Hydrogel material based on interpenetrating polymer network and preparation method thereof
By preparing an interpenetrating polymer network hydrogel material, combined with astragaloside IV and decellularized extracellular matrix, the limitations of poor mechanical properties of hydrogels and traditional bone grafting methods were overcome, achieving the effects of bone defect repair and angiogenesis.
Patent Information
- Application Number
- CN202511229188.1
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-08-29
- Publication Date
- 2025-11-11
AI Technical Summary
Existing hydrogel materials have poor mechanical properties in bone tissue engineering, which limits their application. Furthermore, traditional bone transplantation methods have limitations such as limited donors and immune rejection.
By preparing hydrogel materials based on interpenetrating polymer networks, and utilizing the crosslinking reaction of astragaloside IV and decellularized extracellular matrix with polyvinyl alcohol and polyethylene glycol diacrylate, combined with freeze-thaw cycle treatment, hydrogels with excellent mechanical properties and bioactivity are formed.
It enhances the mechanical properties and bioactivity of hydrogels, promotes bone defect repair and angiogenesis, and significantly improves the effects of bone tissue regeneration and angiogenesis.
Smart Images
Figure CN120919399A_ABST
Abstract
Description
Technical Field
[0001] This invention relates to the field of biomedical materials technology, specifically to an interpenetrating polymer network-based hydrogel material and its preparation method. Background Technology
[0002] Bone defects are a common clinical problem, especially in cases of trauma, tumor resection, and congenital bone dysplasia. Traditional treatments such as autologous and allogeneic bone transplantation have many limitations, such as limited donors and immune rejection. Therefore, developing a biomaterial that can promote bone regeneration and simultaneously achieve vascularization is of great significance.
[0003] In recent years, with the rapid development of bone tissue physiology and materials science, bone tissue engineering has gradually become a hot topic in medical research. Hydrogels are biomaterials composed of hydrophilic three-dimensional network structures, possessing excellent biocompatibility and plasticity, and are widely used in tissue repair research. However, the poor mechanical properties of hydrogels limit their application in bone tissue engineering. Therefore, constructing hydrogel networks with excellent mechanical properties has become a key research area.
[0004] Decellularized extracellular matrix is obtained by decellularizing bone tissue, preserving its natural components and structure. Astragaloside IV is a monomeric substance extracted from Astragalus membranaceus, which promotes angiogenesis and reduces damage caused by hypoxia. Summary of the Invention
[0005] The purpose of this invention is to provide an interpenetrating polymer network hydrogel material and its preparation method, forming an interpenetrating polymer network hydrogel with excellent mechanical properties and bioactivity, which can be used for bone defect repair and angiogenesis.
[0006] To achieve the above objectives, the technical solution provided by the present invention is as follows:
[0007] This invention is achieved through the following technical solution:
[0008] The first aspect of this application provides a method for preparing an interpenetrating polymer network hydrogel material, comprising the following steps:
[0009] Add decellularized extracellular matrix solution, astragaloside IV, polyethylene glycol diacrylate and photoinitiator sequentially to polyvinyl alcohol solution, and mix thoroughly.
[0010] The mixture was refrigerated overnight and then subjected to a cross-linking reaction under ultraviolet light.
[0011] The cross-linked hydrogel was sequentially soaked in CaCl2 solution, soaked in NaOH solution, and subjected to multiple freeze-thaw cycles to finally obtain the repair material.
[0012] To optimize the above technical solution, the specific limitations also include:
[0013] The concentration of the polyvinyl alcohol solution is 5-20% w / w, and the amount of decellularized extracellular matrix added is 0.8-1.2% w / w.
[0014] The amount of astragaloside IV added is 0.2–0.4 mg / ml.
[0015] The amount of polyethylene glycol diacrylate added is 0.8-1.2 g / ml, and the amount of photoinitiator added is 0.003-0.007 g / ml.
[0016] Furthermore, the cross-linking reaction under ultraviolet light is carried out under 365nm ultraviolet light for 25-35 minutes to form a hydrogel.
[0017] Furthermore, in the CaCl2 solution soaking, the concentration of the CaCl2 solution is 70-90 mmol / L, and the soaking time is 8-12 minutes.
[0018] Furthermore, in the NaOH solution soaking, the concentration of the NaOH solution is 5-7 mol / L, and the soaking time is 25-35 minutes.
[0019] Furthermore, the freeze-thaw cycle treatment involves freezing the hydrogel at -22 to -18°C for 1.5 to 2.5 hours, and then thawing it at 23 to 27°C for 1.5 to 2.5 hours, repeating this process 8 to 12 times.
[0020] The second aspect of this application provides a hydrogel material based on an interpenetrating polymer network, prepared using the method described above.
[0021] The aforementioned repair material is used for bone defect repair and blood vessel regeneration.
[0022] Compared with the prior art, the beneficial effects of the present invention are:
[0023] This invention enhances the mechanical properties of hydrogels and endows them with excellent bioactivity by introducing astragaloside IV (ASIV) and decellularized extracellular matrix (DECM). In this invention, astragaloside IV (ASIV) is used as part of the hydrogel matrix, and through the synergistic effect with polyvinyl alcohol (PVA) and decellularized extracellular matrix (DECM), a hydrogel with excellent mechanical properties and bioactivity is formed.
[0024] This invention utilizes freeze-thaw cycles, a hydrogel matrix, and the synergistic effect of astragaloside IV (ASIV) to form an interpenetrating polymer network hydrogel with excellent mechanical properties and bioactivity. This hydrogel not only surpasses existing technologies in mechanical properties but also exhibits significant advantages in bioactivity, effectively promoting bone defect repair and angiogenesis. The specific synergistic effects are as follows:
[0025] Freeze-thaw cycling enhances the mechanical properties of hydrogels through physical means, enabling them to maintain stable mechanical properties even after multiple compression cycles. This treatment method complements the composition of the hydrogel matrix. PVA and DECM form a more stable cross-linked network during freeze-thaw cycling, thereby significantly improving the toughness and elastic recovery of the hydrogel.
[0026] PVA and DECM in the hydrogel matrix provide a good growth environment for cells, while the addition of ASIV further enhances the bioactivity of the hydrogel. ASIV can promote cell proliferation, adhesion and osteogenic differentiation, and also promote angiogenesis. This synergistic effect not only improves the biocompatibility of the hydrogel, but also enables it to better promote bone tissue regeneration and angiogenesis in vivo.
[0027] Freeze-thaw cycling not only enhances the mechanical properties of the hydrogel but also provides favorable conditions for the release of ASIVs. During freeze-thaw cycling, ASIVs can be more evenly distributed within the hydrogel and gradually released in vivo, thus sustaining their bioactivity and making the hydrogel more effective in promoting bone tissue regeneration and angiogenesis. Attached Figure Description
[0028] Figure 1 : Flowchart of the preparation process of the interpenetrating polymer network hydrogel of the present invention.
[0029] Figure 2 : ESEM image of hydrogel based on interpenetrating polymer network.
[0030] Figure 3 Based on interpenetrating polymer network hydrogel (strain) plot and frequency plot.
[0031] Figure 4 Compression-relaxation curves of hydrogels with different ASIVs based on interpenetrating polymer network hydrogels.
[0032] Figure 5Images of ASI@PVA / DECM printed materials: (a) Computer-aided design of the 3D printed model, including grids, irregular squares, circles, and letters; (b) Printed images of ASI@PVA / DECM (scale bar: 3 mm); (c) Microscopic images of ASI@PVA / DECM printed materials (scale bar: 1 mm); (d) Comparison of the printing nozzle diameter with the diameter of the printed hydrogel sample.
[0033] Figure 6 ALP staining diagram of hydrogel based on interpenetrating polymer network.
[0034] Figure 7 : Scratch test results of hydrogel based on interpenetrating polymer network.
[0035] Figure 8 Experimental diagram of hydrogel tube formation based on interpenetrating polymer network.
[0036] Figure 9 Histological staining map of hydrogel based on interpenetrating polymer network. Detailed Implementation
[0037] The present invention will be further described in detail below through specific embodiments, but it should not be construed as limiting the scope of the subject matter of the present invention to the following embodiments. All technologies implemented based on the above content of the present invention fall within the scope of the present invention.
[0038] Unless otherwise specified, the experimental methods used in the following examples are conventional methods, and the reagents, methods and equipment used are conventional reagents, methods and equipment in this technical field.
[0039] For the sake of brevity, this article only discloses some numerical values and the range of options. However, any lower limit can be combined with any upper limit to form an unspecified range; and any lower limit can be combined with other lower limits to form an unspecified range. Similarly, any upper limit can be combined with any other upper limit to form an unspecified range; the options in the range of options can also be combined arbitrarily.
[0040] The technical solution of the present invention will be further described in detail below with reference to specific embodiments:
[0041] Example 1
[0042] The preparation process of the interpenetrating polymer network hydrogel material in this embodiment includes the following steps:
[0043] 1. Preparation of polyvinyl alcohol solution: Dissolve 10g of polyvinyl alcohol in 100mL of deionized water and stir at 55℃ for 4 hours until the polyvinyl alcohol is completely dissolved to obtain a 10% polyvinyl alcohol solution.
[0044] 2. Preparation of decellularized extracellular matrix solution: Dissolve 10 mL of decellularized extracellular matrix in 100 mL of deionized water to obtain a 10% decellularized extracellular matrix solution.
[0045] 3. Mixing reaction: Mix 10 mL of polyvinyl alcohol solution, 10 mL of decellularized extracellular matrix solution, 1 mg / mL astragaloside IV, 10 g of polyethylene glycol diacrylate and 0.05 g of photoinitiator evenly to obtain a mixed solution.
[0046] 4. Cross-linking reaction: The mixed solution was subjected to a cross-linking reaction under ultraviolet light (365nm) for 30 minutes to form a hydrogel.
[0047] 5. Post-treatment: The cross-linked hydrogel was soaked in 80 mmol / L CaCl2 solution for 10 minutes and then in 6 mol / L NaOH solution for 30 minutes. Then, it was subjected to 10 freeze-thaw cycles (the freeze-thaw cycle was to freeze the hydrogel at -20℃ for 2 hours and then thaw it at 25℃ for 2 hours, and repeat) to finally obtain an interpenetrating polymer network hydrogel based on polyvinyl alcohol and decellularized extracellular matrix.
[0048] Example 2
[0049] The preparation process in this embodiment is basically the same as in Example 1, except that the astragaloside IV in step 3 is 2 mg / ml.
[0050] Example 3
[0051] The preparation process in this embodiment is basically the same as in Example 1, except that the astragaloside IV in step 3 is 3 mg / ml.
[0052] Example 4
[0053] The preparation process in this embodiment is basically the same as in Example 1, except that the astragaloside IV in step 3 is 4 mg / ml.
[0054] Comparative Example 1
[0055] The preparation process in this embodiment is basically the same as in Example 1, except that astragaloside IV is not added in step 3.
[0056] Comparative Example 2
[0057] The preparation process in this embodiment is basically the same as in Example 1, except that the astragaloside IV in step 3 is 5 mg / ml.
[0058] Experimental verification:
[0059] 1. Mechanical property testing: The mechanical properties of the hydrogel are evaluated through compression tests and multiple compression-unloading cycle tests.
[0060] 2. Biocompatibility testing: The biocompatibility of the hydrogel was evaluated through CCK-8 assay, live / dead staining, and cell adhesion assay.
[0061] 3. Osteogenesis and angiogenesis capacity tests: The effects of hydrogels on cell osteogenic and angiogenesis were studied through alkaline phosphatase (ALP) staining, osteogenic mineralization nodule staining test, scratch test and tube formation test.
[0062] 4. In vivo experiments: Using a rat femoral condyle defect model, the effects of hydrogel on promoting bone regeneration and angiogenesis in vivo were evaluated.
[0063]
[0064] Figure 2 The surface shape of the hydrogel material based on interpenetrating polymer network was shown using ESEM images. Figure 3 The strain diagram describes the degree of deformation of the hydrogel in compression or tensile tests, and the frequency diagram describes the loading rate of the hydrogel in dynamic rheological tests. Figure 4 Compression-relaxation curves of hydrogels under different ASIVs are presented, showcasing the rheological properties of the interpenetrating polymer network (IPN) hydrogel material, including storage modulus (G') and loss modulus (G”). The maximum compressive strain, maximum compressive stress, Young's modulus, and toughness parameters in the table show slight differences in mechanical properties among the different groups. The compression-fracture curves of the IPN hydrogel material are also presented, corresponding to the maximum compressive strain and maximum compressive stress parameters in the table, providing a clear visual indication of the differences in mechanical properties among the different groups.
[0065] Figure 6 The effects of ALP-stained images on osteoblast differentiation based on interpenetrating polymer network hydrogel materials were shown, corresponding to the ALP activity in the table, reflecting the differences in ALP activity among different groups. Figure 7 The scratch test images demonstrate the effect of interpenetrating polymer network-based hydrogel materials on endothelial cell migration. The images correspond to the migration distances of endothelial cells in the table, indicating that endothelial cells in different groups have different migration abilities. Figure 8 The tube-forming experimental images demonstrate the effect of interpenetrating polymer network hydrogel materials on the formation of tubular structures in endothelial cells. Corresponding to the number of tubular structures in the table, the differences in tubular structure formation ability among different groups can be visually observed. Figure 9 Histological staining images were used to demonstrate the effects of interpenetrating polymer network-based hydrogel materials on promoting bone regeneration and angiogenesis in vivo, highlighting the differences in in vivo effects among different groups.
[0066] The mechanical property test results of this invention show that the hydrogel has high compressive strength and good elastic recovery ability, and can maintain stable mechanical properties after multiple compression cycles.
[0067] Biocompatibility testing results showed that the hydrogel had no significant toxicity to cells and could significantly promote cell proliferation and adhesion.
[0068] Osteogenesis and angiogenesis capacity tests showed that the hydrogel could significantly promote osteoblast differentiation and mineralization, while also promoting endothelial cell migration and tubular structure formation.
[0069] In vivo experiments showed that the hydrogel can significantly promote the repair of bone defects and angiogenesis, with the defect area covered by regenerated bone tissue, and its morphology closely resembling that of natural bone tissue.
[0070] Experiments showed that the groups with astragaloside IV concentrations of 2–4 mg / mL exhibited the best performance in terms of mechanical properties, bioactivity, osteogenic capacity, and angiogenesis capacity, demonstrating the highest repair efficacy. This result provides important experimental evidence for the development of interpenetrating polymer network hydrogels based on polyvinyl alcohol and decellularized extracellular matrix.
[0071] The results of Comparative Example 2 showed that when the concentration of ASIV increased to 6 mg / ml, the mechanical properties (maximum compressive strain, maximum compressive stress, Young's modulus) and biological activities (ALP activity, osteogenic mineralized nodule area, endothelial cell migration distance, number of tubular structures) of the hydrogel decreased. In addition, the degradation rate and ASI release rate also decreased, which may affect the sustained effect and biocompatibility of the hydrogel in vivo.
[0072] The interpenetrating polymer network hydrogel based on polyvinyl alcohol and decellularized extracellular matrix of the present invention can be used in the preparation of materials for the repair or replacement of musculoskeletal tissues, especially for the repair and replacement of bone, cartilage or ligament defects, and to promote tissue regeneration.
[0073] The above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention in any way. Any simple modifications, equivalent substitutions, and improvements made by those skilled in the art to the above embodiments without departing from the scope of the technical solution of the present invention, based on the technical essence of the present invention, shall still fall within the protection scope of the technical solution of the present invention.
Claims
1. A method for preparing a hydrogel material based on an interpenetrating polymer network, characterized in that, Includes the following steps: Add decellularized extracellular matrix solution, astragaloside IV, polyethylene glycol diacrylate and photoinitiator sequentially to polyvinyl alcohol solution, and mix thoroughly. The mixture was refrigerated overnight and then subjected to a cross-linking reaction under ultraviolet light. The cross-linked hydrogel was sequentially soaked in CaCl2 solution, soaked in NaOH solution, and subjected to multiple freeze-thaw cycles to finally obtain the repair material.
2. The method for preparing hydrogel materials based on interpenetrating polymer networks according to claim 1, characterized in that: The concentration of the polyvinyl alcohol solution is 5-20% w / w, and the amount of decellularized extracellular matrix added is 0.8-1.2% w / w.
3. The method for preparing hydrogel materials based on interpenetrating polymer networks according to claim 1, characterized in that: The amount of astragaloside IV added is 0.2–0.4 mg / ml.
4. The method for preparing hydrogel materials based on interpenetrating polymer networks according to claim 1, characterized in that: The amount of polyethylene glycol diacrylate added is 0.8-1.2 g / ml, and the amount of photoinitiator added is 0.003-0.007 g / ml.
5. The method for preparing hydrogel materials based on interpenetrating polymer networks according to claim 1, characterized in that: The cross-linking reaction under ultraviolet light is carried out under 365nm ultraviolet light for 25-35 minutes to form a hydrogel.
6. The method for preparing hydrogel materials based on interpenetrating polymer networks according to claim 1, characterized in that: In the CaCl2 solution immersion, the concentration of the CaCl2 solution is 70-90 mmol / L, and the immersion time is 8-12 minutes.
7. The method for preparing hydrogel materials based on interpenetrating polymer networks according to claim 1, characterized in that: In the NaOH solution soaking process, the concentration of the NaOH solution is 5-7 mol / L, and the soaking time is 25-35 minutes.
8. The method for preparing hydrogel materials based on interpenetrating polymer networks according to claim 1, characterized in that: The freeze-thaw cycle treatment involves freezing the hydrogel at -22 to -18°C for 1.5 to 2.5 hours, and then thawing it at 23 to 27°C for 1.5 to 2.5 hours. This process is repeated 8 to 12 times.
9. A hydrogel material based on an interpenetrating polymer network, characterized in that: Prepared using the method described in any one of claims 1 to 8.
10. The hydrogel material based on interpenetrating polymer network according to claim 9, characterized in that: The aforementioned repair material is used for bone defect repair and blood vessel regeneration.