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Culture system and application thereof as well as method for culturing cartilage cells

A culture method and chondrocyte technology, applied in bone/connective tissue cells, 3D culture, tissue culture, etc., can solve the problems of difficult integration of scaffold materials, low mechanical strength of composites, immune rejection, etc., to shorten the culture time, Improve the effect of excessive mechanical strength and promote proliferation

Inactive Publication Date: 2017-01-25
GUANGZHOU SALIAI STEMCELL SCI & TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, after the cell-scaffold complex is transplanted into the body, the scaffold may cause immune rejection of the recipient; the integration of the original tissue and the scaffold material is difficult, and the mechanical strength of the composite is low

Method used

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  • Culture system and application thereof as well as method for culturing cartilage cells
  • Culture system and application thereof as well as method for culturing cartilage cells
  • Culture system and application thereof as well as method for culturing cartilage cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] The cartilage tissue was divided into several small pieces and placed in 1× double antibody solution. After weighing, quickly transfer to the ultra-clean bench.

[0044] Replace 1×PBS with 10× double antibody solution, and shake the centrifuge tube containing the cartilage vigorously for 10s. Transfer the cartilage to a 100 mm Petri dish. Discard the solution with a 1ml pipette tip, add new 1×PBS, scrape the inner surface of the cartilage piece with a clean scalpel and tweezers, and discard the PBS.

[0045] Add 10 ml of CM medium to the cleaned cartilage slices, cut the cartilage slices into pieces the size of rice grains, and place them in an incubator to be digested.

[0046] Discard the CM medium and add 15 mL of 1 mg / mL type II collagenase solution. Digest on a shaker in an incubator for 14h (50rpm).

[0047] On the second day, transfer the digestion solution of digested cartilage tissue to a 50 mL centrifuge tube with a 1 mL pipette tip. After counting the ce...

Embodiment 2

[0051] 1. Pretreatment of the culture plate:

[0052] 1) Agarose-coated culture plate: Heat the agarose to dissolve before use, spread it on a 24-well plate, store it at 4°C, and use it after solidification.

[0053] 2) Gelatin matrix-coated culture dish: heat and dissolve the gelatin matrix before use, spread on a 24-well plate, and store at 4°C. Use after solidification.

[0054] 2. Preparation of gel solution

[0055] 1), preparation of chitosan colloid: configure 2% acetic acid solution; weigh 1g of chitosan powder respectively, add 10mL of acetic acid solution to magnetically stir for 1h, after the powder is completely dissolved, let it stand for 12h to discharge air bubbles, and obtain 10% chitosan solution.

[0056] 2) Preparation of sodium alginate colloid: Weigh 0.3g of sodium alginate powder, mix it with 10mL ultrapure water in a Erlenmeyer flask, heat in a water bath at 70°C, stir for 30min at the same time, stand still for 20min, repeat 2 times until completely ...

Embodiment 3

[0064] 1. Pretreatment of the culture plate:

[0065] 1) Agarose-coated culture plate: Heat the agarose to dissolve before use, spread it on a 24-well plate, store it at 4°C, and use it after solidification.

[0066] 2) Gelatin matrix-coated culture dish: heat and dissolve the gelatin matrix before use, spread on a 24-well plate, and store at 4°C. Use after solidification.

[0067] 2. Preparation of gel solution

[0068] 1), preparation of chitosan colloid: configure 2% acetic acid solution; weigh 1g of chitosan powder respectively, add 10mL of acetic acid solution to magnetically stir for 1h, after the powder is completely dissolved, let it stand for 12h to discharge air bubbles, and obtain 10% chitosan solution.

[0069] 2) Preparation of sodium alginate colloid: Weigh 0.3g of sodium alginate powder, mix it with 10mL ultrapure water in a Erlenmeyer flask, heat in a water bath at 70°C, stir for 30min at the same time, stand still for 20min, repeat 2 times until completely ...

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Abstract

The invention relates to the technical field of cell culture and in particular relates to a culture system and application thereof as well as a method for culturing cartilage cells. The culture system provided by the invention can be used for three-dimensional culture of the cartilage cells; a formula is reasonable, so that rapid proliferation of the cells can be promoted; meanwhile, the generation of fiber cartilage cells is avoided, and the ratio of transparent cartilage cells in obtained tissues is relatively high. An experiment shows that the culture system provided by the invention can shorten culture time of the cartilage cells and transparent cartilages can be obtained after the cartilage cells are cultured for 26 days.

Description

technical field [0001] The invention relates to the technical field of cell culture, in particular to a culture system, its application and a method for culturing chondrocytes. Background technique [0002] Articular cartilage damage and osteoarthritis are the most common joint diseases, and joints are easily damaged, sports injuries or aging, the current repair methods for articular cartilage damage mainly include mosaic plastic surgery, microfracture, injection of autologous chondrocytes, and these Treatments that primarily produce fibrocartilage rather than hyaline cartilage may relieve pain and improve joint function, but the long-term results are not unsatisfactory. [0003] A more promising method is to inject a large amount of mesenchymal stem cells into the joint cavity, because the cells can be used as nutritional bioactive factors to initiate regeneration activities and lead to repair of defects. However, the conditions of chondrocyte culture and fixation after tr...

Claims

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Application Information

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IPC IPC(8): C12N5/077
CPCC12N5/0655C12N2501/90C12N2513/00
Inventor 陈海佳葛啸虎王一飞万丽张维敏
Owner GUANGZHOU SALIAI STEMCELL SCI & TECH CO LTD
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