An antibacterial and repair-promoting hydrogel dressing and its preparation method

Abstract

This application belongs to the technical field of dental dressings, and particularly relates to an antibacterial and repair-promoting hydrogel dressing and a preparation method thereof. It is first discovered in this application that compound A has significant antibacterial activity against common oral bacteria. After being prepared into a temperature-sensitive hydrogel dressing, it still has antibacterial activity. And injecting it around the implant can prevent alveolar bone reduction, stimulate vertical bone regeneration in the area around the implant, increase bone mass to improve the stability of the implant, ultimately promote the fusion of the wound between the alveolar bone and the implant, and extend the lifespan of the implant.

Description

Technical Field

[0001] The present invention belongs to the technical field of dental dressings, and particularly relates to an antibacterial and repair-promoting hydrogel dressing and a preparation method thereof. Background Art

[0002] Tooth / dental arch loss is one of the most important diseases in oral clinical work. The loss of natural teeth causes the degradation of the normal chewing function of the stomatognathic system, and at the same time brings changes in the facial appearance, affecting aesthetics. The instability and loss of the occlusal relationship will bring disorders and even lesions in the temporomandibular joint area, seriously affecting the quality of life of patients and having an impact on physiological and psychological functions. Implant denture restoration can greatly improve the chewing efficiency after restoration treatment without damaging adjacent teeth, and has characteristics such as high comfort and long durability that are not possessed by conventional restoration methods, and is increasingly becoming the preferred choice in the process of tooth loss restoration treatment.

[0003] The origin of modern oral implant restoration technology can be traced back to the mid-1950s to 1960s. Professor Branemark from Sweden and his colleagues first proposed the phenomenon of osseointegration and titanium implant materials. That is, after the pure titanium implant is implanted into the body and healed, a tightly bonded interface is generated between the pure titanium implant and the surrounding bone tissue, and there is a direct structural and functional connection. This physiological phenomenon is also the basis for the implant to maintain stability and function in bone tissue. After long-term development, the types of oral implants have tended to be diversified. According to different implantation times and methods, they can be divided into one-stage and two-stage implants; according to the implantation site of the implant, they can be divided into endosseous, zygomatic implants, etc.; and according to the different general shapes of the implant, they can be divided into columnar, conical, root-shaped implants, etc.

[0004] As an implantable medical device, implant materials should have high biocompatibility on the basis of good biosafety and exhibit specific physiological functions. In addition, in view of the special physical and chemical environment in the oral cavity, they must also have excellent mechanical properties and corrosion resistance to meet practical applications. Especially for the clinical situation that the bone mass of the mandible of the yellow race is small and the implant space is insufficient compared with that of Europeans and Americans, and a large number of narrow-diameter implants need to be used, new implant materials should meet the characteristics of "high (mechanical) strength, low (elastic) modulus, and narrow diameter". Such clinical requirements also put forward new requirements and directions for the development and research of implant materials.

[0005] As the cornerstone in the development of dental implants, pure titanium materials are the mainstream materials in current implant systems. With the remarkable progress in materials science and processing technologies, it is of great research significance and practical value to develop and utilize various materials with good properties to further improve the comprehensive performance of dental implants and expand their application scope and scenarios. Currently, new implant materials are emerging in an endless stream, mainly including metal materials represented by pure titanium and its alloys, ceramic materials, and organic polymer materials, etc. These materials have different biological and mechanical property characteristics.

[0006] As bio-inert materials, titanium and its alloys face the challenge of insufficient osteoinductive ability in clinical applications and need to improve their osseointegration performance through multi-dimensional technologies. Constructing micro-nano topological structures on the implant surface can simulate the three-dimensional pore characteristics of natural bone tissue and promote the directional migration and mineral deposition of osteoblasts. Related technologies have been verified in the processing of high-precision components in the aerospace field. Forming a hydroxyapatite coating through plasma spraying technology not only enhances the osteoconductivity of the implant but also activates the osteogenic signaling pathway through the gradient release of calcium and phosphate ions. The technical principle of this technology is similar to the preparation process of thermal protection coatings for spacecraft.

[0007] Chemical grafting technology can immobilize bioactive molecules on the titanium substrate surface. For example, introducing the arginine-glycine-aspartic acid (RGD) polypeptide sequence can regulate cell adhesion behavior by specifically recognizing integrin receptors. Such molecular modification strategies have been successfully applied in the treatment of flexible sensor substrates.

[0008] Temperature-sensitive hydrogel systems utilize the phase transition characteristics of temperature-sensitive materials such as poly(N-isopropylacrylamide) to achieve gelation encapsulation at physiological temperature and precisely control the sustained-release kinetics of bone growth promoters such as bone morphogenetic protein (BMP). This controlled-release mechanism has technical commonalities with the intelligent response characteristics of high-end engineering plastics.

[0009] Temperature-sensitive hydrogel systems can bring many clinical advantages and provide a suitable environment for local bone regeneration around implants. Due to the high water content of temperature-sensitive hydrogel systems, bone growth promoters such as BMP-2 can be easily loaded in a solution state by simple mixing. Due to the temperature-dependent gelation characteristics of this system, it can be conveniently applied to the human body at room temperature. In addition, it neither requires surgery nor additional external factors to form a hydrogel. This temperature-sensitive and injectable property is very convenient for patients. Moreover, the hydrophilic environment and high porosity of the hydrogel system are conducive to cell infiltration, blood vessel formation, and subsequent bone regeneration.

[0010] The special physiological structure and environment of the oral cavity make oral drug administration difficult. Therefore, there is an urgent need for a drug carrier to solve the problem of oral drug administration. Polyoxyethylene-polyoxypropylene copolymer (Poloxamer 407) or Carbomer can form a thermosensitive biogel with good mechanical properties and mucoadhesive characteristics. This biogel can be designed as a drug supply platform for an oral implantable drug delivery system. Topical drug administration is usually used to treat local diseases such as periodontitis, oral ulcers, and other oral diseases. The main advantage of this drug delivery method is that it can directly deliver bioactive agents to the diseased site, maintain the required drug concentration for a relatively long period of time, and ensure good retention at the drug administration site. Below the sol-gel transition temperature, the fluidity and compression characteristics of the preparation are extremely low. In contrast, under conditions above the sol-gel transition temperature, these preparations exhibit a wide range of viscoelastic, mechanical, and mucoadhesive properties, which will help their application in specific parts of the oral cavity. High elasticity and mucoadhesion will make the preparation containing Poloxamer 407 an effective platform for controlling local oral drug delivery. Currently, thermosensitive gel dressings have been widely used in the treatment of various periodontal diseases, and there are a variety of preparations. For example, studies on the in-situ gel of moxifloxacin hydrochloride for the treatment of periodontitis, injectable in-situ curcumin gel for the treatment of periodontal pockets, thermosensitive gel for the treatment of oral herpes infections, and clinical cases of thermosensitive gel as a scaffold encapsulation system.

[0011] Professor Xu Wenfang of Shandong University conducted research on L-isoserine tripeptide derivatives as aminopeptidase inhibitors and obtained a series of L-isoserine tripeptide derivatives. The relevant research results were published in the Journal of Enzyme Inhibition and Medicinal Chemistry, and the article title is Design, synthesis and biological evaluation of novel L-isoserine tripeptide derivatives as aminopeptidase N inhibitors, which is translated into Chinese as: Design, Synthesis and Biological Activity Evaluation of Novel L-Isoserine Tripeptide Derivatives as Aminopeptidase N Inhibitors.

[0012] The article focuses on the research of novel L-isoserine tripeptide derivatives as aminopeptidase N (APN) inhibitors, covering aspects such as design, synthesis, and biological activity evaluation, aiming to develop novel anticancer drugs. Taking compound 14b as a lead, novel L-isoserine tripeptide derivatives were designed and synthesized. Except for compound 16j, other target compounds showed inhibitory activity. In terms of in vitro APN inhibitory activity, compound 16l had the strongest inhibitory activity , slightly better than , the tyrosine group may enhance the interaction with APN through hydrogen bonds, and the phenyl group at the R1 position is more beneficial for improving the activity than the benzyl group. The inhibitory activities of the compounds in Series 2 are similar to those in Series 1, but the water solubility is significantly reduced. The results of the experiments at the cellular level are basically consistent with those at the enzyme level, but the inhibitory activity of compound 16l against A549 cells is weak, which may be related to the differences in the binding characteristics of APN in different species.

[0013] In terms of anti-proliferative activity, some compounds showed anti-proliferative effects similar to or even better than Bestatin on different tumor cell lines, but compound 16l did not show outstanding activity in the anti-proliferative experiment. Moreover, the compounds that showed good performance in the APN inhibition experiment did not show more prominent anti-proliferative activity on cells with high APN expression than on cells with low APN expression, suggesting that the anti-proliferative mechanism of these compounds may be different from the mechanism of inhibiting APN. Through molecular docking, it was found that the binding mode of compound 16l was similar to that of Bestatin. Its L-isoserine part coordinated with the zinc ion of APN, the phenyl part of the phenylalanine and tyrosine residues inserted into the S1 pocket of APN, and the leucine part inserted into the S1' pocket, and formed hydrogen bonds with multiple amino acid residues, enhancing the binding affinity. However, the chloramphenicol amine residue of compound 16j did not enter the S1' pocket, resulting in the difference in activity.

[0014] Among the compounds synthesized in the article, 16j has the chemical formula , with a molecular weight of 426.18 and the chemical name 2-(3-amino-2-hydroxypropanamido)-3-hydroxy-3-(4-nitrophenyl) propionyl leucine. The chemical structure is as follows:

[0015] .

[0016] However, there is no report on other activities or uses of the above-mentioned 16j structure (hereinafter simply referred to as compound A) in the existing literature and patents. Summary of the Invention

[0017] As a stomatologist, the inventor has conducted in-depth research on dressings for promoting local bone regeneration around implants. The research found that compound A can be used as the main component of the dressing to prepare a temperature-sensitive hydrogel dressing for promoting local bone regeneration around implants, thereby promoting the repair of alveolar bone wounds after implantation. Unexpectedly, it was found that compound A has significant antibacterial effects and remarkable antibacterial effects on common oral bacteria.

[0018] This application discloses for the first time the use of compound A for preparing a dressing for promoting local bone regeneration around implants.

[0019] The chemical structure of the said compound A is as follows:

[0020] .

[0021] The implant material is one of titanium, titanium alloy, and ceramic.

[0022] The implant material is one of surface-modified titanium, titanium alloy, and ceramic.

[0023] This application further discloses the use of compound A for preparing a dressing for inhibiting the growth of bacteria around implants.

[0024] The dressing is a hydrogel dressing.

[0025] The mass percentage concentration of compound A in the dressing is 0.1% - 5%.

[0026] The mass percentage concentration of compound A in the dressing is 1.5%.

[0027] The mass percentage concentration of compound A in the dressing is 2%.

[0028] The dressing is a temperature-sensitive hydrogel dressing.

[0029] The temperature-sensitive hydrogel dressing is injected around the implant through a syringe, playing the role of inhibiting bacteria, promoting the growth of alveolar bone, promoting the fusion of the wound between it and the implant, and prolonging the lifespan of the implant.

[0030] Poloxamer 407 is used as the hydrogel matrix in the temperature-sensitive hydrogel dressing.

[0031] The preparation method of the temperature-sensitive hydrogel dressing is as follows:

[0032] 1) Dissolve compound A in phosphate buffer solution;

[0033] 2) Take poloxamer 407 and add it to the solution obtained in step 1), stirring while adding. After poloxamer 407 is completely dissolved, make up the volume with phosphate buffer solution;

[0034] 3) Take the gel solution obtained in step 2), dispense it into vials, add rubber stoppers, and crimp aluminum caps to obtain the product.

[0035] Advantages of the present invention: This application first discovers that compound A has significant antibacterial activity against common oral bacteria. After being prepared into a temperature-sensitive hydrogel dressing, it still has antibacterial activity. And injecting it around the implant can prevent alveolar bone reduction, stimulate vertical bone regeneration in the area around the implant, increase bone mass to improve the stability of the implant, and ultimately promote the fusion of the wound between the alveolar bone and the implant, prolonging the lifespan of the implant.

[0036] Abbreviation description:

[0037] TSB medium: Trypticase Soy Broth (TSB);

[0038] BHI medium: Brain Heart Infusion Medium;

[0039] RPMI 1640 medium: Medium No. 1640 formulated by Roswell Park Memorial Institute (RPMI);

[0040] MH medium: Mueller - Hinton medium. Description of the Drawings

[0041] Appendix Figure 1 : Promoting effect of hydrogel dressings in Examples 3 - 5 and control dressings on alveolar bone formation around titanium implants (n = 4)

[0042] Appendix Figure 2 : Effect of hydrogel dressings in Examples 3 - 5 and control on vertical growth height of bone around titanium implants (n = 4) Detailed Description of the Invention

[0043] The present invention will be further described below in conjunction with specific embodiments, but the embodiments do not limit the present invention in any form. Unless otherwise specified, the reagents, methods, and equipment used in the present invention are conventional reagents, methods, and equipment in the technical field.

[0044] Unless otherwise specified, the reagents and materials used in the following examples are all commercially available.

[0045] Example 1 Preparation of Compound A Hydrogel Dressing

[0046] 1) Take 0.1 g of Compound A and dissolve it in 70 ml of phosphate buffer solution (pH 7.4, 100 mM).

[0047] 2) Take 15.0 g of poloxamer 407 and add it to the solution obtained in step 1), stirring while adding. After poloxamer 407 is completely dissolved, add it to phosphate buffer solution (pH 7.4, 100 mM) and make up the volume to 100 ml;

[0048] 3) Take the gel solution obtained in step 2), dispense it into vials, 0.5 ml per vial, add rubber stoppers, and crimp aluminum caps to obtain.

[0049] Control Example:

[0050] According to the preparation process of Example 1, without adding Compound A, prepare a 15% poloxamer 407 hydrogel dressing as a comparative example.

[0051] Preparation of hydrogels with different concentrations of Compound A in Examples 2 - 6

[0052] According to the preparation process of Example 1, only change the addition amount of Compound A in step 1), and sequentially prepare hydrogel dressings containing 0.5%, 1.0%, 1.5%, 2.0%, and 5.0% respectively, which are defined as the hydrogel dressings of Examples 2 - 6 in sequence.

[0053] Antibacterial experiment of Compound A in Example 7 (implemented with reference to Chinese Patent CN113679632B)

[0054] 7.1 Experimental method

[0055] (1) Strain culture

[0056] Incubate Candida albicans ATCC SC5314 in RPMI 1640 medium at 37°C with a humidity of 80%. under the conditions of;

[0057] Incubate Staphylococcus aureus ATCC 6538 in TSB medium at 37°C. under the conditions of;

[0058] Incubate Methicillin - resistant Staphylococcus aureus 18908 (MRSA 18908) in TSB medium at 37°C. under the conditions of;

[0059] Incubate Porphyromonas gingivalis ATCC 33277 in BHI medium added with haemin (5 μg / mL) and menadione (1 μg / mL) at 37°C under strict anaerobic conditions;

[0060] Incubate Streptococcus mutans Clarke ATCC UA159 in brain - heart infusion agar medium at 37°C. under the conditions of;

[0061] Incubate Streptococcus sanguis ATCC 10556 in brain - heart infusion agar medium at 37°C. under the conditions of.

[0062] (2) Sample

[0063] Dissolve compound A in distilled water and adjust the concentration to 10 mg / mL. A total of 20 mL is prepared and stored in a refrigerator at 4 °C for later use.

[0064] (3) Determination of minimum inhibitory concentration (MIC)

[0065] With reference to the regulations of the Clinical and Laboratory Standards Institute (CLSI), the microbroth dilution method is implemented to determine the minimum inhibitory concentration (MIC) of compound A against different microorganisms. This method is a recognized and effective method by the Clinical and Laboratory Standards Institute (CLSI), with a wide and common application range. It can quickly detect the drug interaction effect within a short time, and reliable conclusions can be obtained through further analysis of experimental data. The results are stable and highly reproducible.

[0066] For Candida albicans ATCC SC5314, use normal saline to adjust the concentration of the test strain to and then dilute the bacterial suspension to in RPMI 1640 medium to prepare the test bacterial solution. Add 200 μL of the test bacterial solution to the first well of a flat-bottom 96-well plate, and then add 100 μL of the test bacterial solution to each subsequent well. Add 2 μL of the test compound stock solution (the concentration of compound A in the well is 100 μg / mL) to the first well, pipette and mix well in the first well, take 100 μL of the homogeneous suspension, add it to the second well, pipette and mix well, and repeat until the last well. Discard the excess bacterial solution. Incubate the 96-well plate at 37 °C for 24 hours. The MIC is defined as the minimum drug concentration that can inhibit the growth of microorganisms observed with the naked eye. This experiment is repeated three times, and econazole is selected as the positive control.

[0067] For Staphylococcus aureus ATCC 6538 and methicillin-resistant Staphylococcus aureus 18908, use MH medium to adjust the concentration of the test strain to to prepare the test bacterial solution. Add 200 μL of the test bacterial solution to the first well of a flat-bottom 96-well plate, and then add 100 μL of the test bacterial solution to each subsequent well. Add 2 μL of the test compound stock solution (the concentration of compound A in the well is 100 μg / mL) to the first well, pipette and mix well in the first well, take 100 μL of the homogeneous suspension, add it to the second well, pipette and mix well, and repeat until the last well. Discard the excess bacterial solution. Incubate the 96-well plate at 37 °C for 24 hours. The MIC is defined as the minimum drug concentration that can inhibit the growth of microorganisms observed with the naked eye. This experiment is repeated three times, and vancomycin is used as the positive control.

[0068] For Streptococcus mutans ATCC UA159 and Streptococcus sanguinis ATCC 10556, the test strains were adjusted to a concentration of using BHI medium to prepare the test bacterial suspension. Take 200 μL of the test bacterial suspension and add it to the first well of a flat-bottom 96-well plate. Thereafter, add 100 μL of the test bacterial suspension to each subsequent well. Add 2 μL of the test compound stock solution (the concentration of compound A in the well is 100 μg / mL) to the first well, pipette and mix well in the first well. Take 100 μL of the homogeneous suspension, add it to the second well, pipette and mix well, and repeat until the last well. Discard the excess bacterial liquid. Incubate the 96-well plate at 37 °C, under the conditions for 24 hours. The MIC is defined as the minimum drug concentration that can inhibit microbial growth by visual observation. This experiment was repeated three times, and vancomycin was selected as the positive control.

[0069] For Porphyromonas gingivalis ATCC33277, hemin (5 μg / mL) and vitamin K (1 μg / mL) were added to BHI medium, and the test strain was adjusted to a concentration of to prepare the test bacterial suspension. Take 200 μL of the test bacterial suspension and add it to the first well of a flat-bottom 96-well plate. Thereafter, add 100 μL of the test bacterial suspension to each subsequent well. Add 2 μL of the test compound stock solution (the concentration of compound A in the well is 100 μg / mL) to the first well, pipette and mix well in the first well. Take 100 μL of the homogeneous suspension, add it to the second well, pipette and mix well, and repeat until the last well. Discard the excess bacterial liquid. Incubate the 96-well plate at 37 °C under strict anaerobic conditions for 24 hours. The MIC is defined as the minimum drug concentration that can inhibit microbial growth by visual observation. This experiment was repeated three times, and chlorhexidine was selected as the positive control.

[0070] 7.2 Antibacterial experiment results

[0071] Table 1 Minimum inhibitory concentration MIC values of compound A

[0072] .

[0073] Table 1 shows the antibacterial results of compound A against common oral bacteria. From the results in the table, it can be seen that compound A has significant antibacterial effects on Candida albicans, Staphylococcus aureus, MRSA, Streptococcus mutans, Streptococcus sanguinis, and Porphyromonas gingivalis, and the MIC values are between 12.5 - 50 μg / mL.

[0074] Example 8 Gelation of temperature-sensitive hydrogel dressings in Examples 1 - 6 and Comparative Examples

[0075] When the temperature changes, the aqueous solution of poloxamer 407 hydrogel will undergo a thermosensitive phase transition. The solution remains in a solution state at room temperature. After the temperature rises, gelation occurs and the gel viscosity increases sharply. When the temperature rises to 37 °C, the viscosities of the hydrogel dressings obtained in the comparative example and Examples 1-6 all change greatly. The gelling state was confirmed by the tilting method under body temperature conditions.

[0076] In addition, the loaded drug can change the hydrophobic / hydrophilic ratio, so the gelation characteristics will be affected by the loaded drug. Therefore, loading compound A will affect the temperature-sensitive characteristics of poloxamer 407 hydrogel. To confirm the gelation characteristics of poloxamer 407 hydrogel loaded with compound A at body temperature, the gelation temperatures of the hydrogel dressings obtained in Examples 1-6 and the comparative example were studied. The results show that there is no significant change in the gelation temperature of the hydrogel dressings obtained in Examples 1-6 compared with the poloxamer 407 hydrogel without loaded compound A (comparative example).

[0077] The hydrogel dressings obtained in Examples 1-6 and the comparative example were gradually heated (heating rate: 0.5 °C / min) under continuous stirring (rotation speed: 100 revolutions per minute). The temperature was set by a constant temperature water bath, and the temperature inside the sample was controlled using a precision thermometer (accuracy: ±0.1 °C). When the stir bar (length 25 mm, diameter 6 mm) stopped moving due to gelation, the temperature displayed at this time was recorded as the temperature for the transition from solution to gel, that is, the gelation temperature.

[0078] Table 2 Gelation temperatures of the hydrogel dressings obtained in Examples 1-6 and the hydrogel dressing of the comparative example

[0079] 。

[0080] Example 9 In situ bone formation study of the hydrogel dressings obtained in Examples 1-6 and the comparative example by single injection

[0081] Male C57BL / 6 mice at 6 weeks of age and weighing 18-20 g were randomly divided into eight different groups, with 5 mice in each group. The hydrogel dressings obtained in Examples 1-6 and the comparative example and normal saline were given respectively. After shaving each mouse, 200 μL of the hydrogel dressings or normal saline with different formulations was subcutaneously injected at the back. After 8 weeks, the mice were sacrificed by carbon dioxide asphyxiation, and then specimens of each group were collected at and around the injection site. The specimens were carefully taken out, fixed in 10% neutral buffered formalin solution for 24 hours, and then transferred to 70% ethanol.

[0082] The bone formation ability of the hydrogel dressings obtained in Examples 1-6 and the comparative example was evaluated in situ by single injection.

[0083] Eight weeks after injection, the generated bone was evaluated by soft X-ray and micro-computed tomography (μ-CT). Irregular spherical bone formation was observed at the injection site.

[0084] According to the formula, the increased bone tissue volume = the bone tissue volume observed in the dressing-administered group - the bone tissue volume observed in the control group, in cubic millimeters. The experimental data are as follows:

[0085] Table 3 Increased bone tissue volume of experimental mice in each group

[0086] .

[0087] As the concentration of Compound A in the hydrogel dressing increased, the formed bone tissue gradually became larger. However, when the concentration increased to 2% or more, that is, when the concentration increased to 5%, compared with the hydrogel dressing with a concentration of 2%, the formed bone tissue did not increase significantly. No bone-like tissue was observed in the control group.

[0088] Eight weeks after injection, all the formed hydrogels completely disappeared. No inflammatory reaction was observed in any experimental group. These non-toxic reactions indicate that the hydrogel dressings obtained in Examples 1-6 have excellent biocompatibility. The surrounding cells can infiltrate well into the hydrogel dressing, and the continuously released Compound A from the hydrogel dressing contributes to cell survival, differentiation, and bone tissue formation.

[0089] Example 10 Oral titanium implant experiment in beagle dogs

[0090] 10.1 Implantation of titanium implants:

[0091] All surgeries were performed under general anesthesia and local anesthesia under sterile conditions, using a mixture of cimetidine hydrochloride, cefazolin sodium, dexmedetomidine, and tiletamine / zolazepam. Anesthesia was maintained using isoflurane and 100% pure oxygen.

[0092] After extracting the mandibular teeth from the first premolars to the first molars of two beagle dogs respectively, the incision sites were sutured with 4-0 nylon thread. After 8 weeks, four bone defects with a height of 3 mm and a length of 16 mm were created on the left and right mandibles using a water-cooled rotary dental drill. A total of 16 titanium implants were implanted, 2 implants at each bone defect site, and 8 implants in each dog. The distance between the implants was approximately 4 mm, and 4 mm of the upper part of the implant was exposed outside the alveolar bone. In the control group (four implants), 250 μL of the hydrogel dressing prepared in the comparative example was used at each bone defect site. After gelling, the wound was immediately sutured with 4-0 nylon thread. In the experimental group, 250 μL of the compound A hydrogel dressing prepared in Examples 3-5 was used to treat the bone defects where the remaining twelve implants were located, with four implants in each example group. After the polymer solution gelled under the action of body temperature, the wound was immediately sutured with 4-0 nylon thread.

[0093] Amoxicillin and meloxicam were orally administered for 7 consecutive days after the operation, the former twice a day and the latter once a day. During the experiment, the oral cavity was rinsed with 2% chlorhexidine gluconate every day to maintain oral hygiene.

[0094] 10.2 Clinical observation

[0095] The surgical procedures of the animals were successful. The health status of the animals was checked every day, the dogs were active normally, and the healing process proceeded naturally. Tooth extraction, alveolar ridge trimming, and titanium implant placement were completed according to the research protocol.

[0096] The experimental design is summarized in Table 4. A defect with a height of 3 mm and an area of 4×16 mm was created on the beagle dog mandible using a water-cooled rotary dental drill. After placing the titanium implant, the freshly prepared hydrogel dressing of Examples 3-5 was spread on it to completely cover the bone defect and the titanium implant. After a few seconds, it was confirmed that a solid gel was formed. Subsequently, the exposed tissue was sutured. Within 12 weeks after treatment, all jaw quadrants healed completely, and no wound healing failure or inflammatory reaction occurred.

[0097] Table 4 Experimental design of titanium implants in beagle dogs

[0098] 。

[0099] 10.3 Imaging observation:

[0100] At the 4th week and the 12th week after the operation, two-dimensional images of the mandible were monitored using a portable X-ray machine. For three-dimensional analysis, each specimen was scanned in a cone-beam acquisition mode by micro-CT. A CT analysis program was used to measure the volume and mineral density of the newly formed bone. Three-dimensional surface rendering images were made using Mimics 14.0 version imaging software.

[0101] After tooth loss, alveolar ridge bone resorption and reduction of alveolar bone height are expected to occur. These conditions may reduce the stability of implants, shorten the implant lifespan, and cause difficulties for patients in secondary implant placement. In more severe cases, due to the reduction of alveolar bone height, secondary implant placement may become impossible. The compound A hydrogel dressing prepared in this application is expected to prevent alveolar bone reduction and stimulate vertical bone regeneration around the implant site, increasing bone mass to improve implant stability.

[0102] After treating the critical defect sites around the implant with the hydrogel dressings of Examples 3 - 5, the changes in alveolar bone defects over time were monitored by intraoral X - rays. In the control group, over time, slight bone resorption and implant exposure occurred around the implant site. Compared with the control group, significant bone regeneration was observed in the hydrogel dressing groups loaded with compound A prepared in Examples 3 - 5. The exposed threads of the implant were completely covered by new bone, and the degree of coverage of the threads by new bone increased with the increase in the content of compound A in the dressing.

[0103] 10.4 Histological observation:

[0104] After 12 weeks of treatment, the effects of the hydrogel dressings of Examples 3 - 5 on bone regeneration were evaluated by X - rays and micro - computed tomography (CT).

[0105] For histological analysis, the mandible including the implant was removed using a high - speed water - cooled diamond saw and an EXAKT cutting system.

[0106] The degree of bone regeneration was studied by X - rays and micro - CT scans. There were significant differences in the height of the bone tissue around the four groups of implants. In fact, the exposed area in the control group was rarely covered by regenerated bone. However, after treatment with the hydrogel dressings of Examples 3 - 5, a significant amount of bone regeneration was shown around the implant wall, and it increased with the increase in the content of compound A in the dressing. Most areas of the implant threads in the hydrogel dressing groups of Examples 3 - 5 were wrapped by vertically growing bone tissue. Compared with the control group, the bone volume in the hydrogel dressing groups of Examples 3 - 5 increased significantly. The bone volumes in the regions of interest (measuring bone volume from the innermost point of the implant thread to 0.5 mm from the starting point) for the hydrogel dressings of Examples 3 - 5 were 34.86 mm³, 35.74 mm³, and 35.81 mm³ respectively, and the bone tissue completely covered the threads. While the bone volume in the control group was only 23.73 mm³, and the implant threads were still observable. In summary, compared with the control group, the bone regeneration effects of the hydrogel dressings of Examples 3 - 5 were increased by 1.47 times, 1.50 times, and 1.55 times respectively. In addition, there were also differences in bone mineral density (measured from the innermost point of the implant thread to 0.5 mm from the starting point) between the hydrogel dressings of Examples 3 - 5 and the control group. The specific experimental data are as follows:

[0107] Table 5 Promoting effects of the hydrogel dressings of Examples 3-5 and the control group on alveolar bone formation around titanium implants

[0108] 。

[0109] 10.5 Histomorphometric analysis:

[0110] All specimens were dehydrated through a series of alcohol with gradient concentrations. After dehydration, the specimens were embedded in acrylic resin. Using a high-speed water-cooled diamond saw and an EXAKT cutting system, longitudinal section samples containing tissues and implants were obtained.

[0111] Histomorphometric analysis was performed using an optical microscope and an Image-Pro Plus image analysis system. The central sections of each implant were used for histomorphometric analysis. The induced bone height (mm) was measured at 0.5 mm, 1.0 mm, and 2.0 mm away from the implant.

[0112] Histomorphometric analysis was carried out to evaluate the degree of regenerated bone. Quantitative evaluation showed that there were significant differences in the vertical bone growth of the bone tissues around the implants in the four test groups. The vertical bone increment was measured at each point 0.5 mm, 1.0 mm, and 2.0 mm away from the implant wall. The bone regeneration along the implant wall from the defect interface in the control group was limited. However, the bone height at all measurement points in the hydrogel dressing groups of Examples 3-5 was significantly higher than that in the control group.

[0113] The measured bone vertical growth height in the hydrogel dressing groups of Examples 3-5 was 0.33 - 0.44 mm from the surface of the implant wall to 2.0 mm away from the implant. The bone height at each point in the hydrogel dressing groups of Examples 3-5 was much higher than that in the control group. The specific experimental data are shown in Table 6.

[0114] Table 6 Effects of the hydrogel dressings of Examples 3-5 and the control group on the vertical bone growth height around titanium implants

[0115] 。

[0116] The above embodiments are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications made without departing from the spirit and principle of the present invention shall be equivalent replacement methods and are all included in the protection scope of the present invention.

Claims

1. Use of compound A for preparing a hydrogel dressing for promoting local bone tissue regeneration around titanium implants, characterized in that: The mass concentration of the compound A in the hydrogel dressing is 1.0%-2.0%, and the chemical structure of the compound A is as follows: 。 2. The use according to claim 1, characterized in that The compound A is used for preparing a hydrogel dressing for inhibiting bacterial growth around implants.

3. The use according to claim 1, characterized in that The dressing is a hydrogel dressing.

4. The use according to claim 2, characterized in that The mass percentage concentration of compound A in the dressing is 1.5%.

5. The use according to claim 2, characterized in that The mass percentage concentration of compound A in the dressing is 2%.

6. The use according to any one of claims 3 to 5, characterized in that: The dressing is a temperature-sensitive hydrogel dressing.

7. The use according to claim 6, characterized in that The preparation method of the dressing is as follows: Step 1) dissolving compound A in phosphate buffer; Step 2) Poloxamer 407 is added to the solution obtained in step 1) while stirring, and after the poloxamer 407 is completely dissolved, the solution is added to the phosphate buffer and the volume is adjusted to the full amount; Step 3) Take the gel solution obtained in step 2) and dispense it into vials, add rubber stoppers, and roll aluminum caps to obtain the product.

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