] group

[0041] Each of the above groups was placed at a temperature of 39°C, CO 2 They were cultured in an incubator with a concentration of 5% for 8 days, during which the cultu...

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Abstract

The invention discloses a preparation for enhancing differentiation capacity of chicken skeletal muscle myoblast and application thereof, and belongs to the technical field of biological engineering. The preparation provided by the invention is composed of a muscle derived fibroblast extractive, pyruvate and calcium salt. The muscle derived fibroblast extractive is obtained from the steps of: conducting ultrasonic cracking on chicken muscle derived fibroblast, isolating to obtain a protein solution less than 10KDa, and conducting vacuum drying. Calcium salt and pyruvate can be calcium pyruvate. The preparation provided by the invention has the characteristics of simple and easily available composition and simple and effective usage, and can significantly improve the fusion rate of chicken skeletal muscle myoblast and promote the increase of skeletal muscle fiber diameter and expression of functional contractile protein. The preparation provided by the invention can be used for culturing myoblast; and the fusion rate of the myoblast is increased by 32.01%, the average number of nuclei in myotubes is increased by 91.63%, and average diameter of myotubes is increased by 45.59%.

Description

technical field [0001] The invention relates to a preparation for improving the differentiation ability of chicken skeletal muscle myoblasts and an application thereof, belonging to the technical field of bioengineering. Background technique [0002] Muscle tissue widely exists in animals and is one of the earliest tissues formed during embryonic development. It is a good model for studying basic life phenomena such as cell fate determination, cell differentiation, and morphogenesis. However, so far, the elucidation of the mechanism of muscle development and regeneration is only the tip of the iceberg. Although the establishment of an in vivo myogenic model simulates the muscle regeneration process well, it also adds complexity to the study of related mechanisms. Therefore, the establishment of an efficient and stable skeletal muscle cell culture system in vitro will further elucidate the development and regeneration of skeletal muscle. mechanism lays the foundation. [00...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/077
Inventor 张宇李海刘忠华连正兴
Owner NORTHEAST AGRICULTURAL UNIVERSITY
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