Improved proliferation culture medium for maintaining functions of adipose tissue-derived stromal cells

A technology for proliferating medium and stromal cells, which is applied in the field of improved medium for maintaining the function of adipose-derived mesenchymal stem cells, can solve the problems of inability to meet the requirements of cell culture meat quantity and function, decline in proliferation ability, loss of differentiation ability, etc. Differentiation potential, efficient differentiation, and the effect of accelerating cell proliferation

Pending Publication Date: 2021-03-26
NANJING AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Livestock and poultry adipose-derived mesenchymal stem cells have a continuous decline in their proliferative ability in the process of expanding culture in vitro, and the differentiation efficiency of cells in the later stage of passage decreases or even completely loses their differentiation ability
Sinc...

Method used

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  • Improved proliferation culture medium for maintaining functions of adipose tissue-derived stromal cells
  • Improved proliferation culture medium for maintaining functions of adipose tissue-derived stromal cells
  • Improved proliferation culture medium for maintaining functions of adipose tissue-derived stromal cells

Examples

Experimental program
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Effect test

Embodiment 1

[0039]Example 1 Isolation of porcine adipose-derived mesenchymal stem cells and expanded culture in vitro:

[0040] (1) Single cell isolation: freshly slaughtered piglets were soaked in 75% (volume percentage) ethanol for one minute, and the neck subcutaneous adipose tissue was taken under aseptic conditions and stored in basal culture medium. Under sterile conditions, rinse with PBS buffer containing high concentrations of penicillin and streptomycin 3 times, cut off the visible blood vessels and connective tissue in the adipose tissue, and cut the adipose tissue into 1mm 3 About small pieces, add 0.1% (mass percentage) type Ⅰ collagenase and 3% (volume percentage) penicillin-streptomycin double antibody digestion solution to digest for 90min (37°C, shaking in a water bath), and the digestion is over Then use an equal volume of complete culture solution to neutralize the digestive solution, filter through 100 μm and 40 μm cell sieves, and centrifuge at 1500 rpm for 10 minutes...

Embodiment 2

[0043] Example 2 Detection of adipogenic differentiation ability of adipose-derived mesenchymal stem cells

[0044] (1) Induced differentiation of muscle stem cells: Take the control growth medium obtained in Example 1 and the P5 generation of porcine adipose-derived mesenchymal stem cells obtained by adding bFGF to the growth medium and continue to culture until the cells are completely confluent. The DMEM / F12 differentiation medium containing 10% fetal bovine serum composed of methasone 1 μM, 3-isobutyl-1-methylxanthine 0.5 mM and insulin 10 μg / mL was changed every two days for 5 days. On the 6th day of differentiation, replace with DMEM / F12 maintenance medium containing 10% fetal bovine serum containing 10 μg / mL insulin for 2 days; on the 8th day of differentiation, replace with 10% (volume percentage) fetal bovine serum DMEM / F12 complete medium for 2 days.

[0045] (2) Oil red O detection of lipid droplet accumulation: Take cells on the 10th day of induction of adipogenic...

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Abstract

The invention provides an improved culture medium for in-vitro enlarged culture of cultured meat seed cells, namely adipose tissue-derived stromal cells. The improved culture medium is a growth culture medium added with bFGF, the total number of cells obtained by culturing for 15 days by adopting the improved culture medium is 6 times of the total number of cells obtained in a control growth culture medium, and more importantly, the adipose tissue-derived stromal cells are massively amplified in vitro and can still keep relatively high differentiation capacity after the required cell number isreached. Therefore, the improved culture medium is helpful for obtaining a large number of functional adipose tissue-derived stromal cells in vitro, and better meets the requirements of cultured meatproduction on the number of seed cells and stem cell functions.

Description

technical field [0001] The invention belongs to the technical field of stem cells and animal cell cultured meat, and in particular relates to an improved medium for maintaining the function of adipose-derived mesenchymal stem cells. Background technique [0002] Cultured meat is meat obtained by in vitro culture using livestock and poultry stem cells. It does not need to go through animal breeding, and directly uses cells to produce meat in a factory. It has been reported as an alternative to traditional livestock farming that reduces the negative impacts of existing livestock production systems on the environment and animal welfare. Currently, most research on cell-cultured meat has focused on muscle cells and their tissues, and while fat makes up only a small portion of meat, it is a key factor affecting flavor, texture, nutrition and appearance. Therefore, the production and incorporation of fats in cell-cultured meat products is of paramount importance. [0003] Cultur...

Claims

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Application Information

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IPC IPC(8): C12N5/0775
CPCC12N5/0667C12N2501/115
Inventor 周光宏宋文娟李惠侠丁世杰刘裴裴
Owner NANJING AGRICULTURAL UNIVERSITY
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