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Serum-free conditioned medium hydrogel preparation for promoting wound healing and preparation method of serum-free conditioned medium hydrogel preparation

A wound healing and hydrogel technology, applied in biochemical equipment and methods, culture process, tissue culture, etc., can solve the problems of high cost, unfavorable promotion and application, slow effect, etc., achieve low cost, reduce deterioration, and promote wound repair Effect

Inactive Publication Date: 2017-04-26
THE THIRD AFFILIATED HOSPITAL OF GUANGZHOU MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, the commonly used treatment method is to cultivate stem cells and treat wound preparations, but this method is slow in effect and high in cost, which is not conducive to large-scale promotion and application

Method used

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  • Serum-free conditioned medium hydrogel preparation for promoting wound healing and preparation method of serum-free conditioned medium hydrogel preparation
  • Serum-free conditioned medium hydrogel preparation for promoting wound healing and preparation method of serum-free conditioned medium hydrogel preparation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] Example 1 EPC cell culture fluid sodium alginate hydrogel (I)

[0023] 1) Extraction of human umbilical cord blood endothelial progenitor cells: extract 60ml of umbilical cord blood from healthy newborns with a syringe, anticoagulate with sodium heparin, dilute with PBS at a volume ratio of 1:1, and slowly add Ficoll (1.077) lymphocyte separation solution containing a volume ratio of 2:1 In the centrifuge tube, use the density gradient centrifugation method, centrifuge at 2000rpm in a horizontal centrifuge for 25 minutes, and carefully absorb the mononuclear cell layer with a pipette. Then wash the cells with an equal volume of PBS, centrifuge at 2000rpm for 8 minutes, and inoculate the cells on a 25cm cell coated with human fibronectin. 2 Add an appropriate amount of endothelial cell culture medium (containing 10% fetal bovine serum) to the culture flask for culture, and culture at a saturated humidity, 5% carbon dioxide concentration, and a constant temperature of 37°...

Embodiment 2

[0028] Example 2 EPC cell culture solution sodium alginate hydrogel (Ⅱ)

[0029] The endothelial progenitor cells were subcultured according to the steps of 1) and 2) in Example 1. After culturing the cells to the third generation, when the cells grow to 70-80% confluence, place the cells in 0.5% oxygen concentration, saturated humidity, and 5% carbon dioxide concentration conditions to induce hypoxia for 36 hours, after that, replace the cell culture medium with no The serum basal culture medium was restored to 21% oxygen concentration, and cultured for 72 hours under saturated humidity and 5% carbon dioxide concentration. After centrifugal filtration, the culture medium was collected, and the sodium alginate hydrogel was prepared by the in-situ release method.

Embodiment 3

[0030] Example 3 EPC cell culture solution sodium alginate hydrogel (Ⅲ)

[0031] The endothelial progenitor cells were subcultured according to the steps of 1) and 2) in Example 1. After culturing the cells to the 5th generation, when the cells grow to 70-80% confluence, place the cells in 1.5% oxygen concentration, saturated humidity, and 5% carbon dioxide concentration conditions to induce hypoxia for 24 hours, after that, replace the cell culture medium with no The serum basal culture solution was restored to 21% oxygen concentration, and cultured for 48 hours under saturated humidity and 5% carbon dioxide concentration. After centrifugal filtration, the culture medium was collected, and the sodium alginate hydrogel was prepared by the in-situ release method.

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Abstract

The invention discloses a sodium alginate hydrogel for promoting wound healing and a preparation method of the sodium alginate hydrogel. The sodium alginate hydrogel is prepared from a serum-free medium of endothelial progenitor cells. Specifically, the preparation method comprises the following steps: 1) separating cells in a mononuclear cell layer, inoculating a culture flask with the cells and conducting primary culture, completely changing a medium on the 4th day and changing the medium once every other 2-3 days; 2) conducting subculture when the cells grow to 70-80% fusion; 3) culturing the cells to 3rd-5th generations, changing the medium and conducting hypoxia inducing; 4) conducting re-oxygenation culture on the cells obtained from induced culture after the serum-free medium is changed; and 5) conducting centrifuging and filtering, and collecting a medium, so that the serum-free conditioned cell medium, namely the sodium alginate hydrogel, is prepared. The preparation scheme is convenient and easy to operate, low in cost and high in efficiency; for wound surfaces, the sodium alginate hydrogel has a good therapeutic effect; and the sodium alginate hydrogel can be preserved for a long time.

Description

technical field [0001] The invention relates to a preparation for promoting skin wound healing, in particular to a serum-free conditioned culture solution hydrogel for promoting wound healing. Background technique [0002] Diabetes has become a chronic disease that seriously threatens human health after tumors and cardiovascular diseases. The biggest threat of diabetes to patients comes from its complications. Among them, diabetic wound has become a serious complication that seriously threatens the survival and quality of life of diabetic patients. It is characterized by refractory, susceptible to infection, and high amputation rate. complex. At present, diabetic wounds are mainly limited to traditional treatment, and the clinical treatment effect is not satisfactory. The effect of some newly developed drugs such as single growth factors is still relatively limited, and there is a lack of simple, economical, safe and efficient new drugs and biological treatments. Therefore...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K35/44A61K9/06A61P3/10A61P17/02A61P25/00C12N5/071A61K31/734
CPCA61K35/44A61K9/0002A61K9/06A61K31/734C12N5/0692C12N2500/90A61K2300/00
Inventor 李爽
Owner THE THIRD AFFILIATED HOSPITAL OF GUANGZHOU MEDICAL UNIVERSITY
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