Anisotropic hydrogel fiber coating modified titanium material and preparation method thereof

Abstract

The invention discloses an anisotropic hydrogel fiber coating modified titanium material and a preparation method thereof, and belongs to the technical field of medical instrument surface coating preparation. The anisotropic hydrogel fiber coating modified titanium material comprises a titanium substrate, an adhesion layer and a hydrogel fiber coating, wherein the adhesion layer and the hydrogel fiber coating are compounded on the titanium substrate; the adhesion layer is compounded on the titanium substrate, and the adhesion layer is positioned between the titanium substrate and the hydrogel fiber coating; the hydrogel fiber coating is prepared from methacryloyl gelatin, silk fibroin and nano-hydroxyapatite through electrostatic spinning. The prepared modified titanium material can keep high water content, viscoelasticity and hydrophilicity, meanwhile, a stable micro-orientation structure and a hydrogel fiber structure with good cell compatibility are obtained, endogenous cells can be collected in situ, osteogenic differentiation of the cells is effectively promoted, and the osteogenic differentiation rate of the cells is increased. The method has a wide application prospect in preparation of bone graft materials and bone repair materials.

Description

Technical Field

[0001] This application belongs to the technical field of the preparation of surface coatings for medical devices, and particularly relates to an anisotropic hydrogel fiber-coated modified titanium material and a preparation method thereof. Background Art

[0002] Titanium and its alloys are widely used in bone repair implants due to their excellent mechanical properties, biocompatibility, and corrosion resistance. However, the bio-inertness of the titanium implant surface greatly limits the integration performance between the implant and the surrounding bone tissue, easily causing aseptic loosening or displacement of the implant and increasing the risk of secondary revision. Therefore, bionic functional modification of the titanium implant surface to make the surface structure and properties of titanium more similar to those of natural bone tissue, thereby promoting good bone integration, is one of the research hotspots in biomedical titanium materials. Hydrogels are considered to be ideal bone tissue engineering scaffold materials that can promote the repair and regeneration of bone defect sites. Combining the strong mechanical properties of metal implants as load-bearing components with the bioactivity of hydrogels may promote the success of implanted metal materials and the bio-integration of the hard and soft tissue interfaces. Currently, the composite of hydrogels onto the surface of titanium implants has become a research hotspot in the field of biomedical materials.

[0003] The prior art with the application publication number CN 113384750 A discloses a titanium alloy surface composite hydrogel coating for reducing bone tissue wear, which constructs a PVA / PAA / GO / PDA composite hydrogel coating on the titanium alloy surface through a method combining chemical assembly and sol-gel, improving the friction resistance and corrosion resistance of the material surface.

[0004] However, the products obtained by modifying the surface of titanium alloys with existing composite hydrogel coatings have the following problems: one is the poor bonding force with the base titanium material, resulting in easy coating peeling; the other is the large difference from the natural bone tissue structure, affecting the adaptation of the bone tissue repair process. Summary of the Invention

[0005] This application discloses an anisotropic hydrogel fiber-coated modified titanium material and a preparation method thereof, aiming to solve the technical problems of poor bonding force and poor adaptation of bone tissue repair effect in the modification of the surface of titanium alloys with existing composite hydrogel coatings.

[0006] To achieve the above object, the technical solution of this application is:

[0007] The first aspect of this application provides an anisotropic hydrogel fiber-coated modified titanium material, including: a titanium substrate and an adhesion layer and a hydrogel fiber coating compounded on the titanium substrate;

[0008] The adhesion layer is compounded on the titanium substrate, and the adhesion layer is located between the titanium substrate and the hydrogel fiber coating;

[0009] The hydrogel fiber coating is prepared by electrospinning of methacryloyl gelatin, silk fibroin and nano-hydroxyapatite.

[0010] Preferably in combination with the first aspect, the mass ratio of methacryloyl gelatin, silk fibroin and nano-hydroxyapatite is: 14:6:0.5 - 1.5.

[0011] The second aspect of the present application provides a method for preparing the anisotropic hydrogel fiber-coated modified titanium material described in the first aspect. The preparation method includes:

[0012] Put the pretreated titanium substrate into the hydrochloric acid dopamine solution to obtain AT-PDA;

[0013] Add methacryloyl gelatin, silk fibroin and nano-hydroxyapatite to an organic solvent to prepare a spinning solution;

[0014] Use AT-PDA as the receiving plate, perform electrospinning with the spinning solution, take it out and immerse it in an ethanol solution of a photoinitiator, and then irradiate it with ultraviolet light to obtain the anisotropic hydrogel fiber-coated modified titanium material.

[0015] Preferably in combination with the second aspect, the organic solvent is one or more of hexafluoroisopropanol, acetic acid, formic acid, and trifluoroacetic acid.

[0016] Preferably in combination with the second aspect, the concentration of the spinning solution is 50 - 250 mg / mL.

[0017] Preferably in combination with the second aspect, when performing electrospinning, the spinning voltage is 5 - 20 KV.

[0018] Preferably in combination with the second aspect, when performing electrospinning, the rotation speed is 1000 - 3000 rpm, and the receiving distance is 5 - 40 cm.

[0019] Preferably in combination with the second aspect, when performing electrospinning, the flow rate of the spinning solution is 0.001 - 0.1 mL / min.

[0020] Preferably in combination with the second aspect, the selected photoinitiator is one of I2959 and LAP.

[0021] The third aspect of the present application provides the application of the anisotropic hydrogel fiber-coated modified titanium material described in the first aspect or the anisotropic hydrogel fiber-coated modified titanium material prepared by the preparation method described in the second aspect in the preparation of bone graft materials and bone repair materials.

[0022] Compared with the prior art, the advantages or beneficial effects of the embodiments of the present application at least include:

[0023] The anisotropic hydrogel fiber-coated modified titanium material provided by the present application is prepared by electrospinning methacryloyl gelatin, silk fibroin and nano-hydroxyapatite into a hydrogel fiber coating and compounding it on the surface of a titanium substrate. On the one hand, while maintaining high water content and viscoelasticity, a hydrogel fiber structure with a stable micro-orientation structure and good cell compatibility can be obtained, so that the anisotropic characteristics in bone tissue can be simulated and excellent mechanical properties can be achieved; on the other hand, the strong binding force between the hydrogel fiber coating and the titanium substrate can be maintained, the stability of the overall structure can be maintained during the implantation process, and excellent hydrophilicity can be maintained at the same time; on the third hand, through the dual synergistic mechanism of physical guidance and bioactive substances, endogenous cells can be recruited in situ, the migration ability and orderly aggregation of cells can be improved, and the adhesion and osteogenic differentiation of osteocytes can be promoted, thus accelerating the bone regeneration process. Description of the Drawings

[0024] In order to more clearly illustrate the technical solutions of the embodiments of the present application, the drawings required for the description of the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments recorded in the present application. For those of ordinary skill in the art, other drawings can be obtained based on these drawings without creative efforts.

[0025] Figure 1 It is a process flow chart of the pretreated titanium substrate provided by the embodiment of the present application;

[0026] Figure 2 It is a process flow chart of the preparation of the modified titanium material provided by the embodiment of the present application;

[0027] Figure 3 It is the scanning electron micrograph of Ti, AT-Ti, AT-PDA, AT-P / GS, AT-P / GSH, AT-P / GSH / / provided by the embodiment of the present application;

[0028] Figure 4 It is the fiber diameter distribution diagram of AT-P / GS provided by the embodiment of the present application;

[0029] Figure 5 It is the fiber diameter distribution diagram of AT-P / GSH provided by the embodiment of the present application;

[0030] Figure 6 It is the AT-P / GSH / / fiber diameter distribution diagram provided by the embodiment of the present application;

[0031] Figure 7 It is the binding force diagram of AT-P / GS, AT-P / GSH, AT-P / GSH / / provided by the embodiment of the present application;

[0032] Figure 8 The bond strength diagrams of AT-P / GS, AT-P / GSH, and AT-P / GSH provided in the embodiments of the present application / / ;

[0033] Figure 9 The water contact angles of Ti, AT-Ti, AT-PDA, AT-P / GS, AT-P / GSH, and AT-P / GSH provided in the embodiments of the present application / / ;

[0034] Figure 10 The quantitative cytotoxicity diagrams of Ti, AT-Ti, AT-PDA, AT-P / GS, AT-P / GSH, and AT-P / GSH provided in the embodiments of the present application / / ;

[0035] Figure 11 The cytotoxicity result diagrams of Ti, AT-Ti, AT-PDA, AT-P / GS, AT-P / GSH, and AT-P / GSH provided in the embodiments of the present application / / ;

[0036] Figure 12 The cell migration diagrams of Ti, AT-P / GS, AT-P / GSH, and AT-P / GSH provided in the embodiments of the present application / / ;

[0037] Figure 13 The cell recruitment performance test diagrams of Ti, AT-P / GS, AT-P / GSH, and AT-P / GSH provided in the embodiments of the present application / / ;

[0038] Figure 14 The osteogenic performance test diagrams of Ti, AT-P / GS, AT-P / GSH, and AT-P / GSH provided in the embodiments of the present application / / ; Detailed implementation manners

[0039] Next, the technical solutions in the embodiments of the present application will be clearly and completely described in conjunction with the accompanying drawings in the embodiments of the present application. Obviously, the described embodiments are part of the embodiments of the present application, rather than all of the embodiments. Based on the embodiments in the present application, all other embodiments obtained by those of ordinary skill in the art without creative efforts shall fall within the protection scope of the present application.

[0040] In the following description of this embodiment, the term "and / or" is used to describe the association relationship of associated objects, indicating that there can be three relationships. For example, A and / or B can represent: A exists alone, B exists alone, and the situation where A and B exist simultaneously. Where A and B can be singular or plural. The character " / " generally indicates that the associated objects before and after are in an "or" relationship.

[0041] In the following description of this embodiment, the term "at least one" means one or more, and "a plurality" means two or more. "At least one of the following" or its similar expressions refer to any combination of these items, including any combination of single item(s) or plural item(s). For example, "at least one of a, b or c", or, "at least one of a, b and c" can both represent: a, b, c, a - b (i.e., a and b), a - c, b - c, or a - b - c, where a, b, and c can be single or multiple respectively.

[0042] Those skilled in the art should understand that in the following description of the embodiments of this application, the sequence numbers do not imply the order of execution. Some or all of the steps can be executed in parallel or sequentially, and the execution order of each process should be determined by its function and internal logic, and should not constitute any limitation to the implementation process of the embodiments of this application.

[0043] The terms used in the embodiments of this application are only for the purpose of describing specific embodiments, and are not intended to limit this application. The singular forms "a" and "the" used in the embodiments of this application and the appended claims are also intended to include the plural forms, unless the context clearly indicates otherwise.

[0044] It should be noted that all raw materials and reagents in the embodiments of this application are purchased on the market or prepared by conventional methods well-known to those skilled in the art.

[0045] In a first aspect, an embodiment of this application provides an anisotropic hydrogel fiber-coated modified titanium material, including: a titanium substrate, and an adhesion layer and a hydrogel fiber coating compounded on the titanium substrate;

[0046] The adhesion layer is compounded on the titanium substrate, and the adhesion layer is located between the titanium substrate and the hydrogel fiber coating;

[0047] The hydrogel fiber coating is made by electrospinning of methacryloyl gelatin, silk fibroin, and nano-hydroxyapatite.

[0048] On the one hand, a hydrogel fiber structure that can maintain high water content and viscoelasticity while obtaining a stable micro-oriented structure and good cell compatibility can be obtained, thereby simulating the anisotropic characteristics in bone tissue and having excellent mechanical properties. On the other hand, the strong bonding force between the hydrogel fiber coating and the titanium substrate can be maintained, the stability of the overall structure can be maintained during the implantation process, and excellent hydrophilicity can be maintained at the same time. On the third hand, through the dual synergistic mechanism of physical guidance and bioactive substances, endogenous cells can be recruited in situ, the migration ability and orderly aggregation of cells can be improved, and the adhesion and osteogenic differentiation of osteocytes can be promoted, thereby accelerating the bone regeneration process.

[0049] It should be noted that the specific sources of methacryloyl gelatin and silk fibroin in the embodiments of the present application are not particularly limited, and can be obtained through commercial channels or synthesized by methods known in the art. For example, the preparation method adopted in the embodiments of the present application is as follows:

[0050] (1) Preparation method of silk fibroin:

[0051] Prepare 2 L of 0.2 M anhydrous sodium carbonate (Na 2 CO 3 ) solution and boil it; put 20 g of silkworm cocoons into the boiling anhydrous Na 2 CO 3 solution and boil for 20 minutes. Take out the boiled cocoon silk, wash it with deionized water, and then put the cocoon silk into the newly prepared 2 L boiling 0.2 M anhydrous Na 2 CO 3 solution again, and repeat the above steps 3 times; put the obtained silk floss into an oven at 60 °C and dry it overnight to obtain cocoon silk;

[0052] Weigh the dried cocoon silk, prepare an appropriate volume of 9.3 M lithium bromide (LiBr) solution according to the bath ratio of 1:20 (m / v), heat the LiBr solution to 60 °C, put the dried cocoon silk into it, and react for 4 hours until the cocoon silk is completely dissolved to form an SF solution. To remove LiBr in the solution, dialyze it in a dialysis bag with a molecular weight cut-off of 8 - 14 kDa for 3 days, and then freeze-dry it to obtain a purified SF sample.

[0053] (2) Preparation method of methacryloyl gelatin:

[0054] At 50 °C, add 5 g of gelatin to 50 ml of phosphate buffer solution (PBS, pH = 7.4) and stir until completely dissolved. Add 3 ml of methacrylic anhydride (MA) under light protection and react for 2 h, then add 200 mL of preheated PBS to terminate the reaction. To remove the unreacted MA, dialyze it in a dialysis bag with a molecular weight cut-off of 8 - 14 kDa at 50 °C for 3 - 5 days, and store it under light protection after freeze-drying to obtain it.

[0055] In the embodiments of the present application, the mass ratio of methacryloyl gelatin, silk fibroin and nano-hydroxyapatite is: 14:6:0.5 - 1.5. Among them, by controlling the addition amount of each material, the hydrogel fiber coating can be given excellent osteogenic performance and mechanical properties.

[0056] It should be noted that under ultraviolet light (UV), methacryloyl gelatin (GelMA) can form a stable covalent cross-linked structure. This cross-linked structure not only provides high water content and soft mechanical properties, but also maintains good cell compatibility, thus promoting cell adhesion and proliferation; silk fibroin (SF) provides the necessary mechanical strength and elasticity, and promotes cell adhesion and proliferation through its unique fiber structure, further improving the bioactivity of the implant. At the same time, the surface of the silk fibroin fiber has nucleation sites of hydroxyapatite, which is beneficial to subsequent mineralization; nano-hydroxyapatite (nHAPs) is similar to the natural bone mineral composition and has excellent biocompatibility and bioactivity. In the bionic coating, nHAPs not only enhances the mechanical strength of the material, but its micro-nano structure also promotes the adhesion and osteogenic differentiation of bone cells, thus accelerating the bone regeneration process.

[0057] In the second aspect, the embodiments of the present application also provide a preparation method of the anisotropic hydrogel fiber-coated modified titanium material described in the first aspect. The preparation method includes:

[0058] Put the pretreated titanium substrate into the hydrochloric acid dopamine solution to obtain AT-PDA;

[0059] Add methacryloyl gelatin, silk fibroin and nano-hydroxyapatite into an organic solvent to prepare a spinning solution;

[0060] Use AT-PDA as the receiving plate, perform electrospinning with the spinning solution, take it out and immerse it in an ethanol solution of a photoinitiator, and then irradiate it with ultraviolet light to obtain the anisotropic hydrogel fiber-coated modified titanium material.

[0061] It should be noted that in order to exclude the interference of impurities and to better promote the combination of the hydrogel fiber coating and the titanium substrate, the titanium substrate can be pretreated. The present application does not make special limitations on the method of the titanium substrate. The process adopted in the present application is: cut the titanium foil into square sheets with a side length of 1 cm, place it in an ultrasonic cleaner, wash it with ethanol and deionized water (each for half an hour), and then place it in an oven to dry for standby; immerse titanium (Ti) in a 5M sodium hydroxide solution (NaOH) at 80 °C for 24 h, neutralize it with 0.05M hydrochloric acid (HCl) for 1 h, and then wash it with ethanol and deionized water to obtain the pretreated titanium substrate.

[0062] It should be noted that the preparation process of the electrospinning technology adopted in this application is simple, with low cost, and the bonding strength between the prepared coating and the substrate titanium material is high.

[0063] In the embodiments of the present application, the organic solvent is preferably one or more of hexafluoroisopropanol, acetic acid, formic acid, and trifluoroacetic acid. Among them, these organic solvents can efficiently dissolve methacryloyl gelatin, silk fibroin, and nano-hydroxyapatite, ensure the uniform dispersion of raw materials, and can quickly volatilize during the electrospinning process, which helps the formation and solidification of fibers.

[0064] In the embodiments of the present application, the concentration of the spinning solution is preferably 50 - 250 mg / ml. Among them, by controlling the concentration of the spinning solution, it can be ensured that the spinning solution can stably form a jet under the action of electric field force and be successfully solidified into fibers, improving the spinning efficiency and the quality of the generated fibers.

[0065] In the embodiments of the present application, when electrospinning, the spinning voltage is preferably 5 - 20 KV, the rotation speed is preferably 1000 - 3000 rpm, the receiving distance is preferably 5 - 40 cm, the flow rate is preferably 0.001 - 0.1 mL / min, and the rotation speed is preferably 500 - 3000 rpm. Among them, by controlling the conditions of electrospinning, a stable micro-oriented structure can be generated, improving the migration ability and orderly aggregation of cells, and promoting the adhesion and osteogenic differentiation of osteoblasts.

[0066] In the embodiments of the present application, the selected photoinitiator is preferably one of I2959 and LAP; the ultraviolet light wavelength is preferably 365 - 405 nm. Among them, by immersing the electrospun fibers in the photoinitiator and then irradiating with ultraviolet light, they can be crosslinked and solidified into hydrogel fibers. Through the prepared hydrogel fiber coating, the active recruitment of endogenous cells and signal molecules can be realized, avoiding the dose dependence and individual differences of exogenous factors, reducing the clinical risk, and improving the bone repair efficiency.

[0067] The third aspect of the present application provides the application of the anisotropic hydrogel fiber-coated modified titanium material described in the first aspect or the anisotropic hydrogel fiber-coated modified titanium material prepared by the preparation method described in the second aspect in the preparation of bone graft materials and bone repair materials. Among them, based on the above modified titanium material having a hydrogel fiber structure that can maintain high water content and viscoelasticity while obtaining a stable micro-oriented structure and good cell compatibility, it has excellent mechanical properties, excellent hydrophilicity, and can effectively promote the osteogenic differentiation of stem cells, and has broad application prospects in the preparation of bone graft materials and bone repair materials.

[0068] The technical solutions of the present application will be further elaborated below in conjunction with specific embodiments.

[0069] Example 1

[0070] This embodiment provides a preparation method of A1-anisotropic hydrogel fiber-coated modified titanium material (AT-P / GSH / / ), which is prepared according to the process flow chart shown in Figure 2 and specifically includes:

[0071] S101: Carry out according to the process flow of pretreating titanium shown in Figure 1 : Immerse titanium (Ti) in 5M sodium hydroxide solution (NaOH) at 80 °C for 24 h, neutralize it with 0.05M hydrochloric acid (HCl) for 1 h, and then wash it with ethanol and deionized water to obtain pretreated titanium (AT-Ti); dissolve dopamine hydrochloride in a buffer solution of 10 mM Tris-HCl (pH 8.5) at a concentration of 2 mg / mL, place it in the prepared dopamine hydrochloride solution and incubate it in the dark for 8 h, wash it 3 times with deionized water, and dry it in the dark at room temperature to obtain AT-PDA;

[0072] S102: Disperse 350 mg of methacryloyl gelatin, 150 mg of silk fibroin, and 25 mg of nano-hydroxyapatite in 5 mL of HFIP, stir at room temperature for 24 h, and prepare a spinning solution;

[0073] S103: Put the electrospinning solution into a 10 mL syringe, place AT-PDA on the receiving drum of the electrospinning machine, with a rotation speed of 2000 rpm, a fixed voltage of 12 kV, use a syringe pump to pump out the solution at a speed of 6 mL / h, a receiving distance of 25 cm, obtain oriented fibers by adjusting the collection rotation speed, take them out and soak them in a 0.5% (wt%) I2959 ethanol solution for 30 min, and expose them under ultraviolet light at 365 nm and 7.0 MW / cm 2 for 15 min to obtain A1-anisotropic hydrogel fiber-coated modified titanium material (AT-P / GSH / / ).

[0074] Example 2

[0075] This embodiment provides a preparation method of A2-anisotropic hydrogel fiber-coated modified titanium material. The component ratio, preparation operation, and process parameters are basically the same as those in Example 1, except that the electrospinning parameters in this embodiment are different. The rotation speed is 1000 rpm, and the fixed voltage is 20 kV, to obtain A2-anisotropic hydrogel fiber-coated modified titanium material.

[0076] Example 3

[0077] The preparation method, component ratio, preparation operation, and process parameters of the A3-anisotropic hydrogel fiber-coated modified titanium material in this example are basically the same as those in Example 1, except that the electrospinning parameters in this example are different. The rotation speed is 3000 rpm, and the fixed voltage is 20 kV, obtaining the A3-anisotropic hydrogel fiber-coated modified titanium material.

[0078] Meanwhile, to verify the comprehensive performance of the modified titanium materials prepared in the above examples, this application provides the following comparative examples for detailed elaboration.

[0079] Comparative Example 1

[0080] The preparation method, component ratio, preparation operation, and process parameters of the B1-modified titanium material in this comparative example are basically the same as those in Example 1, except that in this comparative example, the electrospinning solution is only made by dispersing 350 mg of methacryloyl gelatin and 150 mg of silk fibroin in 5 mL of HFIP, obtaining the B1-modified titanium material (AT-P / GS).

[0081] Comparative Example 2

[0082] The preparation method, component ratio, preparation operation, and process parameters of the B2-modified titanium material in this comparative example are basically the same as those in Example 1, except that the conditions for electrospinning in this comparative example are: the fixed voltage is 12 kV, the solution is pumped out at a speed of 6 mL / h using an injection pump, the receiving distance is 25 cm, and by adjusting the collection rotation speed to 500 rpm, isotropic fibers are obtained, obtaining the B2-modified titanium material (AT-P / GSH).

[0083] The properties of the modified titanium materials prepared in Examples 1-3 of this application are basically the same. The A1-anisotropic hydrogel fiber-coated modified titanium material (AT-P / GSH / / ) prepared in Example 1 is used for further comparative testing.

[0084] 1. Microscopic morphology detection of the hydrogel coating surface:

[0085] To verify the appearance morphology of the raw materials used to prepare the modified titanium materials in the examples of this application, the raw materials added in the examples are tested by scanning electron microscopy, and the results are Figures 3 - 6 as shown.

[0086] According to Figures 3 - 6 what is known, the pure Ti sample shows a relatively smooth surface morphology; AT-Ti presents a sponge-like nanostructure with many nanoscale cross grooves; the surface of AT-PDA shows the enrichment of dopamine particles, proving the successful loading of dopamine particles; the AT-P / GS group presents a smooth fiber structure with a diameter of about 0.55 μm; the AT-P / GSH group and AT-P / GSH / / With the addition of nHAPs, the fiber diameter became thicker, among which the AT-P / GSH / / group had the thickest fiber diameter, about 1.5 μm, and had an obvious orientation structure. This may be due to the adsorption of nHAPs on the fiber wall, resulting in an increase in diameter. The porous structure generated by the electrospinning process not only provides a larger surface area, enhances the physical bonding with the implant and tissue, but also provides a good environment for cell adhesion and growth. The existence of the pore structure promotes the exchange of nutrients and metabolic wastes, contributing to the rapid integration and functional recovery of tissues. All the above results can indicate the successful preparation of anisotropic electrospun hydrogel fiber-coated modified titanium.

[0087] 2. Detection of the interfacial bonding strength between the hydrogel coating and titanium:

[0088] The bonding strength between the electrospun hydrogel fiber coating and the modified titanium was tested by the lap shear test. Briefly, an overlapping area (10 mm × 10 mm) was formed between each group of titanium or modified titanium, ensuring that the coating surface was overlapped parallel to another titanium sheet. An in-situ mechanical testing system was used to perform the tensile test at a speed of 0.1 mm / s. A significant drop in the traction force was considered as the failure of coating adhesion, and the bonding strength (kPa) was calculated.

[0089] The calculation formula is: S a =F m / A. S a is the bonding strength, A is the area of the overlapping region, and F m is the maximum tensile force required during the test.

[0090] According to Figures 7 - 8 shown, the average bonding strength of the electrospun hydrogel fiber-coated modified titanium group (i.e., A1 - anisotropic hydrogel fiber-coated surface-modified titanium material (AT-P / GSH / / )) for the material reached above 119.08 kPa ± 4.75 (n = 3). This is not only due to the chemical bonding between dopamine and GelMA, but also the existence of the topological structure formed after alkali heat treatment enhances the adhesion strength between the electrospun hydrogel fiber coating and the substrate.

[0091] 3. Hydrophilicity test of the electrospun hydrogel fiber coating:

[0092] The hydrophilicity of each group of electrospun hydrogel fiber coatings was measured by the water contact angle. The moisture on the surface of each group of hydrogels was wiped with filter paper, placed on a glass slide, the glass slide was placed on the stage, the camera was started, and immediately photographed after dropping the same volume of water droplets. The contact angle was measured by software to define the baseline and the side tangent line.

[0093] According to Figure 9 shown, at the water droplet and AT-P / GSH / / In the initial stage of contact with the sample surface, a high contact angle of about 72.5 ± 3.3° can be observed. However, over time, the sample shows strong water absorption ability, and within 1 minute, the water contact angle drops to about 48.2 ± 0.6°, which is beneficial to improving the adhesion of the hydrogel fiber coating on the material surface.

[0094] 4. Biocompatibility test:

[0095] The CCK-8 method and live / dead cell staining were used to evaluate the cytotoxicity of different samples on mouse bone marrow mesenchymal stem cells.

[0096] The CCK-8 results showed that there was no difference in cell proliferation among different groups ( Figure 10 ). The results of live / dead cells are as Figure 11 shown. The AT-P / GSH / / group had very little red fluorescence compared with other groups, and the cells showed directional arrangement, indicating that the oriented fiber coating significantly promoted cell survival, proliferation and provided cell growth guidance, showing the best performance.

[0097] 5. Cell migration experiment:

[0098] The chemotactic effect of the anisotropic hydrogel fiber-coated surface-modified titanium materials prepared in this application on cells was measured by transwell migration assay.

[0099] According to Figure 12 shown, the migration order of cells is as follows: AT-P / GSH / / >AT-P / GSH>AT-P / GS>Ti, indicating that the oriented fiber coating (AT-P / GSH / / group) improves the migration ability and ordered aggregation of cells through a dual mechanism of physical guidance and addition of bioactive substances.

[0100] 6. Determination of cell osteogenic differentiation:

[0101] Early osteogenic differentiation was measured by the ALP kit method, mid-osteogenic differentiation was measured by the Sirius red kit method, and late osteogenic differentiation was measured by the alizarin red kit method.

[0102] According to Figure 13 shown, the MSCs cells in the Ti group showed a slender morphology, a small diffusion area, and no obvious extension of filopodia. In contrast, AT-P / GS, AT-P / GSH, AT-P / GSH / / The cells of the group had a larger spreading area and more pseudopodia. By quantitatively analyzing the average spreading area of the cells in each group, it was found that the cells in the AT-P / GSH / / group had the largest spreading area, which was more conducive to cell adhesion and growth. This result suggests that the addition of bionic anisotropy and bioactive substances may have a significant impact on cell morphology, thereby affecting cell adhesion.

[0103] The result graph of ALP is as Figure 14 shown. After culturing for 4 and 7 days, the ALP activity of the AT-P / GSH / / group was significantly higher than that of the pure Ti group and other groups. Secondly, the results of the collagen secretion test showed that after culturing for 14 days, the collagen secretion level of the AT-P / GSH / / group was significantly higher than that of the pure Ti group and other treatment groups. Thirdly, after culturing for 21 days, the number of mineralized nodules was analyzed by alizarin red staining. The mineralization results showed a similar trend to the ALP and collagen secretion tests. More mineralized nodules were produced by MSCs cultured on the AT-P / GSH / / group than on the pure Ti group and the AT-P / GS group. The above test results all confirmed that the modification of titanium with anisotropic hydrogel fiber coating can effectively promote the osteogenic differentiation of stem cells.

[0104] Therefore, in this application, methacryloyl gelatin, silk fibroin, and nano-hydroxyapatite are electrospun into a hydrogel fiber coating and compounded on a titanium substrate. The material prepared in this application can maintain a high water content and viscoelasticity while obtaining a stable micro-oriented structure and a hydrogel fiber structure with good cell compatibility, thereby simulating the anisotropic characteristics in bone tissue and having excellent mechanical properties, a firm bonding force, and excellent hydrophilicity. It can effectively promote the osteogenic differentiation of stem cells and has broad application prospects in the preparation of bone graft materials and bone repair materials.

[0105] The various embodiments in this specification are described in a progressive manner. The same or similar parts between the various embodiments can be referred to each other, and each embodiment focuses on the differences from other embodiments.

[0106] The above embodiments are only used to illustrate the technical solutions of this application, rather than to limit this application; although this application has been described in detail with reference to the foregoing embodiments, those of ordinary skill in the art should understand that they can still modify the technical solutions recorded in the foregoing embodiments, or perform equivalent replacements on some or all of the technical features; and these modifications or replacements do not make the essence of the corresponding technical solutions deviate from the scope of the technical solutions of this application.

Claims

1. An anisotropic hydrogel fiber coating modified titanium material, characterized in that: include: A titanium substrate and an adhesive layer and a hydrogel fiber coating composited on the titanium substrate; The adhesive layer is compounded on the titanium substrate, and the adhesive layer is located between the titanium substrate and the hydrogel fiber coating; The hydrogel fiber coating is prepared by electrostatic spinning of methacryloyl gelatin, silk fibroin and nano-hydroxyapatite.

2. The anisotropic hydrogel fiber coating modified titanium material according to claim 1, characterized in that: The mass ratio of the methacryloyl gelatin, silk fibroin and nano-hydroxyapatite is 14:6:0.5-1.

5.

3. A method for preparing anisotropic hydrogel fiber coating modified titanium material according to any one of claims 1 to 2, characterized in that: The preparation method comprises: The pretreated titanium substrate is placed in a dopamine hydrochloride solution to obtain AT-PDA; Methacryloyl gelatin, silk fibroin and nano-hydroxyapatite are added into an organic solvent to prepare a spinning solution; The AT-PDA is used as a receiving plate, and the spinning solution is used for electrostatic spinning. After being taken out, it is immersed in an ethanol solution of a photoinitiator, and then exposed to ultraviolet light to obtain the anisotropic hydrogel fiber coating modified titanium material.

4. The method for preparing anisotropic hydrogel fiber coating modified titanium material according to claim 3, characterized in that: The organic solvent is one or more of hexafluoroisopropanol, acetic acid, formic acid and trifluoroacetic acid.

5. The method for preparing anisotropic hydrogel fiber coating modified titanium material according to claim 3, characterized in that: The concentration of the spinning solution is 50-250 mg / mL.

6. The method for preparing anisotropic hydrogel fiber coating modified titanium material according to claim 3, characterized in that: When the electrostatic spinning is performed, the spinning voltage is 5-20KV.

7. The method for preparing anisotropic hydrogel fiber coating modified titanium material according to claim 3, characterized in that: During the electrostatic spinning, the rotation speed is 1000-3000 rpm and the receiving distance is 5-40 cm.

8. The method for preparing anisotropic hydrogel fiber coating modified titanium material according to claim 3, characterized in that: During the electrospinning, the flow rate of the spinning solution is 0.001-0.1 mL / min.

9. The method for preparing anisotropic hydrogel fiber coating modified titanium material according to claim 3, characterized in that: The selected photoinitiator is one of I2959 and LAP.

10. Use of the anisotropic hydrogel fiber coating modified titanium material according to any one of claims 1 to 2 or the anisotropic hydrogel fiber coating modified titanium material prepared by the preparation method according to any one of claims 3 to 9 in the preparation of bone transplant materials and bone repair materials.

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