Method for producing a medical implant and use thereof

Abstract

The application provides a preparation method of a medical implant and application thereof, and relates to the technical field of medical implant preparation. The medical implant is prepared by forming a strontium-doped TiO2 coating on the surface of a base material by a micro-arc oxidation method, and then forming a TiO2 mesoporous array layer with a honeycomb structure on the surface by an anodic oxidation method. In the technical scheme, micropores and mesoporous arrays are simultaneously introduced into the medical implant, so that the specific surface area of the medical implant is increased. The structure can improve cell differentiation and cell activity of the medical implant when the medical implant is applied in vivo. The obtained medical implant has excellent biocompatibility and great application potential in the loading of macromolecular proteins or drugs.

Description

Technical Field

[0001] This invention relates to the field of medical implant preparation technology, and in particular to a method for preparing a medical implant and its application. Background Technology

[0002] In the medical field, titanium and titanium alloys are commonly used materials for hard tissue implants, finding wide application in artificial joints, artificial bones, spinal braces, intramedullary nails, dental implants, and skull implants. Titanium and titanium alloys possess advantages such as low density, high specific strength, low elastic modulus, and good corrosion resistance. However, direct implantation into the human body presents several challenges, including low bone integration strength, poor bioactivity, and long healing times. To overcome these issues, biomodification of the surface of titanium and titanium alloys is of great significance. Because titanium dioxide has low inherent toxicity, low solubility in water, low reactivity with biomolecules (approaching chemical inertness), and significant anti-inflammatory effects, surface modification of titanium and titanium alloys using oxidation methods can be an effective approach. Micro-arc oxidation is a commonly used method for metal surface oxidation. It is achieved by forming an oxide layer on the metal surface through electrolytic oxidation. The resulting coating can embed trace elements in the electrolyte and form micron-sized pores. However, due to the small specific surface area of ​​the coating, the loading capacity is insufficient when directly loading drugs and proteins, making it difficult to meet the needs of in vivo applications.

[0003] In view of this, it is necessary to design an improved method for preparing medical implants and their applications to solve the above problems. Summary of the Invention

[0004] The purpose of this invention is to provide a method for preparing a medical implant and its application.

[0005] To achieve the above-mentioned objectives, in a first aspect, the present invention provides a method for preparing a medical implant, comprising the following steps:

[0006] S1. Surface pretreatment of the substrate;

[0007] S2. Using micro-arc oxidation, a strontium-doped TiO2 coating is prepared on the surface of the pretreated substrate obtained in step S1; then, a pure Ti transition layer is prepared on the surface of the strontium-doped TiO2 coating using electron beam evaporation.

[0008] S3. The pure Ti transition layer is surface treated by anodizing to form a TiO2 mesoporous array layer on the surface of the pure Ti transition layer, thereby obtaining a medical implant.

[0009] Preferably, in step S3, the step of preparing the TiO2 mesoporous array layer by anodic oxidation is as follows: the anodic oxidation power supply is placed in a mixed electrolyte to perform anodic oxidation treatment on the pure Ti transition layer; the process parameters of the anodic oxidation treatment are as follows: treatment voltage 20V, treatment time 55min.

[0010] Preferably, the mixed electrolyte is composed of the following components: 94.75-99.70 wt% ethylene glycol, 0.20-0.30 wt% ammonium fluoride, and 0.1-5.05 wt% water, and the total mass percentage of the three components is 100%.

[0011] Preferably, in step S2, the strontium-doped TiO2 coating is prepared according to the following steps: using a strontium acetate aqueous solution as the electrolyte, the pretreated substrate as the anode, and a platinum plate as the cathode, electrolytic oxidation is performed to obtain the strontium-doped TiO2 coating on the surface of the substrate; the concentration of the strontium acetate aqueous solution is 3 mol / L.

[0012] Preferably, the process parameters for electrolytic oxidation are as follows: forward current is 3A, reverse current is 1A, pulse frequency is 1000Hz, total duty cycle is 30%, positive and negative duty cycles are both 50%, and processing time is 3min.

[0013] Preferably, in step S2, the steps for preparing the pure Ti transition layer using electron beam evaporation are as follows: the substrate is placed in the cavity of a high-vacuum electron beam evaporation coating machine, and a vacuum is drawn until the system vacuum reaches 5 × 10⁻⁶. -4 After Pa, the evaporation process is initiated, and Ti is used as the target material for deposition. The evaporation process continues until the film thickness reaches 100 nm, completing the deposition and obtaining the desired device.

[0014] Preferably, in step S1, the material of the substrate is at least one of titanium, titanium alloy, tantalum alloy, and stainless steel, and its pretreatment is completed by sanding.

[0015] Secondly, the present invention provides a medical implant, comprising:

[0016] Substrate:

[0017] A composite coating is formed on the surface of the substrate, the composite coating comprising a strontium-doped TiO2 coating and a TiO2 mesoporous array layer covering the surface of the strontium-doped TiO2 coating, the TiO2 mesoporous array layer being formed by oxidation of a pure Ti transition layer, and the strontium-doped TiO2 coating being close to the substrate;

[0018] The TiO2 mesoporous array layer has a honeycomb structure, a thickness of 80-120 nm, and an average surface roughness of Ra 4.63 nm and Rq 6.11 nm.

[0019] Preferably, the pore size of the honeycomb structure in the TiO2 mesoporous array layer is 10-14 nm, and the pores are circular or elliptical.

[0020] Thirdly, the present invention provides an application of a medical implant in enhancing cell activity and / or cell differentiation capacity, and its application in femoral repair.

[0021] The beneficial effects of this invention are:

[0022] 1. The method for preparing medical implants provided by the present invention involves first forming a strontium-doped TiO2 coating on the surface of a substrate using micro-arc oxidation, and then forming a TiO2 mesoporous array layer with a honeycomb structure on its surface using anodic oxidation. This method can produce a medical implant with both micron-sized pores and a mesoporous array, thereby increasing the specific surface area of ​​the medical implant. The appearance of this structure can improve cell differentiation and cell activity when the medical implant is used in vivo.

[0023] 2. The preparation method provided by the present invention uses micro-arc oxidation to prepare a strontium-doped TiO2 coating, which can not only accurately form a coating with a specific structure, but also introduce the active element strontium. The introduction of the active element can enhance the osteogenic activity of the medical implant. At the same time, the layer can also improve the bonding strength between the TiO2 mesoporous array layer and the substrate, thereby improving the overall stability of the medical implant.

[0024] 3. The medical implant provided by this invention has excellent biocompatibility and has great application potential for loading macromolecular proteins or drugs. Attached Figure Description

[0025] Figure 1 This is a flowchart illustrating the specific process for preparing the medical implant proposed in Embodiment 1 of the present invention.

[0026] Figure 2 SEM images of the medical implant prepared in Example 1 of this invention at different magnifications;

[0027] Figure 3 This is an EDS-mapping image of the medical implant obtained in Example 1 of the present invention;

[0028] Figure 4 R is the medical implant prepared in Example 1 of the present invention. a and R q Surface roughness;

[0029] Figure 5 XPS image of the medical implant prepared in Example 1 of this invention;

[0030] Figure 6 The results represent the comprehensive performance characterization of the medical implant obtained in Example 1 of this invention.

[0031] Figure 7 The image shows the results of a live / dead cell staining experiment using the medical implant prepared in Example 1 of this invention.

[0032] Figure 8 The figure shows the results of a cell proliferation experiment using the medical implant prepared in Example 1 of this invention.

[0033] Figure 9 The image shows the results of reactive oxygen species detection using the medical implant prepared in Example 1 of this invention.

[0034] Figure 10 The figure shows the results of a protein adhesion experiment using the medical implant prepared in Example 1 of this invention.

[0035] Figure 11 The image shows the result of a micro-CT scan of the medical implant prepared using Example 1 of the present invention.

[0036] Figure 12 This image shows the results of sequential fluorescence staining of the medical implant prepared using Example 1 of the present invention. Detailed Implementation

[0037] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be described in detail below with reference to the accompanying drawings and specific embodiments.

[0038] It should also be noted that, in order to avoid obscuring the present invention with unnecessary details, only the structures and / or processing steps closely related to the present invention are shown in the accompanying drawings, while other details that are not closely related to the present invention are omitted.

[0039] Additionally, it should be noted that the terms “comprising,” “including,” or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such process, method, article, or apparatus.

[0040] On one hand, the present invention provides a medical implant, comprising:

[0041] The substrate is made of at least one of titanium, titanium alloy, tantalum alloy, and stainless steel.

[0042] The composite coating is formed on the surface of the substrate and consists of a strontium-doped TiO2 coating and a TiO2 mesoporous array layer with a honeycomb structure. The TiO2 mesoporous array layer is obtained by oxidation of a pure Ti transition layer, and the strontium-doped TiO2 coating is closely bonded to the substrate.

[0043] In some embodiments, the TiO2 mesoporous array layer has a honeycomb structure with a thickness of 100 nm, an average surface roughness of Ra 4.63 nm and Rq 6.11 nm, and the pore size of the honeycomb structure is 10-14 nm.

[0044] On the other hand, the present invention provides a method for preparing the above-mentioned medical implant, comprising the following steps:

[0045] S1. Surface pretreatment of the substrate;

[0046] S2. Using micro-arc oxidation, a strontium-doped TiO2 coating is prepared on the surface of the pretreated substrate obtained in step S1; then, a pure Ti transition layer is prepared on the surface of the strontium-doped TiO2 coating using electron beam evaporation.

[0047] S3. The pure Ti transition layer is surface treated by anodic oxidation to obtain a TiO2 mesoporous array layer with a honeycomb structure, thus obtaining a medical implant.

[0048] In some embodiments, in step S1, the material of the substrate includes at least one of titanium, titanium alloy, tantalum alloy, and stainless steel, and its surface pretreatment is performed as follows: the substrate is polished for 30 seconds each using silicon carbide (SiC) sandpaper with grits of 400, 800, 1200, and 2000 respectively. After polishing, the obtained samples are cleaned for 5 minutes each in anhydrous ethanol and deionized water to obtain the pretreated substrate.

[0049] In some embodiments, the step of preparing the strontium-doped TiO2 coating using micro-arc oxidation in step S2 is as follows: Using a strontium acetate aqueous solution as the electrolyte, the pretreated substrate as the anode, and a platinum plate as the cathode, electrolytic oxidation is performed to obtain the strontium-doped TiO2 coating on the substrate surface. The forward current is 3A, the reverse current is 1A, the pulse frequency is 1000Hz, the total duty cycle is 30%, and both the positive and negative duty cycles are 50%. The processing time is 3 minutes, and the concentration of the strontium acetate aqueous solution is 3 mol / L. It should be noted that in other embodiments, air cooling is used for temperature reduction during electrolysis, and this can be adjusted as needed; it is not a limitation here.

[0050] In some embodiments, the step of preparing the pure Ti transition layer by electron beam evaporation in step S2 is as follows: placing the substrate in the cavity of a high-vacuum electron beam evaporation coating machine, and evacuating the system to a vacuum level of 5 × 10⁻⁶. -4 After Pa, the evaporation process is started, and Ti is used as the target material for deposition. The evaporation process continues until the film thickness reaches 100 nm, and the deposition is completed. A pure Ti transition layer can be obtained on the surface of the strontium-doped TiO2 coating.

[0051] In some embodiments, the step of preparing the TiO2 mesoporous array layer by anodizing in step S3 is as follows: The anodizing power supply is placed in a mixed electrolyte to oxidize the substrate with the pure Ti transition layer formed on its surface. The process parameters for the anodizing treatment are as follows: treatment voltage 20V, treatment time 55min. The mixed electrolyte consists of the following components: ethylene glycol 94.75-99.70wt%, ammonium fluoride 0.20-0.30wt%, and water 0.1-5.05wt%, with the total mass percentage of the three components being 100%.

[0052] The preparation method and application of the medical implant proposed in this invention will be further explained below with reference to specific embodiments:

[0053] Example 1

[0054] Please see Figure 1 As shown, this embodiment provides a method for preparing a medical implant, including the following steps:

[0055] S1. Using a titanium plate (length × width × height = 10mm × 10mm × 1mm) as the substrate, the titanium plate is polished sequentially using silicon carbide abrasive paper with grits of 400, 800, 1200, and 2000. The polishing time using 400 grit silicon carbide abrasive paper is 30 seconds, and the polishing time using other grit silicon carbide abrasive paper is the same. After polishing, the plate is cleaned with anhydrous ethanol and deionized water for 5 minutes each under ultrasonic conditions to obtain a pretreated titanium plate.

[0056] S2. Using strontium acetate aqueous solution as electrolyte, a pretreated titanium plate as anode, and a platinum plate as cathode, micro-arc oxidation was performed using an IGBT micro-arc oxidation power supply under the following parameters: processing time 180s, pulse frequency 1000Hz, total duty cycle 30%, positive and negative duty cycles 50% respectively, forward current 3A, reverse current 1A. Air cooling was used during the process. The treated sample was then washed again with methanol, anhydrous ethanol, and deionized water for 5min, and dried at room temperature (20-25℃) to obtain a strontium-doped TiO2 coating on the surface of the titanium plate, denoted as MAO-Sr. Next, under vacuum conditions, the substrate was placed in the cavity of a high-vacuum electron beam evaporation coating machine, and the system vacuum was evacuated to 5×10⁻⁶. -4 After Pa, the evaporation program was started, and Ti was used as the target material for deposition. The evaporation process continued until the film thickness reached 100 nm, completing the deposition to uniformly deposit a 100 nm thick pure titanium film on the surface of the strontium-doped TiO2 coating. Then, it was washed sequentially with methanol, anhydrous ethanol, and water for 5 min, dried at room temperature, and then used for later use, denoted as MAO-Sr@VTC. The strontium acetate aqueous solution was obtained by dissolving 123.4 g of strontium acetate in 3 L of water.

[0057] S3. The anodic oxidation power supply is placed in the mixed electrolyte to perform anodic oxidation treatment on the substrate with a pure titanium film on its surface, thereby treating the pure titanium film into a TiO2 mesoporous array layer with a honeycomb structure. The process parameters for anodic oxidation treatment are as follows: treatment voltage 20V, treatment time 55min. After treatment, the substrate is washed sequentially with methanol, anhydrous ethanol, and water for 5min each, and then dried at room temperature to obtain the medical implant, denoted as MAO-Sr@VTC / AO. The mixed electrolyte consists of the following components: ethylene glycol 94.75wt%, ammonium fluoride 0.25wt%, and water 5wt%. The anodic oxidation power supply was purchased from Huangyang Electronics Co., Ltd. (Hangzhou, China). It should be noted that, unless otherwise specified, the reagents and raw materials used in the specific embodiments of this invention can be obtained commercially.

[0058] The SEM images of the materials obtained in each stage of steps S2-S3 of this embodiment are as follows: Figure 2 As shown, where, Figure 2 Figures a1-a3 in the figure show SEM images of MAO-Sr at different magnifications (5k, 10w, 20w). Figure 2 Figures b1-b3 in the figure show the SEM images of MAO-Sr@VTC at different magnifications. Figure 2 Figures c1-c3 show SEM images of MAO-Sr@VTC / AO at different magnifications. Figures a1-a3 show the micro-arc oxidation-formed coating with undulating micron-sized pores, and the coating surface has a large number of pores formed by discharge breakdown. These pores are mostly circular or elliptical, some are bean-shaped, and some contain multiple smaller secondary pores. The average diameter of the secondary pores is 1.3±0.5μm. At high magnification, fine impurities can be observed on the coating surface. Figures b1-b3 show that the micron-sized pores still exist, and pure Ti particles generated by the coating can be observed at the nanoscale. Figures c1-c3 show that in addition to the micron-sized pores, a titanium dioxide mesoporous structure (MPs) formed by anodic oxidation is added. At a magnification of 5000x, the surface morphology of MAO-Sr@VTC, MAO-Sr@VTC / AO and MAO-Sr is not significantly different. At magnifications of 100,000x and 200,000x, the surface roughness of MAO-Sr@VTC / AO is higher, and fine particles can be seen (pure Ti particles in Figure b3 and TiO2 particles in Figure c3).

[0059] EDS-mapping diagrams for MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO are shown below. Figure 3As shown, Sr has been successfully embedded in the coating, confirming its effective doping at the micrometer scale. The R... (The text abruptly ends here, likely due to an incomplete translation or source material.) a Surface roughness such as Figure 4 As shown in Figure A, R q Surface roughness such as Figure 4 As shown in Figure B, the results show that micro-arc oxidation and anodizing can gradually increase the surface roughness of the material.

[0060] XPS graphs for MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO are shown below. Figure 5 As shown in Figure A, the XRD pattern is as follows: Figure 5 As shown in Figure B. From Figure A, it can be seen that in the MAO-Sr XPS plot, the characteristic peaks of the Ti 2p orbitals appear at 458.24 eV (Ti 2p...). 3 / 2 ) and 463.95eV (Ti 2p 1 / 2 The presence of satellite peaks at 470.97 eV, along with these signals, collectively indicates the presence of TiO2 and SrTiO4 phases, suggesting that the oxidation reaction during MAO generates typical titanium-based oxides. In the Sr 3d spectrum, the 3d phases of SrO were observed. 3 / 2 (132.44eV) and 3d 1 / 2 The (134.27eV) peak and the 3d peak of SrTiO3 3 / 2 (133.27eV) and 3d 1 / 2 The (135.02 eV) peak further confirms that Sr exists stably in the coating as an oxide and reflects its specific chemical valence state. For MAO-Sr@VTC, its Ti2p spectrum shows significant changes compared to MAO-Sr, with the two newly added pairs of peaks corresponding to subvalent titanium (Ti). n+ ) of 2p 3 / 2 (454.78eV) and 2p 1 / 2 (460.92 eV), and the 2p of Ti2O3 3 / 2 (457.23eV) and 2p 1 / 2(463.23 eV) indicates that the pure Ti coating and its oxidation products have been successfully formed. While the main peak of the Sr 3d spectrum did not change significantly, a slight leftward shift was observed, indicating a slight adjustment in the chemical environment of Sr. For MAO-Sr@VTC / AO, its Ti2p and Sr 3d spectra re-exhibited peak characteristics similar to MAO-Sr. Figure B shows that, according to the phase search results, MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO are mainly composed of rutile, anatase, Sr, Ti, and Sr(CO3), with rutile, anatase, and Ti having relatively high contents. Phase overlap was observed at 2θ of 25.3°, 27.48°, 35.14°, 38.44°, 62.92°, and 70.56°. Comparison with the PDF card reveals that 25.3° represents anatase. The (101) crystal plane of anatase and the (111) crystal plane of Sr(CO3); 27.48° represents the (110) crystal plane of Rutile and the (012) crystal plane of Sr(CO3); 35.14° represents the (102) crystal plane of Sr and the (100) crystal plane of Ti; 38.44° represents the (112) crystal plane of anatase and the (002) crystal plane of Ti; 62.92° represents the (002) crystal plane of Rutile and the (203) crystal plane of Sr; 70.56° represents the (220) crystal plane of anatase and the (103) crystal plane of Ti. Compared to MAO-Sr@VTC, the MAO-Sr@VTC / AO sample still mainly exhibits the XRD characteristic diffraction peaks of the substrate, without any new or reduced peaks, only a slight impact on peak intensity. This indicates that the main crystal structure peaks still originate from the substrate. Comparing the MAO-Sr and MAO-Sr@VTC / AO coatings, the peak intensity of the MAO-Sr@VTC coating is lower, indicating that the crystal structure on the sample surface becomes more ordered after oxidation treatment, thereby increasing the diffraction peak intensity.

[0061] The overall performance of Ti, MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO is as follows: Figure 6 As shown, Figure 6 Figure A shows the water contact angles of the four materials under dry conditions (left side of Figure A) and wet conditions (right side of Figure A). The results show that Ti has the smallest average contact angle at 74.4°, indicating relatively strong hydrophilicity. MAO-Sr has the largest contact angle at 80.6°, indicating poor surface wettability. The contact angles of MAO-Sr@VTC and MAO-Sr@VTC / AO are 79.9° and 76.7°, respectively, indicating that coating modification can improve surface wettability to some extent. Although it is slightly lower than that of MAO-Sr, it is still higher than that of Ti overall.

[0062] Figure 6 Figure B shows the continuous leaching curves of Sr element in four materials stored in 10 mL PBS solution for 1 day, 3 days, 7 days, 21 days, and 41 days. The results show that the coatings in each group exhibit similar release trends throughout the release period. The release rate is relatively fast in the initial stage, and the release amount in the first three days accounts for more than 50% of the total release amount (up to the 42nd day), showing typical characteristics of rapid initial release. There is no significant difference in the Sr release behavior between the MAO-Sr group and the MAO-Sr@VTC / AO group. Whether it is the release amount at a single time point or the cumulative release amount at each stage, they are basically similar. However, the MAO-Sr@VTC group shows the highest Sr release level throughout the release process. Its cumulative release amount shows a significant gap with the MAO-Sr group and the MAO-Sr@VTC / AO group in the later stage, about 1.5 times and 2 times of them respectively. This phenomenon may be related to the high-temperature environment during the electron beam evaporation coating process. The high-temperature treatment may make the Sr element more likely to migrate to the surface layer, thus increasing its subsequent dissolution rate in the liquid. In contrast, the Sr release of the MAO-Sr@VTC / AO group is the lowest. The possible reason is that during the anodic oxidation process, some Sr elements fail to be stably retained in the coating but migrate and dissolve into the electrolyte, resulting in a decrease in its effective retention amount in the final coating, thereby affecting its release performance.

[0063] Figure 6 Figure C in it shows the elastic modulus (EIT) of MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO. Figure 6 Figure D in it shows the hardness (HIT) of MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO. With the hierarchical enhancement of the coating structure, its elastic modulus (EIT) and hardness (HIT) show an increasing trend, showing as MAO-Sr group < MAO-Sr@VTC group < MAO-Sr@VTC / AO group. This result indicates that with the superposition of coating and anodic oxidation treatments, the mechanical properties of the coating at the microscale are effectively enhanced.

[0064] Figure 6 Figure E in it shows the critical bonding force between the coating and the substrate titanium plate in MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO. However, the nano-scratch data show that the bonding strength between the coating and the Ti substrate shows a decreasing trend step by step, in the order of MAO-Sr group > MAO-Sr@VTC group > MAO-Sr@VTC / AO group. This may be because after the MAO coating undergoes multiple treatments such as high-temperature electron beam coating and electrochemical anodic oxidation, its original bonding interface with the Ti substrate is weakened to a certain extent, thus reducing the overall bonding force.

[0065] Figure 6Figure F shows the physical adhesion of proteins on the surface of the four materials after co-culturing with fetal bovine serum albumin (FBO) for 24 hours. The concentration of surface-adhered proteins in each material was detected by elution after 24 hours of co-incubation with FBO. The results showed that the MAO-Sr@VTC / AO group had the highest surface protein adsorption, which is attributed to the presence of both micron-sized pores formed by micro-arc oxidation and nanotube structures (MPs) formed by anodic oxidation, significantly increasing the specific surface area and surface energy, thus enhancing the physical adsorption capacity of proteins. The protein adsorption of the MAO-Sr@VTC group was slightly lower than that of the MAO-Sr@VTC / AO group, but higher than that of the MAO-Sr group. This is presumably because the large number of pure Ti nanoparticles formed during electron beam evaporation coating increased surface roughness and active sites, improving protein binding capacity. Although the MAO-Sr group had a certain number of micron-sized pores, its adsorption capacity was slightly inferior compared to the synergistic effect of the micro-nano structures. The pure Ti group had the lowest protein adhesion due to its smooth surface.

[0066] Figure 6 Figure G in the figure shows the scratch electron microscope images of MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO at magnifications of 1k, 10w, and 20w, respectively (the red boxes represent the extrusion deformation rather than peeling of the pure Ti coating and the MPs structure, respectively). The results show that after being subjected to scratch loading force, the material exhibits extrusion deformation but still maintains good adhesion to the underlying MAO coating (as shown in the red box in the figure), without obvious delamination or peeling.

[0067] To enable the medical implants prepared in the above embodiments to be used in vivo, this embodiment first verifies their applicability in vitro, as detailed below:

[0068] (1) Live / dead cell staining experiment: Mouse osteoblast progenitor cells MC3T3-E1 were stained with 2×10⁻⁶ cells. 4 Cells / mL were seeded onto the surfaces of Ti, MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO groups in 24-well plates at densities of 1, 3, and 7, respectively, and cultured in a cell culture incubator (37.0℃, 5% CO2) for 1, 3, and 7 days, respectively. Cell proliferation was assessed using CCK-8 assay. Figure 7 Figure A shows that cell proliferation on each material group exhibited a significant time-dependent effect. There was no significant difference in cell viability between the groups at days 1 and 3, but a significant difference emerged at day 7, with the VTC and AO groups showing a marked increase compared to the Ti and MAO-Sr groups. To further differentiate the effects of Sr release and morphological roughness on cell behavior, an additional experiment was designed. This experiment used CCK-8 assays to detect cell proliferation, and the results are shown below. Figure 7Figure B shows that cell viability generally increased on days 1 and 7, with a slight decreasing trend on day 3. The cell viability of Ti samples treated with the coating extract (Sr extract) was higher than that of the untreated Ti group, but still lower than that of the MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO groups. This indicates that under these experimental conditions, the individual contribution of Sr is not as significant as the effect of surface morphology roughness on cell viability.

[0069] MC3T3-E1 cells were cultured for 48 h on Ti plates, MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO substrates. After staining, the cells were stained using the Calcein-AM / PI live / dead cell double staining kit (Servicebio, Wuhan, China). Following staining, the cells were observed and imaged using a fluorescence microscope (CKX53, OLYMPUS, Japan). Live cells showed green fluorescence, and dead cells showed red fluorescence. The live / dead cell staining results are shown below. Figure 7 As shown in Figure C, the results indicate that all groups exhibited good biocompatibility, and there were no significant differences between the groups. The stained cells were digested and analyzed by flow cytometry (Cytomics FC 500, Beckman Coulter, USA). Figure 7 As shown in Figure D, the results are consistent with those observed under a fluorescence microscope, indicating good biocompatibility in all groups. The ratio of live to dead cells on the surface of each sample was obtained from the fluorescence microscope images, and the results are as follows: Figure 7 As shown in Figure E, the results indicate that all groups have good biocompatibility; the relative spreading area of ​​cells within the fluorescence map of phalloidin was statistically analyzed using ImageJ, and the results are as follows. Figure 7 As shown in Figure F, the results show that the cell extension area increases sequentially with the treatment groups; the adhesion and extension detection results of MC3T3-E1 cells after culturing on the surfaces of Ti, MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO materials for 24 h (red represents actin, blue represents the cell nucleus) are shown in Figure F. Figure 7 As shown in Figure G, the results show that the number of cells did not differ significantly between groups, but the area of ​​spread increased.

[0070] (2) Cell proliferation experiment: MC3T3-E1 cells were injected with 2×10⁻⁶ cells. 3Cells were seeded on the surfaces of four groups of materials, namely Ti, MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO, at a density of cells / mL, and cultured in an incubator at 37°C and 5% CO2 for 1, 3, and 7 days. At each time point, a CCK-8 cell proliferation detection kit (GlpBio, USA) was used for detection: after adding the CCK-8 working solution (the cck-8 solution accounted for 10% of the volume of the basal medium) to each well and incubating for 2 h, the optical density (OD) value at 450 nm was measured using an enzyme-labeled instrument. To further evaluate the effects of element content, surface morphology, and roughness in the materials on cell proliferation activity, the above four groups of samples were immersed in the medium for 7 days. Another group of pure Ti samples was set up, and cell culture was carried out using the leaching solution of the MAO-Sr@VTC group. During the same culture period (1, 3, and 7 days), the CCK-8 test results of the cells were also collected and the OD values were recorded to compare and analyze the synergistic effects of element release and surface structure on cell proliferation.

[0071] Alkaline phosphatase ALP and alizarin red ARS staining were detected on the 7th and 14th days of osteogenic induction culture, and the results were as Figure 8 shown in Figure A in. The results showed that in the early stage of osteogenesis, there was no obvious difference in ALP color development among the groups. In the late stage of osteogenesis, ARS staining showed that the area and staining depth of mineralized nodules showed a trend of Ti < MAO-Sr < MAO-Sr@VTC < MAO-Sr@VTC / AO group. However, due to the background red-violet tone on the surface of the AO group after anodic oxidation treatment, the contrast of the ARS staining results was reduced under the naked eye; the semi-quantitative activity analysis results of ALP and the semi-quantitative analysis results of ARS were shown in Figure B - Figure C in Figure 8 respectively. The semi-quantitative results of ALP activity showed that the Ti group showed the lowest enzyme activity, while the MAO-Sr@VTC group was the highest, even exceeding the MAO-Sr@VTC / AO group. This phenomenon may be related to the release of a larger amount of Sr element by the MAO-Sr@VTC group in the early stage of the experiment, and its cumulative release amount was twice that of the MAO-Sr@VTC / AO group. Although the MAO-Sr@VTC / AO group had a higher surface roughness and an ordered MPs structure, it still failed to achieve a higher ALP activity expression. Under the condition that the Sr element release amount of the MAO-Sr@VTC / AO group was slightly lower than that of the MAO-Sr group, the AO group could have a higher level of ALP activity, indicating that its MPs structure had certain advantages in promoting early osteogenic differentiation. Generally speaking, the ALP activity levels of each group were relatively low, which may be related to the differentiation stage of the cells. The semi-quantitative results of ARS showed that the mineralization levels on the surfaces of the four groups of materials increased in turn, and the MAO-Sr@VTC / AO group was significantly better than the MAO-Sr@VTC group; on the 7th day of culture, the western blot results of osteogenesis-related proteins (OPN, RUNX2, and OCN) were as Figure 8 As shown in Figure D, the results showed that the expression levels of three osteogenic-related proteins, RUNX2, OCN, and OPN, gradually increased from the Ti, MAO-Sr, MAO-Sr@VTC to MAO-Sr@VTC / AO groups. Among these, the expression levels of OPN showed less significant difference among the groups than those of RUNX2 and OCN. OCN was significantly upregulated in the MAO-Sr@VTC and MAO-Sr@VTC / AO groups compared to the Ti and MAO groups. The comparison results of the gray values ​​of osteogenic-related proteins (OPN, RUNX2, and OCN) with the internal control are shown in Figure D. Figure 8 Figures E-G in the table show the quantification of the above results; the expression levels of osteogenic related genes (OPN, RUNX2, OCN) are as follows: Figure 8 As shown in Figures H-J, the results are consistent with the protein-related results.

[0072] (3) Cytoskeleton stretching assay: MC3T3-E1 cells were sputtered at a rate of 2×10⁻⁶ mcg / cm³. 4 Cells were seeded at densities of 1000 cells / mL on Ti plates, MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO substrates, and co-cultured for 24 h. Subsequently, cells were fixed with 4% paraformaldehyde (Biosharp, China) for 10 min; then, they were permeabilized with 0.5% Triton X-100 (T50006, Pusitang, China) for 5 min. After permeabilization, TRITC-labeled phalloidin (C2207S, Beyotime, China) and DAPI (50%, Southern Biotech, USA) were added sequentially to each well to stain cell actin and nuclei. The results are shown below. Figure 7 As shown in Figure F, the results indicate that the cell spreading area increases sequentially with the treatment group. After staining, the cells were observed and photographed using a fluorescence microscope, and the spreading area was measured and counted using ImageJ software. The adhesion and spreading results of MC3T3-E1 cells after culturing on the surfaces of Ti, MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO groups for 24 hours are shown below (red indicates actin, blue indicates the cell nucleus). Figure 7 As shown in Figure G, the results show that the number of cells did not differ significantly between groups, but the area of ​​spread increased.

[0073] (4) Qualitative and semi-quantitative experiments on alkaline phosphatase (ALP) and calcium nodules (ARS): MC3T3-E1 cells were subjected to 6×10⁻⁶ saturates. 4Four groups of materials—Ti plates, MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO—were seeded at a density of cells / mL and cultured under osteogenic induction conditions (osteogenic induction medium consisting of 0.1% dexamethasone, 1% ascorbic acid, 1% sodium β-glycerophosphate, 1% ALA-GLN, 1% penicillin-streptomycin, 10% fetal bovine serum, and basal medium) until day 7. The original medium was discarded, and ALP was qualitatively detected using alkaline phosphatase chromogenic reagent (BCIP / NBT) (Absin, China); semi-quantitative ALP was detected using an alkaline phosphatase assay kit (p0321S) provided by Beyotime (China).

[0074] MC3T3-E1 cells were loaded at 6×10 4 The cells / mL were seeded onto the surfaces of four materials: Ti plates, MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO, and cultured for 14 days. Subsequently, the original culture medium was removed, and ARS qualitative detection was performed using the Beyotime osteoblast mineralization nodule staining kit (Alizarin Red S method); calcium nodules were eluted with 10% hexadecylpyridine chloride solution for quantitative detection. Specific results are described in (2). Figure 8 The contents of Figures B-C are shown in the figure.

[0075] (5) RT-PCR detection of osteogenic-related factors: Real-time quantitative PCR (RT-PCR) was used to detect the expression levels of osteogenic-related genes in MC3T3-E1 cells (after osteogenic induction culture for 14 days), including osteocalcin (OCN), osteopontin (OPN), and Runt-related transcription factor 2 (RUNX2). In the experiment, total RNA was extracted from cell samples co-cultured with each group of materials for 14 days using a total RNA column extraction kit (Liji Biotechnology, China). The concentration and purity of RNA samples were evaluated using a nano-volume UV-Vis spectrophotometer (NanoDrop One, Thermo Fisher Scientific, USA).

[0076] Subsequently, cDNA synthesis was completed using reverse transcription and amplification reagents (Takara Bio, Japan). 1 μg of total RNA was used in each reaction system for reverse transcription to generate cDNA. The resulting cDNA was mixed with TB Green, ROX II, and gene-specific primers (specific sequences are shown in Supplementary Table 1) and amplified using a real-time quantitative PCR instrument (ABI 7500, Thermo Fisher Scientific, USA) to analyze the expression levels of target osteogenic-related genes.

[0077] (6) Quantitative detection of osteogenic-related proteins using Western blotting: The expression levels of osteogenic-related proteins OPN, OCN, and RUNX2 were quantitatively detected using Western blotting (after osteogenic induction culture for 14 days). In the experiment, cell samples were lysed at 4°C for 15 min to ensure sufficient protein release, followed by extraction of total protein and quantification of its concentration. Protein samples were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and then transferred to a polyvinylidene fluoride (PVDF) membrane. Non-specific binding sites on the membrane were blocked with 5% skim milk, and the membrane was then cut into appropriate bands according to the protein molecular weight.

[0078] Each target protein (OPN, OCN, RUNX2) was incubated overnight at 4°C with a 1:1000 dilution of primary antibody (Proteintech, China). The next day, the membrane was washed three times for 10 min each with TBS buffer (TBST) containing 1% Tween-20, and then incubated for 2 h at room temperature with HRP-labeled secondary antibody of the same species (1:1000 dilution, Thermo Fisher Scientific, USA). Subsequently, the membrane was developed using enhanced chemiluminescence (ECL) reagent (Thermo, USA), and the protein bands were observed using a gel imaging system (ChemiDoc MP, USA).

[0079] GAPDH, the internal control protein, was diluted 1:50,000 and incubated with the membrane strips overnight at 4°C. On the third day, the washing, secondary antibody incubation, and development steps were repeated. Finally, ImageJ software was used to perform grayscale analysis on each protein band, calculate the relative expression ratio of the target protein to the internal control, and compare the differences between different groups.

[0080] (7) Detection of reactive oxygen species (ROS): RAW264.7 mononuclear macrophages were subjected to a reaction of 6 × 10⁻⁶ ppm. 4 Cells were seeded at a density of 100 cells / mL on the surfaces of four materials: Ti plates, MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO. Oxidative stress was induced using α-high glucose medium (Zhong Qiao Xinzhou, China) containing 300 μM hydrogen peroxide. After 3 days of co-culturing under these conditions, cells were incubated with a 1:1000 diluted DCFH-DA fluorescent probe (S0033S, Beyotime, China) at 37°C for 20 min to detect intracellular reactive oxygen species (ROS) levels. After incubation, the relative expression of intracellular ROS on the surfaces of each material was analyzed using flow cytometry (Cytomics FC 500, Beckman Coulter, USA) to compare the regulatory ability of different surface modifications on oxidative stress levels.

[0081] After co-culturing cells with four groups of materials for 3 days, the residual reactive oxygen species (ROS) level in the cells was measured by flow cytometry. The results are as follows: Figure 5 As shown in Figure A, the results showed that the mean ROS fluorescence intensity in the Ti group was the highest, reaching 9045.33, while the mean ROS in the MAO-Sr, MAO-Sr@VTC, and MAO@VTC / AO groups were 7500.67, 6376, and 4884.33, respectively, decreasing sequentially from group to group. This indicates that the MAO-Sr@VTC / AO group had the best effect in promoting ROS clearance. After co-culturing for 3 days, the OD value was measured at a wavelength of 450 nm, and the results are as follows. Figure 5 As shown in Figure B, the results indicate that the Ti group had the lowest SOD activity, while the MAO-Sr@VTC / AO group had the highest, followed by the MAO-Sr group and then the MAO-Sr@VTC group. After co-culturing for 3 days, the catalase (CAT) activity of cells on the surface of the four groups was tested, and the results are shown in Figure B. Figure 5 As shown in Figure C, the results show that the overall CAT enzyme activity showed an increasing trend, but the MAO-Sr group was slightly higher than the MAO-Sr@VTC group (0.01998 and 0.01917, respectively).

[0082] (8) Catalase activity assay: RAW264.7 cells were subjected to a 6×10⁻⁶ thiocyanate incubator. 4 Cells were seeded at a density of [number] cells / mL on the surfaces of four materials: Ti plates, MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO, and co-cultured for 3 days under standard culture conditions. After culture, cells were lysed at 4°C, and the supernatant was collected by centrifugation for subsequent assays. First, the protein concentration in the cell lysates of each group was determined using the BCA protein concentration assay kit (Thermo Fisher Scientific, USA). Subsequently, the catalase (CAT) activity assay kit (S0051, China) of Beyotime was used to assess the CAT enzyme activity level in each group of cells. Assays were performed at a wavelength of 520 nm, and the absorbance (A) of the samples was recorded. 520 Blank control A 520 And standard curve data, and calculate catalase activity according to the following formula:

[0083] The catalase activity of the sample is calculated as follows: [number of micromoles of hydrogen peroxide consumed] × [dilution factor] / ([reaction time] × [sample volume] × [protein concentration]). The number of micromoles of hydrogen peroxide consumed is calculated as follows: [number of micromoles of residual hydrogen peroxide in blank control] - [number of micromoles of residual hydrogen peroxide in sample]. The dilution factor is 250. The reaction time is the actual reaction time. The sample volume is in μL. The protein concentration is in mg / mL. The blank control group is a reagent mixture without cell lysis buffer.

[0084] (9) Superoxide dismutase activity assay: RAW264.7 cells were subjected to a 6×10⁻⁶ thiocyanate incubator. 4 Cells were seeded at a density of cells / mL on the surfaces of four materials: Ti plates, MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO, and cultured for 3 days under standard conditions. After culture, cells were fully lysed at 4°C and centrifuged, and the supernatant was collected for subsequent detection.

[0085] The total superoxide dismutase (SOD) activity assay kit from Beyotime (S0101S, China) was used to assess the SOD activity in each group of samples. Cell lysis buffer was mixed with the prepared working solution and incubated at 37°C in the dark for 30 min. In the reaction system, WST-8 reacts with superoxide anions (SOD). 2· - The reaction produces an orange-yellow formazan dye, which has a characteristic absorption peak at a wavelength of 450 nm; while SOD can catalyze O 2· - A disproportionation reaction occurs, reducing the formation of formazan dye, therefore A 450 The value was negatively correlated with SOD activity. This was determined by measuring the A value of each sample. 450 Absorbance values ​​can indirectly evaluate SOD activity.

[0086] (10) Protein Adhesion Assay: Cleaned Ti plates, MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO materials were co-incubated with fetal bovine serum at 37°C for 24 h. After incubation, the adsorbed proteins were recovered by eluting the material surfaces with 1% sodium dodecyl sulfate (SDS, Solarbio, China). Subsequently, the concentration of surface-adhesive proteins in each group was determined using a BCA protein quantification kit (YC373312, Thermo Fisher Scientific, USA) to assess the differences in protein adsorption capacity of different material surfaces. The results are as follows: Figure 10 As shown in the figure, the amount of protein adhered by physical means also increases with the continuous improvement of the processing level.

[0087] The above in vitro experimental results show that medical implants enhance cell viability and differentiation capacity. This embodiment further explores the applicability of medical implants in vivo, and the main contents are as follows:

[0088] (1) Micro-CT Scan: Femoral specimens were fixed in 4% paraformaldehyde for 3 days and then analyzed using Micro-CT (NMC-200, Ping Sheng Medical Technology, China) to evaluate bone repair. Scanning parameters were set as follows: radial field of view (FOV) of 100 mm; maximum axial scan range of 250 mm; maximum scan speed of 4 s / bed; reconstructed pixel size of Min.2 μm; spatial resolution <7.5 μm@10% MTF. The original images were reconstructed using the 3D reconstruction software Recon. The implant area was selected as the region of interest (ROI), and the target ROI was analyzed using the data analysis software Avatar. All samples were analyzed within the same region to obtain the required parameter values. Male SD rats weighing approximately 200 g were used as experimental animals. All animals were randomly divided into four groups, corresponding to the Ti, MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO groups, respectively. After one week of acclimatization, the relevant bone defect model was constructed. Before surgery, rats were anesthetized by intraperitoneal injection of 3% sodium pentobarbital (0.5 mL / kg), and their hind limbs were shaved. The limbs were then repeatedly wiped with iodine and thoroughly disinfected. Subsequently, a longitudinal incision was made in the skin and muscle layer of the knee to expose the distal femur. Holes were drilled along the long axis of the femur, and samples from the corresponding experimental group were implanted parallel to the medullary cavity to complete the model construction.

[0089] Specific results are as follows Figure 11 As shown, where, Figure 11 Figures A and B in the image are the sagittal and cross-sectional views of an isolated femur, respectively. Figure 11 Figure C in the image shows a visualized new bone reconstructed in 3D using Micro-CT data (blue in the image). Figure 11 Figures D-F in the table show the statistical analysis of Bone Volume / Tissue Volume (BV / TV), Bone Surface / Tissue Volume (BS / TV), and Tb.Th from micro-CT scans. Figure C shows that the amount of new bone formation increases with the increase in the treatment layer of the group. Figures D-F show that BS / TV, BV / TV, and Tb.Th all increase sequentially with Ti, MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO groups, but Tb.Th does not show a statistically significant difference. (BV / TV is the most direct indicator of bone mineral density; a higher value indicates more bone mass. BS / TV assesses bone remodeling activity; a higher value indicates more active bone remodeling. Tb.Th reflects the average thickness of bone trabeculae; greater thickness indicates more stable bone structure.)

[0090] (2) Sequential fluorescence staining: Rats were sacrificed 6 weeks post-surgery. Four weeks and two weeks before sacrifice, calcein (5 mg / kg) and alizarin red (2.5 mg / kg) were injected intraperitoneally, respectively. Specimens were fixed with 4% PFA and then sequentially dehydrated in 60%-70%-80%-90%-95% ethanol for 24 hours each time. The tissues were then infiltrated with T7200 resin for 20 days. The tissues were placed in an embedding mold, and an appropriate amount of T7200 resin was added, allowing polymerization for 12 hours. The embedded sample blocks were then attached to a glass slide using T4000 adhesive. Using 1200-grit sandpaper, the sample blocks attached to the slide were smoothed to create a uniform surface, exposing the tissue to be observed. The sample blocks with the attached slide were then attached to the upper slide using T7210 precision adhesive. The slides were irradiated with blue light for approximately 10 minutes before removal. Hard tissue sections with a thickness of 200 μm were obtained using a hard tissue microtome. These sections were then ground into hard tissue sections with a thickness of 20-30 μm using a hard tissue grinder. Images were then captured using a confocal microscope (STELLARIS 5, Leica, Germany), and the fluorescence area was quantitatively analyzed using ImageJ software. The results are as follows: Figure 12 As shown, where, Figure 12 Figure A shows a fluorescent double-staining image of calcein (green) and alizarin red (red). Figure 12 Figures B-C show the sequential fluorescence staining analysis of the staining area of ​​Calcein and Alizarin Red around the medical implant, respectively. Figure A shows that at 2 weeks post-operation, the amount of new bone formation indicated by Calcein around the implant was relatively small, with the MAO-Sr group showing the highest bone deposition, although MAO-Sr@VTC / AO was still significantly higher than the other two groups. At 4 weeks post-operation, the amount of new bone formation indicated by Alizarin Red increased sequentially with the treatment level of Ti, MAO-Sr, MAO-Sr@VTC, and MAO-Sr@VTC / AO groups; Figures B and C show the quantitative data of the staining area of ​​Calcein and Alizarin Red around the implant, respectively.

[0091] Comparative Example 1

[0092] The only difference between Comparative Example 1 and the Example is that only micro-arc oxidation treatment is performed, that is, step S3 is omitted. The remaining experimental conditions and parameters are the same as those in Example 1, and will not be repeated here.

[0093] Comparative Example 2

[0094] The only difference between Comparative Example 1 and the Example is that only the anodizing method is performed, that is, step S2 is omitted. The remaining experimental conditions and parameters are the same as those in Example 1, and will not be repeated here.

[0095] Comparative Example 3

[0096] The only difference between Comparative Example 3 and the Example is that the titanium plate is first anodized and then micro-arc oxidized. The other experimental conditions and parameters are the same as those in Example 1, and will not be repeated here.

[0097] During the experiment, it was found that the materials prepared in Comparative Examples 1 to 3 had significantly lower biocompatibility and stability than those in Example 1. This is because: in Example 1, the titanium plate was first subjected to micro-arc oxidation to form a strontium-doped TiO2 coating, and then anodizing was used to form a TiO2 mesoporous array layer with a specific honeycomb structure. This process can form an element-micron-nano composite coating, while Comparative Examples 1 to 2 could not form the aforementioned synergistic composite functional layer. As for Comparative Example 3, which first performed anodizing and then micro-arc oxidation, the cylindrical or honeycomb nanotube structure formed by anodizing was broken down and melted by the high-voltage arc of the subsequent micro-arc oxidation. Although this increased the thickness of the micro-arc oxidation coating, the original nanotube structure was destroyed, resulting in lower biocompatibility than in Example 1.

[0098] The above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit it. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the spirit and scope of the technical solutions of the present invention.

Claims

1. A method for preparing a medical implant, characterized in that, Includes the following steps: S1. Surface pretreatment of the substrate; S2. Using micro-arc oxidation, a strontium-doped TiO2 coating is prepared on the surface of the pretreated substrate obtained in step S1; then, a pure Ti transition layer is prepared on the surface of the strontium-doped TiO2 coating using electron beam evaporation. The preparation method of the strontium-doped TiO2 coating is as follows: using strontium acetate aqueous solution as electrolyte, the pretreated substrate as anode, and a platinum plate as cathode, electrolytic oxidation is performed to obtain the strontium-doped TiO2 coating on the surface of the substrate. The concentration of the strontium acetate aqueous solution is 3 mol / L. The process parameters of electrolytic oxidation are as follows: forward current is 3A, reverse current is 1A, pulse frequency is 1000Hz, total duty cycle is 30%, positive and negative duty cycles are both 50%, and processing time is 3min. The steps for preparing a pure Ti transition layer using electron beam evaporation are as follows: The substrate is placed inside the cavity of a high-vacuum electron beam evaporation coating machine, and a vacuum is drawn until the system vacuum reaches 5 × 10⁻⁶. -4 After Pa, the evaporation process is started, and Ti is used as the target material for deposition. The evaporation process continues until the film thickness reaches 100 nm, and the deposition is completed to obtain the desired device. S3. The pure Ti transition layer is surface treated by anodizing to form a TiO2 mesoporous array layer on the surface of the pure Ti transition layer, thereby obtaining a medical implant.

2. The preparation method according to claim 1, characterized in that, In step S3, the steps for preparing the TiO2 mesoporous array layer by anodic oxidation are as follows: the anodic oxidation power supply is placed in a mixed electrolyte to perform anodic oxidation treatment on the pure Ti transition layer; the process parameters for the anodic oxidation treatment are as follows: treatment voltage 20V, treatment time 55min.

3. The preparation method according to claim 2, wherein the mixed electrolyte comprises the following components: Ethylene glycol 94.75-99.70 wt%, ammonium fluoride 0.20-0.30 wt%, water 0.1-5.05 wt%, and the total mass percentage of the three is 100%.

4. The preparation method according to claim 1, characterized in that, In step S1, the substrate is made of at least one of titanium, titanium alloy, tantalum alloy, and stainless steel, and its pretreatment is completed by sanding.

5. A medical implant prepared by the method according to any one of claims 1-4, characterized in that, include: Substrate: A composite coating is formed on the surface of the substrate, the composite coating comprising a strontium-doped TiO2 coating and a TiO2 mesoporous array layer covering the surface of the strontium-doped TiO2 coating, the TiO2 mesoporous array layer being formed by oxidation of a pure Ti transition layer, and the strontium-doped TiO2 coating being close to the substrate; The TiO2 mesoporous array layer has a honeycomb structure, a thickness of 80-120 nm, and an average surface roughness of Ra 4.63 nm and Rq 6.11 nm.

6. The medical implant according to claim 5, characterized in that, The honeycomb structure in the TiO2 mesoporous array layer has a pore size of 10-14 nm, and the pores are circular or elliptical.

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