Preparation method of gradient photo-crosslinking hydrogel, hydrogel prepared by method and nerve conduit

Abstract

The invention provides a preparation method of gradient photo-crosslinking hydrogel, the hydrogel prepared by the method and a nerve conduit. The method comprises the following steps: step 1, adding a photo-curable hydrogel precursor into a photoinitiator to prepare a hydrogel precursor solution; optionally, adding a bioactive substance or a drug to prepare a drug-loaded hydrogel precursor solution; 2, filling the hydrogel precursor solution or the drug-loaded hydrogel precursor solution prepared in the step 1 into a transparent tube, and carrying out photo-crosslinking through irradiation of a light source to obtain gradient photo-crosslinked hydrogel; wherein a variable gradient gray scale photomask with the transparency of 0-100% is arranged between the transparent tube and the light source. The gradient cross-linked hydrogel provided by the invention can be singly and orderly released, can well realize slow release of bioactive molecules and drugs, and can continuously guide cell migration for a long time when being used as a filler of a nerve conduit during nerve repair.

Description

Technical Field

[0001] The present invention relates to the technical field of biomaterials, and more particularly to a method for preparing a gradient photo-crosslinked hydrogel, the hydrogel prepared by the method, and a nerve conduit prepared by the hydrogel. Background Art

[0002] Hydrogels have extracellular matrix-like properties, which can not only promote cell movement and tissue repair, but also load drugs and bioactive substances. Hydrogels are degradable and can induce cell migration. The degradation time of hydrogels and the migration of induced cells are closely related to the degree of cross-linking. At present, researchers mainly use ultraviolet light or blue light to photo-crosslink and cure photocurable hydrogels. The cross-linking degree of hydrogels irradiated with uniform light is uniform, and the degradation rate of hydrogels with uniform cross-linking degree is the same, which cannot meet the needs of long-term gradual release of growth factors or drugs in fields such as tissue repair. Gradient photo-crosslinked hydrogels can achieve gradient degradation and can control the bioactive substances or drugs added to the hydrogel to achieve a sustained release effect. Therefore, in order to better control the sustained release of loaded drugs and bioactive substances, gradient photo-crosslinked hydrogels have gradually become the focus of research.

[0003] CN103721293A discloses a photo-crosslinked multilayer gradient hydrogel with controllable release of active factors and a preparation method. The gel structure is three layers, and the main components of each layer are modified gelatin and carboxymethyl chitosan. Three modified gelatin and modified carboxymethyl chitosan, a crosslinker and a photoinitiator are respectively configured into three reaction solutions, and the three reaction solutions are injected into a mold in turn, and a multilayer gradient hydrogel is obtained after photo-crosslinking. This multilayer gradient hydrogel can meet the requirements of tissue repair for the gradient structure, composition and controllable release of active factors of the material, but its preparation method is relatively complicated and not conducive to application. Summary of the invention

[0004] To solve the above problems, the present invention provides a method for preparing a gradient photo-crosslinked hydrogel. The hydrogel with a gradient crosslinking degree prepared by the method can be released in a single orderly manner, and can better achieve the sustained release of bioactive molecules and drugs. At the same time, as a filler for a nerve conduit, it can continuously guide cell migration for a long time during nerve repair.

[0005] First, one of the purposes of the present invention is to provide a method for preparing a gradient photocrosslinked hydrogel, the method comprising the following steps:

[0006] Step 1: firstly add a photocurable hydrogel precursor into a photoinitiator to prepare a hydrogel precursor solution; optionally, add a bioactive substance or a drug to prepare a drug-loaded hydrogel precursor solution;

[0007] Step 2: putting the hydrogel precursor solution or the drug-loaded hydrogel precursor solution prepared in step 1 into a transparent tube, and performing photo-crosslinking by irradiating a light source to obtain a gradient photo-crosslinked hydrogel;

[0008] Among them, the present invention is provided with a variable gradient grayscale light mask with a transparency of 0 to 100% between the transparent tube and the light source. The hydrogel obtained by photocrosslinking has a continuous gradient crosslinking structure, and the crosslinking degree of the hydrogel gradually increases from small to large transparency. The hydrogel starts to degrade from the part with a smaller crosslinking degree. The bioactive substances or drugs filled in the hydrogel can be released along the direction of gradually increasing crosslinking degree, realizing single-item orderly release and inducing cells to migrate from the proximal end to the distal end.

[0009] Preferably, the light source is an ultraviolet light or a blue light source.

[0010] Preferably, in step 1, the preparation conditions of the hydrogel precursor solution are: temperature of 50-60° C., heating for 10-30 min.

[0011] Preferably, in step 1, the concentration of the photocurable hydrogel in the hydrogel precursor solution is 5 to 35 wt %; more preferably 10 to 20 wt %.

[0012] Preferably, in step 1, the photocurable hydrogel precursor is selected from one or a combination of methacrylated gelatin solid, methacrylated chitosan, methacrylated silk fibroin, and methacrylated hyaluronic acid.

[0013] Preferably, in step one, the photoinitiator is selected from one or a combination of phenyl (2,4,6-trimethylbenzoyl) phosphate lithium salt, 2-hydroxy-2-methyl-1-[4-(2-hydroxyethoxy)phenyl]-1-propanone, 2-hydroxy-2-methyl-1-phenylpropanone, 1-hydroxycyclohexyl phenyl ketone, and 2,4,6-trimethylbenzoyl-diphenylphosphine oxide.

[0014] Preferably, in step 1, the bioactive substance or drug is selected from one or a combination of nerve growth factor, neurotrophic factor and deferoxamine.

[0015] Preferably, in step 1, the concentration of the bioactive substance or drug in the drug-loaded hydrogel precursor solution is 0 to 200 μg / mL; more preferably 50 to 150 μg / mL.

[0016] Secondly, the second object of the present invention is to provide a gradient photo-crosslinked hydrogel obtained by the preparation method of one of the objects of the present invention.

[0017] Again, the third object of the present invention is to provide a nerve guide, comprising the gradient photo-cross-linked hydrogel of the second object of the present invention.

[0018] Specifically, gradient photo-cross-linked hydrogels are used to fill nerve conduits, which can continuously guide cell migration for a long time during nerve repair.

[0019] Furthermore, in the present invention, the nerve conduit is prepared by the following method:

[0020] Step A, adding a high molecular polymer solution into a syringe and installing it on an electrospinning machine propeller to perform electrospinning to obtain high molecular polymer fibers. After the solvent evaporates, an electrospun fiber membrane is obtained; preferably, the thickness of the electrospun fiber membrane is 250 to 400 μm.

[0021] Step B: cutting the electrospun fiber membrane obtained in step A into the same length as the gradient photo-crosslinked hydrogel, then wrapping the electrospun fiber membrane in the gradient photo-crosslinked hydrogel, bonding the gaps with a polymer solution, and after drying, obtaining a gradient photo-crosslinked hydrogel-filled nerve conduit.

[0022] Preferably, in step A, the electrospinning conditions are:

[0023] The distance between the needle and the receiving device is 10 to 25 cm;

[0024] The propeller rate is 0.5 to 1.5 mL / h;

[0025] The ambient temperature is 20-30℃;

[0026] The voltage between the two electrodes of the electrospinning device is 15-25 kV;

[0027] The spinning time is 10 to 12 hours.

[0028] Preferably, in step A, the high molecular polymer is one or a combination of polycaprolactone, polylactic acid-glycolic acid, type I collagen, and methacryloyl gelatin.

[0029] Preferably, in step A, the mass concentration of the polymer solution is 6 to 15 wt %; more preferably 10 to 15 wt %.

[0030] Compared with the prior art, the beneficial effects of the present invention are:

[0031] 1. The preparation method of the hydrogel provided by the present invention has a simple device and a simple process. A gradient grayscale sheet with a variable range is used as a photomask to achieve gradient transmittance, thereby obtaining a hydrogel with a gradient cross-linking degree, which can be degraded in a gradient manner, thereby solving the problem of short-term burst release of growth factors.

[0032] 2. The present invention provides a simple and convenient method for filling nerve conduits with hydrogels, which can control the light transmission range and illumination time of the light-transmitting sheet according to specific application requirements. The gradient cross-linked drug-loaded hydrogel has controllable gel concentration and controllable drug release duration, has good biocompatibility, can meet the needs of nerve repair, and realize unidirectional orderly release and long-term sustained release of drugs. BRIEF DESCRIPTION OF THE DRAWINGS

[0033] Figure 1 Schematic diagram of the device for preparing gradient cross-linked hydrogel in the present invention. The gradient photomask is placed on the light generator to act as a gradient barrier for the transmitted light. Among them, 1. light generator; 2. gradient photomask; 3. gradient light intensity transmitted after being shielded by the photomask; 4. gradient cross-linked hydrogel;

[0034] Figure 2 The photomask plates with different transparency ranges in gradients prepared in Example 1 of the present invention;

[0035] Figure 3 The photomask plates with different transparency ranges in gradients prepared in Example 2 of the present invention;

[0036] Figure 4 The photomask plates with different transparency ranges in gradients prepared in Example 3 of the present invention;

[0037] Figure 5 This is a SEM image of the gradient cross-linked hydrogel prepared in Example 3 of the present invention;

[0038] Figure 6 This is a growth factor release curve of the nerve conduit prepared in Example 1 of the present invention. DETAILED DESCRIPTION

[0039] The present invention is described in detail below in conjunction with specific drawings and embodiments. It is necessary to point out that the following embodiments are only used to further illustrate the present invention and cannot be understood as limiting the scope of protection of the present invention. Some non-essential improvements and adjustments made to the present invention by those skilled in the art based on the content of the present invention still fall within the scope of protection of the present invention.

[0040] Example 1

[0041] The following is used to illustrate the preparation of gradient photo-crosslinked drug-loaded hydrogel.

[0042] Step 1: Solution preparation

[0043] Take an appropriate amount of methacryloyl gelatin GelMA (EFL; EFL-GM-60), dissolve it in 2.5wt% photoinitiator phenyl (2,4,6-trimethylbenzoyl) lithium phosphate LAP at 50°C, keep it for 20 minutes, and prepare a GelMA precursor solution with a mass fraction of 10wt%; dissolve nerve growth factor NGF in the precursor solution to obtain a hydrogel precursor solution loaded with 100μg / mL nerve growth factor NGF.

[0044] Step 2:

[0045] Construction of hydrogel mold: Take a transparent tube with an inner diameter of 1.5 mm and cut it into small tubes with a length of 1.5 cm to serve as a hydrogel molding mold.

[0046] Construction of the light-transmitting sheet: Use Word to draw a rectangle with a length of 5 cm and a width of 1 cm, and select the internal filling to be 20-100%; after printing it on film paper, cut out the rectangle along the edge and use it as a photomask template (such as Figure 2 as shown) and placed on the blue light generator.

[0047] The drug-loaded hydrogel precursor solution prepared in step 1 is slowly injected into the transparent tube. During this process, bubbles are avoided as much as possible in the tube. The drug-loaded hydrogel precursor solution is injected about 1 cm. The transparent tube containing the drug-loaded hydrogel precursor solution is placed parallel to the gradient extension direction of the photomask (such as Figure 1 As shown), turn on the 405 nm blue light generator and irradiate for 8 seconds to obtain a gradient photo-crosslinked drug-loaded hydrogel.

[0048] The following is used to illustrate the preparation of a nerve conduit filled with gradient photo-crosslinked drug-loaded hydrogel.

[0049] Step A: Preparation of electrospun fiber membrane

[0050] Polycaprolactone (Sigma-Aldrich; 24980-41-4) was dissolved in a mixed solvent of dichloromethane and N,N-dimethylformamide (mass ratio of 8:2), and stirred at room temperature to prepare a polycaprolactone solution with a mass fraction of 12wt%; the polycaprolactone solution was added to a syringe and fixed on the propeller of the electrospinning machine, the distance between the needle and the receiving device was set to 15cm, the propeller rate was set to 1mL / h, the ambient temperature was controlled at 30°C, the voltage between the two electrodes of the electrospinning device was set to 20kV, and after spinning for 12h, the polycaprolactone fibers were collected, and the residual solvent was evaporated in a fume hood to obtain an electrospun fiber membrane with an orderly orientation in the inner layer, and the fiber membrane thickness was 300μm.

[0051] Step B: Preparation of gradient photo-crosslinked drug-loaded hydrogel-filled nerve conduits

[0052] The fiber membrane prepared in step A is cut into the same length as the gradient photo-crosslinked drug-loaded hydrogel prepared in step 2, and the cut fiber membrane is tightly wrapped in the gradient photo-crosslinked drug-loaded hydrogel. The gaps are bonded with the 12wt% polycaprolactone solution of step A, and then dried in a fume hood to obtain a gradient photo-crosslinked drug-loaded hydrogel-filled nerve conduit.

[0053] Figure 6 The growth factor release curve of the nerve conduit prepared in this example, GN is the gradient cross-linked GelMA hydrogel loaded with NGF wrapped in fiber membrane; GNP is the proximal end of the gradient cross-linked GelMA hydrogel loaded with NGF wrapped in fiber membrane; GNM is the middle end of the gradient cross-linked GelMA hydrogel loaded with NGF wrapped in fiber membrane; GND is the distal end of the gradient cross-linked GelMA hydrogel loaded with NGF wrapped in fiber membrane. Figure 6 It can be seen that the proximal part initially releases from 1 to 5 days, while there is no large amount of NGF release in the middle and distal ends; the middle end releases from 2 to 9 days, and the distal end releases from 4 to 11 days. It can be seen that the gradient hydrogel loaded with NGF can achieve a unidirectional ordered slow release effect that changes with the cross-linking strength of the hydrogel.

[0054] Example 2

[0055] The following is used to illustrate the preparation of gradient photo-crosslinked drug-loaded hydrogel.

[0056] Step 1: Solution preparation

[0057] Take an appropriate amount of methacryloyl hyaluronic acid HAMA (EFL; EFL-HAMA-400K)), dissolve it in 2.5wt% photoinitiator phenyl (2,4,6-trimethylbenzoyl) lithium phosphate LAP at 60°C, keep it for 20 minutes, and prepare a HAMA precursor solution with a mass fraction of 15wt%.

[0058] Step 2:

[0059] Construction of hydrogel mold: Take a transparent tube with an inner diameter of 1.2 mm and cut it into small tubes with a length of 1.2 cm to serve as a hydrogel molding mold.

[0060] Construction of the light-transmitting sheet: Use Word to draw a rectangle of 5 cm in length and 1 cm in width, and select 10-80% for the internal filling; Print it on film paper, and cut out the rectangle with scissors. After printing on film paper, cut out the rectangle along the edge and use it as a photomask template (such as Figure 3 as shown) and placed on the blue light generator.

[0061] The hydrogel precursor solution prepared in step 1 is slowly injected into the transparent tube. During this process, bubbles are avoided as much as possible in the tube. The hydrogel precursor solution is injected about 1 cm. The transparent tube containing the hydrogel precursor solution is placed parallel to the gradient extension direction of the photomask (such as Figure 1 As shown), turn on the 405 nm blue light generator to irradiate for 15 seconds to obtain a gradient photo-crosslinked hydrogel;

[0062] The following is used to illustrate the preparation of a nerve conduit filled with a gradient photo-crosslinked hydrogel.

[0063] Step A: Preparation of electrospun fiber membrane

[0064] The polylactic acid-glycolic acid copolymer (Jinan Daigang Biotechnology Co., Ltd.; DG-50DLG150) was dissolved in trifluoroethanol and stirred at room temperature for 24 hours to prepare a polylactic acid-glycolic acid copolymer solution with a mass fraction of 10wt%; the polylactic acid-glycolic acid copolymer solution was added into a syringe and fixed on the propeller of an electrospinning machine, the distance between the needle and the receiving device was set to 15cm, the propeller rate was set to 1mL / h, the ambient temperature was controlled at 25°C, the voltage between the two electrodes of the electrospinning device was set to 17kV, and after spinning for 12h, the polylactic acid-glycolic acid fibers were collected, and the residual solvent was evaporated in a fume hood to obtain an electrospun fiber membrane with an orderly orientation in the inner layer, and the fiber membrane thickness was 300μm.

[0065] Step B: Preparation of gradient photocrosslinked hydrogel-filled nerve conduits

[0066] The fiber membrane prepared in step A is cut into the same length as the gradient photo-crosslinked hydrogel prepared in step 2, and the cut fiber membrane is tightly wrapped in the gradient photo-crosslinked hydrogel. The gaps are bonded with the 10wt% polylactic acid-glycolic acid solution of step A, and then dried in a fume hood to obtain a gradient photo-crosslinked drug-loaded hydrogel-filled nerve conduit.

[0067] Example 3

[0068] The following is used to illustrate the preparation of gradient photo-crosslinked drug-loaded hydrogel.

[0069] Step 1: Solution preparation

[0070] Methacrylylated silk protein SilMA (EFL; EFL-SilMA-001)) was dissolved in 2.5wt% photoinitiator phenyl (2,4,6-trimethylbenzoyl) lithium phosphate LAP at 55°C for 25min to prepare a SilMA precursor solution with a mass fraction of 20wt%. Nerve growth factor NGF was dissolved in the precursor solution to obtain a hydrogel precursor solution loaded with 150μg / mL nerve growth factor NGF.

[0071] Step 2:

[0072] Construction of hydrogel mold: Take a transparent tube with an inner diameter of 1.5 mm and cut it into small tubes with a length of 1.5 cm as a hydrogel molding mold;

[0073] Construction of the light-transmitting sheet: Use Word to draw a rectangle with a length of 5 cm and a width of 1 cm, and select the internal filling to be 20-80%; after printing it on film paper, cut out the rectangle along the edge and use it as a photomask template (such as Figure 4 shown);

[0074] The hydrogel precursor solution prepared in step 1 is slowly injected into the transparent tube. During this process, bubbles are avoided as much as possible in the tube. The hydrogel precursor solution is injected about 1 cm. The transparent tube containing the hydrogel precursor solution is placed parallel to the gradient extension direction of the photomask (such as Figure 1 As shown), turn on the 405 nm blue light generator and irradiate for 8 seconds to obtain a gradient photo-crosslinked drug-loaded hydrogel.

[0075] Figure 5 This is a SEM image of the hydrogel prepared in this example after freeze-drying. As can be seen from the figure, when the corresponding photomask transparency decreases from left to right during photocrosslinking, the pore size in the corresponding hydrogel gradually increases from left to right. Judging from the pore size, it can be seen that when the photomask transparency is larger, the pore size is smaller, and the degree of crosslinking is greater.

[0076] The following is used to illustrate the preparation of a nerve conduit filled with gradient photo-crosslinked drug-loaded hydrogel.

[0077] Step A: Preparation of electrospun fiber membrane

[0078] Type I collagen (Solarbio; C8060) and methacryloyl gelatin GelMA (EFL; EFL-GM-60) were dissolved in trifluoroethanol at a mass ratio of 5:5, and stirred at room temperature to prepare a mixed solution of type I collagen and GleMA with a mass fraction of 10wt%; the mixed solution of type I collagen and GleMA was added into a syringe and fixed on the propeller of an electrospinning machine, the distance between the needle and the receiving device was set to 15cm, the propeller rate was set to 1mL / h, the ambient temperature was controlled at 25℃, the voltage between the two electrodes of the electrospinning device was set to 17kV, and after spinning for 10h, the type I collagen and GleMA fibers were collected, and the residual solvent was evaporated in a fume hood to obtain an electrospun fiber membrane with an orderly orientation in the inner layer, and the fiber membrane thickness was 200μm.

[0079] Step B: Preparation of gradient photo-crosslinked drug-loaded hydrogel-filled nerve conduits

[0080] The fiber membrane prepared in step A is cut into the same length as the gradient photo-cross-linked drug-loaded hydrogel prepared in step 2, and the cut fiber membrane is tightly wrapped in the gradient photo-cross-linked drug-loaded hydrogel. The gaps are bonded with a mixed solution of 10wt% type I collagen and GleMA in step A, and then dried in a fume hood to obtain a gradient photo-cross-linked drug-loaded hydrogel-filled nerve conduit.

[0081] Example 4

[0082] The preparation method of the gradient photo-crosslinked drug-loaded hydrogel in this embodiment is the same as that in embodiment 3, except that:

[0083] Step 1: Solution preparation

[0084] A GelMA precursor solution with a mass fraction of 12 wt% was prepared; the photoinitiator was 2-hydroxy-2-methyl-1-[4-(2-hydroxyethoxy)phenyl]-1-propanone; and a hydrogel precursor solution loaded with 100 μg / mL VEGF and 100 μg / mL NGF was prepared.

[0085] Step 2:

[0086] Construction of the light-transmitting sheet: The internal filling is selected to be a photomask template of 10 to 90%.

[0087] Irradiate with a 405 nm ultraviolet light generator for 15 seconds to obtain a gradient photo-crosslinked drug-loaded hydrogel.

[0088] The preparation method of the nerve conduit of this embodiment is the same as that of embodiment 3, except that:

[0089] Step A: Preparation of electrospun fiber membrane

[0090] A solution with a mass ratio of polycaprolactone to gelatin of 4:1 and a mass fraction of 10 wt% was prepared to prepare an electrospun fiber membrane with an orderly oriented inner layer. The fiber membrane had a thickness of 300 μm.

[0091] Example 5

[0092] The preparation method of the gradient photo-crosslinked drug-loaded hydrogel in this embodiment is the same as that in embodiment 3, except that:

[0093] Step 1: Solution preparation

[0094] A GelMA precursor solution with a mass fraction of 10 wt% was prepared; the photoinitiator was phenyl (2,4,6-trimethylbenzoyl) lithium phosphate LAP; and a hydrogel precursor solution loaded with 100 μg / mL NGF and 2 mg / ml deferoxamine DFO was prepared.

[0095] Step 2:

[0096] Construction of the light-transmitting sheet: The internal filling is selected to be a photomask template of 20-70%.

[0097] Irradiate with a 405 nm ultraviolet light generator for 20 seconds to obtain a gradient photo-crosslinked drug-loaded hydrogel.

Claims

1. A method for preparing a gradient photocrosslinked hydrogel, characterized in that: The method comprises the following steps: Step 1: firstly add a photocurable hydrogel precursor into a photoinitiator to prepare a hydrogel precursor solution; optionally, add a bioactive substance or a drug to prepare a drug-loaded hydrogel precursor solution; Step 2: putting the hydrogel precursor solution or the drug-loaded hydrogel precursor solution prepared in step 1 into a transparent tube, and performing photo-crosslinking by irradiating a light source to obtain a gradient photo-crosslinked hydrogel; Wherein, in the step 2, a variable gradient grayscale light mask with a transparency of 0 to 100% is arranged between the transparent tube and the light source.

2. The method for preparing the gradient photo-crosslinked hydrogel according to claim 1, characterized in that: In the step 1, The photocurable hydrogel precursor is selected from one or a combination of methacrylated gelatin solid, methacrylated chitosan, methacrylated silk fibroin, and methacrylated hyaluronic acid; and / or, The photoinitiator is selected from one or a combination of phenyl (2,4,6-trimethylbenzoyl) phosphate lithium salt, 2-hydroxy-2-methyl-1-[4-(2-hydroxyethoxy)phenyl]-1-propanone, 2-hydroxy-2-methyl-1-phenylpropanone, 1-hydroxycyclohexyl phenyl ketone, and 2,4,6-trimethylbenzoyl-diphenylphosphine oxide.

3. The method for preparing the gradient photo-crosslinked hydrogel according to claim 1, characterized in that: In the step 1, The preparation conditions of the hydrogel precursor solution are: temperature of 50-60° C., heating for 10-30 min; and / or, The concentration of the photocurable hydrogel in the hydrogel precursor solution is 5 to 35 wt %, preferably 10 to 20 wt %.

4. The method for preparing a gradient photo-crosslinked hydrogel according to claim 1, characterized in that: In the step 1, The bioactive substance or drug is selected from one or a combination of nerve growth factor, neurotrophic factor, and deferoxamine; and / or, The concentration of the bioactive substance or drug in the drug-loaded hydrogel precursor solution is 0 to 200 μg / mL, preferably 50 to 150 μg / mL.

5. The method for preparing the gradient photo-crosslinked hydrogel according to claim 1, characterized in that: In the step 2, the light source is ultraviolet light or blue light.

6. A gradient photo-crosslinked hydrogel, characterized in that: The method is prepared by any one of claims 1 to 5.

7. A nerve conduit, characterized in that Comprising the gradient cross-linked hydrogel as described in claim 6.

8. A method for preparing a nerve conduit as claimed in claim 7, characterized in that: The preparation method comprises the following steps: Step A, adding a high molecular polymer solution into a syringe and installing it on an electrospinning machine propeller to perform electrospinning to obtain high molecular polymer fibers, and after the solvent evaporates, obtaining an electrospun fiber membrane; Step B: cutting the electrospun fiber membrane obtained in step A into the same length as the gradient photo-crosslinked drug-loaded hydrogel, then wrapping the electrospun fiber membrane in the gradient photo-crosslinked hydrogel, bonding the gaps with a polymer solution, and after drying, obtaining a gradient photo-crosslinked hydrogel-filled nerve conduit.

9. The preparation method according to claim 8, characterized in that: In step A, the electrospinning conditions are: The distance between the needle and the receiving device is 10 to 25 cm; The propeller rate is 0.5 to 1.5 mL / h; The ambient temperature is 20-30℃; The voltage between the two electrodes of the electrospinning device is 15-25 kV; The spinning time is 10 to 12 hours.

10. The preparation method according to claim 8, characterized in that: The high molecular polymer is one or a combination of polycaprolactone, polylactic acid-glycolic acid, type I collagen, and methacryloyl gelatin; and / or, The mass concentration of the high molecular polymer solution is 6 to 15 wt %, preferably 10 to 15 wt %.

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