3D printing of high-strength supramolecular hydrogel and its application in bone defect repair

Abstract

The application discloses a 3D printing high-strength supramolecular hydrogel and application thereof in bone defect repair. The high-strength supramolecular hydrogel is formed by hydrogen bond, ion interaction and hydrophobic association non-covalent cross-linking of a positively charged high-molecular polymer, a negatively charged functional monomer and a photoinitiator, and has a dynamic network structure. The hydrogel has a structure similar to an extracellular matrix, has self-healing, viscoelasticity and dynamics, and is beneficial to cell adhesion, proliferation, migration and differentiation. The hydrogel has superfast gelation, realizes second-level gelation solidification, and can accurately 3D print a microporous structure due to triple forces of strong hydrogen bond, electrostatic interaction and hydrophobic interaction. The hydrogel has good biodegradability, and the degradation rate can be designed by adjusting ingredients and cross-linking density, so that the degradation rate is matched with the speed of new bone formation, and finally, the scaffold material is completely replaced by new bone tissue.

Description

Technical Field

[0001] This invention relates to a 3D-printed high-strength supramolecular hydrogel and its application in bone defect repair, belonging to the field of hydrogel biomaterials technology. Background Technology

[0002] Bone tissue is the body's primary weight-bearing organ, undertaking vital functions for various life activities. Currently, the repair of large or critical-sized bone defects, especially those in weight-bearing areas, caused by traumatic fractures, degenerative diseases, and bone tumors remains a significant challenge in clinical treatment. However, commonly used bone defect repair materials, such as autologous bone, metal prostheses, bioceramics, and polyetheretherketone (PEEK), suffer from problems such as immune rejection, mechanical property mismatch, lack of bioactivity, and non-degradability. Furthermore, the stability and integration of bone repair materials with surrounding native bone tissue under tensile, compressive, shear, and torsional loads are crucial for bone function recovery, but currently used biomaterials often fail to simultaneously meet the requirements for repairing defects in weight-bearing bone areas.

[0003] Hydrogels, as a novel biomaterial, not only have a structure and function similar to the natural extracellular matrix (ECM), providing mechanical support and promoting the exchange of nutrients, oxygen and waste in cells, but also possess unique physicochemical properties such as good biocompatibility, degradability, viscoelasticity and adjustable mechanical properties.

[0004] Supramolecular hydrogels form dynamic network structures through reversible non-covalent interactions between macromolecules and small molecules (such as hydrogen bonds, electrostatic interactions, hydrophobic interactions, and host-guest interactions). They do not require chemical covalent cross-linking, thus preserving their biocompatibility. Furthermore, they effectively dissipate energy, thereby improving the strength and toughness of the hydrogel and exhibiting a structure and properties more closely resembling those of the extracellular matrix (ECM). In summary, supramolecular hydrogels can provide new design ideas for the development of biomimetic high-strength bone repair hydrogels inspired by ECM, laying a theoretical foundation for the application of high-strength hydrogel materials in bone tissue engineering, and possessing significant clinical value and translational prospects.

[0005] Although hydrogels are widely used in soft tissue engineering such as cartilage and skin, their inherent low mechanical strength (typically less than 1 MPa) and tendency to swell in aqueous environments severely limit their application in load-bearing bone repair. To improve mechanical properties, researchers have developed various strategies, such as dual-network structures and nanocomposite hydrogels.

[0006] Existing technology (CN202310335681.6) discloses a method for preparing hydrogel scaffolds using 3D printing technology, but the strength of the prepared hydrogels is less than 10 MPa, far below the requirements for human load-bearing bones (cortical bone can withstand compressive strengths of 100-200 MPa). Furthermore, existing high-strength hydrogels typically rely on chemical cross-linking; the cytotoxicity of cross-linking agents, as well as the mechanical property degradation and structural instability caused by hydrogel swelling, remain key bottlenecks hindering their clinical translation.

[0007] In summary, the existing technology (especially the existing technology CN202310335681.6) has the following main problems, which limit the application of hydrogels in bone defect repair, especially in weight-bearing areas:

[0008] Insufficient mechanical properties: Existing hydrogels generally have low compressive strength and modulus (usually <10MPa), which cannot provide the necessary mechanical support for load-bearing bone defects, and are prone to collapse and deformation after implantation.

[0009] High swelling rate: In an aqueous environment, traditional hydrogels will swell significantly, which will further reduce their mechanical properties and may compress surrounding tissues, triggering an inflammatory response.

[0010] Biocompatibility risks: Chemical cross-linking agents widely used to improve strength may remain and produce cytotoxicity, affecting the tissue repair process.

[0011] Lack of biomimetic structure and function: Many hydrogels lack the dynamic and adaptive properties of ECM, which is not conducive to cell migration, proliferation and differentiation, and has poor osteointegration ability.

[0012] In view of the shortcomings of the prior art, the present invention aims to provide a high-strength supramolecular hydrogel to achieve the repair of load-bearing bone tissue. Summary of the Invention

[0013] The purpose of this invention is to provide a high-strength supramolecular hydrogel to solve the problems of poor mechanical strength, high swelling rate in aqueous environment, and toxicity caused by crosslinking agents in existing bone hydrogels. The hydrogel of this invention has an extracellular matrix-like structure, possesses self-healing, viscoelasticity, and dynamic properties, which are beneficial to cell adhesion, proliferation, migration, and differentiation. The hydrogel of this invention has ultra-fast gelation properties, achieving gelation and curing in seconds. Thanks to its strong triple forces of hydrogen bonding, electrostatic interaction, and hydrophobic interaction, it can accurately 3D print microporous structures.

[0014] This invention utilizes a supramolecular polymerization strategy to develop a novel high-strength supramolecular hydrogel that mimics ECM in terms of material composition, structural design, and biological function, thereby achieving the repair of load-bearing bone tissue. By inducing molecular self-assembly through triple non-covalent interactions (hydrogen bonds, ionic bonds, and hydrophobic interactions) and coupling inorganic nanomaterials through electrostatic interactions, a microphase-separated hydrogel with a nanodomain structure is obtained.

[0015] The present invention provides a high-strength supramolecular hydrogel, which is formed by positively charged polymers, negatively charged functional monomers and photoinitiators through hydrogen bonding, ionic interactions and hydrophobic association non-covalent cross-linking to form a dynamic network structure.

[0016] The high-strength supramolecular hydrogel of the present invention also includes bioactive nanoparticles, wherein the bioactive nanoparticles are selected from at least one of nano-clay, hydroxyapatite, calcium phosphate and silica.

[0017] The hydrogel of this invention has ultra-high mechanical properties: through the triple non-covalent synergistic effect of hydrogen bonding, ion interaction and hydrophobic association, and combined with bioactive nanoparticles (such as nano-clay), a stable network structure with microphase separation is constructed, so that the compressive modulus of the hydrogel is close to 200 MPa, which far exceeds the existing reported hydrogel materials and can match the mechanical requirements of load-bearing bones.

[0018] The hydrogel of this invention has excellent anti-swelling properties: the dense physical cross-linked network effectively inhibits excessive swelling in the aqueous environment, maintaining the long-term stability of implant size and mechanical properties.

[0019] The hydrogel of this invention exhibits excellent biocompatibility and functionality: it employs a primarily physical cross-linking method, avoiding the toxicity issues associated with chemical cross-linking agents. The hydrogel possesses self-healing, shear-thinning, and rapid photocuring properties, facilitating injection and enabling the fabrication of porous scaffolds with precise microstructures via 3D printing, providing channels for cell ingrowth and nutrient delivery. Furthermore, the bioactive nanoparticles (such as nanoclay and hydroxyapatite) in its composition provide the necessary ionic environment for bone repair, promoting osteogenic formation.

[0020] The dynamic reversible properties of the hydrogel in this invention mimic the natural ECM, which is beneficial for cell behavior regulation. Combined with 3D printing technology, it can achieve personalized customization and precisely repair bone defects of different shapes.

[0021] Preferably, the positively charged polymer is selected from at least one of polyethyleneimine, hyperbranched polyethyleneimine, polylysine, chitosan, polymethacrylic acid, N,N-dimethylaminoethyl ester, polyethyleneimine, and polypropyleneimine.

[0022] The molecular weight of the positively charged polymer can be 30,000-100,000, preferably 40,000-80,000, and more preferably 50,000-70,000.

[0023] In this invention, the positively charged polymer is the core framework and functional center constituting the high-strength supramolecular hydrogel network:

[0024] The positively charged polymer, acting as a cationic polyelectrolyte, provides strong electrostatic interactions (ionic bonds). Its molecular chains are rich in amino groups (-NH2 or -NH-), which protonate in solution, imparting a strong positive charge to the entire molecular chain. These positively charged groups exhibit strong electrostatic attraction with negatively charged functional monomers and negatively charged bioactive nanoparticles, forming dense ionic cross-links. Ionic bonds are strong non-covalent bonds, capable of forming very robust cross-linking points, which explains the hydrogel's near-200 MPa ultra-high compressive modulus. This electrostatic interaction occurs rapidly, allowing the physical cross-linking network to form instantaneously during photo-initiated free radical polymerization of the prepolymer, achieving "second-level" curing and meeting the process requirements of 3D printing.

[0025] The positively charged polymer acts as a hydrogen bond donor, constructing a multi-layered physical cross-linking network. The amine (-NH2) and imine (-NH-) groups on the positively charged polymer chains are excellent hydrogen bond donors, forming extensive hydrogen bond networks with carboxyl groups (-COOH) on negatively charged functional monomers, hydroxyl groups (-OH) on the surface of bioactive nanoparticles, and water molecules. Hydrogen bonds are dynamically reversible; under external force, they break before covalent bonds to dissipate energy, significantly improving the toughness and fatigue resistance of the hydrogel and preventing brittle fracture. Broken hydrogen bonds can reform after contact at the fracture surface, giving the hydrogel self-healing capabilities and extending the material's lifespan.

[0026] The positively charged polymer serves as the macromolecular backbone, forming a physically entangled and microphase-separated structure. This positively charged polymer has a high molecular weight (30,000-100,000), and its long molecular chains can physically entangle with each other. When combined with kinetic monomers of opposite charge, the mutual attraction between the positive and negative charges leads to the co-assembly of the molecular chains, forming localized hydrophobic microregions, which in turn induces the microphase-separated structure. This physical entanglement and microphase-separated structure further enhances the physical cross-linking density of the network, making the hydrogel network more stable and effectively inhibiting excessive swelling in an aqueous environment (anti-swelling property). This microphase-separated structure is similar to the multi-layered structure of natural biological tissues (such as cartilage and tendons), helping to mimic the mechanical microenvironment of the extracellular matrix.

[0027] The positively charged polymer, as a functional component, improves biocompatibility and bioactivity. While providing mechanical support, it can also prevent infection and promote tissue regeneration, achieving "functionalized" repair rather than simply "filling" defects.

[0028] Preferably, the negatively charged functional monomer is selected from at least one of acrylic acid, methacrylic acid, 2-acrylamide-2-methylpropanesulfonic acid, methacrylamide hyaluronic acid, itaconic acid, vinyl phosphate, hydroxyethyl phosphate methacrylate, and 10-methacryloyloxydecyl dihydrophosphate.

[0029] Preferably, the molar ratio of the positively charged polymer to the negatively charged functional monomer is 1:(0.6-2), and the positively charged polymer is expressed in terms of the molar amount of structural units.

[0030] In this invention, the negatively charged functional monomer is another core component constituting the high-strength supramolecular hydrogel network. Its function complements that of the positively charged polymer, jointly constructing a unique network structure that combines high strength and dynamism.

[0031] The negatively charged functional monomer, acting as an anionic polyelectrolyte, forms strong ionic crosslinks with the positively charged polymer. The negatively charged functional monomer contains a large number of carboxyl groups (-COOH) in its molecular structure. In an aqueous solution, these carboxyl groups ionize to generate negatively charged carboxylate ions (-COO₂). - These negatively charged groups interact with the protonated amino groups (-NH3) on the positively charged first monomer (such as polyethyleneimine-PEI) molecular chain. + Strong electrostatic interactions occur, forming ionic bonds. This is the most important and strongest physical cross-linking mechanism in the entire hydrogel network. Numerous and robust ionic bond cross-linking points are the fundamental contributors to the hydrogel's ultra-high compressive strength and modulus (approaching 200 MPa). The attraction between positive and negative charges leads to phase separation of the molecular chains, forming dense ionic cluster microregions, which further enhances the network's rigidity and stability and effectively inhibits swelling.

[0032] The negatively charged functional monomers provide photopolymerizable groups, constructing a stable covalent network framework. These negatively charged functional monomers are all vinyl monomers containing carbon-carbon double bonds. Under photoinitiator and ultraviolet light irradiation, these double bonds undergo free radical copolymerization, forming a polymer backbone linked by covalent bonds. The network formed by covalent polymerization provides a permanent and stable framework for the entire hydrogel, preventing the physical cross-linking network from dissociating and being lost under long-term use or extreme conditions, ensuring the structural integrity and durability of the material. The covalent network "locks" the positively charged polymer and the negatively charged functional monomers together, allowing the physical cross-linking, such as the ionic bonds formed between them, to function stably and persistently.

[0033] The negatively charged functional monomers act as hydrogen bond acceptors, participating in the construction of multiple dynamic physical cross-links. The carboxyl / carboxylate groups on these negatively charged functional monomers are not only sites for ionic bonding but also extremely strong hydrogen bond acceptors. They can form extensive hydrogen bonds with the amine groups (-NH-) on the first monomer, water molecules, and groups on the surface of inorganic nanomaterials. The dynamic breaking and reforming of hydrogen bonds effectively dissipates the mechanical energy applied to the material, greatly improving the hydrogel's toughness, fatigue resistance, and damage tolerance, making it less prone to brittle fracture. The broken hydrogen bonds can rapidly recover after re-contact at the fracture surface, which is one of the important mechanisms for the material's self-healing function.

[0034] The negatively charged functional monomers modulate hydrophobic interactions, stabilizing the microphase separation structure. When combined with the positively charged polymer via ionic bonds, ion pairs are formed, decreasing the polarity of the region and enhancing hydrophobicity, thereby driving hydrophobic association. Hydrophobic interactions, acting as an additional physical cross-linking force, further solidify the microphase separation structure induced by ionic bonds, making the network denser and more stable. The presence of hydrophobic microregions effectively repels water molecules, which is one of the key reasons why the hydrogel exhibits a low swelling rate.

[0035] Preferably, the bioactive nanoparticles are selected from at least one of nano-clay, hydroxyapatite, calcium phosphate, and silica;

[0036] The amount of the bioactive nanoparticles used is 0.5-5% of the total mass of the raw materials.

[0037] In this invention, the bioactive nanoparticles serve as multifunctional crosslinking points and active components, forming a tight hybrid structure with the organic polymer network, which plays a crucial role in enhancing the overall performance of the hydrogel.

[0038] The bioactive nanoparticles, acting as multifunctional physical crosslinking points, significantly enhance mechanical properties. These bioactive nanoparticles typically possess a layered structure with numerous negative charges (such as -OH and -O-) on their surfaces. These negatively charged surfaces can bind to positively charged polymers (such as PEI) through strong electrostatic interactions. Simultaneously, hydroxyl groups and other functional groups on their surfaces can form hydrogen bonds with functional groups on the polymer chains. A single nanoclay layer can simultaneously bind to multiple polymer chains, serving as a highly efficient multifunctional crosslinking point and greatly increasing the crosslinking density of the network. This "one-to-many chain" crosslinking mode effectively disperses stress across the entire inorganic nanosheet, rather than concentrating it at a single crosslinking point, thereby significantly improving the compressive strength, modulus, and toughness of the hydrogel. This is one of the key synergistic factors in achieving an ultra-high modulus of 200 MPa. The rigid nanosheets can physically hinder the relative slippage of polymer chains under external forces, further strengthening the network.

[0039] The bioactive nanoparticles serve as a nano-reinforcing phase, constructing an organic-inorganic hybrid network. These bioactive nanoparticles are uniformly embedded in the polymer three-dimensional network as a nanoscale dispersed phase, forming a unique organic-inorganic hybrid structure. This structure mimics the composition (organic collagen + inorganic nano-apatite crystals) and micro-nano hierarchical structure of natural bone tissue. The combination of the rigid inorganic nanophase and the flexible organic polymer exhibits a synergistic reinforcing effect, enabling the material to possess both high strength and a certain degree of toughness, with mechanical properties far superior to pure polymer hydrogels.

[0040] The bioactive nanoparticles endow the hydrogel with excellent anti-swelling properties. The uniformly dispersed nano-clay sheets act as a physical barrier within the hydrogel network, extending the penetration path of water molecules and effectively delaying water absorption. The extremely high cross-linking density results in a greater elastic recoil force on the polymer network during swelling, strengthening its resistance to network expansion. The hydrogel of this invention maintains dimensional stability and structural integrity in bodily fluid environments, preventing a decline in mechanical properties and pressure on surrounding tissues due to excessive swelling, which is crucial for long-term implantation.

[0041] The bioactive nanoparticles improve rheological properties, making them suitable for 3D printing. The addition of nanoclay significantly alters the rheological properties of the prepolymer. Due to the interaction between its layered structure and polymer chains, the prepolymer typically exhibits significant shear-thinning behavior (viscosity decreases under external force). Under the shear force of the 3D printing nozzle, the viscosity of the prepolymer rapidly decreases, exhibiting good flowability and facilitating extrusion. After extrusion, the shear force disappears, and the viscosity recovers instantly, effectively maintaining the extruded shape, preventing structural collapse, and ensuring printing accuracy. This makes this high-strength hydrogel prepolymer suitable for surface projection photopolymerization 3D printing, enabling the fabrication of personalized bone scaffolds with complex microstructures (such as porous interconnected structures), which is crucial for nutrient delivery and bone tissue ingrowth.

[0042] The bioactive nanoparticles provide bioactivity and promote osteogenic repair.

[0043] Preferably, the photoinitiator is selected from at least one of α-ketoglutaric acid, 2,2-diethoxyacetophenone, and lithium phenyl (2,4,6-trimethylbenzoyl)phosphate;

[0044] The amount of the photoinitiator can be 0.5-1% of the mass of the negatively charged functional monomer.

[0045] The supramolecular hydrogel of this invention has a water content of 50-70%, a compression modulus of 50-200 MPa, a swelling rate of ≤5%, and self-healing properties.

[0046] The present invention also provides a method for preparing the supramolecular hydrogel, comprising the following steps:

[0047] S1 includes the following steps S1-1 or S1-2:

[0048] S1-1. The positively charged polymer, the negatively charged functional monomer, and the photoinitiator are mixed in a solvent to form a prepolymer solution;

[0049] S1-2. The polymer matrix, the functional monomer, the bioactive nanoparticles, and the photoinitiator are mixed in a solvent to form a prepolymer solution;

[0050] S2. The prepolymer solution is subjected to deoxygenation and degassing treatment;

[0051] S3. The prepolymer liquid undergoes a polymerization reaction under ultraviolet light irradiation to obtain the high-strength supramolecular hydrogel.

[0052] Preferably, the solvent includes physiological saline or deionized water, and the mass ratio of the solvent to the positively charged polymer is 1:(6-10).

[0053] The positively charged polymer is added in the form of an aqueous solution with a mass concentration of 10-50%.

[0054] Preferably, the wavelength of the ultraviolet light is 350-405 nm, and the light intensity is 50-200 mW / cm². 2 ;

[0055] The polymerization reaction takes 10-30 minutes.

[0056] The high-strength supramolecular hydrogel provided by this invention is suitable for preparing bone defect repair materials, and shows great potential, especially in the repair of bone defects in load-bearing parts.

[0057] The high-strength supramolecular hydrogel serves as an implantable bone repair scaffold. It can be precisely fabricated into a personalized implantable scaffold with a specific shape and internal porous structure using surface projection curing 3D printing technology. The porous structure has a porosity of 70–80%, a main pore diameter of 200–500 μm, and connecting pores ≥100 μm.

[0058] Traditional hydrogel scaffolds have low strength (<10MPa) and are prone to collapse and deformation after implantation in weight-bearing areas (such as long bones of the limbs and jawbone), leading to surgical failure. The hydrogel of this invention has a compressive modulus of up to 200MPa, which can effectively withstand physiological loads, provide temporary mechanical support for bone defects, maintain the defect space, and create a stable mechanical microenvironment.

[0059] Based on the patient's CT data, a scaffold that closely matches the shape of the bone defect can be 3D printed, enabling personalized repair and improving surgical precision and repair outcomes.

[0060] The interconnected porous structure constructed by 3D printing facilitates the migration, spread, and proliferation of osteoblasts and vascular endothelial cells, while also facilitating the transport of nutrients and metabolic waste, accelerating new bone formation and vascularization, and achieving bone integration.

[0061] The supramolecular hydrogel of this invention has good biodegradability. Its degradation rate can be designed by adjusting the composition and crosslinking density to match the rate of new bone formation, ultimately achieving complete replacement of the scaffold material by new bone tissue.

[0062] This invention relates to a supramolecular hydrogel as a minimally invasive injectable filler for bone defects. Utilizing the shear-thinning and rapid photocuring properties of the hydrogel prepolymer, it can be used as a minimally invasive injectable filler.

[0063] For irregular or difficult-to-access bone defects, the prepolymer solution can be directly injected into the defect cavity using a syringe, greatly reducing surgical trauma. After injection, it can be cured in situ within tens of seconds by using an endoscope-mounted fiber optic cable or percutaneous irradiation with 405nm visible light, perfectly filling defects of any shape and avoiding the mismatch between traditional prefabricated scaffolds and defect morphology. The rapid curing characteristic effectively prevents material migration within the body, ensuring the accuracy of the repair placement. Attached Figure Description

[0064] Figure 1 This refers to the solubility of methacrylic acid in aqueous and pure water of polyethyleneimine and the effect after polymerization.

[0065] Figure 2 These are the XPS elemental scanning results of the hydrogel prepared in Example 1 of this invention.

[0066] Figure 3 The results are the compressive strength test results of the hydrogel prepared in Example 1 of this invention.

[0067] Figure 4 The swelling test results are for the hydrogel prepared in Example 1 of this invention.

[0068] Figure 5 These are the rheological behavior test results of the hydrogel prepared in Example 1 of this invention.

[0069] Figure 6 This invention relates to a porous scaffold with different dimensions produced by hydrogel 3D printing.

[0070] Figure 7 This describes the repair effect of the hydrogel prepared in Example 1 of the present invention on femoral bone defects in rabbits.

[0071] Figure 8 The results and illustrations show the mechanical tensile test results prepared in Example 6 of this invention.

[0072] Figure 9 This is a compressed photograph of the hydrogel sample prepared in Comparative Example 1 of this invention.

[0073] Figure 10 This is a confocal photograph of comparative example 2 of the present invention regarding cell biocompatibility. Detailed Implementation

[0074] Unless otherwise specified, the experimental methods used in the following examples are conventional methods.

[0075] Unless otherwise specified, all materials and reagents used in the following examples are commercially available.

[0076] This invention provides a supramolecular hydrogel that combines ultra-high mechanical strength, excellent anti-swelling properties, rapid photocuring ability, self-healing characteristics, and good bioactivity, which can be used for bone defect repair, especially for the repair of load-bearing bones.

[0077] The invention is characterized by mimicking the "organic-inorganic" hybrid structure and dynamic characteristics of the extracellular matrix of natural bone tissue. Through a "one-step" preparation process, multiple non-covalent synergistic effects are combined with photo-initiated covalent polymerization to construct a high-performance hybrid supramolecular hydrogel system.

[0078] The present invention uses the following raw materials:

[0079] Positively charged polymers: High molecular weight, amino-rich cationic polyelectrolytes (such as polyethyleneimine PEI) are selected as the positively charged core and physical cross-linking points of the network.

[0080] Negatively charged functional monomers: Select anionic vinyl monomers containing carboxyl groups and double bonds (such as methacrylic acid MAA), which serve as both a source of negative charge and a site for photopolymerization.

[0081] Bioactive nanoparticles: Layered nanoclay or nanohydroxyapatite are introduced as multifunctional nano-crosslinking agents and bioactive components.

[0082] The present invention employs the following crosslinking mechanism:

[0083] The hydrogel of this invention possesses a synergistic triple dynamic physical crosslinking and covalent network:

[0084] Strong ionic bonds: The electrostatic interaction between the positively charged amino groups of the polymer matrix and the negatively charged carboxyl groups of the functional monomers constitutes the source of network strength.

[0085] Hydrogen bonding and hydrophobic interactions: Extensive hydrogen bonding and hydrophobic association between the components effectively dissipate energy, giving the material toughness and self-healing properties.

[0086] Covalent bonds: The covalent network formed by the polymerization of functional monomers induced by ultraviolet light provides a permanent framework and structural stability for the entire system.

[0087] Nano-enhancing effect: Bioactive nanoparticles, acting as rigid nano-crosslinking points, greatly improve crosslinking density and mechanical properties, and significantly inhibit swelling.

[0088] Through the aforementioned materials and crosslinking agent mechanism, this invention solves the bottleneck problem that traditional hydrogels cannot be used in load-bearing components, specifically as follows:

[0089] Breakthrough in mechanical properties: The compressive modulus is close to 200MPa, and the strength is comparable to that of cortical bone, which can meet the load-bearing requirements.

[0090] High structural stability: Excellent resistance to swelling, enabling it to maintain its shape and mechanical integrity over a long period of time in a body fluid environment.

[0091] Excellent processing performance: It has the ability to cure quickly, self-heal and 3D printable, making it easy to operate and apply in clinical settings.

[0092] Active biological function: The components are biomimetic, have good biocompatibility, and can promote osteoblast differentiation and guide new bone formation, actively participating in the repair process.

[0093] Example 1: Preparation of high-strength hydrogel

[0094] 5g of methacrylic acid and 10g of polyethyleneimine aqueous solution (20wt%, molecular weight of polyethyleneimine is 70000) were mixed evenly. Then, 0.1g of nanoclay (XLS) was evenly dispersed in 4g of physiological saline and added to the above mixture. Finally, 50μL of photoinitiator DEAP was added. The mixture was ultrasonically sheared and vibrated to remove air bubbles. The mixture was then dropped into a PTFE mold and irradiated under 365nm ultraviolet light (power is 150W) for 10 minutes to obtain a hydrogel sample. The sample was soaked in physiological saline for 48h to remove unpolymerized monomers and obtain a high-strength hydrogel material with nanoclay and a water content of about 65%.

[0095] In this embodiment, the image of the mixture of methacrylic acid and polyethyleneimine aqueous solution is shown below. Figure 1 As shown, methacrylic acid was simultaneously mixed with pure water. The solubility of methacrylic acid in polyethyleneimine aqueous solution and pure water, and the effect after polymerization, are as follows. Figure 1 As shown, methacrylic acid separates into layers in pure water, but is clear and transparent in the aqueous solution of polyethyleneimine, and turns milky white after polymerization. This indicates that there is a strong ionic interaction between methacrylic acid and polyethyleneimine, resulting in obvious phase separation domains after polymerization.

[0096] The XPS elemental scan results of the hydrogel in this embodiment are as follows: Figure 2 As shown, the hydrogel contains phosphorus, silicon, and magnesium, providing favorable conditions for bone repair.

[0097] The rheological behavior of the hydrogel in this embodiment was tested as follows:

[0098] Photopolymerization: Oscillatory rheological tests were performed using an MR302 rheometer to characterize the photogelation process of the prepolymer. 5 mL of the hydrogel prepolymer was added to a sterile petri dish. An appropriate amount of sample was loaded onto the parallel plate of the rheometer (a quartz transparent plate was used to ensure UV transmittance). Time scanning was then performed under conditions of γ = 1% (within the linear viscoelastic region) and f = 1 Hz, continuously recording the storage modulus G' and loss modulus G'". After acquiring data for 10 seconds, UV irradiation was turned on, and monitoring continued until the curve stabilized. The gelation transition time was defined as the intersection of G' and G'' (G' = G'), tanδ = 1.

[0099] Self-healing: Perform step strain cycling: γ_low = 1% (120s) and γ_high = 200% (100s), f = 1Hz, cycle 3-5 times, results are as follows. Figure 5 As shown, it can be seen that in the initial stage of photocuring, the loss modulus (G”) > the storage modulus (G’), exhibiting a liquid state. After 10s of ultraviolet light irradiation, G’ = G”, and then G’ continues to be greater than G”, exhibiting solid characteristics. This indicates that the hydrogel prepolymer of the present invention has rapid photocuring properties. Furthermore, the hydrogel modulus can still recover under low strain and high strain cycles, indicating that the hydrogel has self-healing properties under physical dynamic crosslinking.

[0100] Example 2

[0101] Mix 5g of methacrylic acid and 10g (20wt%) of polyethyleneimine aqueous solution evenly, then add 4g of physiological saline, and finally add 50μL of photoinitiator DEAP. Mix evenly by ultrasonic shearing and vibration to remove air bubbles, and drop it into a PTFE mold. Irradiate under 365nm ultraviolet light (power 150W) for 10 minutes to obtain a hydrogel sample. Soak in physiological saline for 48h to remove unpolymerized monomers to obtain a high-strength hydrogel material without nanoclay.

[0102] The compressive strength of the hydrogels prepared in Examples 1 and 2 was tested.

[0103] The hydrogel was prepared into cylindrical specimens (10 mm in diameter, 5 mm in height, wet state) and equilibrated in physiological saline for three days. Testing was performed using a universal testing machine with a loading speed of 20 mm / min. -1 When the strain is increased to 80% or the specimen becomes unstable / failed, the compressive modulus E is taken as the slope of the linear fitting within the strain range of 2-10%.

[0104] The results are as follows Figure 3As shown, the high-strength hydrogel prepared in Example 1 has a compressive modulus close to 200 MPa and a compressive strength of 120 MPa. Compared with the hydrogel in Example 2 without the addition of inorganic nano-clay, both the compressive modulus and strength have increased, indicating that the coupling and cross-linking of inorganic nanosheets further improves the mechanical properties of the hydrogel.

[0105] The swelling properties of the hydrogels in Examples 1 and 2 were tested. Cylindrical samples (10 mm in diameter, 5 mm in height) were prepared from the hydrogels and placed in physiological saline. The samples were weighed daily for two weeks. The swelling ratio (SR) was calculated using the following formula:

[0106]

[0107] Where W0 is the initial weight of the hydrogel, W t It represents the weight at each point in time.

[0108] The results are as follows Figure 4 As shown, the addition of inorganic nano-clay XLS improves the hydrophilicity of the material and slightly increases the swelling rate, but the swelling rate after equilibrium is still less than 3%.

[0109] Example 3: 3D printing of porous scaffolds that match bone tissue structure

[0110] 5g of methacrylic acid and 10g (20wt%) of polyethyleneimine aqueous solution were mixed evenly, and then 4g of physiological saline containing 0.1g of XLS nanoclay was added. Finally, 40μL of photoinitiator LAP was added. The mixture was ultrasonically sheared and vibrated to remove air bubbles. The mixture was then dropped into a surface projection photopolymerization 3D printer and irradiated under 405nm ultraviolet light to print a porous scaffold. The obtained high-strength hydrogel sample was cleaned with alcohol to remove residual polymer in the porous structure. Finally, it was soaked in physiological saline for 48h to remove unpolymerized monomers to obtain a high-strength porous hydrogel scaffold with a structure matching bone defects.

[0111] like Figure 6 As shown, the hydrogel of the present invention can be 3D printed into porous scaffolds with different sizes, having a porosity of 70-80%, a main pore diameter of 200-500 μm, and connecting pores ≥100 μm.

[0112] Example 4: High-strength hydrogel scaffold for repairing femoral defects in rabbits

[0113] With ethical approval, preoperative CT scans of the femur were performed on New Zealand rabbits under anesthesia. DICOM was imported into Mimics / 3DSlicer for segmentation and three-dimensional reconstruction to obtain models of the femur and defect area. Individualized porous hydrogel scaffold models were then established based on the defect geometry.

[0114] Under sterile conditions, a photopolymerization 3D printer was used to complete the precise in vitro printing process. The surgery created a standardized segmental defect through a lateral femoral approach, and a hydrogel scaffold (Example 3) was placed and precisely fitted. Internal fixation was performed to ensure axial stability. The incision was routinely irrigated and closed, and antibiotics and analgesia were administered. The patient was fed for 6 months postoperatively and a follow-up CT scan was performed.

[0115] The results are as follows Figure 7 As shown in the figure, compared with the control group, implantation of the high-strength porous hydrogel scaffold provided by the present invention significantly promoted the repair of bone defects.

[0116] Example 5: Modifying the functional monomer

[0117] The difference from Example 1 is that methacrylic acid is replaced with an equimolar amount of acrylic acid.

[0118] like Figure 8 As shown, after replacing methacrylic acid with acrylic acid, the system changes from a high-strength, high-modulus hydrogel system to a low-strength, low-modulus hydrogel system with high flexibility. The tensile elongation at break exceeds 700%. The stress-strain curve was obtained by performing a single tensile test at a speed of 20 mm / min using a universal testing machine, which shows that the hydrogel system designed in this invention has a wide range of adjustable mechanical properties.

[0119] Comparative Example 1: Replacing the polymer matrix and methacrylic acid monomer

[0120] The difference between this comparative example and Example 1 is that polyethyleneimine is replaced with polyvinyl alcohol, and methacrylic acid monomer is replaced with methacrylamide gelatin. Specifically, 5g of polyvinyl alcohol (2899) is dissolved in 45g of hot water, heated and mechanically stirred for 2 hours, and then 1g of methacrylamide gelatin and 10μL of photoinitiator are added. After mixing evenly and removing air bubbles, the mixture is poured into a mold and frozen-thawed three times at -20°C. Finally, photocrosslinking is performed to synthesize a double-network hydrogel. This gel cannot form ionic bonds and is crosslinked only through hydrogen bonds.

[0121] like Figure 9 As shown, the high-strength hydrogel prepared by polyethyleneimine, methacrylic acid and nano-clay in Example 1 can withstand the weight of an adult without deformation or damage. The double-network hydrogel prepared by uncharged polyvinyl alcohol and methacrylamide gelatin exhibits low modulus and softness, which once again proves the important influence of strong ionic interactions on mechanical properties proposed in this invention.

[0122] Comparative Example 2: Using a chemical cross-linking agent

[0123] The difference between this comparative example and Example 1 is that, while retaining all components, 1 wt% of N,N'-methylenebisacrylamide (MBA) was added as a chemical crosslinking agent.

[0124] Depend on Figure 10 The confocal image of cell biocompatibility shown indicates that the addition of chemical cross-linking agents reduces biocompatibility.

Claims

1. A high-strength supramolecular hydrogel, comprising a dynamic network structure formed by positively charged polymers, negatively charged functional monomers, and photoinitiators through non-covalent cross-linking via hydrogen bonding, ionic interactions, and hydrophobic association; No bifunctional or multifunctional chemical crosslinking agents are added during the preparation of the high-strength supramolecular hydrogel, and the crosslinking network of the high-strength supramolecular hydrogel does not contain interchain covalent crosslinking points formed by bifunctional or multifunctional chemical crosslinking agents. The positively charged polymer is polyethyleneimine with a molecular weight of 30,000-100,000. The negatively charged functional monomer is methacrylic acid; The high-strength supramolecular hydrogel also includes bioactive nanoparticles; The bioactive nanoparticles are selected from at least one of nano-clay, hydroxyapatite, calcium phosphate, and silicon dioxide. The amount of the bioactive nanoparticles used is 0.5-5% of the total mass of the raw materials.

2. The supramolecular hydrogel according to claim 1, characterized in that: The photoinitiator is selected from at least one of α-ketoglutaric acid, 2,2-diethoxyacetophenone, and lithium phenyl (2,4,6-trimethylbenzoyl)phosphate; The amount of photoinitiator used is 0.5-1% of the mass of the negatively charged functional monomer.

3. The supramolecular hydrogel according to claim 1 or 2, characterized in that: The supramolecular hydrogel has a water content of 50-70%, a compressive modulus of 50-200 MPa, a swelling rate of ≤5%, and self-healing properties. The molar ratio of the positively charged polymer to the negatively charged functional monomer is 1:(0.6-2), and the positively charged polymer is expressed in terms of the molar amount of structural units.

4. A method for preparing the high-strength supramolecular hydrogel according to any one of claims 1-3, comprising the following steps: S1. The positively charged polymer, the negatively charged functional monomer, the bioactive nanoparticles, and the photoinitiator are mixed in a solvent to form a prepolymer solution; S2. The prepolymer solution is subjected to deoxygenation and degassing treatment; S3. The prepolymer liquid undergoes a polymerization reaction under ultraviolet light irradiation to obtain the high-strength supramolecular hydrogel.

5. The preparation method according to claim 4, characterized in that: The solvent includes physiological saline or deionized water, and the mass ratio of the solvent to the positively charged polymer is 1:(6-10). The positively charged polymer is added in the form of an aqueous solution with a mass concentration of 10-50%.

6. The preparation method according to claim 4 or 5, characterized in that: The ultraviolet light has a wavelength of 350-405 nm and a light intensity of 50-200 mW / cm². 2 ; The polymerization reaction takes 10-30 minutes.

7. The use of the high-strength supramolecular hydrogel according to any one of claims 1-3 in the preparation of bone defect repair materials; The bone defect repair material is a bone repair scaffold, and the porous bone repair scaffold is made of the high-strength supramolecular hydrogel by 3D printing. The porosity of the bone repair scaffold is 70-80%, the main pore diameter is 200-500 μm, and the connecting pores are ≥100 μm.

8. A bone repair scaffold, characterized in that, The high-strength supramolecular hydrogel described in any one of claims 1-3 is 3D printed and has a porous structure with a porosity of 70-80%, a main pore diameter of 200-500 μm, and connecting pores ≥100 μm.

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