A kind of hollow open gelatin cell microcarrier and its preparation method and application

A technology of cell microcarriers and microcarriers, applied in prosthetics, medical science, etc., can solve the problems of insufficient space reserved for newborn tissues, unfavorable high-density delivery of therapeutic cells, and low efficiency of delivery of cells, and achieve good biodegradability and cell compatibility, excellent biocompatibility, and good application prospects

Inactive Publication Date: 2017-04-12
SUN YAT SEN UNIV
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  • Abstract
  • Description
  • Claims
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Problems solved by technology

[0003] However, the biodegradable microcarriers studied in the past are all microspherical carriers. In vivo injection experiments of the osteoblast / microcarrier / hydrogel composite system have shown that the efficiency of this system for transporting cells is relatively low, and cells can only attach to The surface of the carrier grows in a single layer, and most of the space in the composite system is occupied by a cell-free carrier or hydrogel, which is not conducive to a large number of high-density delivery of therapeutic cells, and there is insufficient space reserved for new tissues
Although the use of porous microspheres instead of solid microspheres can partially solve this problem, because this material needs to meet the needs of injection therapy, the internal mesh structure of this porous microsphere is required to be very small, and such a small network Pores make it difficult for cells to access the center of the microcarrier sphere

Method used

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  • A kind of hollow open gelatin cell microcarrier and its preparation method and application
  • A kind of hollow open gelatin cell microcarrier and its preparation method and application
  • A kind of hollow open gelatin cell microcarrier and its preparation method and application

Examples

Experimental program
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Embodiment 1

[0060] Embodiment 1 Preparation of hollow open gelatin cell microcarriers of the present invention

[0061] 1. Preparation method

[0062] S1. Preparation of gelatin and beeswax microspheres by double emulsion method

[0063] S11. Weigh 4mg of gelatin, dissolve in 40mL of pure water, and heat to 80°C;

[0064] S12. Add 4 mg of beeswax to the gelatin solution of S11 at a rotational speed of 700 rpm, and stir for 5 minutes at a rotational speed of 700 rpm;

[0065] S13. Pour the solution obtained in S12 into edible peanut oil preheated at 60°C, and turn it at 400rpm for 15min;

[0066] S14. Pour the solution obtained in S13 into -20°C pre-cooled ice ethanol, let it stand for 15 minutes, then wash it with dioxane / acetone solution three times, put it into a 60°C oven for drying, and obtain gelatin beeswax microspheres;

[0067] The morphology of the obtained gelatin beeswax microspheres observed under the light microscope is as attached figure 1 shown.

[0068] S2. Gelatin be...

Embodiment 2

[0083] Example 2 Layer-by-layer Self-Assembly Modified Surface Biocompatibility of Hollow Open Gelatin Microcarriers

[0084] In this example, the surface biocompatibility of the hollow open gelatin cell microcarriers prepared in Example 1 was modified to further improve the surface biocompatibility of the microcarriers.

[0085] 1. Weigh 80 mg of hollow open gelatin cell microcarriers, place them in 75% ethanol solution, immerse and sterilize for 4 hours, wash with sterile PBS three times, and place them in a 12-well plate;

[0086] 2. Prepare a mixed solution containing 0.2 mg / mL casein and 4 mg / mL heparin as a negatively charged macromolecule assembly solution, and prepare a mixed solution containing 1.0 mg / mL collagen and 10 g / mL basic fibroblast growth factor as a positively charged macromolecule Assembly solution;

[0087] 3. Add 4mL casein / heparin solution to the 12-well plate where the microcarriers are placed, soak for 15 minutes, so that a layer of casein / heparin ma...

Embodiment 3

[0090] Example 3 Inoculation of bone marrow mesenchymal stem cells with hollow open gelatin microcarriers

[0091] 1. Method

[0092] (1) Place the hollow open gelatin microcarrier obtained in Example 2 in a centrifuge tube (EP tube);

[0093] (2) Digest P3 generation bone marrow mesenchymal stem cells with 0.25% trypsin, count the cells, add 0.5mL of 500,000 cells / mL cell suspension into the EP tube with hollow open gelatin microcarriers, and then Place the EP tube at 37°C, 5% CO 2 Cultivate in the incubator for 8 hours;

[0094] (3) Add 0.5mL of 500,000 cells / mL cell suspension into the EP tube equipped with hollow open gelatin microcarriers, and then place the EP tube at 37°C, 5% CO 2 Cultivate in the incubator for 8 hours;

[0095] (4) Transfer the hollow open gelatin cell microcarriers to a 6-well plate, and use basal medium (α-MEM, 10% FBS, 100 U / mL penicillin, 100 g / mL streptomycin, 0.25 g / mL amphoteric Mycin B) was cultured, and the medium was changed every other ...

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Abstract

The invention discloses a hollow open gelatin cell micro-carrier and a preparation method and an application thereof. The preparation method includes that using gelatin and bee wax as raw materials, using a double-emulsion method to prepare a gelatin-bee wax micro-ball, remodeling in ethanol / water solution, cross-linking in glutaraldehyde / ethanol solution, and extruding out a small bee wax ball from the gelatin micro-ball to obtain the hollow open gelatin micro-carrier. The hollow open gelatin cell micro-carrier uses the bee wax as pore-foaming agent and uses the double-emulsion method to prepare the gelatin micro-ball, after heating and drying ethanol / water, a dense shell layer is formed at the surface of the gelatin, and glutaraldehyde / ethanol solution is added to enable the surface to cross-link; force is exerted to extrude the micro-ball after being dried, and the small bee wax ball in the gelatin micro-ball is stressed to enable the cross-linked layer of the surface to break to obtain the hollow open shell structure gelatin micro-carrier; the micro-carrier is featured with low density and high specific surface area and enables the large-scale cell culture to be possible; the hollow open gelatin cell micro-carrier is excellent in biological degradability and biocompatibility and capable of being used as stuffing repairing material for the bone defect in the tissue engineering.

Description

technical field [0001] The invention belongs to the technical field of biological materials of tissue engineering. More specifically, it relates to a hollow open gelatin cell microcarrier and its preparation method and application. Background technique [0002] Tissue engineering and regenerative medicine technology is a new means of regeneration and repair of tissues and organs. It is an interdisciplinary subject based on chemistry, materials science, life science and engineering. And after organ transplantation, a third means of treatment for tissue and organ defect or failure. The three key factors in tissue engineering are the cultivation of seed cells, the preparation of tissue scaffolds, and the formation of cell scaffold complexes, the core of which is the cell scaffold complex composed of cells and biomaterials. Cell microcarriers are cell composite scaffolds that have been developed in the past twenty years and have been widely used. It does not change the charact...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61L27/56A61L27/22
Inventor 龚逸鸿庄良婷曹智楠唐颖舒曼李裕民李燕蒋庆
Owner SUN YAT SEN UNIV
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