Renewable boundary lubricating natural polyphenol anti-inflammatory hydrogel, and preparation method and application thereof

Abstract

The application belongs to the technical field of biomedical anti-inflammatory gels, and discloses a renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel as well as a preparation method and application thereof. The hydrogel has a hydrophilic viscoelastic island structure, the base material is gallic acid grafted gelatin blended with hyaluronic acid, and the interior is loaded with celecoxib liposomes. The joint cavity is injected and gelled through enzyme cross-linking technology. The hyaluronic acid and the liposomes form a hydration lubricating layer at the cartilage-gel interface. After the interface is worn, the interior liposomes and hyaluronic acid are exposed to supplement lubrication to form sustainable lubrication. The liposomes are broken to release celecoxib in response to frictional shear, which cooperates with gallic acid at the interface to resist inflammation and repair cartilage. The application develops an injectable hydrogel based on gallic acid, hyaluronic acid, gelatin and celecoxib liposomes to treat early osteoarthritis. Mechanical lubrication and inflammation resistance are combined to effectively overcome the problems of large-dose toxicity and short lubrication period caused by single treatment, and the application has important clinical application significance.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of biomedical anti-inflammatory gels, and particularly relates to a renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel as well as a preparation method and application thereof. BACKGROUND

[0002] Osteoarthritis is a chronic degenerative disease characterized by progressive degeneration of articular cartilage, synovial inflammation and abnormal bone remodeling. According to WHO statistics, osteoarthritis has a high incidence, and there are more than 300 million patients worldwide, among which more than 80% of the knee joint inflammation of the middle-aged and elderly population over 65 years old is the susceptible population, and the trend is becoming younger. Studies have shown that obesity, sports injury and genetic factor mutation can all cause the age of onset to be advanced. The normal joint friction coefficient is close to that of ice surface (μ≈0.03), and the joint lubrication function depends on the key lubrication molecules of synovial fluid and cartilage surface layer, and its failure is the core pathological mechanism of osteoarthritis. At the same time, due to the failure of joint lubrication, the friction coefficient increases by more than 10 times, accelerating the wear of cartilage, forming a vicious cycle of "tissue damage-inflammation-lubrication decrease-secondary damage-inflammation aggravation".

[0003] At present, the main clinical osteoarthritis treatment methods are oral non-steroidal anti-inflammatory drugs and intra-articular injection of hyaluronic acid, but the exogenous hyaluronic acid has a short half-life and a non-durable mechanical lubrication effect, and the oral non-steroidal anti-inflammatory drugs have a large demand dose and a risk of gastrointestinal toxicity and cardiovascular damage.

[0004] Therefore, it is of great market prospect to develop a new drug for treating osteoarthritis. SUMMARY

[0005] The application aims to solve the problems in the prior art, and provides a renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel as well as a preparation method and application thereof.

[0006] In order to solve the technical problem, the technical scheme of the application is as follows: the renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel comprises gallic acid, hyaluronic acid, gelatin, celecoxib liposomes and a solvent, and the weight percentage contents of the gallic acid, the hyaluronic acid, the gelatin and the celecoxib liposomes are 1-2 wt%, 0.3-0.8 wt%, 8-12 wt% and 1-5 wt% respectively, and the solvent makes up the balance, the celecoxib liposomes are composed of lecithin, cholesterol, distearoyl phosphatidyl ethanolamine-polyethylene glycol and celecoxib, and the weight ratio of the lecithin, the cholesterol, the distearoyl phosphatidyl ethanolamine-polyethylene glycol and the celecoxib is 15-25:2-6:1-5:1-5.

[0007] Preferably, the weight percentage contents of the gallic acid, the hyaluronic acid, the gelatin and the celecoxib liposomes are 1.2 wt%, 0.5 wt%, 10 wt% and 2 wt% respectively, and the solvent makes up the balance.

[0008] Preferably, the weight ratio of the lecithin, cholesterol, distearoylphosphatidyl ethanolamine-polyethylene glycol and celecoxib is 20:4:2:2.

[0009] Preferably, the solvent is a sodium chloride solution, a phosphate buffer or pure water.

[0010] Preferably, the method for preparing the renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel comprises the following steps for preparing the renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel described above:

[0011] Step 1: activation of gallic acid; mix ultrapure water and N,N-dimethylformamide uniformly and sufficiently to release heat, the volume ratio of the ultrapure water to the N,N-dimethylformamide is 1.2-1.6:1, add gallic acid and N-hydroxysuccinimide or 1-ethyl-(3-dimethylaminopropyl) carbodiimide, then place in a 4-6℃ environment under stirring at 300-800 rpm for 4-12 h, and protect the whole process of activation under a nitrogen atmosphere, to obtain activated gallic acid, the concentration of gallic acid in the activated gallic acid is 2-3wt%, the concentration of N-hydroxysuccinimide is 2.5-3.5wt%, and the concentration of 1-ethyl-(3-dimethylaminopropyl) carbodiimide is 2-2.5wt%;

[0012] Step 2: preparation of gallic acid modified gelatin; mix a 5wt% gelatin solution with the activated gallic acid of step 1, the volume ratio is 1-2:1, react at 40℃ for 12-24 h under a nitrogen atmosphere, the stirring speed is 800-1000 rpm, collect the reaction solution, remove unreacted molecules by dialysis at room temperature for 6-10 days, and collect by freeze-drying to obtain gallic acid modified gelatin GAG;

[0013] Step 3: preparation of natural polyphenol anti-inflammatory hydrogel; dissolve the gallic acid modified gelatin GAG and hyaluronic acid in a solvent, the weight ratio of the gallic acid modified gelatin to the hyaluronic acid is 8-12:0.3-0.8, the dissolution temperature is <55℃, add 0.5wt% transaminase to the above mixed system, gel at 37℃ for 3-5 min, remove unreacted molecules by dialysis at room temperature for 4-14 days, and collect by freeze-drying to obtain the natural polyphenol anti-inflammatory hydrogel GAGgel;

[0014] Step 4: preparation of celecoxib liposomes; dissolve lecithin, cholesterol, distearoylphosphatidyl ethanolamine-polyethylene glycol and celecoxib in chloroform, the weight ratio is 15-25:2-6:1-5:1-5; remove chloroform by rotary evaporation at 37℃, and use solvent hydration, obtain celecoxib liposome dispersion CLP by ultrasonic crushing, and concentrate the dispersion to a final concentration of 1-5wt% by ultrafiltration centrifugation;

[0015] Step 5: Preparation of renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel; dissolve the natural polyphenol anti-inflammatory hydrogel GAGgel into the celecoxib liposome dispersion CLP and mix well, the dissolution temperature is <45℃, then add 0.5wt% transaminase, 3~5min at 37℃ to obtain the renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel GAGgel@CLP, wherein the weight percentage contents of gallic acid, hyaluronic acid, gelatin and celecoxib liposome are 1~2wt%, 0.3~0.8wt%, 8~12wt%, 1~5wt% respectively, and the balance is solvent, and the solvent is sodium chloride solution, phosphate buffer or pure water.

[0016] Compared with the prior art, the application has the following advantages:

[0017] (1) The application provides a renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel, which has a hydrophilic viscoelastic island structure, and the substrate is gallic acid grafted gelatin blended with hyaluronic acid, and the inside is loaded with celecoxib liposomes, which are injected into the joint cavity to form a gel through enzyme crosslinking technology. Hyaluronic acid and liposomes form a hydrated lubricating layer at the cartilage-gel interface, and the inside liposomes and hyaluronic acid are exposed after interface wear to form sustainable lubrication. The liposomes respond to frictional shear to break and release celecoxib to resist inflammation progression and repair cartilage in cooperation with gallic acid at the interface. The application develops an injectable hydrogel based on gallic acid, hyaluronic acid, gelatin and celecoxib liposomes to treat early osteoarthritis, which combines mechanical lubrication and inflammation resistance, effectively overcoming the problems of large dose toxicity and short lubrication period caused by single treatment, and has important clinical application significance.

[0018] (2) The celecoxib liposomes in the application contain a large number of phosphatidyl hydrophilic groups on the surface, which can tightly bind with water molecules to form a hydrated film. Water molecules can move relatively freely in the hydrated film, thereby producing a lubricating effect at the interface. At the same time, the celecoxib liposomes are exposed to the cartilage surface in response to friction, thereby achieving the purpose of anti-inflammation.

[0019] (3) The application uses enzyme crosslinked gallic acid gelatin as a substrate to coat high-concentration liposomes and hyaluronic acid. Hyaluronic acid and liposomes form a hydrated lubricating layer at the cartilage-gel interface, and the inside liposomes and hyaluronic acid are exposed after interface wear to form renewable lubrication.

[0020] (4) The renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel prepared by the application has good anti-inflammatory function and lubricating effect. The hydrogel first exposes gallic acid to scavenge active oxygen to avoid oxidative stress, and then responds to frictional shear to release the sequential treatment strategy of anti-inflammatory drugs. This strategy is in line with the pathogenesis of early osteoarthritis, especially traumatic osteoarthritis. The hydrogel has good biocompatibility and injectability, and is suitable for the treatment of early osteoarthritis, especially traumatic osteoarthritis.

[0021] (5) The preparation method of the application has simple process and low cost, uses transaminase as a crosslinking agent to avoid the cytotoxicity of traditional photocuring agents, improves injectability, uses liposome to load non-steroidal anti-inflammatory drug celecoxib to synergistically inject gallic acid-gelatin in situ, greatly reduces the dosage of the drug and improves the utilization rate, and avoids gastrointestinal damage and cardiovascular hazards caused by oral administration. BRIEF DESCRIPTION OF DRAWINGS

[0022] Figure 1 A process flow chart of the preparation method of the renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel of the application;

[0023] Figure 2 A nuclear magnetic resonance hydrogen spectrum of gallic acid modified gelatin GAG in Example 1 of the application 1 HNMR and Fourier infrared spectrum results;

[0024] Figure 3 A transmission electron microscope image and particle size and distribution chart of celecoxib liposome dispersion CLP in Example 2 of the application;

[0025] Figure 4 Scanning electron microscope images, mechanical property charts and swelling charts of the natural polyphenol anti-inflammatory hydrogel GAGgel and the renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel GAGgel@CLP in Examples 1 and 3 of the application;

[0026] Figure 5 Friction coefficient charts of the natural polyphenol anti-inflammatory hydrogel GAGgel and the renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel GAGgel@CLP in Examples 1 and 3 of the application;

[0027] Figure 6 Cytotoxicity charts and hemolysis detection charts of the natural polyphenol anti-inflammatory hydrogel GAGgel and the renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel GAGgel@CLP in Examples 1 and 3 of the application;

[0028] Figure 7 Flow cytometry charts of the natural polyphenol anti-inflammatory hydrogel GAGgel and the renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel GAGgel@CLP in Examples 1 and 3 of the application. DETAILED DESCRIPTION

[0029] The application is described in detail below with reference to the accompanying drawings and specific examples, but the application is not limited to these examples. The application covers any alternatives, modifications, equivalent methods and solutions made within the spirit and scope of the application. In order to make the public have a thorough understanding of the application, specific details are described in the following examples of the application, but the description of these details is also completely understood by those skilled in the art without the description of the application.

[0030] The application provides a renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel, which has a hydrophilic viscoelastic island structure, gallic acid molecules are coupled to the side chain of gelatin through acylation, and hyaluronic acid and liposomes form a hydrated lubricating layer at the cartilage-gel interface to play a lubricating role.

[0031] The natural polyphenol compound gallic acid provided by the application can efficiently scavenge ROS to resist oxidative stress, and target the NF-κB / MAPK pathway to inhibit the inflammatory cascade reaction; gelatin contains an arginine-glycine-aspartic acid sequence, which is beneficial to cell adhesion and growth, and the presence of a large number of amino groups also provides the possibility for gallic acid modification; celecoxib is a non-steroidal anti-inflammatory drug, which specifically inhibits COX-2 enzyme activity and reduces the generation of prostaglandin E2; liposomes are assembled from components such as phospholipids and cholesterol, and have similar composition, drug loading capacity and hydrated lubricating potential to cell membranes.

[0032] The application discloses a renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel, which comprises gallic acid, hyaluronic acid, gelatin, celecoxib liposomes and a solvent, and the weight percentage of the gallic acid, the hyaluronic acid, the gelatin and the celecoxib liposomes is 1-2 wt%, 0.3-0.8 wt%, 8-12 wt% and 1-5 wt% respectively, and the solvent makes up the balance, and the celecoxib liposomes are composed of lecithin, cholesterol, distearoyl phosphatidyl ethanolamine-polyethylene glycol and celecoxib, and the weight ratio of the lecithin, the cholesterol, the distearoyl phosphatidyl ethanolamine-polyethylene glycol and the celecoxib is 15-25:2-6:1-5:1-5.

[0033] Preferably, the weight percentage of the gallic acid, the hyaluronic acid, the gelatin and the celecoxib liposomes is 1.2 wt%, 0.5 wt%, 10 wt% and 2 wt% respectively, and the solvent makes up the balance.

[0034] Preferably, the weight ratio of the lecithin, the cholesterol, the distearoyl phosphatidyl ethanolamine-polyethylene glycol and the celecoxib is 20:4:2:2.

[0035] Preferably, the solvent is a sodium chloride solution, a phosphate buffer or pure water.

[0036] AsFigure 1 As shown, preferably, the preparation method of the renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel, for preparing the above-mentioned renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel, comprises the following steps:

[0037] Step 1: activation of gallic acid; mix ultrapure water and N,N-dimethylformamide uniformly and sufficiently to release heat, the volume ratio of ultrapure water to N,N-dimethylformamide is 1.2-1.6:1, add gallic acid and N-hydroxysuccinimide or 1-ethyl-(3-dimethylaminopropyl) carbonyldiimide, then place in a 4-6℃ environment under stirring at 300-800 rpm for 4-12 h, and the whole process is protected by nitrogen atmosphere, to obtain activated gallic acid, the concentration of gallic acid in the activated gallic acid is 2-3wt%, the concentration of N-hydroxysuccinimide is 2.5-3.5wt%, and the concentration of 1-ethyl-(3-dimethylaminopropyl) carbonyldiimide is 2-2.5wt%;

[0038] Step 2: preparation of gallic acid modified gelatin; mix a 5wt% gelatin solution with the activated gallic acid of step 1, the volume ratio is 1-2:1, react at 40℃ for 12-24 h under nitrogen atmosphere, the stirring speed is 800-1000 rpm, collect the reaction solution, remove unreacted molecules by dialysis at room temperature for 6-10 days, and collect by freeze-drying to obtain gallic acid modified gelatin GAG;

[0039] Step 3: preparation of natural polyphenol anti-inflammatory hydrogel; dissolve the gallic acid modified gelatin GAG and hyaluronic acid in a solvent, the weight ratio of gallic acid modified gelatin to hyaluronic acid is 8-12:0.3-0.8, the dissolving temperature is <55℃, add 0.5wt% transaminase to the above-mentioned mixed system, gel at 37℃ for 3-5 min, remove unreacted molecules by dialysis at room temperature for 4-14 days, and collect by freeze-drying to obtain natural polyphenol anti-inflammatory hydrogel GAGgel;

[0040] Step 4: preparation of celecoxib liposomes; dissolve lecithin, cholesterol, distearoylphosphatidyl ethanolamine-polyethylene glycol and celecoxib in chloroform, the weight ratio is 15-25:2-6:1-5:1-5; remove chloroform by rotary evaporation at 37℃, and use solvent hydration, and obtain celecoxib liposome dispersion CLP by ultrasonic crushing, and concentrate the dispersion to a final concentration of 1-5wt% by ultrafiltration centrifugation;

[0041] Step 5: Preparation of renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel; dissolve the natural polyphenol anti-inflammatory hydrogel GAGgel into the celecoxib liposome dispersion CLP and mix well, the dissolving temperature is less than 45℃, then add 0.5wt% transaminase, and obtain the renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel GAGgel@CLP under 37℃ for 3~5min, wherein the weight percentage contents of gallic acid, hyaluronic acid, gelatin and celecoxib liposome are 1~2wt%, 0.3~0.8wt%, 8~12wt%, 1~5wt% respectively, and the rest is solvent, which is sodium chloride solution, phosphate buffer or pure water.

[0042] Preferably, the application of the renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel, the above-mentioned renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel is used for the treatment of osteoarthritis.

[0043] Example 1: Preparation of natural polyphenol anti-inflammatory hydrogel GAGgel;

[0044] Activation of gallic acid: mix 38ml ultrapure water and 25ml N,N-dimethylformamide uniformly and release heat sufficiently, then add gallic acid and N-hydroxysuccinimide or 1-ethyl-(3-dimethylaminopropyl) carbonyldiimide, the concentrations are 2.5wt%, 2.7wt% and 2.3wt% respectively. Under the condition of stirring at 600 rpm, activate in 4℃ environment for 12h, and carry out nitrogen atmosphere protection during the whole activation process.

[0045] Preparation of gallic acid modified gelatin GAG: mix the gelatin solution with a concentration of 5wt% with the above-activated gallic acid, the volume ratio is 2:1, react at 40℃ for 10 hours under nitrogen atmosphere, and the stirring speed is 800rpm.

[0046] Collect the reaction solution, remove the unreacted molecules by dialysis at room temperature for 6~10, and collect by freeze-drying.

[0047] As shown in Figure 2 , Figure 2 Fig. 1, (a) and (b) are the FT-IR spectra of blank gelatin and gallic acid modified gelatin respectively. 1 HNMR results, wherein Figure 2 In (b), the characteristic peak at 7.0 ppm can be observed, which is generated by the hydrogen adjacent to the benzene ring of gallic acid, proving the success of the modification. Figure 2 In (c), the Fourier infrared spectrum results, a wide peak can be observed at 3500cm -1 , which is generated by the stretching vibration of phenolic hydroxyl group, further proving the successful modification of gallic acid to gelatin.

[0048] Solvent selection 0.9% sodium chloride solution, the above gallic acid modified gelatin (10wt%) and hyaluronic acid ((0.5wt%) are dissolved into 0.9% sodium chloride solution, the dissolution temperature is <55℃. 0.5wt% transaminase is added to the above mixed system, and the gel is formed at 37℃ for 3~5min to obtain the natural polyphenol anti-inflammatory hydrogel GAGgel.

[0049] As shown in Figure 4 , Figure 4 (a) left side of the figure is the scanning electron microscope image of the natural polyphenol anti-inflammatory hydrogel GAGgel, and the hydrogel shows a continuous porous structure with a pore size of 5~50μm, which provides sufficient internal space for cell growth and penetration and drug transport. Figure 4 As shown in Figure 4 (b), the black line of the natural polyphenol anti-inflammatory hydrogel GAGgel is the mechanical property, and the three-dimensional network of the hydrogel system is doped with hyaluronic acid to form a semi-interpenetrating special structure with linear molecules, and the strong water activity of the hyaluronic acid molecules will shorten the distance between the polymer molecular chains, enhance the hydrogen bond effect, and make the storage modulus of the natural polyphenol anti-inflammatory hydrogel GAGgel increase to 10kPa.

[0050] As shown in Figure 5 , the friction behavior of the natural polyphenol anti-inflammatory hydrogel GAGgel is shown. The gel block is loaded in a liquid pool with a size of 3cm*3cm*0.5cm, and the probe is loaded with a diameter of 8mm polyethylene ball under a load of 1N for linear reciprocating friction test. Figure 5 As shown in , the natural polyphenol anti-inflammatory hydrogel GAGgel described in the application reduces the friction coefficient from 0.23 (blank group) to 0.1 (example 1), which has excellent lubricating performance.

[0051] Figure 6 As shown in Figure 6 (a), the cells still maintain more than 90% survival rate after being incubated with the natural polyphenol anti-inflammatory hydrogel GAGgel, Figure 6 (b) shows that there is no obvious hemolysis phenomenon after the natural polyphenol anti-inflammatory hydrogel GAGgel is co-incubated with 2% packed red blood cells, which shows that the natural polyphenol anti-inflammatory hydrogel GAGgel has good biocompatibility.

[0052] As shown in Figure 7As shown in the lipopolysaccharide constructed Raw 264.7 macrophage inflammation model, compared with the blank group, LPS inflammation group and inflammation + GAGgel treatment, the natural polyphenol anti-inflammatory hydrogel GAGgel co-incubated significantly promoted the down-regulation of M1 type macrophage ratio and avoided inflammatory polarization.

[0053] Example 2: Preparation of celecoxib liposome dispersion CLP for knee joint injection;

[0054] The celecoxib liposome is prepared by the film hydration method. The lecithin, cholesterol, distearoyl phosphatidyl ethanolamine-polyethylene glycol (DSPE-PEG2000) and celecoxib are dissolved in chloroform with a mass ratio of 20:4:2:2. The chloroform is removed by rotary evaporation at 37°C and hydrated with 0.9% sodium chloride solution. The celecoxib liposome dispersion CLP is obtained by ultrasonic crushing. The dispersion is concentrated to a final concentration of 1-5 wt% by ultrafiltration centrifugation.

[0055] As shown in FIG. 1, Figure 3 , Figure 3 (a) is a transmission electron microscope image of the celecoxib liposome dispersion CLP, Figure 3 (b) is the particle size distribution of the celecoxib liposome dispersion CLP.

[0056] Example 3: Preparation of renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel GAGgel@CLP;

[0057] The above natural polyphenol anti-inflammatory hydrogel GAGgel (10 wt%) and hyaluronic acid (0.5 wt%) are dissolved into the celecoxib liposome dispersion CLP and mixed well. The dissolution temperature is <45°C. 0.5 wt% transaminase is added to the above mixed system, and the gel is formed at 37°C for 3-5 min.

[0058] As shown in FIG. 2, Figure 4 , Figure 4 (a) is an electron microscope image of the renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel GAGgel@CLP, Figure 4 (b) is the mechanical property of the renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel GAGgel@CLP.

[0059] Compared with Figure 4 GAGgel and GAGgel@CLP in FIG. 2(b), the liposome is uniformly incorporated into the hydrogel as a nanoparticle, which can disperse stress when stressed, thereby avoiding the destruction of the hydrogel structure due to local stress concentration, so that the storage modulus of the renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel GAGgel@CLP is further improved to 12 kPa, which can meet the mechanical support requirements of the cartilage surface.

[0060] As shown in FIG. 3,Figure 5 As shown in the figure, the friction behavior of the renewable boundary lubricating natural polyphenol anti-inflammatory hydrogel GAGgel@CLP is shown. The gel block is loaded in a 3cm*3cm*0.5cm size liquid pool, and the probe is loaded with a 8mm diameter polyethylene ball under a 1N load for linear reciprocating friction test. As shown in the figure, the renewable boundary lubricating natural polyphenol anti-inflammatory hydrogel GAGgel@CLP reduces the friction coefficient from 0.23 (blank group) to 0.02 (Example 3), with significant lubricating performance and joint lubricating potential.

[0061] As shown in the figure, Figure 6 As shown in the figure, Figure 6 In (a), the cells still maintain more than 85% survival rate after co-incubation with the renewable boundary lubricating natural polyphenol anti-inflammatory hydrogel GAGgel@CLP, Figure 6 In (b), the renewable boundary lubricating natural polyphenol anti-inflammatory hydrogel GAGgel@CLP shows no obvious hemolysis after co-incubation with 2% packed red blood cells, showing good biocompatibility of the renewable boundary lubricating natural polyphenol anti-inflammatory hydrogel GAGgel@CLP.

[0062] As shown in the figure, Figure 7 As shown in the figure, in the lipopolysaccharide constructed Raw 264.7 macrophage inflammation model, compared with the blank group, LPS inflammation group, inflammation + GAGgel treatment and inflammation + GAGgel@CLP treatment, the renewable boundary lubricating natural polyphenol anti-inflammatory hydrogel GAGgel@CLP co-incubation significantly promotes the down-regulation of M1 type macrophage ratio and avoids inflammatory polarization, and is suitable for the treatment of osteoarthritis, especially traumatic osteoarthritis.

[0063] Through experimental verification, the renewable boundary lubricating natural polyphenol anti-inflammatory hydrogel of the present application has anti-inflammatory lubricating effect in the range of 1~2wt% gallic acid, 0.3~0.8wt% hyaluronic acid, 8~12wt% gelatin, 1~5wt% celecoxib liposome and solvent to make up the rest.

[0064] The application provides a renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel, which has a hydrophilic viscoelastic sea-island structure, a substrate of gallic acid grafted gelatin blended with hyaluronic acid, and internal encapsulation of celecoxib liposomes, and is injected into a joint cavity to form a gel through an enzyme crosslinking technology; the hyaluronic acid and the liposomes form a hydrated lubricating layer at a cartilage-gel interface, and the internal liposomes and the hyaluronic acid are exposed after interface wear to form sustainable lubrication; the liposomes are broken in response to frictional shear to release celecoxib to resist inflammation progression and repair cartilage in cooperation with gallic acid at the interface; the application develops an injectable hydrogel for treating early-stage osteoarthritis based on gallic acid, hyaluronic acid, gelatin and celecoxib liposomes, and combines mechanical lubrication and inflammation resistance, thereby effectively overcoming problems such as large-dose toxicity and short lubrication period caused by single treatment, and having important clinical application significance.

[0065] The celecoxib liposomes contain a large number of phosphatidyl hydrophilic groups on the surface, the groups can be combined with water molecules to form a hydrated film, the water molecules can move relatively freely in the hydrated film, thereby generating a lubricating effect at the interface, and the celecoxib liposomes are exposed to the cartilage surface in response to friction, thereby achieving the anti-inflammatory purpose.

[0066] The application adopts enzyme crosslinked gallic acid gelatin as a substrate to coat high-concentration liposomes and hyaluronic acid, and the hyaluronic acid and the liposomes form a hydrated lubricating layer at a cartilage-gel interface, and the internal liposomes and the hyaluronic acid are exposed after interface wear to form renewable lubrication.

[0067] The renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel prepared in the application has good anti-inflammatory function and lubricating effect, and the hydrogel first exposes gallic acid to remove active oxygen to avoid oxidative stress, and then releases anti-inflammatory drugs in response to frictional shear in a sequential treatment strategy, which is in line with the pathogenesis of early-stage osteoarthritis, especially traumatic osteoarthritis, and the hydrogel has good biocompatibility and injectability, and is suitable for the treatment of early-stage osteoarthritis, especially traumatic osteoarthritis.

[0068] The preparation method of the application has simple process and low cost, adopts transaminase as a crosslinking agent to avoid cytotoxicity caused by traditional photocuring agents, and improves injectability, adopts liposomes to load non-steroidal anti-inflammatory drug celecoxib to cooperate with gallic acid-gelatin to greatly reduce the dosage of the drug and improve the utilization rate, and avoid gastrointestinal damage and cardiovascular hazards caused by oral administration.

[0069] The preferred embodiments of the application are described in detail above, but the application is not limited to the above-described embodiments, and various changes can be made within the knowledge of those skilled in the art without departing from the purpose of the application.

[0070] Many other changes and modifications can be made to the application without departing from the spirit and scope of the application. It is to be understood that this application is not limited to particular embodiments described, and is intended in its broadest sense. The scope of the application is to be defined by the appended claims.

Claims

1. A renewable boundary-lubricating natural polyphenol anti-inflammatory hydrogel, characterized in that: The natural polyphenol anti-inflammatory hydrogel comprises gallic acid, hyaluronic acid, gelatin, celecoxib liposome and solvent, the weight percentage of the gallic acid, the hyaluronic acid, the gelatin and the celecoxib liposome is 1-2 wt%, 0.3-0.8 wt%, 8-12 wt%, 1-5 wt% respectively, and the solvent makes up the balance, the celecoxib liposome is composed of lecithin, cholesterol, distearoylphosphatidylethanolamine-polyethylene glycol and celecoxib, and the weight ratio of the lecithin, the cholesterol, the distearoylphosphatidylethanolamine-polyethylene glycol and the celecoxib is 15-25:2-6:1-5:1-5; the natural polyphenol anti-inflammatory hydrogel has a hydrophilic viscoelastic sea-island structure, the base material is gallic acid grafted gelatin blended with hyaluronic acid, and the celecoxib liposome is internally encapsulated.

2. The renewable boundary lubricating natural polyphenol anti-inflammatory hydrogel according to claim 1, characterized by: The weight percentage of the gallic acid, the hyaluronic acid, the gelatin and the celecoxib liposome is 1.2 wt%, 0.5 wt%, 10 wt% and 2 wt% respectively, and the solvent makes up the balance.

3. The renewable boundary lubricating natural polyphenol anti-inflammatory hydrogel according to claim 1, characterized in that: The weight ratio of the lecithin, the cholesterol, the distearoylphosphatidylethanolamine-polyethylene glycol and the celecoxib is 20:4:2:

2.

4. The renewable boundary lubricating natural polyphenol anti-inflammatory hydrogel according to claim 1, characterized in that: The solvent is a sodium chloride solution, a phosphate buffer or pure water.

5. A method for the preparation of a renewable boundary lubricating natural polyphenol anti-inflammatory hydrogel, characterized by, The method for preparing the natural polyphenol anti-inflammatory hydrogel comprises the following steps: Step 1: activation of gallic acid; ultrapure water and N,N-dimethylformamide are uniformly mixed and sufficiently exothermed, the volume ratio of the ultrapure water to the N,N-dimethylformamide is 1.2-1.6:1, gallic acid and N-hydroxysuccinimide or 1-ethyl-(3-dimethylaminopropyl) carbodiimide are added, then the mixture is activated under stirring at 300-800 rpm in an environment at 4-6 DEG C for 4-12 h, and the whole process is protected by nitrogen atmosphere, thereby obtaining activated gallic acid, wherein the concentration of the gallic acid in the activated gallic acid is 2-3 wt%, the concentration of the N-hydroxysuccinimide is 2.5-3.5 wt%, and the concentration of the 1-ethyl-(3-dimethylaminopropyl) carbodiimide is 2-2.5 wt%; Step 2: preparation of gallic acid modified gelatin; a 5 wt% gelatin solution is mixed with the activated gallic acid of step 1, the volume ratio is 1-2:1, the reaction is carried out at 40 DEG C for 12-24 h under nitrogen atmosphere, the stirring speed is 800-1000 rpm, the reaction solution is collected, dialysis is carried out at room temperature for 6-10 days to remove unreacted molecules, and then freezing drying is carried out to obtain gallic acid modified gelatin GAG; Step 3: preparation of natural polyphenol anti-inflammatory hydrogel; the gallic acid modified gelatin GAG and the hyaluronic acid are dissolved in a solvent, the weight ratio of the gallic acid modified gelatin to the hyaluronic acid is 8-12:0.3-0.8, the dissolving temperature is less than 55 DEG C, 0.5 wt% transaminase is added to the above-mentioned mixed system, gelation is carried out at 37 DEG C for 3-5 min, dialysis is carried out at room temperature for 4-14 days to remove unreacted molecules, and then freezing drying is carried out to obtain the natural polyphenol anti-inflammatory hydrogel GAGgel; Step 4: Preparation of Celecoxib Liposomes; dissolve egg phospholipid, cholesterol, distearoyl phosphatidyl ethanolamine-polyethylene glycol and celecoxib in chloroform with a weight ratio of 15-25:2-6:1-5:1-5; remove chloroform at 37℃ by rotary evaporation, and use solvent hydration, and obtain celecoxib liposome dispersion CLP by ultrasonic crushing; concentrate the dispersion to a final concentration of 1-5wt% by ultrafiltration centrifugation; Step 5: Preparation of Renewable Boundary Lubrication Natural Polyphenol Anti-inflammatory Hydrogel; dissolve natural polyphenol anti-inflammatory hydrogel GAGgel into celecoxib liposome dispersion CLP and mix well, with a dissolution temperature <45℃, then add 0.5wt% transaminase, and obtain renewable boundary lubrication natural polyphenol anti-inflammatory hydrogel GAGgel@CLP at 37℃ for 3-5min, wherein the weight percentage contents of gallic acid, hyaluronic acid, gelatin and celecoxib liposomes are 1-2wt%, 0.3-0.8wt%, 8-12wt%, 1-5wt% respectively, and the rest is solvent, which is sodium chloride solution, phosphate buffer or pure water.

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