CN115068678A

CN115068678A - HAase/NIR (hydroxyapatite/NIR) responsive titanium implant as well as preparation method and application thereof

Info

Publication number: CN115068678A
Application number: CN202210687482.7A
Authority: CN
Inventors: 王丹阳; 汤宇; 杨维虎; 蔡开勇
Original assignee: Chongqing University; Second Affiliated Hospital Army Medical University
Current assignee: Chongqing University; Second Affiliated Hospital Army Medical University
Priority date: 2022-06-17
Filing date: 2022-06-17
Publication date: 2022-09-20

Abstract

The invention discloses a HAase/NIR responsive titanium implant and a preparation method and application thereof. The technical scheme of the invention is that the modified titanium-based implant is prepared, and the response release of ciprofloxacin is realized by using trace antibiotic and combining a relatively safe mild PTT treatment strategy, so that the titanium-based implant is endowed with good postoperative anti-infection performance and a function of promoting bone injury repair. The invention constructs a photo-thermal nano-drug delivery system local drug delivery system, greatly reduces the use of antibiotics, combines a mild PTT therapy, constructs an implant surface which can release drugs in a responsive way and promote bone regeneration, can achieve good and biosafety antibacterial effect, and can promote bone tissue repair and regeneration.

Description

HAase/NIR (ultraviolet ray/near infrared) responsive titanium implant as well as preparation method and application thereof

Technical Field

The invention belongs to the technical field of human body implant materials, and particularly relates to an HAase/NIR responsive titanium implant and a preparation method and application thereof.

Background

With the improvement of living standard of people, the improvement of social medical conditions and the increase of aging degree of population, the clinical use amount of the dental and orthopedic implant is increased year by year. Titanium and its alloy are widely used as bone repair implant for orthopedic surgery due to its excellent biocompatibility, mechanical properties and corrosion resistance. In the later stages of titanium implant treatment, the titanium oxide layer begins to absorb ions, proteins and polysaccharides, and then osteoblasts and other immune and inflammatory cells begin to migrate to the bone implant surface, resulting in tight bone attachment. A titanium implant-bone host interface is created, which means that bone attachment is allowed and results in bone anchoring. This process, known as osseointegration, is a key factor in the success of implants to treat these severe orthopedic disorders. However, in clinical applications, bacterial infections, particularly drug-resistant bacterial infections, after implantation of titanium implants can lead to implant failure. Moreover, the inherent biological inertia of the surface of the titanium implant causes insufficient integration capability with the surrounding bone, thereby causing the problem of loosening of the implant.

Staphylococcus aureus (s. aureus) is the most common pathogen in titanium implant infections, and Methicillin-resistant Staphylococcus aureus (MRSA) infections are also on the rise today. In recent years, Photo-thermal therapy (PTT) based on Near Infrared (NIR) has been highlighted for research on treatment of bacterial infections on the surface of implants. Photothermal sterilization can be explained as irreversible damage to proteins or enzymes in microbial cell membranes caused by local high temperature under near infrared, resulting in leakage of cell contents, further influencing physiological activities of bacteria, and even killing the bacteria. However, to achieve effective killing of bacteria, higher temperatures need to be generated if only near-infrared based photothermal therapy is used, at the expense of damage to surrounding normal cells and tissues.

Disclosure of Invention

In order to effectively and sustainably solve the problems of the background art, there is an urgent need to design and develop a titanium implant having good antibacterial and osseointegration capabilities and low side effects so as to achieve infection resistance and promote bone repair after implantation.

In order to achieve the purpose, the invention adopts the technical scheme that the HAase/NIR responsive titanium implant, and the preparation method and the application thereof are provided. The preparation method of the HAase/NIR responsive titanium implant comprises the following steps:

s1: pretreating a titanium substrate, and then forming a hydroxyapatite microstructure on the pretreated titanium substrate to obtain an implant substrate;

s2: preparing hollow mesoporous polydopamine nanoparticles;

s3: ciprofloxacin is loaded on the hollow mesoporous polydopamine nanoparticles prepared by the S2 to obtain drug-loaded nanoparticles;

s4: anchoring the drug-loaded nanoparticles to the implant substrate;

s5: and (3) coating the modified hyaluronic acid on the surface of the implant substrate treated by the S4 to obtain the modified hyaluronic acid.

On the basis of the technical scheme, the invention can be further improved as follows.

Further, the method for pretreating the titanium substrate is micro-arc oxidation.

Further, the formation of the hydroxyapatite microstructure on the titanium substrate comprises the following steps:

s1: constructing an electrodeposition device by taking a titanium rod as a cathode, a pretreated titanium substrate as an anode and a mixed solution of calcium acetate and beta-sodium glycerophosphate as an electrolyte, and then reacting for 5-8 min under the voltage of 400-450V;

s2: and (3) putting the titanium substrate treated by the S1 into alkali liquor, and reacting for 5-8 hours at 180-200 ℃ to obtain the titanium substrate.

Further, the concentration of calcium acetate in the mixed solution of calcium acetate and sodium beta-glycerophosphate is 0.4M, and the concentration of sodium beta-glycerophosphate is 0.05M; the alkaline solution is sodium hydroxide solution with pH value of 9.

Further, the hollow mesoporous polydopamine nanoparticle is prepared by the following steps:

dissolving poly (ethylene glycol) -block-poly (propylene glycol) -block-poly (ethylene glycol), 3',5,5' -tetramethylbenzidine, tris (hydroxymethyl) aminomethane hydrochloride and dopamine hydrochloride in an ethanol solution according to the dosage ratio of 0.3-0.4 g: 0.4-0.45 ml: 80-100 mg: 50-80 mg, reacting at room temperature for 20-30 h, and then extracting by using a mixed solution of acetone and ethanol to remove poly (ethylene glycol) -block-poly (propylene glycol) -block-poly (ethylene glycol) and 3,3',5,5' -tetramethylbenzidine to obtain the compound.

Further, the drug-loaded nanoparticles are prepared by the following steps:

dispersing ciprofloxacin hydrochloride and hollow mesoporous polydopamine nanoparticles into an N-2-hydroxyethyl piperazine-N' -2-ethanesulfonic acid aqueous solution together, and stirring for reacting for 6 hours to obtain the compound.

Further, the drug-loaded nanoparticles are anchored to the implant substrate by:

s1: dissolving dopamine hydrochloride in Tris hydrochloric acid buffer solution with the pH value of 8.5, then soaking the implant substrate into the solution, and reacting overnight at room temperature in a dark place;

s2: dispersing the drug-loaded nanoparticles into phosphate buffer solution with the pH value of 7.5 according to the concentration of 100 mu g/mL, then putting the implant substrate treated by S1 into the solution, and reacting for 24h at room temperature in a dark place to obtain the drug-loaded nanoparticles.

Further, the modified hyaluronic acid is prepared by the following steps:

dissolving hyaluronic acid in phosphate buffer solution with pH value of 5.0, adding 1-ethyl- (3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide into the solution, and stirring for reaction at room temperature for 30 min; adding dopamine hydrochloride into the mixed solution, keeping the pH of the mixed solution at 5.0, reacting at room temperature for 12 hours, and dialyzing in phosphate buffer salt solution with pH of 5.0 and double distilled water in sequence to obtain the dopamine hydrochloride aqueous solution; the mass ratio of the hyaluronic acid to the 1-ethyl- (3-dimethylaminopropyl) carbodiimide to the N-hydroxysuccinimide to the dopamine hydrochloride is 0.5: 0.8-0.85: 0.4-0.5: 0.5.

Further, the modified hyaluronic acid is coated on the surface of the implant substrate by means of spin coating.

The invention has the beneficial effects that:

1. according to the invention, the hydroxyapatite microstructure is firstly generated on the titanium substrate, so that not only can the surface of the titanium substrate be regulated and controlled, but also the hydroxyapatite has a regulating and controlling effect on the osteogenic differentiation of mesenchymal stem cells, wherein calcium and phosphorus elements can be released from the material and participate in the regulation process of bone tissue metabolism.

2. The hollow mesoporous polydopamine nanoparticles have high porosity and can efficiently load drugs such as ciprofloxacin and the like. In addition, the polydopamine nanoparticles have the advantages of high photothermal conversion efficiency, good biocompatibility, easiness in modification, low toxicity and the like, local high temperature generated during infrared irradiation can be generated along with Reactive Oxygen Species (ROS) responsiveness, and the polydopamine nanoparticles have certain killing power on bacteria; the hollow mesoporous polydopamine nanoparticles are used as drug carriers, a novel antibiotic packaging and transmitting strategy can be provided, and the antibacterial activity of the drug-loaded particles can be enhanced and the toxic and side effects can be reduced by combining the packaged antibiotics.

3. Ciprofloxacin (CIP) is a common fluoroquinolone antibiotic with broad-spectrum antibacterial activity, has an obvious killing effect on gram-positive strains, is loaded on hollow mesoporous polydopamine nanoparticles, and can remarkably improve the bacterial killing effect by combining the reactive oxygen generation capacity of the polydopamine nanoparticles at high temperature.

4. After the drug-loaded nanoparticles are anchored on the implant substrate, the hydrophilic hyaluronic acid coating is coated on the surface of the implant substrate, so that the early adhesion of staphylococcus aureus can be effectively inhibited, and the implant substrate can be used as an antibacterial drug responsive release unit and a subsequent cell bioactive unit. Under normal conditions, the drug can be well blocked in the material; when bacterial infection is present, bacterial secreted hyaluronidase (HAase) accelerates the degradation of the coating, thereby effecting an enzyme-responsive release of the drug. More importantly, the prepared material can accelerate the release of the medicine from the material under the triggering of near infrared light irradiation, and simultaneously can activate photo-thermal nanoparticles, the generated local thermotherapy can destroy the integrity of bacteria, and the two kill the surrounding bacteria in a synergistic manner.

5. The modified titanium-based implant is prepared, trace antibiotics are used in combination with a relatively safe mild PTT treatment strategy, responsive release of ciprofloxacin is achieved, and the titanium-based implant is endowed with good postoperative anti-infection performance and a function of promoting bone injury repair. The invention constructs a photo-thermal nano-drug delivery system local drug delivery system, greatly reduces the use of antibiotics, combines a mild PTT therapy, constructs an implant surface which can release drugs in a responsive way and promote bone regeneration, can achieve good and biosafety antibacterial effect, and can promote bone tissue repair and regeneration.

Drawings

FIG. 1 is an SEM image of Ti, Ti-M and Ti-M-H;

FIG. 2 is an SEM image of Ti-M-H-PDA @ CIP on a different scale;

FIG. 3 is an SEM image of Ti-M-H-PDA @ CIP-HAc on a different scale;

FIG. 4 is an XPS survey of Ti and Ti-M-H-PDA @ CIP-HAc;

FIG. 5 is the results of the responsive release of ciprofloxacin under hyaluronidase in different samples;

FIG. 6 shows the results of ciprofloxacin release under near infrared light for different samples;

FIG. 7 is the results of the responsive release of ciprofloxacin in Staphylococcus aureus for the different samples;

FIGS. 8 and 9 show the antimicrobial effect of different samples against Staphylococcus aureus;

FIGS. 10 and 11 show the antimicrobial effect of different samples against methicillin-resistant Staphylococcus aureus;

FIG. 12 shows the activity of the cells at 7 days after the surface inoculation of rat bone marrow mesenchymal stem cells on different samples;

FIG. 13 is a graph showing the alkaline phosphatase level of cells at 7 days after rat bone marrow mesenchymal stem cells were surface-seeded on different samples;

FIG. 14 is a graph showing collagen secretion trends of cells when rat bone marrow mesenchymal stem cells were seeded on the surfaces of different samples for 14 days;

FIG. 15 shows the ability of mineralization to form calcium deposits when rat bone marrow mesenchymal stem cells are seeded on the surface of different samples for 21 days;

FIGS. 16 and 17 show the antimicrobial effect of different samples at the implantation site;

FIG. 18 shows the osteogenic effect of different samples at the implantation site.

Detailed Description

The following examples are provided to illustrate specific embodiments of the present invention.

Example 1

A HAase/NIR responsive titanium implant, prepared by:

(1) taking a clean titanium foil, carrying out micro-arc oxidation pretreatment on the clean titanium foil, then constructing an electrodeposition device by taking a clean titanium rod as a cathode, the pretreated titanium foil as an anode and a mixed solution of calcium acetate and beta-sodium glycerophosphate as an electrolyte, and carrying out electrolytic reaction for 6min under the voltage of 420V, wherein the concentration of the calcium acetate in the electrolyte is 0.4M, and the concentration of the beta-sodium glycerophosphate is 0.05M; and putting the titanium foil after the electrolytic reaction into a sodium hydroxide solution with the pH value of 9, and carrying out an alkali-thermal reaction for 6h at 190 ℃ to form a hydroxyapatite microstructure on the titanium foil, thereby obtaining the implant substrate.

(2) The hollow mesoporous polydopamine nanoparticle is prepared by a one-pot method, and the specific method comprises the following steps:

0.36g of poly (ethylene glycol) -block-poly (propylene glycol) -block-poly (ethylene glycol) ((ethylene glycol))

F127) 0.417mL of 3,3',5,5' -tetramethylbenzidine, 90mg of tris (hydroxymethyl) aminomethane hydrochloride and 60mg of dopamine hydrochloride are dissolved in an ethanol solution (a mixture of 65mL of tribasic water and 60mL of ethanol), the mixture is reacted at room temperature for 24 hours, and then reaction liquid is collected, and then a mixed solution of acetone and ethanol (volume ratio of 1:1) is adopted for extraction to remove poly (ethylene glycol) -block-poly (propylene glycol) -block-poly (ethylene glycol) and 3,3',5,5' -tetramethylbenzidine, so as to obtain the compound.

(3) The method for loading ciprofloxacin by using the prepared hollow mesoporous polydopamine nanoparticle comprises the following steps:

and (2) dispersing 40 mu g of ciprofloxacin hydrochloride and 2mg/mL of hollow mesoporous polydopamine nanoparticles (1mL) in an N-2-hydroxyethyl piperazine-N' -2-ethanesulfonic acid aqueous solution together for drug loading, reacting for 6h under magnetic stirring, and loading ciprofloxacin into the nanoparticles through pi-pi accumulation to obtain the drug-loaded nanoparticles.

(4) Anchoring the drug-loaded nanoparticles to an implant substrate, the specific method is as follows:

s1: dissolving dopamine hydrochloride into Tris hydrochloric acid buffer solution (PBS buffer solution) with the pH value of 8.5 according to the concentration of 2mg/mL, then soaking the implant substrate into the solution, and carrying out dark reaction at room temperature overnight to construct a dopamine coating on the surface of the implant substrate;

(5) The hyaluronic acid is modified by the following specific method:

dissolving 0.5g of hyaluronic acid in phosphate buffered saline (PBS buffer) at pH 5.0 to dissolve it sufficiently; subsequently, 0.815g of 1-ethyl- (3-dimethylaminopropyl) carbodiimide and 0.49g N-hydroxysuccinimide were added to the hyaluronic acid solution, and the mixed solution was stirred for 30 min; then, 0.5g of dopamine hydrochloride was added to the mixed solution to keep the pH of the solution at 5.0, the mixture was reacted at room temperature for 12 hours, and after the reaction was completed, the product was purified by dialysis in phosphate buffered saline at pH 5.0 and in double distilled water (MWCO 3500) for 3 days, respectively.

(6) Coating the modified hyaluronic acid on an implant substrate anchored with drug-loaded nanoparticles by adopting a spin coating mode to obtain the drug-loaded nanoparticle; the spin coating process specifically comprises the following steps:

preparing the modified hyaluronic acid into a solution of 5 mg/mL; placing the implant substrate anchored with the drug-loaded nanoparticles on a spin coater, dropping 30 mu L of modified hyaluronic acid solution on the implant substrate, and repeating the steps of spreading the membrane for 5 times by adopting a rotation speed of 1000rpm for 3 seconds and a rotation speed of 3000rpm for 20 seconds each time, thereby obtaining the modified hyaluronic acid coating.

Example 2

A HAase/NIR responsive titanium implant, prepared by:

(1) taking a clean titanium foil, carrying out micro-arc oxidation pretreatment on the clean titanium foil, then constructing an electrodeposition device by taking a clean titanium rod as a cathode, the pretreated titanium foil as an anode and a mixed solution of calcium acetate and beta-sodium glycerophosphate as an electrolyte, and carrying out electrolytic reaction for 8min under the voltage of 400V, wherein the concentration of the calcium acetate in the electrolyte is 0.4M, and the concentration of the beta-sodium glycerophosphate is 0.05M; and putting the titanium foil after the electrolytic reaction into a sodium hydroxide solution with the pH value of 9, and carrying out alkali-thermal reaction for 5h at the temperature of 200 ℃ to form a hydroxyapatite microstructure on the titanium foil, thereby obtaining the implant substrate.

0.3g of poly (ethylene glycol) -block-poly (propylene glycol) -block-poly (ethylene glycol) ((ethylene glycol))

F127) 0.4mL of 3,3',5,5' -tetramethylbenzidine, 80mg of tris (hydroxymethyl) aminomethane hydrochloride and 50mg of dopamine hydrochloride are dissolved in an ethanol solution (a mixture of 65mL of distilled water and 60mL of ethanol), reacted at room temperature for 20 hours, and then a reaction liquid is collected, and then poly (ethylene glycol) -block-poly (propylene glycol) -block-poly (ethylene glycol) and 3,3',5,5' -tetramethylbenzidine are removed by extraction with a mixed solution of acetone and ethanol (volume ratio 1:1), so as to obtain the compound.

(3) The method for loading ciprofloxacin by using the prepared hollow mesoporous polydopamine nano particles comprises the following steps:

(4) Anchoring the drug-loaded nanoparticles to the implant substrate by the specific method as follows:

(5) The hyaluronic acid is modified by the following specific method:

dissolving 0.5g of hyaluronic acid in phosphate buffered saline (PBS buffer) at pH 5.0 to dissolve it sufficiently; subsequently, 0.8g of 1-ethyl- (3-dimethylaminopropyl) carbodiimide and 0.45g N-hydroxysuccinimide were added to the hyaluronic acid solution, and the mixed solution was stirred for 30 min; then, 0.5g of dopamine hydrochloride was added to the mixed solution to keep the pH of the solution at 5.0, the mixture was reacted at room temperature for 12 hours, and after the reaction was completed, the product was purified by dialysis (MWCO ═ 3500) in phosphate buffered saline at pH 5.0 and in double distilled water for 3 days, respectively, to obtain the compound.

preparing the modified hyaluronic acid into a solution of 5 mg/mL; placing the implant substrate anchored with the drug-loaded nanoparticles on a spin coater, dripping 30 mu L of modified hyaluronic acid solution on the implant substrate, and repeating the steps of laying the film for 5 times, wherein the film is rotated at the rotating speed of 1000rpm for 3 seconds and at the rotating speed of 3000rpm for 20 seconds each time.

Example 3

A HAase/NIR responsive titanium implant, prepared by:

(1) taking a clean titanium foil, carrying out micro-arc oxidation pretreatment on the clean titanium foil, then constructing an electrodeposition device by taking a clean titanium rod as a cathode, the pretreated titanium foil as an anode and a mixed solution of calcium acetate and beta-sodium glycerophosphate as an electrolyte, and carrying out electrolytic reaction for 5min under the voltage of 450V, wherein the concentration of the calcium acetate in the electrolyte is 0.4M, and the concentration of the beta-sodium glycerophosphate is 0.05M; and putting the titanium foil after the electrolytic reaction into a sodium hydroxide solution with the pH value of 9, and carrying out alkali-thermal reaction for 8h at 180 ℃ to form a hydroxyapatite microstructure on the titanium foil, thereby obtaining the implant substrate.

0.4g of poly (ethylene glycol) -block-poly (propylene glycol) -block-poly (ethylene glycol) ((ethylene glycol))

F127) 0.45mL of 3,3',5,5' -tetramethylbenzidine, 100mg of tris (hydroxymethyl) aminomethane hydrochloride and 80mg of dopamine hydrochloride are dissolved in an ethanol solution (a mixture of 65mL of tribasic water and 60mL of ethanol), reacted at room temperature for 20 hours, and then a reaction liquid is collected, and then poly (ethylene glycol) -block-poly (propylene glycol) -block-poly (ethylene glycol) and 3,3',5,5' -tetramethylbenzidine are removed by extraction with a mixed solution of acetone and ethanol (volume ratio 1:1), thus obtaining the target compound.

(5) The hyaluronic acid is modified by the following specific method:

dissolving 0.5g of hyaluronic acid in phosphate buffered saline (PBS buffer) at pH 5.0 to dissolve it sufficiently; subsequently, 0.85g of 1-ethyl- (3-dimethylaminopropyl) carbodiimide and 0.5g N-hydroxysuccinimide were added to the hyaluronic acid solution, and the mixed solution was stirred for 30 min; then, 0.5g of dopamine hydrochloride was added to the mixed solution to keep the pH of the solution at 5.0, the mixture was reacted at room temperature for 12 hours, and after the reaction was completed, the product was purified by dialysis (MWCO ═ 3500) in phosphate buffered saline at pH 5.0 and in double distilled water for 3 days, respectively, to obtain the compound.

Analysis of results

The performance of the HAase/NIR responsive titanium implant prepared in example 1 is illustrated.

The surface SEM appearances of the pure titanium foil (Ti), the titanium foil (Ti-M) after the micro-arc oxidation treatment and the titanium foil (Ti-M-H) after the alkali-thermal reaction are respectively shown in figures 1a, 1b and 1 c. As can be seen from FIG. 1, the micro-arc oxidation treatment forms a smooth micron-sized stacked porous structure on the surface of the titanium material, and the alkaline heat treatment forms a plurality of hydroxyapatite particle deposits on the basis of the previous step. The surface area of the material is increased after micro-arc oxidation and alkali heat treatment, and the contact area between the material and particles is increased.

SEM images after drug-loaded nanoparticles were anchored to the implant substrate (Ti-M-H-PDA @ CIP) are shown in fig. 2, with scales of 10 μ M and 1 μ M for fig. 2d1 and fig. 2d2, respectively. SEM images of the implant substrate after coating with modified hyaluronic acid (Ti-M-H-PDA @ CIP-HAc) are shown in FIG. 3, with scales of 10 μ M and 1 μ M for FIGS. 3e1 and 3e2, respectively. As can be seen from fig. 2 and 3, the drug-loaded nanoparticles are uniformly distributed on the surface of the implant substrate.

XPS detection is carried out on Ti and Ti-M-H-PDA @ CIP-HAc samples, and the result is shown in FIG. 4, wherein the amplified part is the characteristic peak of the F element in the XPS energy spectrum of the surface of the Ti-M-H-PDA @ CIP-HAc sample. As can be seen from the figure, the Ti-M-H-PDA @ CIP-HAc sample had a similar peak pattern to Ti, and characteristic peaks of Ca (2P) and P (2P) elements, and the surface formed TiO due to micro-arc oxidation ₂ The content of the O element in the ceramic layer sample is obviously improved compared with that of a pure Ti group. XPS results show that the deposit formed is hydroxyapatite. The characteristic peak of the F element on the sample Ti-M-H-PDA @ CIP-HAc at a position of 688eV is subjected to fitting analysis, the existence of the characteristic element-F element contained in ciprofloxacin on the material is proved, and the successful loading of the medicine is proved.

The release effect of the drugs of different samples is examined, and the results are shown in figures 5-7. Wherein, fig. 5 is the result of the response release of the ciprofloxacin of the sample under the hyaluronidase, fig. 6 is the result of the release of the ciprofloxacin of the sample under the near infrared light, and fig. 7 is the result of the response release of the ciprofloxacin of the sample in staphylococcus aureus. As can be seen from FIGS. 5-7, the sample surface modified hyaluronic acid coating can be degraded by HAase secreted by Staphylococcus aureus, so that the responsive release of ciprofloxacin is realized, and the release of the drug can be accelerated through photothermal conversion.

The antibacterial effect of the Ti-M-H-PDA @ CIP-HAc sample is verified through in vitro experiments by taking staphylococcus aureus and methicillin-resistant staphylococcus aureus as model bacteria, and the results are shown in fig. 8-11, wherein fig. 8 and 9 are the experimental results of staphylococcus aureus, and fig. 10 and 11 are the experimental results of methicillin-resistant staphylococcus aureus. As can be seen from the figure, Ti-M-H-PDA @ CIP-HAc has good antibacterial performance, and can achieve better antibacterial effect by combining with PTT treatment.

In vitro experiments verify the influence of cell activity when rat bone marrow mesenchymal stem cells (BMSCs) are inoculated on the surface of a Ti-M-H-PDA @ CIP-HAc sample for 7 days, the alkaline phosphatase level of 7 days, the collagen secretion trend of 14 days and the capability of forming calcium deposition by mineralization in 21 days, and the results are shown in FIGS. 12-15. As can be seen from the figure, the higher cell viability was exhibited on the surface of the Ti-M-H-PDA @ CIP-HAc sample, with good biocompatibility. Under the condition of no matter whether near infrared light is irradiated, the Ti-M-H-PDA @ CIP-HAc sample shows higher alkaline phosphatase activity, collagen secretion amount and the tendency of promoting mineralization, and the effect of promoting bone differentiation is obvious.

A rat bone defect infection model is constructed for in-vivo experimental verification, and the antibacterial effect of the material Ti-M-H-PDA @ CIP-HAc on an implanted part is detected 7 days later, and the result is shown in FIGS. 16-17. As can be seen from the figure, Ti-M-H-PDA @ CIP-HAc showed good antibacterial properties similar to the in vivo test results, and a better antibacterial effect was achieved in combination with PTT treatment.

A rat bone defect infection model was constructed for in vivo experimental verification, and the material Ti-M-H-PDA @ CIP-HAc was examined for bone-promoting properties at the implantation site by HE staining and MASSON staining after 7 days, with the results shown in FIG. 18. As can be seen from the figure, a larger area of new bone tissue appears around the Ti-MH-PDA @ CIP-HAc group implant, the fusion condition of the implant and the host is better, the large area of the cavity around the implant is reduced, and the Ti-M-H-PDA @ CIP-HAc NIR ⁺ The group showed an advantage.

The analysis proves that the Ti-M-H-PDA @ CIP-HAc material shows good antibacterial effect and capability of promoting regeneration of bone tissues in-vivo and in-vitro experiments.

While the present invention has been described in detail with reference to the embodiments, it should not be construed as limited to the scope of the patent. Various modifications and changes may be made by those skilled in the art without inventive step within the scope of the appended claims.

Claims

1. A preparation method of a HAase/NIR responsive titanium implant is characterized by comprising the following steps:

s2: preparing hollow mesoporous polydopamine nanoparticles;

s4: anchoring the drug-loaded nanoparticles to the implant substrate;

2. The method of preparing a HAase/NIR responsive titanium implant according to claim 1, wherein: the method for pretreating the titanium substrate is micro-arc oxidation.

3. The method of preparing a HAase/NIR responsive titanium implant according to claim 1, wherein the formation of the hydroxyapatite microstructure on the titanium substrate comprises the steps of:

4. The method of preparing a HAase/NIR responsive titanium implant according to claim 3, wherein: the concentration of calcium acetate in the electrolyte is 0.4M, and the concentration of beta-sodium glycerophosphate is 0.05M; the alkali liquor is sodium hydroxide solution with pH value of 9.

5. The method for preparing a HAase/NIR responsive titanium implant according to claim 1, wherein the hollow mesoporous polydopamine nanoparticle is prepared by the following steps:

6. The method of preparing a HAase/NIR responsive titanium implant according to claim 1, wherein the drug-loaded nanoparticles are prepared by:

7. The method of claim 1, wherein the drug-loaded nanoparticles are anchored to the implant substrate by:

8. The method of preparing a HAase/NIR responsive titanium implant according to claim 1, wherein the modified hyaluronic acid is prepared by the steps of:

dissolving hyaluronic acid in phosphate buffer solution with pH of 5.0, adding 1-ethyl- (3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide into the solution, and stirring to react for 30min at room temperature; adding dopamine hydrochloride into the mixed solution, keeping the pH value of the mixed solution at 5.0, reacting for 12 hours at room temperature, and dialyzing in phosphate buffer salt solution with the pH value of 5.0 and double distilled water in sequence to obtain the dopamine hydrochloride; the mass ratio of the hyaluronic acid to the 1-ethyl- (3-dimethylaminopropyl) carbodiimide to the N-hydroxysuccinimide to the dopamine hydrochloride is 0.5: 0.8-0.85: 0.4-0.5: 0.5.

9. The HAase/NIR responsive titanium implant prepared by the preparation method of any one of claims 1 to 8.

10. Use of the HAase/NIR responsive titanium implant according to claim 9 for the preparation of antibacterial and osteogenesis-promoting materials.