A sandwich structured hydrogel cartilage replacement material and a method for preparing the same

Abstract

The present application belongs to the technical field of hydrogel preparation, and relates to a sandwich structure hydrogel cartilage replacement material and a preparation method thereof. The present application uses grape seed protein, hyaluronic acid and other biocompatible polymers as raw materials, and prepares the cartilage replacement hydrogel material by adopting a biomimetic concept. The obtained hydrogel has a sandwich structure of a drag reduction layer, a strong and tough layer and a mineralization layer. The surface of the drag reduction layer is introduced with a hydrophobic layer and phospholipid molecules, so as to endow the hydrogel with excellent friction performance and long-term wear resistance. In the strong and tough layer, a variety of non-covalent bonds are synergistically crosslinked, so that the obtained hydrogel has very high compressive strength. The mineralization layer contains hydroxyapatite nanoparticles, so that the obtained hydrogel has excellent bone induction potential. The sandwich structure hydrogel of the present application has a water content of 65%, a compressive strength of 71 MPa, and a friction coefficient of about 0.01. After 50,000 friction cycles, the friction coefficient remains unchanged, and the cell viability is as high as 100%, thereby providing strength and long-lasting durability guarantee for the material implantation into a living body.

Description

Technical Field

[0001] The present invention belongs to the technical field of hydrogel preparation, and in particular relates to a sandwich structure hydrogel cartilage substitute material and a preparation method thereof. Background Art

[0002] Osteoarthritis is a chronic disease characterized by dysfunction in joint lubrication, shock absorption, and load-bearing functions. Over 150 million people in my country suffer from osteoarthritis, and tens of millions of patients with bone defects require surgical treatment each year. With an aging population and increasing obesity, this number is increasing annually, significantly impacting human health. The pathogenesis of arthritis primarily stems from the aging of articular cartilage. Since articular cartilage is non-regenerative, the development of high-performance articular cartilage replacement materials is a challenge in both the medical and materials fields. Existing cartilage replacement materials are primarily based on rigid materials, such as titanium alloys and ultra-high molecular weight polyethylene. While these materials offer excellent compressive strength, they can cause severe surface wear and generate large amounts of wear debris during use due to the lack of lubrication at the sliding interface and stress concentration, which can impact joint life. Natural cartilage, on the other hand, is a special, soft and slippery material with a high moisture content of over 60%, a compressive strength exceeding 50 MPa, a coefficient of friction approximately 0.01, and excellent osteoinductive properties with underlying bone. Hydrogels have become the most promising cartilage replacement materials because of their mechanical properties, water content, cell compatibility and lubricating behavior similar to those of cartilage.

[0003] Although existing technologies have made many advances in the field of cartilage replacement hydrogels, and the individual properties of the resulting materials are close to those of cartilage, the overall performance is still far behind that of natural cartilage. For example, improving the mechanical properties of hydrogels usually requires reducing the water content, while a low friction coefficient requires a high degree of hydration of the polymer chains. This inherent contradiction makes the construction of artificial cartilage hydrogel materials extremely challenging. In addition, the bottom of the cartilage replacement material must have osteoinductive potential, and when introducing materials with osteoinductive potential such as hydroxyapatite and bioglass, the surface roughness and friction coefficient are often increased. However, natural cartilage can effectively integrate these seemingly contradictory properties. This is mainly because natural cartilage has an anisotropic multi-scale nanocomposite structure, while traditional hydrogel materials are isotropic materials. For example, the superficial layer of cartilage, that is, the external area in contact with synovial fluid, is mainly composed of hyaluronic acid, lubricants and phosphatidylcholine lipids. The highly hydrated phosphatidylcholine head group can reduce friction through a hydration lubrication mechanism, and achieve long-term wear resistance through the gradual release of lipids during wear; the middle layer has a higher concentration of collagen fibrils to provide load-bearing capacity; the bottom calcification area achieves integration with the subchondral tissue.

[0004] Therefore, how to effectively integrate the seemingly contradictory properties of high water content, wear resistance, high strength and osteoinduction potential to synthesize materials with structure and performance close to natural cartilage, while achieving super wear resistance, high strength and osteoinduction potential is a technical challenge faced by the field of hydrogel technology in the preparation of cartilage substitute materials. Summary of the Invention

[0005] To address the problems and shortcomings of the existing technology, the primary objective of this invention is to provide a sandwich-structured hydrogel cartilage replacement material. This material mimics the multi-layered structure of cartilage, using biocompatible polymers such as grape seed protein, hyaluronic acid, polyvinyl alcohol, chitosan, and cellulose as raw materials. The hydrogel adopts a biomimetic approach to prepare a cartilage replacement hydrogel material. The hydrogel comprises a sandwich structure consisting of a drag-reducing layer, a toughening layer, and a mineralized layer.

[0006] Another object of the present invention is to provide a method for preparing a sandwich structure hydrogel cartilage substitute material, which comprises four steps: preparation of a precursor hydrogel, surface modification, loading of lecithin, and bottom mineralization.

[0007] To achieve the above object, the present invention is implemented through the following technical solutions:

[0008] A sandwich-structured hydrogel cartilage replacement material, characterized in that the hydrogel has a sandwich structure of a drag-reducing layer, a toughening layer, and a mineralized layer; a hydrophobic layer and phospholipid molecules are introduced on the surface of the drag-reducing layer, giving the hydrogel excellent friction performance and long-term wear resistance. After 50,000 cycles, its friction coefficient is still 0.01, and no obvious wear is observed on the sliding surface; due to the synergistic cross-linking effect of multiple non-covalent bonds in the toughening layer, the compressive strength is as high as 71MPa; the mineralized layer contains hydroxyapatite nanoparticles, which have bone induction potential; in addition, the hydrogel has a high water content and excellent biocompatibility, with a water content of up to 65% and a cell viability of 100%.

[0009] A method for preparing a sandwich structure hydrogel cartilage substitute material, characterized by comprising the following steps:

[0010] S01. Preparation of Precursor Hydrogel

[0011] 1) Heat 10-20 parts of polyvinyl alcohol, 1-5 parts of chitosan, and 80-90 parts of deionized water at 90°C for 6 hours to obtain a uniform solution, then inject the resulting solution into a mold and allow it to stand at room temperature to remove bubbles. Place the solution in a -40°C to -20°C freezer for 8-12 hours, transfer it to room temperature for 2-6 hours, and perform 2-7 freeze-thaw cycles to obtain a polyvinyl alcohol-chitosan pregel.

[0012] 2) freeze-drying the pregel to remove water to obtain a polyvinyl alcohol-chitosan aerogel;

[0013] 3) The aerogel is immersed in an aqueous solution of a polymer material capable of forming ionic bonds with chitosan for 1 to 12 hours to produce a high-strength precursor hydrogel A with multiple non-covalent coordinated crosslinks, which constitutes the tough layer of the target sandwich structure hydrogel.

[0014] S02. Surface modification

[0015] The high-strength precursor hydrogel A obtained in step S01 is freeze-dried to obtain an aerogel, and then the top of the aerogel is immersed in a mixed solution of short-chain alkyl acyl chloride in dichloromethane and dimethyl sulfoxide; after reacting at room temperature for 2 to 24 hours, the residual acyl chloride is removed by washing with deionized water to obtain an aerogel with C8 to C12 hydrophobic alkyl chains on the surface;

[0016] The alkyl chain of acyl chloride has a hydrophobic surface, and when it is attached to the surface of the hydrogel, it greatly reduces the water absorption capacity of the gel, indicating the success of the hydrophobic modification.

[0017] S03. Loaded with lecithin

[0018] The hydrophobically modified side of the aerogel obtained in step S02 is immersed in a lecithin aqueous solution with a molar concentration of 1 to 20 mM for 12 to 24 hours, and then washed with deionized water to remove residual lecithin to obtain a lecithin-loaded hydrogel. The lecithin-loaded layer is the drag-reducing layer.

[0019] S04. Bottom mineralization

[0020] The non-hydrophobically modified side of the lecithin-loaded hydrogel obtained in step S03 is immersed in a K2HPO4 aqueous solution for 50 to 150 seconds, then immersed in a CaCl2 aqueous solution for 50 to 150 seconds, and the cycle is repeated 2 to 8 times. Finally, the hydrogel is incubated in situ at 37°C for 8 to 24 hours to form a mineralized layer, and the surface residual substances are washed with deionized water to obtain a hydrogel with a sandwich structure.

[0021] Furthermore, the time for standing at room temperature to remove bubbles in step S01 is 2-5 hours.

[0022] Furthermore, the mass fraction of the aqueous solution of the polymer material in step S01 is 0.1% to saturation; the polymer material is any one of hyaluronic acid, sodium hyaluronate, alginic acid, grape seed protein, carboxymethyl cellulose, and polyacrylic acid.

[0023] Furthermore, the acyl chloride in step S02 is one or more of octanoyl chloride, lauroyl chloride, stearyl chloride, and phenylacetyl chloride.

[0024] Furthermore, the volume ratio of the two solvents, dichloromethane and dimethyl sulfoxide, used to prepare the mixed solution in step S02 is 1:1 to 1:9; the molar ratio of the obtained aerogel to the acyl chloride is 1:1 to 1:10; and the density of the mixed solution is greater than the density of the pregel to ensure that the hydrophobic modification occurs only on the surface.

[0025] Furthermore, the lecithin in step S03 is a small molecule active agent containing cations, anions and hydrophobic chains, specifically one or both of phosphatidylcholine and hydrogenated soybean phosphatidylcholine.

[0026] Furthermore, in step S04, the molar concentration of the K2HPO4 aqueous solution is 0.1-0.6 mM, and the molar concentration of the CaCl2 aqueous solution is 0.1-0.8 mM.

[0027] The hydrogel obtained in this invention features a sandwich structure consisting of a drag-reducing layer, a toughening layer, and a mineralized layer. Based on the relationship between material structure and performance, the inventors utilized a biomimetic approach to prepare a sandwich-structured hydrogel. The resulting hydrogel exhibits ultra-low tribological properties, excellent compressive strength, and osteoinductive potential. Simultaneously achieving ultra-wear resistance, high strength, and osteoinductive potential has long been a technical challenge in the field of cartilage replacement materials. Inspired by the superlubricity of natural cartilage, the inventors incorporated zwitterionic phosphatidylcholine (lecithin) into the hydrogel. The highly hydrated phosphatidylcholine head group reduces surface friction on the hydrogel through a water-lubricating mechanism. To further achieve this friction-reducing effect, the inventors used acyl chloride to hydrophobically modify the hydrogel surface before loading the phosphatidylcholine (lecithin). Compared to direct lecithin loading, the hydrophobic treatment reduced the friction coefficient by more than twofold. Secondly, to ensure that the hydrogel can properly integrate with the underlying bone after introduction into the body, the inventors proposed mineralizing hydroxyapatite, a material with ultra-high osteoinductive potential, into the hydrogel. However, mineralization may reduce the hydrogel's water content and increase its surface roughness, resulting in an increase in the hydrogel's friction coefficient, which fails to meet the requirements of cartilage replacement materials (as shown in the data of Comparison 6 in Table 1). Based on this, the inventors' team designed a sandwich hydrogel. One side of the hydrogel is hydrophobically modified and loaded with lecithin to achieve ultra-low friction, while the other side is mineralized with hydroxyapatite to achieve osteoinductive properties. A strong and tough layer in the middle provides strength support. This allows the introduction of hydroxyapatite while maintaining its water content and avoiding negative impacts on the hydrogel's friction properties.

[0028] The sandwich structure hydrogel obtained by the present invention has a water content of up to 65%, a compressive strength of up to 71 MPa, and a friction coefficient of about 0.01. After 50,000 friction cycles, its friction coefficient remains unchanged, and no obvious wear marks are found on the worn surface, indicating good wear resistance.

[0029] Beneficial effects of the present invention:

[0030] Compared with the prior art, the present invention has the following outstanding substantive features and significant improvements.

[0031] (1) The inventors of this invention, based on the relationship between material structure and performance, used the biomimetic concept to prepare a sandwich-structured hydrogel. The resulting hydrogel has a sandwich structure consisting of a drag-reducing layer, a toughening layer, and a mineralized layer. The introduction of a hydrophobic layer and phospholipid molecules on the surface of the drag-reducing layer gives the hydrogel excellent friction properties and long-term wear resistance; the synergistic cross-linking effect of multiple non-covalent bonds in the toughening layer gives the resulting hydrogel high compressive strength; the mineralized layer contains hydroxyapatite nanoparticles, which gives the resulting hydrogel excellent osteoinductive potential.

[0032] (2) The sandwich-structured hydrogel obtained by the present invention has ultra-low tribological properties, excellent compressive strength and osteoinduction potential. The obtained sandwich-structured hydrogel has a water content of about 60%, a compressive strength of up to 71 MPa, a friction coefficient of about 0.01, and its friction coefficient remains unchanged after 50,000 friction cycles. The cell viability is as high as 100%, meeting the requirements for cartilage-substitute hydrogel materials. This provides strength and long-term durability for the material to be implanted in the body. The inventors summarized the performance parameters of the hydrogels obtained in Examples 16-20 of the present invention and compared them with the relevant literature data of existing cartilage-substitute hydrogel materials, as shown in Table 1:

[0033] .

[0034] (3) The preparation method of the sandwich structure hydrogel of the present invention is simple, and no toxic or harmful substances are used in the preparation process. The results of the cell viability test show that the sandwich structure hydrogel material obtained by the present invention has excellent biocompatibility, avoiding the adverse effects that may be caused to surrounding biological tissues after implantation into the organism.

[0035] (4) The sandwich structure hydrogel of the present invention has excellent osteoinductive potential, supports efficient osteogenesis, and provides a new idea for promoting biological cell growth and bone integration.

[0036] (5) The present invention not only provides a method for preparing a sandwich-structured hydrogel, but also provides a new method for expanding the preparation of hydrogel materials. In addition, during the preparation process, it also provides a new idea for integrating the contradictory properties of hydrogel materials, which is of great significance to the research, development and application of hydrogel materials in the field of artificial cartilage. BRIEF DESCRIPTION OF THE DRAWINGS

[0037] Figure 1 This is a schematic diagram of the synthesis of the sandwich structure hydrogel of the present invention;

[0038] Among them, 1 is the drag reduction layer; 2 is the toughening layer; and 3 is the mineralized layer. Implementation Method

[0039] The technical solution of the present invention will be described in detail below with reference to the accompanying drawings, but the implementation of the present invention is not limited thereto. Example 1

[0040] Preparation of precursor hydrogel

[0041] 10 parts polyvinyl alcohol and 1 part chitosan were weighed and added to 90 parts deionized water. The solution was dissolved at 90°C for 6 hours to form a homogeneous solution. The solution was then poured into a mold and allowed to stand for a period of time to remove air bubbles. The solution was then placed in a -20°C freezer for 12 hours and then thawed at room temperature for 2 hours. This cycle was repeated twice to synthesize a physically cross-linked polyvinyl alcohol-chitosan pregel. The polyvinyl alcohol-chitosan pregel was dehydrated in a freeze dryer and then immersed in a saturated sodium hyaluronate aqueous solution for 2 hours to obtain precursor hydrogel A1. Example 2

[0042] Preparation of precursor hydrogel

[0043] 20 parts polyvinyl alcohol and 5 parts chitosan were weighed and added to 80 parts deionized water. The solution was dissolved at 90°C for 6 hours to form a uniform solution. The solution was then poured into a mold and allowed to stand for a period of time to remove air bubbles. The solution was then placed in a -40°C freezer for 8 hours and then thawed at room temperature for 6 hours. This cycle was repeated three times to synthesize a physically cross-linked polyvinyl alcohol-chitosan pregel. The polyvinyl alcohol-chitosan pregel was dehydrated in a freeze dryer and then immersed in a 0.1% sodium alginate aqueous solution for 2 hours to obtain the precursor hydrogel A2. Example 3

[0044] Preparation of precursor hydrogel

[0045] 14 parts polyvinyl alcohol and 3 parts chitosan were weighed and added to 86 parts deionized water. The solution was dissolved at 90°C for 6 hours to form a uniform solution. The solution was then poured into a mold and allowed to stand for a period of time to remove air bubbles. The solution was then placed in a freezer at -30°C for 10 hours and then thawed at room temperature for 3 hours. This cycle was repeated four times to synthesize a physically cross-linked polyvinyl alcohol-chitosan pregel. The polyvinyl alcohol-chitosan pregel was dehydrated in a freeze dryer and then immersed in a 12% (mass fraction) carboxymethyl cellulose aqueous solution for 3 hours to obtain the precursor hydrogel A3. Example 4

[0046] Preparation of precursor hydrogel

[0047] 16 parts polyvinyl alcohol and 2 parts chitosan were weighed and added to 84 parts deionized water. The solution was dissolved at 90°C for 6 hours to form a uniform solution. The solution was then poured into a mold and allowed to stand for a period of time to remove air bubbles. The solution was then frozen in a freezer at -35°C for 9 hours and then thawed at room temperature for 4 hours. This cycle was repeated three times to synthesize a physically cross-linked polyvinyl alcohol-chitosan pregel. The polyvinyl alcohol-chitosan pregel was then dehydrated in a freeze dryer and immersed in a 50% (mass fraction) polyacrylic acid aqueous solution for 12 hours to obtain the precursor hydrogel A4.

[0048] The properties of the precursor hydrogel A4 obtained in Example 4 were tested. The test results showed that the precursor hydrogel obtained in Example 4 had a water content of 63%, a compressive strength of 51 MPa, a friction coefficient of 0.12, and a cell survival rate of 113% in biocompatibility testing. However, after 50,000 friction cycles, the worn surface showed obvious wear marks, a large amount of debris around the wear surface, and a polymer loss of over 10%. The friction performance was poor and the hydrogel lacked osteoinductive properties. Example 5

[0049] Preparation of precursor hydrogel

[0050] 18 parts of polyvinyl alcohol and 4 parts of chitosan were weighed by weight, added to 82 parts of deionized water, dissolved at 90°C for 6 hours to form a uniform solution, injected into a mold, and allowed to stand for a period of time to remove bubbles. Freeze in a freezer at -25°C for 11 hours, then transfer to room temperature to thaw for 5 hours, and repeat this cycle four times to synthesize a physically cross-linked polyvinyl alcohol-chitosan pregel. Grape seed protein was extracted using the extraction method disclosed in Chinese patent application CN201810129805.4. 30 parts of grape seed protein were weighed by weight and dissolved in 70 parts of deionized water to obtain a grape seed protein aqueous solution. The polyvinyl alcohol-chitosan pregel was dehydrated in a freeze dryer and then immersed in the grape seed protein aqueous solution for 4 hours to obtain the precursor hydrogel A5. Example 6

[0051] Preparation of aerogel A1-h with hydrophobic alkyl chains on its surface

[0052] First, the precursor hydrogel A1 obtained in Example 1 was completely dehydrated in a freeze dryer, and then its top was immersed in a mixed solution of dichloromethane and dimethyl sulfoxide containing lauroyl chloride (the volume ratio of the two solvents dichloromethane and dimethyl sulfoxide was 1:9), where the molar ratio of aerogel to acyl chloride was 1:1, and the reaction was carried out at room temperature for 2 hours. The alkyl chain of acyl chloride has a hydrophobic surface, and when it attaches to the surface of the hydrogel, it greatly reduces the water absorption capacity of the gel, indicating the success of the hydrophobic modification. The above mixed solution is characterized by a density greater than the density of the pre-gel (1.0 g / cm 3) to ensure that the hydrophobic modification occurs only on the surface. After the reaction, the hydrogel was washed 5-8 times with deionized water to remove residual acyl chloride, yielding aerogel A1-h with hydrophobic alkyl chains on its surface. Example 7

[0053] Preparation of aerogel A2-h containing hydrophobic alkyl chains on its surface

[0054] First, the precursor hydrogel A2 obtained in Example 2 was completely dehydrated in a freeze dryer. The top of the hydrogel was then immersed in a mixture of dichloromethane and dimethyl sulfoxide (DMSO) containing octanoyl chloride (the volume ratio of dichloromethane to DMSO was 1:1). The molar ratio of aerogel to acyl chloride was 1:4, and the reaction was allowed to proceed at room temperature for 4 hours. After the reaction, the hydrogel was washed 5-8 times with deionized water to remove residual acyl chloride, yielding aerogel A2-h with hydrophobic alkyl chains on its surface. Example 8

[0055] Preparation of aerogel A3-h containing hydrophobic alkyl chains on its surface

[0056] First, the cylindrical precursor hydrogel A3 obtained in Example 3 was completely dehydrated in a freeze dryer. The top of the hydrogel was then immersed in a mixture of dichloromethane and dimethyl sulfoxide (DMSO) containing a hard acyl chloride (the volume ratio of dichloromethane to DMSO was 1:6). The molar ratio of aerogel to acyl chloride was 1:10. The reaction was allowed to proceed at room temperature for 24 hours. After the reaction, the hydrogel was washed 5-8 times with deionized water to remove any residual acyl chloride, yielding aerogel A3-h with hydrophobic alkyl chains on its surface. Example 9

[0057] Preparation of aerogel A4-h containing hydrophobic alkyl chains on its surface

[0058] First, the cylindrical precursor hydrogel A4 obtained in Example 4 was completely dehydrated in a freeze dryer. The top of the hydrogel was then immersed in a mixture of dichloromethane and dimethyl sulfoxide (DMSO) containing phenylacetyl chloride (the volume ratio of dichloromethane to DMSO was 1:5). The molar ratio of aerogel to acyl chloride was 1:8, and the reaction was allowed to proceed at room temperature for 8 hours. After the reaction, the hydrogel was washed 5-8 times with deionized water to remove residual acyl chloride, yielding aerogel A4-h with hydrophobic alkyl chains on its surface. Example 10

[0059] Preparation of aerogel A5-h containing hydrophobic alkyl chains on its surface

[0060] First, the cylindrical precursor hydrogel A5 obtained in Example 5 was completely dehydrated in a freeze dryer. The top of the hydrogel was then immersed in a mixture of dichloromethane and dimethyl sulfoxide (DMSO) containing a hard acyl chloride (the volume ratio of dichloromethane to DMSO was 1:8). The molar ratio of aerogel to acyl chloride was 1:6, and the reaction was allowed to proceed at room temperature for 6 hours. After the reaction, the hydrogel was washed 5-8 times with deionized water to remove residual acyl chloride, yielding aerogel A5-h with hydrophobic alkyl chains on its surface. Example 11

[0061] Preparation of hydrogel containing drag reducing layer

[0062] The hydrophobically modified side of the aerogel A1-h obtained in Example 6 was immersed in a 1 mM phosphatidylcholine aqueous solution for 12 hours, and then washed with deionized water to remove residual reagents to obtain a lecithin-loaded hydrogel, i.e., a hydrogel containing a drag-reducing layer. Example 12

[0063] Preparation of hydrogel containing drag reducing layer

[0064] The hydrophobically modified side of the aerogel A2-h obtained in Example 7 was immersed in a 6 mM hydrogenated soybean phosphatidylcholine aqueous solution for 12 hours, and then washed with deionized water to remove the residual reagent to obtain a lecithin-loaded hydrogel, i.e., a hydrogel containing a drag-reducing layer. Example 13

[0065] Preparation of hydrogel containing drag reducing layer

[0066] The hydrophobically modified side of the aerogel A3-h obtained in Example 8 was immersed in a 20 mM phosphatidylcholine aqueous solution for 12 hours, and then washed with deionized water to remove residual reagents to obtain a lecithin-loaded hydrogel, i.e., a hydrogel containing a drag-reducing layer. Example 14

[0067] Preparation of hydrogel containing drag reducing layer

[0068] The hydrophobically modified side of the aerogel A4-h obtained in Example 9 was immersed in a 1 mM hydrogenated soybean phosphatidylcholine aqueous solution for 24 hours, and then washed with deionized water to remove the residual reagent to obtain a lecithin-loaded hydrogel, i.e., a hydrogel containing a drag-reducing layer. Example 15

[0069] Preparation of hydrogel containing drag reducing layer

[0070] The hydrophobically modified side of the aerogel A5-h obtained in Example 10 was immersed in a 3 mM phosphatidylcholine aqueous solution for 12 hours, and then washed with deionized water to remove residual reagents to obtain a lecithin-loaded hydrogel, i.e., a hydrogel containing a drag-reducing layer. Example 16

[0071] Preparation of hydrogels with sandwich structures

[0072] The non-hydrophobically modified side of the lecithin-loaded hydrogel obtained in Example 11 was immersed in a 0.1 mM K2HPO4 aqueous solution for 110 seconds, then immersed in a 0.2 mM CaCl2 aqueous solution for 110 seconds, and the cycle was repeated 8 times. The mixture was in situ mineralized at 37°C for 8 hours to form a mineralized layer containing hydroxyapatite. The surface residual material was removed by washing with deionized water to obtain a hydrogel with a sandwich structure. The synthesis diagram of the sandwich structure hydrogel obtained in Example 16 is shown in FIG. Figure 1 .

[0073] The properties of the sandwich-structured hydrogel obtained in Example 16 were tested. The test results showed that the sandwich-structured hydrogel had a water content of 60%, a compressive strength of 69 MPa, and a friction coefficient of 0.015. In biocompatibility testing, the cell survival rate reached 120%. After 50,000 friction cycles, the friction coefficient remained less than 0.017, and no obvious signs of wear were observed on the worn surface. This demonstrates that the sandwich-structured hydrogel has good wear resistance and osteoinductive properties. Example 17

[0074] Preparation of hydrogels with sandwich structures

[0075] The non-hydrophobically modified side of the lecithin-loaded hydrogel obtained in Example 12 was immersed in an aqueous solution with a molar concentration of 0.6 mM K2HPO4 for 50 seconds, and then immersed in a 0.5 mM CaCl2 aqueous solution for 50 seconds, and the cycle was repeated twice. The hydrogel was in situ mineralized at 37°C for 8 hours to form a mineralized layer containing hydroxyapatite. The surface residual substances were removed by washing with deionized water to obtain a hydrogel with a sandwich structure.

[0076] The properties of the sandwich-structured hydrogel obtained in Example 17 were tested. The test results showed that the sandwich-structured hydrogel obtained in Example 17 had a water content of 65%, a compressive strength of 71 MPa, a friction coefficient of 0.01, and a cell survival rate of 108% in biocompatibility testing. After 50,000 friction cycles, the friction coefficient remained less than 0.012, and no obvious signs of wear were observed on the worn surface. This demonstrates that the sandwich-structured hydrogel has good wear resistance and osteoinductive properties. Example 18

[0077] Preparation of hydrogels with sandwich structures

[0078] The non-hydrophobically modified side of the lecithin-loaded hydrogel obtained in Example 13 was immersed in an aqueous solution with a molar concentration of 0.3 mM K2HPO4 for 150 seconds, and then immersed in a 0.1 mM CaCl2 aqueous solution for 150 seconds, and the cycle was repeated 4 times. The product was mineralized in situ at 37°C for 8 hours to form a mineralized layer containing hydroxyapatite. The surface residual substances were removed by washing with deionized water to obtain a hydrogel with a sandwich structure.

[0079] The properties of the sandwich-structured hydrogel obtained in Example 18 were tested. The test results showed that the sandwich-structured hydrogel obtained in Example 18 had a water content of 61%, a compressive strength of 58 MPa, and a friction coefficient of 0.016. In biocompatibility testing, the cell survival rate reached 110%. After 50,000 friction cycles, the friction coefficient remained less than 0.018, and no obvious signs of wear were observed on the worn surface. This demonstrates that the sandwich-structured hydrogel has good wear resistance and osteoinductive properties. Example 19

[0080] Preparation of hydrogels with sandwich structures

[0081] The non-hydrophobically modified side of the lecithin-loaded hydrogel obtained in Example 14 was immersed in an aqueous solution with a molar concentration of 0.4 mM K2HPO4 for 80 seconds, and then immersed in a 0.5 mM CaCl2 aqueous solution for 80 seconds, and the cycle was repeated 4 times. The product was in situ mineralized at 37°C for 24 hours to form a mineralized layer containing hydroxyapatite. The surface residual substances were removed by washing with deionized water to obtain a hydrogel with a sandwich structure.

[0082] The properties of the sandwich-structured hydrogel obtained in Example 19 were tested. The test results showed that the sandwich-structured hydrogel obtained in Example 19 had a water content of 62%, a compressive strength of 66 MPa, and a friction coefficient of 0.014. In biocompatibility testing, the cell survival rate reached 115%. After 50,000 friction cycles, the friction coefficient remained less than 0.016, and no obvious signs of wear were observed on the worn surface. This demonstrates that the sandwich-structured hydrogel has good wear resistance and osteoinductive properties. Example 20

[0083] Preparation of hydrogels with sandwich structures

[0084] The non-hydrophobically modified side of the lecithin-loaded hydrogel obtained in Example 14 was immersed in an aqueous solution with a molar concentration of 0.5 mM K2HPO4 for 60 seconds, and then immersed in an aqueous solution of 0.8 mM CaCl2 for 60 seconds, and the cycle was repeated 5 times. The product was in situ mineralized at 37°C for 24 hours to form a mineralized layer containing hydroxyapatite. The surface residual substances were washed with deionized water to obtain a hydrogel with a sandwich structure.

[0085] The properties of the sandwich-structured hydrogel obtained in Example 20 were tested. The test results showed that the sandwich-structured hydrogel obtained in Example 20 had a water content of 65%, a compressive strength of 62 MPa, a friction coefficient of 0.018, and a cell survival rate of 119% in biocompatibility testing. After 50,000 friction cycles, the friction coefficient remained less than 0.02, and no obvious signs of wear were observed on the worn surface. This demonstrates that the sandwich-structured hydrogel has good wear resistance and osteoinductive properties.

[0086] In order to facilitate comparison of the properties of the sandwich structure hydrogel material obtained in the present invention with the properties of the pregel, the performance data of the pregel obtained in Example 4 and the performance data of the sandwich structure hydrogel materials obtained in Examples 16-20 are summarized in Table 2:

[0087] .

[0088] As can be seen from Table 2, after the pregel is prepared into a sandwich structure hydrogel in the present invention, the obtained sandwich structure hydrogel still has a high water content, greatly improved compressive strength, significantly reduced friction coefficient, excellent wear resistance, and excellent osteoinduction potential.

[0089] The inventors of the present invention employed this ingenious method to integrate the conflicting properties of hydrogel materials into a multi-layered sandwich hydrogel, thereby enhancing the strength of the hydrogel while maintaining its water content. This method imbues the material with osteoinductive potential while also imparting excellent tribological properties. Furthermore, the preparation method is simple, achieving the desired effect through a simple immersion process. The resulting hydrogel exhibits superior properties to the comprehensive performance of existing cartilage-substitute hydrogel materials (see Tables 1 and 2 for details), which is of great significance for expanding the application of hydrogels in the field of artificial cartilage.

[0090] The foregoing description is merely a preferred embodiment of the present invention and is not intended to limit the scope of the present invention. Any technical solution implemented within the scope of the present invention, or any solution that can be modified and varied by a person skilled in the art using the above-disclosed method, falls within the scope of protection of the present invention.

Claims

1. A method for preparing a sandwich structure hydrogel cartilage substitute material, characterized in that: The hydrogel has a sandwich structure of a drag-reducing layer, a tough layer, and a mineralized layer; a hydrophobic layer and phospholipid molecules are introduced into the surface of the drag-reducing layer; and multiple non-covalent bonds are cooperatively cross-linked in the tough layer; The mineralized layer contains hydroxyapatite nanoparticles; The preparation method comprises the following steps: S01. Preparation of Precursor Hydrogel 1) Heat 10-20 parts of polyvinyl alcohol, 1-5 parts of chitosan, and 80-90 parts of deionized water at 90°C for 6 hours to obtain a uniform solution, then inject the resulting solution into a mold and allow it to stand at room temperature to remove bubbles. Place the solution in a -40°C to -20°C freezer for 8-12 hours, transfer it to room temperature for 2-6 hours, and perform 2-7 freeze-thaw cycles to obtain a polyvinyl alcohol-chitosan pregel. 2) freeze-drying the pregel to remove water to obtain a polyvinyl alcohol-chitosan aerogel; 3) Soaking the aerogel in an aqueous solution of a polymer material capable of forming ionic bonds with chitosan for 1 to 12 hours to produce a high-strength precursor hydrogel A with multiple non-covalent cross-links, which constitutes the tough layer of the target sandwich structure hydrogel; S02. Surface modification The high-strength precursor hydrogel A obtained in step S01 is freeze-dried to obtain an aerogel, and then the top of the aerogel is immersed in a mixed solution of short-chain alkyl acyl chloride in dichloromethane and dimethyl sulfoxide; after reacting at room temperature for 2 to 24 hours, the residual acyl chloride is removed by washing with deionized water to obtain an aerogel with C8 to C12 hydrophobic alkyl chains on the surface; S03. Loaded with lecithin Soak the hydrophobically modified side of the aerogel obtained in step S02 in a lecithin aqueous solution with a molar concentration of 1 to 20 mM for 12 to 24 hours, then wash with deionized water to remove residual lecithin to obtain a lecithin-loaded hydrogel. This lecithin-loaded layer is the drag-reducing layer; S04. Bottom mineralization The non-hydrophobically modified side of the lecithin-loaded hydrogel obtained in step S03 is immersed in a K2HPO4 aqueous solution for 50 to 150 seconds, then immersed in a CaCl2 aqueous solution for 50 to 150 seconds, and the cycle is repeated 2 to 8 times. Finally, the hydrogel is incubated in situ at 37°C for 8 to 24 hours to form a mineralized layer, and the surface residual substances are washed with deionized water to obtain a hydrogel with a sandwich structure. The time for standing at room temperature to remove bubbles in step S01 is 2-6 hours; The mass fraction of the aqueous solution of the polymer material in step S01 is 0.1% to saturation; the polymer material is any one of hyaluronic acid, sodium hyaluronate, alginic acid, grape seed protein, carboxymethyl cellulose, and polyacrylic acid; The short-chain alkyl acyl chloride in step S02 is one or more of octanoyl chloride, lauroyl chloride, stearyl chloride, and phenylacetyl chloride; The hydrophobic alkyl chain in step S02 is a C8~C12 hydrophobic alkyl chain; The volume ratio of the two solvents, dichloromethane and dimethyl sulfoxide, used to prepare the mixed solution in step S02 is 1:1 to 1:9; the molar ratio of the obtained aerogel to the acyl chloride is 1:1 to 1:10; the density of the mixed solution is greater than the density of the pregel; The lecithin in step S03 is a small molecule active agent containing cations, anions and hydrophobic chains, specifically one or both of phosphatidylcholine and hydrogenated soybean phosphatidylcholine.

2. The method for preparing the sandwich structure hydrogel cartilage substitute material according to claim 1, characterized in that: The molar concentration of the K2HPO4 aqueous solution in step S04 is 0.1~0.6 mM, and the molar concentration of the CaCl2 aqueous solution is 0.1~0.8 mM.

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