CN111748120A - Polydopamine-doped glucan hydrogel porous scaffold, and preparation method and application thereof - Google Patents
Polydopamine-doped glucan hydrogel porous scaffold, and preparation method and application thereof Download PDFInfo
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- 238000002360 preparation method Methods 0.000 title claims abstract description 20
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- 239000000499 gel Substances 0.000 claims abstract description 26
- 229920001690 polydopamine Polymers 0.000 claims abstract description 22
- CTENFNNZBMHDDG-UHFFFAOYSA-N Dopamine hydrochloride Chemical compound Cl.NCCC1=CC=C(O)C(O)=C1 CTENFNNZBMHDDG-UHFFFAOYSA-N 0.000 claims abstract description 14
- 229960001149 dopamine hydrochloride Drugs 0.000 claims abstract description 14
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- CUGZWHZWSVUSBE-UHFFFAOYSA-N 2-(oxiran-2-ylmethoxy)ethanol Chemical compound OCCOCC1CO1 CUGZWHZWSVUSBE-UHFFFAOYSA-N 0.000 claims abstract description 3
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- 238000003756 stirring Methods 0.000 claims description 9
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- 239000007787 solid Substances 0.000 claims description 6
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- 239000003242 anti bacterial agent Substances 0.000 claims description 2
- 239000003963 antioxidant agent Substances 0.000 claims description 2
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- 206010052428 Wound Diseases 0.000 description 3
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- 238000004132 cross linking Methods 0.000 description 3
- LOKCTEFSRHRXRJ-UHFFFAOYSA-I dipotassium trisodium dihydrogen phosphate hydrogen phosphate dichloride Chemical compound P(=O)(O)(O)[O-].[K+].P(=O)(O)([O-])[O-].[Na+].[Na+].[Cl-].[K+].[Cl-].[Na+] LOKCTEFSRHRXRJ-UHFFFAOYSA-I 0.000 description 3
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- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 1
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 1
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Abstract
The invention relates to a polydopamine-doped glucan hydrogel porous scaffold, a preparation method and application thereof, and the polydopamine-doped glucan hydrogel porous scaffold is used for in-vitro 3D culture and wound repair of cells, and is prepared by the following method: the method comprises the steps of dissolving glucan by using a sodium hydroxide solution to prepare a glucan solution, adding dopamine hydrochloride into the sodium hydroxide solution to perform oxidative polymerization to generate a polydopamine solution, uniformly mixing the polydopamine solution and the polydopamine solution according to a certain proportion, adding a cross-linking agent ethylene glycol glycidyl ether, and placing the mixture in a constant temperature cabinet at 37 ℃ for 12 hours to form gel.
Description
Technical Field
The invention relates to the field of biological scaffolds, in particular to a polydopamine-doped glucan hydrogel porous scaffold, and a preparation method and application thereof.
Background
Glucan is a microbial polysaccharide consisting of glucose as a monosaccharide secreted by certain microorganisms during growth, wherein glucose units are connected with each other through glycosidic bonds. The glucan has good biocompatibility and degradability, but the glucan gel has weak adhesion capability and is easy to fall off from organisms in practical application, so that further application of the glucan is hindered.
Researches find that proteins secreted by marine life mussels through foot glands and having super-strong adhesion performance can be firmly adhered to the surfaces of various materials in humid environments such as seawater, and polydopamine is a mussel bionic material and has a structure similar to that of mussel adhesion proteins and super-strong adhesion performance.
Hydrogel scaffolds are applied to the biomedical field, and need to have not only excellent biocompatibility but also certain mechanical properties and biological functions to meet different medical fields and clinical requirements. The demand for developing a simple, quick and low-cost multifunctional hydrogel porous scaffold is still very urgent, and the technical problem to be solved when the current hydrogel porous scaffold is applied to clinical practice is urgent.
Disclosure of Invention
Aiming at the defects in the prior art, the invention aims to provide a polydopamine-doped glucan hydrogel porous scaffold, and a preparation method and application thereof.
In order to achieve the purpose, the invention provides a preparation method of a polydopamine-doped glucan hydrogel porous scaffold, which comprises the steps of dissolving solid dopamine hydrochloride in a 1.2M sodium hydroxide aqueous solution, stirring the solid dopamine hydrochloride to perform oxidation reaction at normal temperature to prepare a dopamine-hydroxide aqueous solution containing 30mg/mL of hydrochloric acid, dissolving glucan in a 1.2M sodium hydroxide aqueous solution, stirring the solution at normal temperature to perform oxidation reaction to prepare a solution containing 15% (w/v) of glucan, mixing and stirring the dopamine-hydroxide aqueous solution and the glucan aqueous solution, and adding a cross-linking agent in a volume ratio of 1: 10 to the mixed solution, wherein the volume ratio of the dopamine-hydroxide aqueous solution to the glucan aqueous solution is 1-5: 5.
Further, the crosslinking agent is ethylene glycol glycidyl ether.
The invention also provides the polydopamine-doped glucan hydrogel porous scaffold prepared by the preparation method.
The invention also provides application of the polydopamine-doped glucan hydrogel porous scaffold, and application of the polydopamine-doped glucan hydrogel porous scaffold in preparation of cell scaffolds.
As an application mode of the invention, the pore space of the cell scaffold is 5-30 μm.
The invention also provides application of the polydopamine-doped glucan hydrogel porous scaffold, and application of the polydopamine-doped glucan hydrogel porous scaffold in preparation of skin repair gel.
As an application mode of the present invention, the skin repair gel is loaded with one or more of a cell growth factor, an antibacterial agent, a humectant, a preservative, an antioxidant, an emulsifier, a thickener, a sweetener, a mucilage, an aromatic and a flavoring agent.
The invention has the following advantages: the glucan has good biocompatibility and degradability, the porous glucan gel generated after gelling can provide growth sites for cells, and the polysaccharide material can provide certain nutrient substances for the growth of the cells, so that the porous glucan gel is an ideal cell scaffold material, has good application prospects in cell 3D culture and wound repair, the Polydopamine (PDA) can obviously enhance the adhesion property of the material due to the fact that the polydopamine has functional groups (catechol groups) similar to foot muscle protein of mussel, the catechol groups of the polydopamine can interact with various surfaces through hydrogen bonds, static interaction and pi-pi accumulation, so that the gel has obvious tissue adhesion properties, and after being doped into the glucan gel, the adhesion of the cells in the gel and the separation of gel carriers on organisms can be promoted, and in addition, the mechanical property of the glucan gel can be enhanced through the doping of the polydopamine, the hydrogel porous scaffold has the advantages that the pore size of the gel is adjusted, the advantages of glucan and polydopamine are combined, the hydrogel porous scaffold with excellent performance and strong practicability is expected to be constructed, and the hydrogel porous scaffold constructed by the invention has wide application prospects in the biomedical fields of cell 3D culture, wound repair and the like.
Drawings
FIG. 1 is the storage modulus of dextran/polydopamine hydrogel scaffolds;
FIG. 2 is a scanning electron micrograph of a dextran/polydopamine hydrogel scaffold;
FIG. 3 shows LIVE/DEAD staining analysis results after co-culturing fibroblasts with dextran/dopamine hydrogel;
fig. 4 is a photograph of wound repair in rats for the blank group (sterilized PBS) and the hydrogel group (D4).
Detailed Description
The present invention will be further described in detail with reference to examples and effect examples, but the scope of the present invention is not limited thereto.
Example 1: preparation of dextran/polydopamine porous hydrogel cell scaffold
The preparation method comprises the following steps:
1) the dopamine hydrochloride sodium hydroxide aqueous solution is prepared by dissolving 30g of dopamine hydrochloride solid in 1L of 1.2M sodium hydroxide aqueous solution and carrying out oxidation reaction under the air condition (25 ℃, and magnetic stirring for 24 hours);
2) dissolving dextran with sodium hydroxide solution (1.2M) to prepare dextran solution with concentration of 15% (w/v);
3) 2mL of dopamine sodium hydroxide hydrochloride aqueous solution in step 1, 5mL of dextran solution in step 2, and 3mL of 1.2M sodium hydroxide solution were mixed together and stirred at room temperature.
4) And (3) adding 1.0mL of crosslinking agent EGDE into the mixed solution prepared in the step (3), uniformly stirring, placing in a 37 ℃ thermostat, and crosslinking for 12h to form gel, which is recorded as D2.
Example 2: preparation of dextran/polydopamine porous hydrogel cell scaffold
The preparation method comprises the following steps:
1) the dopamine hydrochloride sodium hydroxide aqueous solution is prepared by dissolving 30g of dopamine hydrochloride solid in 1L of 1.2M sodium hydroxide aqueous solution and carrying out oxidation reaction under the air condition (25 ℃, and magnetic stirring for 24 hours);
2) dissolving dextran with sodium hydroxide solution (1.2M) to prepare dextran solution with concentration of 15% (w/v);
3) 3mL of dopamine sodium hydroxide hydrochloride aqueous solution in step 1, 5mL of dextran solution in step 2 and 2mL of 1.2M sodium hydroxide solution were mixed together and stirred at room temperature.
4) And (3) adding 1.0mL of crosslinking agent EGDE into the mixed solution prepared in the step (3), uniformly stirring, placing in a 37 ℃ thermostat, and crosslinking for 12h to form gel, which is recorded as D3.
Example 3: preparation of dextran/polydopamine porous hydrogel cell scaffold
The preparation method comprises the following steps:
1) the dopamine hydrochloride sodium hydroxide aqueous solution is prepared by dissolving 30g of dopamine hydrochloride solid in 1L of 1.2M sodium hydroxide aqueous solution and carrying out oxidation reaction under the air condition (25 ℃, and magnetic stirring for 24 hours);
2) dissolving dextran with sodium hydroxide solution (1.2M) to prepare dextran solution with concentration of 15% (w/v);
3) 5mL of dopamine hydrochloride aqueous sodium hydroxide solution in the step 1 and 5mL of dextran solution in the step 2 are mixed together and stirred uniformly at room temperature.
4) And (3) adding 1.0mL of crosslinking agent EGDE into the mixed solution prepared in the step (3), uniformly stirring, placing in a 37 ℃ thermostat, and crosslinking for 12h to form gel, which is recorded as D4.
Example 4: mechanical property test of glucan/polydopamine hydrogel scaffold
The evaluation steps are as follows:
a control was set up and dextran solution was prepared at 15% (w/v) concentration by dissolving dextran with sodium hydroxide solution (1.2M), 5mL of dextran solution and 5mL of 1.2M sodium hydroxide solution were mixed together, 1.0mL of cross-linking agent EGDE was added, stirred well and placed in a 37 ℃ incubator and cross-linked for 12h to form a gel, noted D1.
Mechanical property tests of the gels with different dopamine doping amounts of four groups of D1, D2, D3 and D4 by using a rheometer show that the storage modulus of the composite gel is remarkably increased from 780Pa to 1600Pa along with the increase of the doping amount of polydopamine, and the results are shown in figure 1. The surface morphology of the composite gel is tested by a scanning electron microscope, the pore diameter of each gel is uniform, the pore diameter can be adjusted by the doping amount of polydopamine, and is 5-30 micrometers, as shown in fig. 2, the pore diameter gradually decreases with the increase of the doping amount of polydopamine, the pore diameter of the gel in a control group is the largest, the whole structure is loose, and the pore diameter of D2 is 15-25 micrometers; d3 pore diameter between 10-20 microns; the pore diameter of D4 is between 3-10 microns, so that it can be applied to scaffolds of different cell types, simulate different cell environments, and have more various applications.
Example 5: co-culture of dextran/polydopamine porous hydrogel cell scaffold and mouse fibroblast
The evaluation steps are as follows:
a control was set up and dextran solution was prepared at 15% (w/v) concentration by dissolving dextran with sodium hydroxide solution (1.2M), 5mL of dextran solution and 5mL of 1.2M sodium hydroxide solution were mixed together, 1.0mL of cross-linking agent EGDE was added, stirred well and placed in a 37 ℃ incubator and cross-linked for 12h to form a gel, noted D1.
Dividing the hydrogel of D1 and D4 into 32 parts, wherein the dry weight of each part is about 5-7 mg, putting the parts into a pore plate, disinfecting the parts with 75% alcohol for 12h, soaking and washing the parts with Phosphate Buffered Saline (PBS) for 1-2 days, and changing the solution 2-3 times per day.
The confocal dish was inoculated with a quantity of mouse fibroblasts (NIH3T3) in DMEM medium at 5% CO2After incubation for 24h at 37 ℃ under the gas condition, D1 and D4 hydrogel samples are added, and the states of fibroblasts in the confocal dish are evaluated by cell live/dead staining after the fibroblasts and the hydrogel samples are cultured for 1 day, 3 days and 5 days respectively. Cells in the experiment were changed medium every two days.
FIG. 3 shows the results of the cell pseudopodia showing adhesion extension in the fiber, and the cells are in various forms such as triangular, polygonal and fusiform. This means that the hydrogel porous scaffold constructed by the present inventors is suitable for cell adhesion proliferation and growth. Compared with the pure glucan hydrogel (D1), the dopamine-doped hydrogel (D4) is denser in cells after 5 days of culture, which indicates that the doping of dopamine can effectively promote the adhesion and proliferation of cells, and the hydrogel scaffold provides a new possibility for 3D culture of cells.
Example 6: polydopamine doped dextran hydrogel scaffolds for wound repair
The evaluation steps are as follows:
a control was set up and dextran solution was prepared at 15% (w/v) concentration by dissolving dextran with sodium hydroxide solution (1.2M), 5mL of dextran solution and 5mL of 1.2M sodium hydroxide solution were mixed together, 1.0mL of cross-linking agent EGDE was added, stirred well and placed in a 37 ℃ incubator and cross-linked for 12h to form a gel, noted D1.
SD rats of 4 months of age were randomly divided into 3 groups, and after anaesthesia with pentobarbital, a circular wound of 8mm diameter was constructed on the skin of the midline of the back of the rats. The experimental group was attached with 8 mm-sized D4 hydrogel sheet at the middle of the back of the rat, and the blank group was attached with 8 mm-sized D1 hydrogel sheet at the wound. The observation time points after operation were 0, 7, 14, 21 days, and after the wound healing was recorded by photographing, rats were euthanized by pentobarbital anesthesia.
The results in fig. 4 show that the dopamine-doped hydrogel group (D4) was effective in promoting and accelerating wound healing in rats after 21 days of culture, compared to the control group (D1). The dopamine-doped hydrogel constructed in the invention provides a new possibility for clinical medical wound dressings. And the hydrogel scaffold can be further loaded with some growth factors, antibacterial drugs and the like, so that the material has potential application value in repairing skin wounds.
Finally, it must be said here that: the above embodiments are only used for further detailed description of the technical solutions of the present invention, and should not be understood as limiting the scope of the present invention, and the insubstantial modifications and adaptations made by those skilled in the art according to the above descriptions of the present invention are within the scope of the present invention.
Claims (7)
1. A preparation method of a polydopamine-doped glucan hydrogel porous scaffold is characterized by comprising the following steps: dissolving dopamine hydrochloride solid in 1.2M sodium hydroxide aqueous solution, stirring at normal temperature to perform oxidation reaction to prepare dopamine hydrochloride aqueous solution containing 30mg/mL, dissolving glucan in 1.2M sodium hydroxide aqueous solution, stirring at normal temperature to perform oxidation reaction to prepare glucan solution containing 15% (w/v), mixing and stirring the dopamine hydrochloride aqueous solution and the glucan solution, adding a cross-linking agent which is 1: 10 in volume ratio to the mixed solution, wherein the volume ratio of the dopamine hydrochloride aqueous solution to the glucan solution is 1-5: 5.
2. A preparation method of a polydopamine-doped glucan hydrogel porous scaffold is characterized by comprising the following steps: the cross-linking agent is ethylene glycol glycidyl ether.
3. A polydopamine-doped dextran hydrogel porous scaffold, characterized in that the hydrogel porous scaffold is a gel prepared by the preparation method of the polydopamine-doped dextran hydrogel porous scaffold according to claim 1 or 2.
4. Use of a polydopamine doped dextran hydrogel porous scaffold according to claim 3 for the preparation of a cell scaffold.
5. The use of the polydopamine-doped glucan hydrogel porous scaffold according to claim 4, wherein the pores of the cell scaffold are 5-30 μm.
6. Use of a polydopamine doped dextran hydrogel porous scaffold according to claim 3 for the preparation of a skin repair gel.
7. The use of a polydopamine-doped dextran hydrogel porous scaffold according to claim 6, wherein said skin repair gel is loaded with one or more of a cell growth factor, an antibacterial agent, a humectant, a preservative, an antioxidant, an emulsifier, a thickener, a sweetener, a mucilage, an aroma and a flavoring agent.
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| CN113528424A (en) * | 2021-08-25 | 2021-10-22 | 中国人民解放军陆军特色医学中心 | Photosensitive biological material porous scaffold and application thereof |
| CN113842494A (en) * | 2021-09-10 | 2021-12-28 | 西北大学 | Injectable hemostatic crystal gel for promoting tissue regeneration and preparation method and application thereof |
| CN114230817A (en) * | 2021-11-09 | 2022-03-25 | 东华大学 | Hydrogel dressing for repairing chronic wounds of diabetic feet and preparation method thereof |
| CN115006585A (en) * | 2022-06-01 | 2022-09-06 | 楼天女 | Wound dressing with good hemostatic performance and preparation method thereof |
| CN115316405A (en) * | 2022-09-14 | 2022-11-11 | 昆山博青生物科技有限公司 | Deodorizing bactericide and preparation method thereof |
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| CN113336967A (en) * | 2021-05-13 | 2021-09-03 | 福州大学 | PH-responsive bactericidal-self-cleaning hydrogel and preparation method and application thereof |
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| CN113842494A (en) * | 2021-09-10 | 2021-12-28 | 西北大学 | Injectable hemostatic crystal gel for promoting tissue regeneration and preparation method and application thereof |
| CN114230817A (en) * | 2021-11-09 | 2022-03-25 | 东华大学 | Hydrogel dressing for repairing chronic wounds of diabetic feet and preparation method thereof |
| CN115006585A (en) * | 2022-06-01 | 2022-09-06 | 楼天女 | Wound dressing with good hemostatic performance and preparation method thereof |
| CN115316405A (en) * | 2022-09-14 | 2022-11-11 | 昆山博青生物科技有限公司 | Deodorizing bactericide and preparation method thereof |
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