Anti-ectopic ossification artificial temporomandibular joint prosthesis modified by multiple controlled-release drug-loaded coating and preparation method thereof

Abstract

The invention discloses an anti-ectopic ossification artificial temporomandibular joint prosthesis modified by multiple controlled-release drug-loaded coatings and a preparation method thereof. The preparation method comprises the following steps: carrying out surface treatment on a bionic artificial temporomandibular joint prosthesis; dopamine is adopted to assist co-deposition on the surface of the bionic artificial temporomandibular joint porous titanium prosthesis to sequentially form a DF-HA self-assembled nanosphere coating and a meloxicam liposome drug-loaded coating; and grafting RGD on the surface of the meloxicam liposome drug-loading coating by adopting chemical natural deposition to obtain the meloxicam liposome drug-loading coating. On one hand, the active drug-loading meloxicam liposome is used for modifying the artificial temporomandibular joint prosthesis, and on the other hand, the RGD-DF-HA layered drug-loading is used for modifying the artificial temporomandibular joint prosthesis, so that long-term slow release and programmed release of drugs are realized, synergistic anti-inflammation is realized, and the purpose of treating and preventing ectopic ossification after artificial joint replacement is realized; long-term stability and early prevention and intervention of artificial joint replacement are facilitated, the service life of the artificial joint prosthesis is prolonged, and the life quality of a patient is improved.

Description

Technical Field

[0001] The present invention belongs to the technical field of temporomandibular joints, and in particular relates to an anti-heterotopic ossification artificial temporomandibular joint prosthesis modified with multiple controlled-release drug-loaded coatings and a preparation method thereof. Background Art

[0002] The temporomandibular joint (TMJ) is the only movable joint in the head and face, responsible for important activities such as mouth opening and closing, chewing, speaking, and swallowing. Temporomandibular joint replacement has gradually replaced autologous bone grafting as the primary treatment for advanced diseases such as osteoarthritis, ankylosing spondylosis, and tumors. Long-term follow-up has shown that surgery can significantly relieve pain, improve mouth opening, and improve quality of life. However, there are still issues with limited lateral and protrusive mandibular movement and heterotopic ossification around the joint. Studies have found that the loss of attachment of the lateral pterygoid muscle and the inability to reattach to the prosthesis contribute to insufficient muscle dynamics. Previous studies have investigated two generations of artificial joint prostheses, respectively improving prosthetic morphology to accommodate Chinese anatomical characteristics and increasing the functional attachment area for the lateral pterygoid muscle to promote ingrowth of the muscle and prosthesis. To further address the issue of heterotopic ossification around the prosthesis, mechanistic studies could provide important insights for the development of third-generation drug-eluting artificial joint prostheses.

[0003] The incidence of heterotopic ossification (HO) around prosthetic joints ranges from 15% to 90%. It can cause pain and loss of joint function, necessitating secondary surgery for prosthetic revision, which compromises the long-term effectiveness of the prosthesis and increases patient suffering and socioeconomic burdens. Because its mechanisms remain unclear, effective preventive measures are currently lacking, making it a challenging clinical problem that urgently needs to be addressed. Studies have shown that HO is closely linked to an inflammatory response triggered by mechanical stimulation and surgical trauma. Inflammatory factors activate stem cells in muscle tissue to undergo osteoblastic differentiation, leading to HO. Although nonsteroidal anti-inflammatory drugs (NSAIDs), such as diclofenac and meloxicam, are currently widely used in orthopedics for postoperative prevention, they cannot completely prevent the formation of HO. Studies have shown that mechanical stimulation directly promotes osteogenesis and also exacerbates the inflammatory response. However, the role of mechanical stimulation generated during prosthetic joint movement in the development of HO remains largely unexplored. To this end, it is necessary to first clarify the interrelationship between mechanical stimulation and inflammatory responses in ossification and the key regulatory molecules involved. Next, new inhibitory targets can be identified to enhance the effects of NSAIDs, and finally, their therapeutic efficacy can be confirmed through topical application. This has important theoretical and practical value for the prevention and treatment of heterotopic ossification of artificial joints and the reduction of secondary surgical revisions. Summary of the Invention

[0004] The main purpose of the present invention is to provide a method for preparing an anti-heterotopic ossification artificial temporomandibular joint prosthesis modified with a multiple controlled-release drug-loaded coating, which is achieved by loading meloxicam liposomes in combination with DF-HA into a bionic artificial temporomandibular joint prosthesis.

[0005] Another object of the present invention is to provide an artificial temporomandibular joint prosthesis modified with multiple controlled-release drug-loaded coatings to resist heterotopic ossification, which is prepared by the above-mentioned method for preparing an artificial temporomandibular joint prosthesis modified with multiple controlled-release drug-loaded coatings to resist heterotopic ossification.

[0006] To achieve the above object, the present invention adopts the following technical solutions:

[0007] A first aspect of the present invention provides a method for preparing an artificial temporomandibular joint prosthesis modified with a multiple controlled-release drug-loaded coating to resist heterotopic ossification, comprising the following steps:

[0008] (1) Surface treatment of porous titanium prosthesis of bionic artificial temporomandibular joint;

[0009] (2) using a dopamine-assisted co-deposition process to deposit a diclofenac (DCF)-grafted hyaluronic acid (HA) (DF-HA) self-assembled nanosphere coating on the surface of the bionic artificial temporomandibular joint porous titanium prosthesis after the surface treatment in step (1);

[0010] (3) depositing a meloxicam liposome drug-loaded coating on the surface of the DF-HA self-assembled nanosphere coating formed in step (2) using a dopamine-assisted co-deposition process;

[0011] (4) Grafting arginine-glycine-aspartic acid (RGD) onto the surface of the meloxicam liposome drug-loaded coating layer described in step (3) by chemical natural deposition to obtain the product.

[0012] Preferably, in step (1), the bionic artificial temporomandibular joint porous titanium prosthesis is ultrasonically cleaned with acetone, ethanol and deionized water in sequence once, each time for 10 minutes, and air-dried to remove surface impurities.

[0013] Preferably, in step (2), a 3-5 mg / mL dopamine Tris HCl solution and a 3-5 mg / mL hyaluronic acid-polyethylene glycol-diclofenac (HA-PEG-DCF) aqueous solution are mixed in a volume ratio of 1:1-3, and immediately added to a 24-well plate containing the bionic artificial temporomandibular joint porous titanium prosthesis, and soaked in a dark environment for 18-30 hours to form a DF-HA self-assembled nanosphere coating by dopamine-assisted co-deposition, rinsed with deionized water, and blown dry.

[0014] Preferably, in step (2), the method for preparing the HA-PEG-DCF aqueous solution comprises:

[0015] (a) Weigh 62 mg of diclofenac-N-hydroxysuccinimide (DCF-NHS) and dissolve it in 25 mL of N,N-dimethylformamide (DMF). Stir until fully dissolved to obtain a DCF-NHS mixture.

[0016] (b) Weigh 62 mg of amino-polyethylene glycol-hyaluronic acid (NH2-PEG-HA) and dissolve it in 25 mL of deionized water. Stir until fully dissolved, and add the DCF-NHS mixture described in step (a) dropwise while stirring to obtain a mixture of NH2-PEG-HA and DCF-NHS.

[0017] (c) The mixture of NH2-PEG-HA and DCF-NHS in step (b) is heated at 40°C for 2-3 hours to clarify the solution, then stirred at room temperature for 24-36 hours, and finally dialyzed using an 8000-14000 dialysis bag for 3 days and freeze-dried to obtain a white cotton-like product.

[0018] More preferably, in step (2), the synthesis method of DCF-NHS comprises:

[0019] (2-1) DCF (296 mg, 1 mmol), NHS (126.6 mg, 1.1 mmol) and 13 mL of anhydrous dioxane were added to a 100 mL round-bottom flask and incubated under a N2 atmosphere for 25 min;

[0020] (2-2) Cool a round-bottom flask to 0°C in an ice bath, weigh DCC (227 mg, 1.1 mmol) and 10 mL of anhydrous dioxane, shake to dissolve, and inject the DCC mixture into the round-bottom flask using a 20 mL syringe while stirring. Continuously flow N2 for 5 min, seal the flask with parafilm, and stir for 12 h.

[0021] (2-3) Add n-hexane and ethyl acetate in a volume ratio of 7:3 to the plate and observe whether the reaction is complete or successful;

[0022] (2-4) Remove the byproduct DCU of DCC by filtration and continuously wash with a large amount of anhydrous dioxane until the liquid in the filtration flask is clear and transparent;

[0023] (2-5) Remove the solvent anhydrous dioxane by rotary evaporation for 2-3 h until the surface of the round-bottom flask is completely covered with white product;

[0024] (2-6) Using isopropanol for crystallization, first add 3–5 mL of isopropanol to a round-bottom flask and sonicate until all the white product on the surface of the round-bottom flask is dissolved in the isopropanol. Then, heat the open round-bottom flask at 82 °C for 3–4 h. During the heating process, add 3–7 mL of isopropanol until the solution becomes clear and free of solids.

[0025] (2-7) Cool the round-bottom flask at room temperature until a large amount of crystals form at the bottom of the round-bottom flask;

[0026] (2-8) The round-bottom flask was placed in a -20 °C refrigerator for 2 days for crystallization, and filtered to obtain a white product.

[0027] More preferably, in step (2), the synthesis method of NH2-PEG-HA comprises:

[0028] (2-a) Weigh 25 mg of hyaluronic acid and dissolve it in 10 mL of ultrapure water. Stir continuously until the HA is dissolved.

[0029] (2-b) Add 16 mg of EDC, 17 mg of NHS, and 5 mL of ultrapure water and activate for 1 h;

[0030] (2-c) Weigh 100 mg of amino-functionalized polyethylene glycol (NH2-PEG-NH2) (MW = 2000), dissolve it in 10 mL of ultrapure water, and then add it dropwise to the activation solution of step (2-b);

[0031] (2-d) Triethylamine was used to adjust the pH to 9;

[0032] (2-e) The mixture was stirred at 43°C for 24 h, dialyzed using a 8000-14000 dialysis bag for 3 days, and lyophilized into a white cotton-like substance. The reaction was protected from light during the reaction.

[0033] Preferably, in step (3), a 3-5 mg / mL dopamine Tris HCl solution and meloxicam liposomes are co-deposited with the aid of dopamine to form a drug-loaded coating.

[0034] Preferably, in step (4), the meloxicam liposome drug-loaded coating in step (3) is immersed in an RGD solution (1 mg / mL in PBS) and incubated at room temperature for 3-5 hours to form an RGD coating, and then rinsed with PBS to remove the RGD that is not firmly adsorbed.

[0035] The second aspect of the present invention provides an artificial temporomandibular joint prosthesis with resistance to heterotopic ossification modified by multiple controlled-release drug-loaded coatings, which is obtained by the preparation method of the artificial temporomandibular joint prosthesis with resistance to heterotopic ossification modified by the multiple controlled-release drug-loaded coatings, and includes a bionic artificial temporomandibular joint porous titanium prosthesis and a DF-HA self-assembled nanosphere coating, a meloxicam liposome coating and an RGD coating chemically modified thereon in sequence.

[0036] Compared with the prior art, the present invention proposes an artificial temporomandibular joint prosthesis modified with a multiple controlled-release drug-loaded coating to resist heterotopic ossification. On the one hand, the artificial temporomandibular joint prosthesis is modified with active drug-loaded meloxicam liposomes, and on the other hand, the artificial temporomandibular joint prosthesis is modified with RGD-DF-HA layered drug loading, thereby achieving long-term sustained release and programmed release of drugs, synergistically anti-inflammatory, and achieving long-term sustained release of small drug molecules, thereby achieving the purpose of treating and preventing heterotopic ossification after artificial joint replacement. Specifically, it has the following beneficial effects:

[0037] 1. The modified system of the present invention includes an active drug-loaded meloxicam liposome and a DF-HA self-assembled nanosphere system. In vitro and in vivo experiments have confirmed that the constructed multiple controlled-release drug-loaded coating has good biocompatibility and safety. It can be implanted in the body and release the drug into the surrounding tissues, thereby achieving long-term inhibition of inflammation and heterotopic ossification around artificial temporomandibular joint prostheses, thereby achieving the important goals of early prevention and treatment, intervention, increasing the service life of artificial joint prostheses, and improving the quality of life of patients.

[0038] 2. The anti-heterotopic ossification artificial temporomandibular joint prosthesis modified with multiple controlled-release drug-loaded coatings in the present invention can form a gradient release of anti-inflammatory drugs, directly acting on adjacent tissues, which is superior to oral administration and more in line with clinical application needs. It can better achieve and maintain the anti-heterotopic ossification effect of the artificial temporomandibular joint prosthesis for a long time.

[0039] 3. The anti-heterotopic ossification artificial temporomandibular joint prosthesis obtained in the present invention has a sustained-release effect, which meets the long-term anti-inflammatory needs after temporomandibular joint replacement, improves its anti-inflammatory and anti-heterotopic ossification effects, and is beneficial to the long-term stability of artificial joint replacement. BRIEF DESCRIPTION OF THE DRAWINGS

[0040] Figure 1 Schematic diagram of the preparation process of the anti-heterotopic ossification artificial temporomandibular joint prosthesis modified with multiple controlled-release drug-loaded coatings in the examples.

[0041] Figure 2 These are the performance characterization results of the meloxicam liposomes in the examples.

[0042] Figure 3 The effects of the aqueous components in meloxicam liposomes on drug loading capacity, appearance and performance characterization results are shown in the examples.

[0043] Figure 4 1 is the H NMR spectrum result of DF-HA self-assembled nanospheres in the embodiment.

[0044] Figure 5 This is the nuclear magnetic hydrogen spectrum characterization result of HA-PEG-DCF in the example.

[0045] Figure 6This is a scanning electron microscope image of the microsphere structure formed by co-precipitation of PDA and DF-HA self-assembled nanospheres on the porous titanium surface in the example.

[0046] Figure 7 The PCR and Western blot experiments in the examples characterize the protective effects of RGD, DF-HA and meloxicam liposomes on ectopic ossification of myoblasts under mechanical stimulation conditions.

[0047] Figure 8 The PCR and Western blot experiments in the examples characterize the protective effects of RGD-DF-HA and meloxicam liposomes on myoblast ectopic ossification under mechanical and inflammatory factor stimulation. DETAILED DESCRIPTION

[0048] In order to more fully understand and demonstrate the technical solutions, objectives, and advantages of the present invention, the following is a further detailed and complete description of the technical effects produced by the present invention in conjunction with the accompanying drawings and specific embodiments. Obviously, the described embodiments are only some of the embodiments of the present invention, and not all of them. It should be pointed out that for those skilled in the art, other embodiments obtained without departing from the concept of the present invention are all within the scope of protection of the present invention.

[0049] like Figure 1 As shown, the following embodiment provides an anti-heterotopic ossification artificial temporomandibular joint prosthesis modified with a multiple controlled-release drug-loaded coating, comprising a bionic artificial temporomandibular joint porous titanium prosthesis and a DF-HA self-assembled nanosphere coating, a meloxicam liposome coating, and an RGD coating sequentially chemically modified thereon. The preparation method comprises:

[0050] (1) Surface treatment of porous titanium prosthesis of bionic artificial temporomandibular joint;

[0051] (2) using a dopamine-assisted co-deposition process to deposit a diclofenac (DCF)-grafted hyaluronic acid (HA) (DF-HA) self-assembled nanosphere coating on the surface of the bionic artificial temporomandibular joint porous titanium prosthesis after the surface treatment in step (1);

[0052] (3) depositing a meloxicam liposome drug-loaded coating on the surface of the DF-HA self-assembled nanosphere coating formed in step (2) using a dopamine-assisted co-deposition process;

[0053] (4) Grafting RGD onto the surface of the meloxicam liposome drug-loaded coating in step (3) by chemical natural deposition.

[0054] In some embodiments, in step (1), the bionic artificial temporomandibular joint porous titanium prosthesis is ultrasonically cleaned with acetone, ethanol and deionized water in sequence once, each time for 10 minutes, and air-dried to remove surface impurities.

[0055] In some embodiments, in step (2), 3-5 mg / mL dopamine Tris HCl solution and 3-5 mg / mL hyaluronic acid-polyethylene glycol-diclofenac (HA-PEG-DCF) aqueous solution are mixed in a volume ratio of 1:1-3, and immediately added to a 24-well plate containing a bionic artificial temporomandibular joint porous titanium prosthesis, and soaked in a dark environment for 18-30 hours to form a DF-HA self-assembled nanosphere coating by dopamine-assisted co-deposition, rinsed with deionized water, and blown dry.

[0056] In some embodiments, in step (2), the method for preparing the HA-PEG-DCF aqueous solution comprises:

[0057] (a) Weigh 62 mg of diclofenac-N-hydroxysuccinimide (DCF-NHS) and dissolve it in 25 mL of DMF. Stir until fully dissolved to obtain a DCF-NHS mixture.

[0058] (b) Weigh 62 mg of amino-polyethylene glycol-hyaluronic acid (NH2-PEG-HA) and dissolve it in 25 mL of deionized water. Stir until fully dissolved, and add the DCF-NHS mixture in step (a) dropwise while stirring to obtain a mixture of NH2-PEG-HA and DCF-NHS.

[0059] (c) The mixture of NH2-PEG-HA and DCF-NHS in step (b) is heated at 40°C for 2-3h to clarify the solution, then stirred at room temperature for 24-36h, and finally dialyzed using an 8000-14000 dialysis bag for 3 days and freeze-dried to obtain a white cotton-like product.

[0060] In some embodiments, in step (2), the synthesis method of DCF-NHS comprises:

[0061] (2-1) DCF (296 mg, 1 mmol), NHS (126.6 mg, 1.1 mmol) and 13 mL of anhydrous dioxane were added to a 100 mL round-bottom flask and incubated under a N2 atmosphere for 25 min;

[0062] (2-2) Cool a round-bottom flask to 0°C in an ice bath, weigh DCC (227 mg, 1.1 mmol) and 10 mL of anhydrous dioxane, shake to dissolve, and inject the DCC mixture into the round-bottom flask using a 20 mL syringe while stirring. Continuously flow N2 for 5 min, seal the flask with parafilm, and stir for 12 h.

[0063] (2-3) Add n-hexane and ethyl acetate in a volume ratio of 7:3 to the plate and observe whether the reaction is complete or successful;

[0064] (2-4) Remove the byproduct DCU of DCC by filtration and continuously wash with a large amount of anhydrous dioxane until the liquid in the filtration flask is clear and transparent;

[0065] (2-5) Remove the solvent anhydrous dioxane by rotary evaporation for 2-3 h until the surface of the round-bottom flask is completely covered with white product;

[0066] (2-6) Using isopropanol for crystallization, first add 3–5 mL of isopropanol to a round-bottom flask and sonicate until all the white product on the surface of the round-bottom flask is dissolved in the isopropanol. Then, heat the open round-bottom flask at 82 °C for 3–4 h. During the heating process, add 3–7 mL of isopropanol until the solution becomes clear and free of solids.

[0067] (2-7) Cool the round-bottom flask at room temperature until a large amount of crystals form at the bottom of the round-bottom flask;

[0068] (2-8) The round-bottom flask was placed in a -20 °C refrigerator for 2 days for crystallization, and filtered to obtain a white product.

[0069] In some embodiments, in step (2), the synthesis method of NH2-PEG-HA comprises:

[0070] (2-a) Weigh 25 mg of hyaluronic acid and dissolve it in 10 mL of ultrapure water. Stir continuously until the HA is dissolved.

[0071] (2-b) Add 16 mg of EDC, 17 mg of NHS, and 5 mL of ultrapure water and activate for 1 h;

[0072] (2-c) Weigh 100 mg of amino-functionalized polyethylene glycol (NH2-PEG-NH2) (MW = 2000), dissolve it in 10 mL of ultrapure water, and then add it dropwise to the solution in step 2;

[0073] (2-d) Triethylamine was used to adjust the pH to 9;

[0074] (2-e) Continue stirring at 43°C for 24 hours. Dialyze the solution using an 8000-14000 dialysis bag for 3 days. Freeze-dry the solution to a white, cotton-like substance. (Protect from light during the reaction.)

[0075] In some embodiments, in step (3), a 3-5 mg / mL dopamine Tris HCl solution is co-deposited with meloxicam liposomes via dopamine-assisted co-deposition to form a drug-loaded coating.

[0076] In some embodiments, in step (4), the meloxicam liposome drug-loaded coating in step (3) is immersed in an RGD solution (1 mg / mL in PBS) and incubated at room temperature for 3-5 hours to form an RGD coating, and then rinsed with PBS to remove the RGD that is not firmly adsorbed.

[0077] Example 1: Modification and performance evaluation of active meloxicam-loaded liposome porous titanium stent

[0078] 1) Meloxicam was added to the dialyzed liposomes at different drug-to-lipid ratios (0.05, 0.1, 0.2, and 0.3) to obtain actively loaded meloxicam liposomes.

[0079] 2) The particle size and PDI of drug-loaded liposomes were determined by dynamic light scattering.

[0080] 3) The preparation efficiency was detected by horizontal gel electrophoresis, and the encapsulation efficiency and drug loading of the liposomes were calculated.

[0081] 4) Calcium salt liposomes with high encapsulation efficiency, drug loading capacity and low leakage under different drug-to-lipid ratios are preferred.

[0082] 5) 4 mg / mL dopamine solution was co-deposited with meloxicam liposomes onto the porous titanium scaffold.

[0083] 6) The sustained-release pattern of the drug was determined by in vitro release rate test and enzyme-labeled assay.

[0084] The results are as follows Figure 2 and 3 As shown, the active drug-loaded meloxicam liposomes were successfully constructed, and their particle size distribution met the requirements. Biological transmission electron microscopy showed that the liposomes were successfully loaded with drugs, and the loaded drugs were in a "bowl-shaped" structure.

[0085] Example 2: Modification and performance evaluation of RGD-DF-HA layered drug-loaded porous titanium scaffold

[0086] 1) Preparation of materials for grafting diclofenac: First, diclofenac-N-hydroxysuccinimide (DCF-NHS) was synthesized, followed by chemical synthesis of amino-polyethylene glycol-hyaluronic acid (NH2-PEG-HA); finally, hyaluronic acid-polyethylene glycol-diclofenac (HA-PEG-DCF) was synthesized by substitution.

[0087] 2) Preparation of multilayer coating on the surface of porous titanium stent: drug-loaded porous titanium modified functional area was constructed by nano-self-assembly deposition of dopamine assisted by meloxicam liposome co-deposition.

[0088] 3) The outermost layer is grafted with RGD coating by natural deposition, left to stand for 4 hours and then washed.

[0089] 4) The prepared samples were subjected to nuclear magnetic resonance hydrogen spectrum, infrared spectrum, scanning electron microscopy and EDS detection to confirm the material synthesis effect.

[0090] The test results are as follows Figure 4-6As shown, the nuclear magnetic hydrogen spectrum showed that the HA-PEG-DCF compound was successfully constructed, and the scanning electron microscopy results further confirmed that the drug was loaded on the surface of the porous titanium scaffold through nano-autonomous deposition, and then RGD was autonomously loaded on the surface to complete the multi-layer drug modification.

[0091] Example 3: Biological functions of multi-layer controlled release drug delivery system

[0092] (1) PCR detection of the biological effects of meloxicam liposomes and RGD-DF-HA drug delivery system on anti-inflammatory and osteogenesis inhibition

[0093] 1) After myoblasts grow to an appropriate density, subsequent drug stimulation is performed.

[0094] 2) 24 hours after transfection, stimulate with IL-1β for 24 hours and collect samples with Trizol.

[0095] 3) RNA was extracted using the Trizol method.

[0096] 4) Reverse transcription of RNA into cDNA.

[0097] 5) RT-qPCR was used to detect the osteogenic tendency and myogenic stability of C2C12 myoblasts in different groups.

[0098] (2) Western Blot detection of the biological effects of meloxicam liposomes and RGD-DF-HA drug delivery system on anti-inflammatory and osteogenesis inhibition

[0099] 1) Wait until the chondrocytes grow to an appropriate density before adding drugs for stimulation.

[0100] 2) 24 hours after transfection, the cells were stimulated with TNF-α for 48 hours and the cell lysate was collected.

[0101] 3) Add 5X protein loading buffer and boil to denature.

[0102] 4) Preparation of SDS-PAGE polyacrylamide gel.

[0103] 5) Then, 30 μg of total protein was loaded into each well and electrophoresed at 80V for the upper gel and 120V for the lower gel.

[0104] 6) 280mA, 90min, transfer membrane.

[0105] 7) Primary and secondary antibody incubation.

[0106] The results are as follows Figure 7 and 8As shown, imaging confirmed the osteogenic tendency and myogenic stability of C2C12 myoblasts in different groups. RT-qPCR and Western Blot results showed that meloxicam liposomes and RGD-DF-HA drug delivery system have a good effect in inhibiting the osteogenic differentiation of myoblasts, which is beneficial to maintaining the homeostasis of muscle cells.

[0107] In summary, the meloxicam liposomes and RGD-DF-HA drug delivery system provided by the present invention have good biological functions, can effectively inhibit the osteogenic feedback of myoblasts in an inflammatory environment, promote the homeostasis of muscle cells, and have good application potential in maintaining function after artificial joint replacement.

[0108] The above are merely preferred embodiments of the present invention and are not intended to limit the present invention. Those skilled in the art will readily appreciate that various modifications and variations of the present invention are possible. Any modifications, equivalent substitutions, or improvements made within the spirit and principles of the present invention are intended to be within the scope of protection of the present invention.

Claims

1. A method for preparing an artificial temporomandibular joint prosthesis modified with a multiple controlled-release drug-loaded coating to resist heterotopic ossification, characterized in that: The following steps are involved: (1) Surface treatment of porous titanium prosthesis of bionic artificial temporomandibular joint; (2) A dopamine-assisted co-deposition process was used to deposit a DF-HA self-assembled nanosphere coating on the surface of the bionic artificial temporomandibular joint porous titanium prosthesis after surface treatment; (3) using a dopamine-assisted co-deposition process to deposit a meloxicam liposome drug-loaded coating on the surface of the DF-HA self-assembled nanosphere coating; (4) Grafting RGD onto the surface of the meloxicam liposome drug-loaded coating by chemical natural deposition to obtain the product.

2. The method for preparing the anti-heterotopic ossification artificial temporomandibular joint prosthesis according to claim 1, characterized in that: In step (1), the bionic artificial temporomandibular joint porous titanium prosthesis is ultrasonically cleaned with acetone, ethanol and deionized water in sequence once, each time for 10 minutes, and air-dried to remove surface impurities.

3. The method for preparing the anti-heterotopic ossification artificial temporomandibular joint prosthesis according to claim 1, characterized in that: In step (2), a 3-5 mg / mL dopamine Tris HCl solution and a 3-5 mg / mL HA-PEG-DCF aqueous solution are mixed in a volume ratio of 1:1-3, and immediately added to a 24-well plate containing the bionic artificial temporomandibular joint porous titanium prosthesis, and immersed in a dark environment for 18-30 hours to form a DF-HA self-assembled nanosphere coating by dopamine-assisted co-deposition, rinsed with deionized water, and blown dry.

4. The method for preparing the anti-heterotopic ossification artificial temporomandibular joint prosthesis according to claim 3, characterized in that: In step (2), the preparation method of the HA-PEG-DCF aqueous solution comprises: (a) DCF-NHS was dissolved in DMF and stirred until fully dissolved to obtain a DCF-NHS mixture with a DCF-NHS to DMF mass volume ratio of 62:25 mg / mL. (b) NH2-PEG-HA was dissolved in deionized water and stirred until fully dissolved, and the DCF-NHS mixture was added dropwise while stirring to obtain a mixture of NH2-PEG-HA and DCF-NHS, wherein the mass volume ratio of NH2-PEG-HA to deionized water was 62:25 mg / mL; (c) The mixture of NH2-PEG-HA and DCF-NHS is heated at 40°C for 2-3 hours to clarify the solution, and then stirred at room temperature for 24-36 hours. The mixture is then dialyzed using an 8000-14000 dialysis bag for 3 days and lyophilized to obtain a white cotton-like product.

5. The method for preparing the anti-heterotopic ossification artificial temporomandibular joint prosthesis according to claim 4, characterized in that: In step (2), the synthesis method of DCF-NHS comprises: (2-1) 296 mg, 1 mmol DCF, 126.6 mg, 1.1 mmol NHS, and 13 mL of anhydrous dioxane were added to a 100 mL round-bottom flask and incubated under N2 atmosphere for 25 min. (2-2) Cool a round-bottom flask to 0°C in an ice bath. Weigh 227 mg, 1.1 mmol of DCC and 10 mL of anhydrous dioxane. Dissolve the mixture by shaking. Pour the DCC mixture into the round-bottom flask using a 20 mL syringe while stirring. Continuously flow N2 for 25 min. Seal the flask with parafilm and stir for 12 h. (2-3) Add n-hexane and ethyl acetate in a volume ratio of 7:3 to the plate and observe whether the reaction is complete or successful; (2-4) Remove the byproduct DCU of DCC by filtration and continuously wash with a large amount of anhydrous dioxane until the liquid in the filtration flask is clear and transparent; (2-5) Remove the solvent anhydrous dioxane by rotary evaporation for 2-3 h until the surface of the round-bottom flask is completely covered with white product; (2-6) Using isopropanol for crystallization, first add 3–5 mL of isopropanol to a round-bottom flask and sonicate to dissolve all the white product on the surface of the round-bottom flask in the isopropanol. Then, heat the open round-bottom flask at 82 °C for 3–4 h, adding 3–7 mL of isopropanol during the heating process until the solution becomes clear and contains no solids. (2-7) Cool the round-bottom flask to room temperature until a large amount of crystals form at the bottom of the flask; (2-8) The round-bottom flask was placed at -20 °C for crystallization for 2 days and filtered to obtain a white product.

6. The method for preparing the anti-heterotopic ossification artificial temporomandibular joint prosthesis according to claim 4, characterized in that: In step (2), the synthesis method of NH2-PEG-HA comprises: (2-a) Weigh 25 mg of HA and dissolve it in 10 mL of ultrapure water. Stir continuously until the HA dissolves. (2-b) Add 16 mg of EDC, 17 mg of NHS, and 5 mL of ultrapure water and activate for 1 h; (2-c) Weigh 100 mg of NH2-PEG-NH2 (MW = 2000), dissolve it in 10 mL of ultrapure water, and then add it dropwise to the activation solution; (2-d) adjusting the pH to 9 with triethylamine; (2-e) The mixture was stirred at 43°C for 24 h, dialyzed using a 8000-14000 dialysis bag for 3 days, and lyophilized into a white cotton-like substance. The reaction was protected from light during the reaction.

7. The method for preparing the anti-heterotopic ossification artificial temporomandibular joint prosthesis according to claim 1, characterized in that: In step (3), a 3-5 mg / mL dopamine Tris HCl solution is co-precipitated with meloxicam liposomes through dopamine assistance to form a drug-loaded coating.

8. The method for preparing the anti-heterotopic ossification artificial temporomandibular joint prosthesis according to claim 1, characterized in that: In step (4), the meloxicam liposome drug-loaded coating is immersed in a 1 mg / mL RGD solution in PBS and incubated at room temperature for 3-5 hours to form an RGD coating, and then rinsed with PBS.

9. An artificial temporomandibular joint prosthesis modified with multiple controlled-release drug-loaded coatings to resist heterotopic ossification, characterized in that: The invention is obtained by the preparation method of the artificial temporomandibular joint prosthesis modified with multiple controlled-release drug-loaded coatings according to any one of claims 1 to 8, comprising a bionic artificial temporomandibular joint porous titanium prosthesis and a DF-HA self-assembled nanosphere coating, a meloxicam liposome coating and an RGD coating chemically modified thereon in sequence.

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