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Cross-linked hydrogel for muscle stem cell culture as well as preparation method and application of cross-linked hydrogel

A technology for stem cell culture and cross-linked hydrogel, which is applied in the field of preparation of new enzymatic-physical method double-cross-linked network hydrogel, can solve the problems of strong adhesion of muscle stem cells and short retention time of growth factors, and achieve improved Effect of high biocompatibility and mechanical strength

Active Publication Date: 2021-05-11
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, growth factors have a short residence time in the medium
Therefore, there is still no good solution for natural hydrogel scaffolds that are non-toxic, have good mechanical stability, and have strong adhesion to muscle stem cells, which can promote the maintenance of stemness of muscle stem cells.

Method used

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  • Cross-linked hydrogel for muscle stem cell culture as well as preparation method and application of cross-linked hydrogel
  • Cross-linked hydrogel for muscle stem cell culture as well as preparation method and application of cross-linked hydrogel
  • Cross-linked hydrogel for muscle stem cell culture as well as preparation method and application of cross-linked hydrogel

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] Example 1: Preparation of Enzymatic-Physical Double Network Hydrogel of Collagen / ε-PL / Heparan Sulfate Proteoglycan / Alginate

[0049] according to figure 1 Preparation of hydrogel: Add 10 g of collagen to 100 mL of 0.02 mol / L acetic acid solution, stir and dissolve to obtain a collagen solution. Add 15g of alginate to the collagen solution, stir until the alginate solution is obtained to obtain a collagen / alginate solution; then add 20 μg of heparan sulfate proteoglycan to the prepared solution to obtain a collagen / alginate / heparan sulfate proteoglycan solution; add the ε-PL whose molar ratio of amino group and carboxyl group in the alginate is 1:1 to the solution, stir evenly to obtain the first slurry prepared by physical cross-linking; Add 0.1 g of TG enzyme to the slurry, and stir evenly to obtain the second slurry prepared by enzyme crosslinking. The slurry was put into a mold, cross-linked at 37°C for 12 hours, and demolded to obtain an enzymatic-physical double n...

Embodiment 2

[0050] Example 2: Preparation of Enzymatic-Physical Double Network Hydrogel of Collagen / ε-PL / Heparan Sulfate Proteoglycan / Alginate

[0051] according to figure 1 Preparation of hydrogel: Add 15 g of collagen to 100 mL of 0.05 mol / L acetic acid solution, stir and dissolve to obtain a collagen solution. Add 25g of alginate to the collagen solution, stir until the alginate solution is obtained to obtain a collagen / alginate solution; then add 50 μg of heparan sulfate proteoglycan to the prepared solution to obtain a collagen / alginate / heparan sulfate proteoglycan solution; add the ε-PL whose molar ratio of amino group and carboxyl group in the alginate is 2:1 to the solution, stir evenly to obtain the first slurry prepared by physical cross-linking; Add 1.5 g of TG enzyme into the slurry, stir evenly, and obtain the second slurry prepared by enzyme crosslinking. The slurry was put into a mold, cross-linked at 37°C for 36 hours, and demolded to obtain an enzymatic-physical double...

Embodiment 3

[0052] Example 3: Adsorption of Hydrogel Growth Factors

[0053] Adsorption experiment of growth factors: wash the enzymatically-physically cross-linked double network hydrogel (the hydrogel of Example 1 and Example 2) with PBS, and soak the obtained hydrogel in 75% ethanol for 20 Minutes, followed by repeated immersion in sterile deionized water for 5 minutes, the operation of washing ethanol with sterile water was carried out three times to remove all residual ethanol (FoodHydrocolloids, 2017, 72, 210-218), and then the hydrogel was transferred to a cell containing growth In the solution of factor vitamin C (0.05μg / mL) and bFGF (10ng / mL), it was obtained after swelling for 24 hours. Finally, the content of bFGF (450nm) and vitamin C (536nm) in the remaining solution was detected by enzyme-linked immunosorbent assay (ELISA), and the effect of hydrogel on growth was calculated according to the difference between the initial concentration of bFGF and vitamin C in the solution a...

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Abstract

The invention discloses cross-linked hydrogel for muscle stem cell culture as well as a preparation method and application of the cross-linked hydrogel, and belongs to the technical field of biological food materials. The preparation method comprises the following steps: dissolving collagen to prepare a solution, then adding a certain amount of alginate and heparan sulfate proteoglycan, and uniformly mixing the alginate and heparan sulfate proteoglycan with the collagen solution; and adding epsilon-PL and TG enzyme into the solution, uniformly conducting stirring, and putting slurry into a mold for crosslinking to obtain the hydrogel. According to the hydrogel disclosed by the invention, collagen, polylysine and heparan sulfate proteoglycan are connected through TG enzyme to form covalent crosslinking, and a compact three-dimensional 'egg box' net structure is formed through physical electrostatic interaction between polylysine and alginate. By utilizing the hydrogel, the absorption of the muscle stem cells on nutrient substances can be enhanced, and the growth of the muscle stem cells is facilitated. The bi-crosslinked network hydrogel has the potential to be used as a scaffold for muscle stem cell growth of stem cell culture meat.

Description

technical field [0001] The present invention relates to a cross-linked hydrogel used for culturing muscle stem cells and its preparation method and application, in particular to a novel enzymatic method-physical method double cross-linked network hydrogel preparation method and its application, belonging to the technical field of biological food materials . Background technique [0002] The massive consumption of meat is a major environmental burden, with more than 90% of the world's population eating meat. Stem cell cultured meat is a new technology that uses muscle cells to grow meat in vitro. Currently, the development of cultured meat has been accelerating due to its potential for sustainable production. Compared with traditional meat production, stem cell cultured meat can not only reduce the suffering of animals, but also reduce the environmental pollution caused by animal husbandry (Food & Agriculture Organization of the United Nations, 2006). At the same time, the...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C08J3/075C08J3/24C08L5/04C08L89/00C08L5/10C12N5/0775A23L13/00
CPCC08J3/075C08J3/243C12N5/0668A23L13/00C12N2533/54C12N2533/74C12N2533/70C08J2489/00C08J2305/04C08J2405/10C12N9/1044A23L17/00C12N5/0658C12N2533/30C12N2513/00C12N2527/00C12N2537/10C08J3/246C08J2389/06C08J2405/04
Inventor 胡静尹健汪翔
Owner JIANGNAN UNIV
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