Self-repairing piezoelectric hydrogel as well as preparation method and application thereof

Abstract

The invention belongs to the field of piezoelectric materials, and discloses self-repairing piezoelectric hydrogel as well as a preparation method and application thereof. The preparation method comprises the following steps: dissolving hyaluronic acid in water, adding sodium periodate, and carrying out a light-shielding reaction, dialysis and freeze drying to obtain formylated hyaluronic acid; the preparation method comprises the following steps: preparing a hydroformylated hyaluronic acid solution, adding 1-ethyl-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide to carry out activation reaction, grafting dopamine hydrochloride, dialyzing and freeze-drying to obtain dopamine-hydroformylated hyaluronic acid, and preparing a dopamine-hydroformylated hyaluronic acid solution; mixing the dopamine-aldehyde hyaluronic acid solution, the carboxymethyl chitosan solution and glycine crystals, and stirring at room temperature to form gel, so as to obtain the self-repairing piezoelectric hydrogel. The preparation method is simple, easy to operate and mild in gel forming condition, and the prepared hydrogel has good piezoelectric, self-repairing and adhesion functions and can be applied to preparation of bioelectrical stimulation piezoelectric materials.

Description

Technical Field

[0001] The invention belongs to the field of piezoelectric materials and relates to a self-repairing piezoelectric hydrogel, a preparation method and application thereof. Background Art

[0002] Spinal cord injury (SCI) is a devastating disease that causes partial or complete loss of sensory and / or motor function, severely affecting the patient's quality of life. After SCI, an inhibitory environment gradually forms at the lesion site, leading to impaired axonal regeneration. In the past few decades, many research strategies for the treatment of SCI have been explored, however, to date there is no effective treatment that can repair the damaged spinal cord. Electrical stimulation (ES) has been shown to promote axonal regeneration and upregulate gene expression of brain-derived neurotrophic factor (BDNF) and its receptor tropomyosin receptor kinase C (TrkC) in motor neurons, which is associated with accelerated axonal regeneration. However, conventional electrical stimulation is performed through implanted electrodes, which is traumatic, and the electrodes are in continuous contact with neural tissue, which is prone to cause interface effects, secondary injury, and inflammatory responses. Unlike conventional ES treatment, the use of piezoelectric materials can provide ES without electrodes and external power supplies. Piezoelectric materials can generate electrical stimulation when mechanically compressed and bent. This mechanical deformation caused by physiological movements (such as posture changes) has also been observed in the spinal cord. Once transplanted into the spinal cord, electrical stimulation may be activated through spinal cord deformation without the need for an external power source. However, most existing piezoelectric materials have many problems in the application of spinal cord injuries, such as poor biodegradability, loose fit with the short end of the spinal cord, and mismatch of Young's modulus, which limit their application in vivo.

[0003] At present, the development of tissue engineering represented by hydrogels has brought hope for the repair of spinal cord injuries. The injectability of hydrogels makes it easier for them to reach the site of spinal cord defects, thereby fitting the stumps of the spinal cord and providing an extracellular matrix (ECM) to guide axon regeneration while filling the spinal cord defect. It is worth noting that the life of hydrogels can be shortened by deformation or damage. In recent years, self-healing hydrogels have attracted widespread attention in the field of tissue engineering. The network structure and function of self-healing hydrogels can self-repair after damage, thereby increasing their lifespan. Summary of the invention

[0004] In view of this, the purpose of the present invention is to provide a self-healing piezoelectric hydrogel, a preparation method and application thereof. The preparation method is simple and easy to operate, the gelling conditions are mild, and the prepared hydrogel has good piezoelectric, self-healing and adhesion functions.

[0005] A first aspect of the present invention provides a method for preparing a self-repairing piezoelectric hydrogel, comprising the following steps:

[0006] S1. dissolving hyaluronic acid in water and adding sodium periodate, and obtaining aldehyde-modified hyaluronic acid after light-protected reaction, dialysis and freeze-drying;

[0007] S2. dissolving the aldehyde-formylated hyaluronic acid in water to obtain a aldehyde-formylated hyaluronic acid solution, adding 1-ethyl-(3-dimethylaminopropyl)carbodiimide and N-hydroxysuccinimide to the aldehyde-formylated hyaluronic acid solution for activation reaction, grafting dopamine hydrochloride, dialyzing and freeze-drying to obtain dopamine-aldehyde-formylated hyaluronic acid, and dissolving the dopamine-aldehyde-formylated hyaluronic acid in water to obtain a dopamine-aldehyde-formylated hyaluronic acid solution;

[0008] S3. Mix the dopamine-formylated hyaluronic acid solution, carboxymethyl chitosan solution and glycine crystals, and stir them into gel at room temperature to obtain the self-healing piezoelectric hydrogel.

[0009] Furthermore, in step S1, the usage ratio of hyaluronic acid, sodium periodate and water is 10 mg:7 mg:1 mL.

[0010] Furthermore, in step S1, the light-proof reaction is specifically: after the light-proof reaction for 8-10 hours, ethylene glycol is added to terminate the reaction.

[0011] Furthermore, in step S1, the dialysis is specifically: dialysis with distilled water for 3 days.

[0012] Furthermore, in step S2, the pH of water is 4.8-5.2, and the concentration of the aldehyded hyaluronic acid solution is (1.25-3.75) mg / mL.

[0013] Furthermore, in step S2, the mass ratio of the formaldehyded hyaluronic acid, 1-ethyl-(3-dimethylaminopropyl)carbodiimide, N-hydroxysuccinimide and dopamine hydrochloride is (1.25-3.75):(4.31-12.93):(2.58-7.74):(2.56-7.68).

[0014] Furthermore, in step S2, the dialysis is specifically: dialysis with water at pH 4.8-5.2 for 48 hours, and then dialysis with distilled water for 12 hours.

[0015] Furthermore, in step S3, the concentration of the dopamine-aldehyde-hydrated hyaluronic acid solution is 20 mg / mL; the concentration of the carboxymethyl chitosan solution is 42 mg / mL; and the dosage ratio of the glycine crystals, the dopamine-aldehyde-hydrated hyaluronic acid solution and the carboxymethyl chitosan solution is (50-150) mg:0.5 mL:0.5 mL.

[0016] Furthermore, in step S4, the usage ratio of glycine crystals, the dopamine-aldehyded hyaluronic acid solution and the carboxymethyl chitosan solution is 50 mg: 0.5 mL: 0.5 mL.

[0017] Furthermore, in step S4, the gelation time of the glycine crystals and the DA-HA-CHO / CMCS hydrogel solution is 150-360S.

[0018] The second aspect of the present invention also provides a self-repairing piezoelectric hydrogel prepared according to the above preparation method.

[0019] The third aspect of the present invention also provides a use of the self-repairing piezoelectric hydrogel in the preparation of bioelectric stimulation piezoelectric materials.

[0020] A fourth aspect of the present invention further provides a spinal cord repair product comprising the following independently packaged components:

[0021] Dopamine-aldehyded hyaluronic acid or its solution preparation;

[0022] Carboxymethyl chitosan or its solution preparation;

[0023] and glycine crystals;

[0024] The dopamine-formylated hyaluronic acid is obtained by adding 1-ethyl-(3-dimethylaminopropyl)carbodiimide and N-hydroxysuccinimide to the formylated hyaluronic acid solution for activation reaction, then grafting dopamine hydrochloride, dialysis and freeze-drying.

[0025] When the components are independently packaged dopamine-aldehyded hyaluronic acid, carboxymethyl chitosan and glycine crystals, dopamine-aldehyded hyaluronic acid and carboxymethyl chitosan are added to water to prepare a solution, which is then mixed with glycine crystals and immediately injected into the injured part of the spinal cord.

[0026] When the components are dopamine-aldehyde hyaluronic acid solution preparation, carboxymethyl chitosan solution preparation and glycine crystals, when used, the dopamine-aldehyde hyaluronic acid solution preparation, carboxymethyl chitosan solution preparation and glycine crystals are directly mixed and then immediately injected into the injured part of the spinal cord.

[0027] Compared with the prior art, the present invention has the following technical effects:

[0028] (1) The present invention uses natural macromolecular hyaluronic acid (HA) and chitosan (CS) as base materials, adds dopamine (DA) to enhance adhesion, and forms a reversible covalent cross-linking network through Schiff base reaction. The synthesized hydrogel has good biodegradability, biocompatibility, injectability, adhesion and self-repairing effects.

[0029] (2) The present invention designs a self-healing piezoelectric hydrogel doped with an appropriate amount of glycine crystals, and the prepared self-healing piezoelectric hydrogel not only has a self-healing effect, but also has piezoelectric properties. At the same time, the hydrogel can be endowed with more dynamic Schiff base bonds, thereby enhancing adhesion, making it easier to fit the stump of the spinal cord, and self-healing when damaged, thereby increasing the service life and promoting the repair of the spinal cord. It has excellent application prospects in the field of spinal cord repair materials.

[0030] (3) The preparation method provided by the present invention is simple and effective, easy to operate, and has mild gelling conditions, which is suitable for industrial application. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] These and / or other aspects and advantages of the present invention will become apparent and readily understood from the following description of the preferred embodiments in conjunction with the accompanying drawings, in which:

[0032] Figure 1 The self-healing experimental diagrams of the hydrogels prepared in Example 1, Example 2, Example 3 and the comparative example are shown;

[0033] Figure 2 The ultrasonic piezoelectric test diagrams of the hydrogels prepared in Example 1, Example 2, Example 3 and the comparative example are shown;

[0034] Figure 3 The glass slide adhesion performance graph of the hydrogels prepared in Example 1, Example 2, Example 3 and the comparative example is shown;

[0035] Figure 4 SEM images of the hydrogels of Example 1, Example 2, Example 3 and the comparative example are shown. DETAILED DESCRIPTION

[0036] The technical solution of the present invention is further specifically described below by examples and in conjunction with the accompanying drawings. In the specification, the same or similar reference numerals indicate the same or similar components. The following description of the embodiments of the present invention with reference to the accompanying drawings is intended to explain the overall inventive concept of the present invention and should not be construed as a limitation of the present invention.

[0037] Example 1

[0038] 1) Weigh 1 g of hyaluronic acid and dissolve it in 100 mL of distilled water. Dissolve it in a 40°C water bath to obtain a hyaluronic acid solution. Weigh 0.7 g of sodium periodate and add it to the hyaluronic acid solution. React in the dark for 8 h. Add 2 mL of ethylene glycol to terminate the reaction. Dialyze with distilled water for 3 days and freeze-dry to obtain aldehyde-modified hyaluronic acid (HA-CHO).

[0039] 2) Add hydrochloric acid to distilled water to obtain water with a pH of 4.8-5.2, weigh 0.25 g of aldehyded hyaluronic acid and dissolve it in 200 mL of water with a pH of 4.8-5.2 to obtain a aldehyded hyaluronic acid solution, weigh 0.8625 g of 1-ethyl-(3-dimethylaminopropyl)carbodiimide (EDC) and 0.5175 g of N-hydroxysuccinimide (NHS) and add them to the aldehyded hyaluronic acid solution, react for 15 minutes, then add 0.5125 g of dopamine hydrochloride and react for 8 hours, dialyze with water of about pH 5.0 for 48 hours, dialyze with distilled water for 12 hours, and freeze-dry to obtain dopamine-aldehyded hyaluronic acid (DA-HA-CHO).

[0040] 3) Drop a 100 mg / mL glycine aqueous solution on a glass slide and air-dry to obtain glycine crystals, and prepare a 20 mg / mL dopamine-aldehyded hyaluronic acid solution and a 42 mg / mL carboxymethyl chitosan solution (CMCS) with distilled water.

[0041] 4) 0.5 mL of dopamine-aldehyded hyaluronic acid solution, 0.5 mL of carboxymethyl chitosan solution and 50 mg of glycine crystals were mixed and stirred at room temperature for 240 s to form a gel, thereby obtaining a glycine piezoelectric hydrogel with a glycine crystal content of 50 mg / mL.

[0042] Example 2

[0043] 1) Weigh 1 g of hyaluronic acid and dissolve it in 100 mL of distilled water. Dissolve it in a 40°C water bath to obtain a hyaluronic acid solution. Weigh 0.7 g of sodium periodate and add it to the hyaluronic acid solution. React in the dark for 8 h. Add 2 mL of ethylene glycol to terminate the reaction. Dialyze with distilled water for 3 days and freeze-dry to obtain aldehyde-modified hyaluronic acid (HA-CHO).

[0044] 2) Add hydrochloric acid to distilled water to obtain water with a pH of about 4.8-5.2, weigh 0.25 g of aldehyded hyaluronic acid and dissolve it in 200 mL of water with a pH of 4.8-5.2 to obtain a aldehyded hyaluronic acid solution, weigh 0.8625 g of 1-ethyl-(3-dimethylaminopropyl)carbodiimide (EDC) and 0.5175 g of N-hydroxysuccinimide (NHS) and add them to the aldehyded hyaluronic acid solution, react for 10 minutes, then add 0.5125 g of dopamine hydrochloride and react for 8 hours, dialyze with water with a pH of about 4.8-5.2 for 48 hours, dialyze with distilled water for 12 hours, and freeze-dry to obtain dopamine-aldehyded hyaluronic acid (DA-HA-CHO).

[0045] 3) Drop a 100 mg / mL glycine aqueous solution on a glass slide and air-dry to obtain glycine crystals, and prepare a 20 mg / mL dopamine-aldehyded hyaluronic acid solution and a 42 mg / mL carboxymethyl chitosan solution (CMCS) with distilled water.

[0046] 4) 0.5 mL of dopamine-aldehyded hyaluronic acid solution, 0.5 mL of carboxymethyl chitosan solution and 100 mg of glycine crystals were mixed and stirred at room temperature for 180 s to form a gel, thereby obtaining a glycine piezoelectric hydrogel with a glycine crystal content of 100 mg / mL.

[0047] Example 3

[0048] 1) Weigh 1 g of hyaluronic acid and dissolve it in 100 mL of distilled water. Dissolve it in a 40°C water bath to obtain a hyaluronic acid solution. Weigh 0.7 g of sodium periodate and add it to the hyaluronic acid solution. React in the dark for 8 h. Add 2 mL of ethylene glycol to terminate the reaction. Dialyze with distilled water for 3 days and freeze-dry to obtain aldehyde-modified hyaluronic acid (HA-CHO).

[0049] 2) Add hydrochloric acid to distilled water to obtain water with a pH of about 4.8-5.2, weigh 0.75 g of aldehyded hyaluronic acid and dissolve it in 200 mL of water with a pH of 4.8-5.2 to obtain a aldehyded hyaluronic acid solution, weigh 2.5875 g of 1-ethyl-(3-dimethylaminopropyl)carbodiimide (EDC) and 1.5525 g of N-hydroxysuccinimide (NHS) and add them to the aldehyded hyaluronic acid solution, react for 10 minutes, then add 1.5375 g of dopamine hydrochloride and react for 8 hours, dialyze with water with a pH of about 5.0 for 48 hours, dialyze with distilled water for 12 hours, and freeze-dry to obtain dopamine-aldehyded hyaluronic acid (DA-HA-CHO).

[0050] 3) Drop a 100 mg / mL glycine aqueous solution on a glass slide and air-dry to obtain glycine crystals, and prepare a 20 mg / mL dopamine-aldehyded hyaluronic acid solution and a 42 mg / mL carboxymethyl chitosan solution (CMCS) with distilled water.

[0051] 4) 0.5 mL of dopamine-aldehyded hyaluronic acid solution, 0.5 mL of carboxymethyl chitosan solution and 150 mg of glycine crystals were mixed, and stirred at room temperature for 150 s to form a gel, thereby obtaining a glycine piezoelectric hydrogel with a glycine crystal content of 150 mg / mL.

[0052] Comparative Example (without addition of glycine crystals)

[0053] 1) Weigh 1 g of hyaluronic acid and dissolve it in 100 mL of distilled water. Dissolve it in a 40°C water bath to obtain a hyaluronic acid solution. Weigh 0.7 g of sodium periodate and add it to the hyaluronic acid solution. React in the dark for 8 h. Add 2 mL of ethylene glycol to terminate the reaction. Dialyze with distilled water for 3 days and freeze-dry to obtain aldehyde-modified hyaluronic acid (HA-CHO).

[0054] 2) Add hydrochloric acid to distilled water to obtain water with a pH of about 4.8-5.2, weigh 0.75 g of aldehyded hyaluronic acid and dissolve it in 200 mL of water with a pH of 4.8-5.2 to obtain a aldehyded hyaluronic acid solution, weigh 2.5875 g of 1-ethyl-(3-dimethylaminopropyl)carbodiimide (EDC) and 1.5525 g of N-hydroxysuccinimide (NHS) and add them to the aldehyded hyaluronic acid solution, react for 15 minutes, then add 1.5375 g of dopamine hydrochloride and react for 8 hours, dialyze with water with a pH of about 5.0 for 48 hours, dialyze with distilled water for 12 hours, and freeze-dry to obtain dopamine-aldehyded hyaluronic acid (DA-HA-CHO).

[0055] 3) Prepare 20 mg / mL dopamine-aldehyded hyaluronic acid solution and 42 mg / mL carboxymethyl chitosan solution (CMCS) with distilled water respectively.

[0056] 4) 0.5 mL of dopamine-aldehyded hyaluronic acid solution and 0.5 mL of carboxymethyl chitosan solution were mixed and stirred at room temperature for 360 s to form a gel to obtain DA-HA-CHO / CMCS hydrogel.

[0057] Experimental Example 1

[0058] The self-repairing piezoelectric hydrogels of Examples 1-3 and the comparative example were subjected to self-repairing experiments. The results are as follows: Figure 1 As shown, the hydrogels all have self-healing properties, and the 50 mg / mL glycine piezoelectric hydrogel exhibits better self-healing properties. The hydrogel morphology is complete after repair.

[0059] Experimental Example 2

[0060] The self-healing piezoelectric hydrogels obtained in Examples 1, 2 and 3 and the comparative example DA-HA-CHO / CMCS hydrogel were subjected to ultrasonic piezoelectric tests. The results are as follows: Figure 2 As shown, the control example without adding glycine crystals has no ultrasonic piezoelectric performance, and the piezoelectric voltage of the embodiment increases with the increase of the concentration of glycine crystals added. The 150 mg / mL glycine piezoelectric hydrogel has the highest piezoelectric performance, but there is no significant difference in the piezoelectric performance between the 50 mg / mL and 100 mg / mL glycine piezoelectric hydrogels.

[0061] Experimental Example 3

[0062] The hydrogels of Examples 1-3 and Comparative Examples were subjected to adhesion tests. Figure 3 The results showed that the addition of an appropriate amount of glycine crystals would increase the adhesion strength of the hydrogel, but as the concentration of glycine crystals increased, the adhesion strength of the hydrogel decreased. Therefore, it can be seen that the optimal concentration of glycine crystals added is 50 mg / mL.

[0063] Experimental Example 4

[0064] The hydrogel morphology test of Examples 1-3 and Comparative Example, Figure 4 The results show that the porous structure of the 50 mg / mL glycine piezoelectric hydrogel obtained in Example 1 is more obvious than that in Comparative Example 1, but as the concentration of glycine crystals increases, the porous structure of the hydrogel decreases, and the 150 mg / mL glycine piezoelectric hydrogel has almost no porous structure. This shows that although glycine can give the hydrogel more dynamic Schiff base bonds and promote the formation of cross-linking bonds, adding too many glycine crystals will affect its cross-linking density.

[0065] In summary, 50 mg / mL glycine piezoelectric hydrogel has the most comprehensive performance, that is, it has suitable piezoelectric properties while having the best self-healing ability and adhesion properties.

[0066] A person skilled in the art may make various other specific modifications and combinations based on the technical inspirations disclosed in the present invention without departing from the essence of the present invention, and these modifications and combinations are still within the protection scope of the present invention.

Claims

1. A method for preparing a self-repairing piezoelectric hydrogel, characterized in that: The preparation method comprises the following steps: S1. dissolving hyaluronic acid in water and adding sodium periodate, and obtaining aldehyde-modified hyaluronic acid after light-protected reaction, dialysis and freeze-drying; S2. dissolving the aldehyde-formylated hyaluronic acid in water to obtain a aldehyde-formylated hyaluronic acid solution, adding 1-ethyl-(3-dimethylaminopropyl)carbodiimide and N-hydroxysuccinimide to the aldehyde-formylated hyaluronic acid solution for activation reaction, grafting dopamine hydrochloride, dialyzing and freeze-drying to obtain dopamine-aldehyde-formylated hyaluronic acid, and dissolving the dopamine-aldehyde-formylated hyaluronic acid in water to obtain a dopamine-aldehyde-formylated hyaluronic acid solution; S3. Mix the dopamine-formylated hyaluronic acid solution, carboxymethyl chitosan solution and glycine crystals, and stir them into gel at room temperature to obtain the self-healing piezoelectric hydrogel.

2. The preparation method according to claim 1, characterized in that: In step S1, the usage ratio of hyaluronic acid, sodium periodate and water is 10 mg:7 mg:1 mL.

3. The preparation method according to claim 1, characterized in that: In step S2, the pH of water is 4.8-5.2, and the concentration of the aldehyded hyaluronic acid solution is 1.25-3.75 mg / mL.

4. The preparation method according to claim 3, characterized in that: In step S2, the mass ratio of the aldehyde-modified hyaluronic acid, 1-ethyl-(3-dimethylaminopropyl)carbodiimide, N-hydroxysuccinimide and dopamine hydrochloride is (1.25-3.75):(4.31-12.93):(2.58-7.74):(2.56-7.68).

5. The preparation method according to claim 1, characterized in that: In step S3, the concentration of the dopamine-aldehyde hyaluronic acid solution is 20 mg / mL; the concentration of the carboxymethyl chitosan solution is 42 mg / mL; the usage ratio of glycine crystals, the dopamine-aldehyde hyaluronic acid solution and the carboxymethyl chitosan solution is (50-150) mg:0.5 mL:0.5 mL.

6. The preparation method according to claim 5, characterized in that: In step S3, the usage ratio of glycine crystals, the dopamine-aldehyded hyaluronic acid solution and the carboxymethyl chitosan solution is (50-150) mg:0.5 mL:0.5 mL.

7. The preparation method according to claim 6, characterized in that: In step S3, the stirring time for gelling is 150-360 seconds.

8. A self-repairing piezoelectric hydrogel prepared according to the preparation method according to any one of claims 1 to 7.

9. Use of the self-repairing piezoelectric hydrogel according to claim 8 in preparing bioelectric stimulation piezoelectric materials.

10. A product for spinal cord repair, comprising the following independently packaged components: Dopamine-aldehyded hyaluronic acid or its solution preparation; Carboxymethyl chitosan or its solution preparation; and glycine crystals; The dopamine-formylated hyaluronic acid is obtained by adding 1-ethyl-(3-dimethylaminopropyl)carbodiimide and N-hydroxysuccinimide to the formylated hyaluronic acid solution for activation reaction, then grafting dopamine hydrochloride, dialysis and freeze-drying.

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