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A method for preparing cell membrane sheet using sodium alginate hydrogel

A sodium alginate and cell membrane technology, which is applied in prosthesis, medical science and other directions, can solve the problems of difficulty in obtaining seed cells, limited popularization and application, complicated technical operation, etc., and achieves easy cell adhesion and value-added, good application prospects, and operation steps. simple effect

Inactive Publication Date: 2018-10-02
SUN YAT SEN UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] However, the current CST is mainly achieved by raising cells on the temperature-sensitive surface and then changing the temperature to separate the base from the cells, or by modifying the surface with special proteins, which has defects such as complicated technical operations. Like other tissue engineering methods, the application of cell membranes When using cell sheet technology for tissue reconstruction, it is difficult to obtain seed cells; when the superimposed cell sheet exceeds a certain thickness, it often leads to necrosis of internal cells; the application of cell sheet technology to construct large functional tissue structures is still a challenge for cell sheet technology. Big problem, which limits its popularization and application

Method used

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  • A method for preparing cell membrane sheet using sodium alginate hydrogel
  • A method for preparing cell membrane sheet using sodium alginate hydrogel
  • A method for preparing cell membrane sheet using sodium alginate hydrogel

Examples

Experimental program
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Effect test

Embodiment 1

[0066] Example 1 Oxidation of sodium alginate

[0067] Sodium alginate is a polysaccharide with a cis-o-diol structure. When reacting with sodium periodate, sodium periodate can promote the C-C bond breakage of sodium alginate, and oxidize the adjacent diol structure into a more reactive adjacent aldehyde structure. The introduction of the new functional group aldehyde group provides more means for the surface modification of sodium alginate, which is a commonly used modification method for biomaterials.

[0068] 1. Weigh 0.2 g of sodium alginate and dissolve it in 10 mL of pure water. A certain amount of sodium periodate was added, and the reaction was stirred for 1 h in the dark, and the stirring speed was 300 rpm. After 1 h, add ethylene glycol in the amount of sodium periodate and other substances to the solution, and continue the reaction for 0.5 h. After the reaction was completed, the above solution was added to the dialysis bag for dialysis for 3 days, during which ...

Embodiment 2

[0071] Example 2 Preparation of Sodium Alginate Hydrogel Base Material

[0072] 1. Weigh a certain mass of sodium alginate and an equal mass of OSA80 (prepared in Example 1, oxidized sodium alginate) to form a mixed solution with a total sodium alginate concentration of 2% w / v. At the same time, add Add sodium chloride to make the final concentration of sodium chloride 1 mol / L. After the above solution is mixed evenly, add 0.5 mL per well to a 6-well culture plate, freeze at -80°C overnight, and then freeze-dry in a vacuum freeze dryer. At the same time, prepare materials without adding sodium chloride and freeze them under the same conditions. dry as a control.

[0073] 2. Fix the material on the sample stage, spray it with gold, put it in the vacuum chamber of the cold field emission scanning electron microscope, and observe it at a voltage of 15 kV to obtain the SEM observation picture. Such as figure 2 As shown, where A, B are sodium alginate base materials with poroge...

Embodiment 3

[0075] Example 3 Surface Grafting of COL-I on Sodium Alginate Scaffold

[0076] 1. Submerge the material prepared in Example 2 in dehydrated ethanol completely, then slowly add 2% w / v calcium chloride solution, after the added calcium chloride solution is equal to the volume of dehydrated ethanol, use 2% The calcium chloride solution was used to cross-link and wash the material, and finally, the calcium chloride solution remaining in the material was removed by washing with deionized water.

[0077] 2. After the material was sterilized by soaking in ethanol for 3 hours, it was air-dried naturally, and 0.5 mL of COL-I solution (1 mg / mL) dissolved in 0.2 mmol / L glacial acetic acid was added to each well, and cross-linked in a refrigerator at 4 °C for 24 hours. The bacterial water was washed 3 times to obtain the surface-modified sodium alginate hydrogel, which was the base material for the preparation of cell membranes.

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Abstract

The invention discloses a substrate material which can be used for building a cell sheet and a method for preparing the cell sheet from the substrate material. The method comprises the following steps that firstly, sodium alginate hydrogel is prepared into a sheet by a freeze drying and pore-preparing agent adding method, wherein in order to introduce a necessary reactive group, sodium alginate is required to be chemically modified, i.e. the substrate surface or the main body of the sodium alginate is modified by biomacromolecules, and the sodium alginate can be used for inoculating a cell; secondly, after the cell is inoculated on the substrate material, a culturing environment is adjusted and controlled to promote cell proliferation, generate a large number of extracellular matrixes and form an integrated cell sheet; thirdly, the substrate material, i.e. the sodium alginate is dissolved by using sodium citrate solution with certain concentration to obtain the integrated cell sheet with good biological performances. The technology has the characteristics that the substrate is dissolvable, the cell sheet is suitable for a large amount of cells, the operation is simple and the cost is low; the influence on cells by the traditional method is avoided; the biocompatibility is good.

Description

technical field [0001] The invention belongs to the field of tissue engineering. More specifically, it relates to a method for preparing cell membrane sheet using sodium alginate hydrogel. Background technique [0002] There are three elements in the construction of tissue engineering materials, namely seed cells, scaffold materials, and cytokines. In the construction of traditional tissue engineering materials, digestion with trypsin or proteolytic enzymes is usually required in the process of obtaining seed cells to obtain cell suspensions, but trypsin and proteolytic enzymes can cause damage to cells to a certain extent, resulting in The massive loss and low viability of cells is one of the reasons why many cells change shape and age gradually with the increase of passage number. At the same time, the cells are directly compounded with the scaffold material, the cell utilization rate is low, and it is difficult to form dense bone tissue. And it will also make the cell'...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C08J9/42C08J9/28C08J9/26C08J3/24C08L5/04C08B37/04A61L27/36A61L27/52A61L27/56A61L27/50
Inventor 龚逸鸿曹智楠李燕罗语溪李裕民李喜梅蒋庆
Owner SUN YAT SEN UNIV
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