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Preparation method and application method of double-network hydrogel for three-dimensional cell culture

A three-dimensional culture, hydrogel technology, applied in general culture methods, cell culture supports/coatings, biochemical equipment and methods, etc., can solve the difficulty of taking into account cell compatibility, mechanical strength and long-term stable culture at the same time. , can not promote cell spreading, poor cell adhesion and other problems, to achieve the effect of promoting cell adhesion and proliferation, improving degradation rate, high mechanical strength

Active Publication Date: 2021-09-28
SOUTHEAST UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

This patented technology allows creators to produce both hydrogels with unique features like being able to attach cells or stimulate their growth without damaging them during surgery while also having excellent physical properties that allow it to be used effectively within various medical applications including drug deliver systems (DDS) and implants.

Problems solved by technology

The technical problem addressed in this patented text relating to improving the properties of hyaluronan polymer matrices such as supporting tissues during 3D cell culturing processes involves controllably creating different shapes from liquid crystals without causing unwanted side effects like reactants being added at random locations within each particle's interior. This requires an effective method of producing large quantities of chemically modified hyalurosaccharons called chondroitin mesh hydrous capsules (CGS).

Method used

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  • Preparation method and application method of double-network hydrogel for three-dimensional cell culture
  • Preparation method and application method of double-network hydrogel for three-dimensional cell culture
  • Preparation method and application method of double-network hydrogel for three-dimensional cell culture

Examples

Experimental program
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Effect test

Embodiment 1

[0032] (1) Preparation of Sortase A Enzymatic Crosslinking Substrate Methacrylated Hyaluronic Acid (HAMA-P)

[0033] Weigh 23.55 mg of HAMA (n(-C=C-)=50 μmol, 1 equiv) and completely dissolve it in 8 mL of ultrapure water, add 16 mL of 300 mM triethanolamine (TEOA) buffer solution (pH 8.0) and mix well, pass through N 2 15min to remove the oxygen in the solution. Subsequently, weigh 65mg P 1 (75μmol, 1.5equiv) or 76mg P 2 (75μmol, 1.5equiv) was added to the above mixture, at room temperature, N 2 The reaction was continuously stirred in the atmosphere for 24h. After the reaction was completed, a large amount of absolute ethanol was added to precipitate the reaction product, centrifuged at 5000 rpm / min for 15 min, and the obtained precipitate was dissolved in ultrapure water again. The above purification steps were repeated twice. Finally, the product was dialyzed against ultrapure water (Mw 3500 Da) for 4 days at room temperature, freeze-dried for 3 days, and stored at -...

Embodiment 2

[0042] The difference between (1) in this embodiment and embodiment 1 is: take 32.5mg P 1 (37.5μmol, 0.75equiv) or 38mg P 2 (3.75μmol, 0.75equiv) was added to the above-mentioned TEOA mixed solution containing HAMA, at room temperature N 2 Reaction in the atmosphere for 24h. HAMA-P 1 / P 2 The double bonds are not completely replaced, and can be further cross-linked by ultraviolet light to form a single-component hyaluronic acid double network hydrogel, which will not be described in this invention. Others are the same as in Example 1.

Embodiment 3

[0044] The difference between (2) in this embodiment and Example 1 is: take 1% (w / v) of HAMA-P and 5% (w / v) of GelMA and dissolve them completely in the DPBS buffer containing 0.5% LAP , adding 80 μM Sortase A enzyme, forming a gel for about 1 min, and then cross-linking with ultraviolet light for 10 s to prepare a double network hydrogel.

[0045] Others are the same as in Example 1.

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Abstract

The invention discloses a preparation method and an application method of double-network hydrogel for three-dimensional cell culture, belonging to the field of biomedical materials. The preparation method comprises the steps that firstly, a methacrylic acid hyaluronic acid (HAMA) conjugate (HAMA-P) grafted with Sortase A enzyme-specific substrate short peptide is synthesized, and then the substrate is subjected to enzymatic crosslinking with Sortase A with a certain concentration, so injectable hyaluronic acid single-network hydrogel can be obtained. The method has the advantages of easily available raw materials, mild reaction conditions, short reaction time and the like. Then, the enzyme-light double-crosslinking hyaluronic acid-gelatin double-network hydrogel is prepared; the enzymatically-crosslinked hyaluronic acid hydrogel serves as a first network to rapidly form gel, and ultraviolet-crosslinked methacrylic acid gelatin (GelMA) hydrogel serves as a second enhanced network. The double-network hydrogel prepared by the invention provides a suitable bracket and a three-dimensional microenvironment for cell adhesion growth, and has good application prospects in the aspects of injectable tissue engineering, 3D printing, three-dimensional cell culture and the like.

Description

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Claims

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Application Information

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Owner SOUTHEAST UNIV
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