Magnetic drug-loaded hydrogel microspheres, preparation method and application thereof
By preparing magnetic drug-loaded hydrogel microspheres loaded with small-molecule anti-inflammatory drugs and large-molecule nerve growth factors, and combining them with external magnetic field intervention, the problems of drug matching and inflexible delivery methods in the treatment of spinal cord injury were solved, and multimodal synergistic repair and precise treatment of spinal cord injury were achieved.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- TSINGHUA UNIVERSITY
- Filing Date
- 2026-02-27
- Publication Date
- 2026-05-29
Smart Images
Figure CN122097546A_ABST
Abstract
Description
Technical Field
[0001] This invention belongs to the field of hydrogel technology, specifically relating to a magnetic drug-loaded hydrogel microsphere, its preparation method, and its application. Background Technology
[0002] Spinal cord injury (SCI) is a highly destructive neurological disease. Its complex pathophysiological cascade and limited endogenous regenerative capacity lead to poor clinical repair outcomes. In recent years, conductive bio-scaffolds have shown promising results in promoting the reconstruction of spinal cord neural circuits. However, electromagnetic interactions are often closely related, and magnetic intervention may be an important way to regulate nerve cell behavior and promote the repair of damaged neural tissue. Furthermore, traditional drug delivery strategies are often limited to loading and sustained-release of single-type drugs, making it difficult to match the dynamic pathological processes of acute inflammatory flare-ups and chronic regenerative inhibition in SCI. On the other hand, the form of material implantation is also receiving increasing attention; meeting injectability requirements and constructing flexible and convenient multiplex drug delivery systems are also urgent problems to be solved. Summary of the Invention
[0003] This application provides a magnetic drug-loaded hydrogel microsphere, its preparation method, and its application, aiming to solve the problems of existing spinal cord injury treatment drugs being unable to match the dynamic pathological processes of acute inflammatory outbreaks and chronic regenerative inhibition in spinal cord injury, as well as the single drug loading and inflexible delivery methods.
[0004] The first aspect of this application provides a magnetic drug-loaded hydrogel microsphere comprising the following raw materials in parts by weight: 1-40 parts of a hydrogel crosslinking precursor, 2-30 parts of an initiator, 0.1-1 parts of magnetic nanoparticles loaded with small molecule drugs, and 0-1 parts of a macromolecule drug.
[0005] According to some embodiments of the magnetic drug-loaded hydrogel microspheres described in this application, the hydrogel crosslinking precursor includes an ionic hydrogel crosslinking precursor and / or a photosensitive hydrogel crosslinking precursor; preferably, the ionic hydrogel crosslinking precursor includes sodium alginate; preferably, the photosensitive hydrogel crosslinking precursor includes at least one of methacrylamide hyaluronic acid, methacrylamide gelatin, methacrylamide polyvinyl alcohol, methacrylamide sodium alginate, methacrylamide silk fibroin, methacrylamide chitosan, methacrylamide glucose, methacrylamide chondroitin sulfate, and methacrylamide diacrylate.
[0006] According to some embodiments of the magnetic drug-loaded hydrogel microspheres described in this application, the initiator includes an ionic initiator and / or a photoinitiator; preferably, the ionic initiator includes calcium chloride; preferably, the photoinitiator includes at least one of lithium phenyl (2,4,6-trimethylbenzoyl)phosphate, IRGACURE 2959, and ruthenium.
[0007] According to some embodiments of the magnetic drug-loaded hydrogel microspheres described in this application, the magnetic nanoparticles in the magnetic nanoparticles loaded with small molecule drugs include at least one of Fe3O4, γ-Fe2O3, MnFe2O4, NaGdF4, and Gd2O3.
[0008] According to some embodiments of the magnetic drug-loaded hydrogel microspheres described in this application, the small molecule drug in the magnetic nanoparticles loaded with small molecule drugs includes at least one of natural small molecule drugs, nonsteroidal anti-inflammatory drugs (NSAIDs), and steroidal anti-inflammatory drugs; preferably, the natural small molecule drug includes curcumin; preferably, the NSAID includes aspirin and / or indomethacin; preferably, the steroidal anti-inflammatory drug includes fluticasone and / or hydroxychloroquine;
[0009] According to some embodiments of the magnetic drug-loaded hydrogel microspheres described in this application, the mass ratio of magnetic nanoparticles to small molecule drugs in the magnetic nanoparticles loaded with small molecule drugs is (45-55):1.
[0010] According to some embodiments of the magnetic drug-loaded hydrogel microspheres described in this application, the macromolecular drug includes at least one of basic fibroblast growth factor (bFGF), epidermal growth factor (EGF), and vascular endothelial growth factor (VEGF).
[0011] According to some embodiments of the magnetic drug-loaded hydrogel microspheres described in this application, a gel medium is also included.
[0012] According to some embodiments of the magnetic drug-loaded hydrogel microspheres described in this application, the gel medium includes phosphate buffer.
[0013] According to some embodiments of the magnetic drug-loaded hydrogel microspheres described in this application, the particle size of the magnetic drug-loaded hydrogel microspheres is 60-75 μm.
[0014] This application also provides a method for preparing the magnetic drug-loaded hydrogel microspheres described in the first aspect of this application, comprising the following steps: (1) The hydrogel crosslinking precursor, initiator, magnetic nanoparticles loaded with small molecule drugs and macromolecule drugs are mixed to obtain an aqueous phase; (2) Droplets were prepared by shearing the aqueous phase with the oil phase and then solidified; the oil phase was removed to obtain the magnetic drug-loaded hydrogel microspheres.
[0015] According to some embodiments of the preparation method of magnetic drug-loaded hydrogel microspheres described in this application, in step (2), the oil phase includes at least one of paraffin oil, edible oil and surfactant in HFE7500.
[0016] According to some embodiments of the preparation method of magnetic drug-loaded hydrogel microspheres described in this application, in step (2), the volume ratio of the aqueous phase to the oil phase is 1:(4-10).
[0017] According to some embodiments of the method for preparing magnetic drug-loaded hydrogel microspheres described in this application, in step (2), the method for preparing droplets includes microfluidics.
[0018] According to some embodiments of the method for preparing magnetic drug-loaded hydrogel microspheres described in this application, the method further includes the step of preparing magnetic nanoparticles loaded with small molecule drugs. Specifically, the following steps are included: Magnetic nanoparticles and a buffer solution containing nanoparticle modifiers are mixed to obtain functionalized magnetic nanoparticle powder; the functionalized magnetic nanoparticle powder is mixed with a small molecule drug solution, filtered, washed, and dried to obtain the magnetic nanoparticles loaded with the small molecule drug.
[0019] According to some embodiments of the preparation method of the magnetic drug-loaded hydrogel microspheres described in this application, the buffer solution includes Tris-HCl buffer solution, the pH of the buffer solution is 8.5, and the concentration of the nanoparticle modifier in the buffer solution is 1-3 mg / ml.
[0020] According to some embodiments of the preparation method of magnetic drug-loaded hydrogel microspheres described in this application, the nanoparticle modifier includes polydopamine.
[0021] According to some embodiments of the preparation method of magnetic drug-loaded hydrogel microspheres described in this application, the mass ratio of the magnetic nanoparticles to the nanoparticle modifier is (4-6):3.
[0022] According to some embodiments of the preparation method of the magnetic drug-loaded hydrogel microspheres described in this application, the concentration of the small molecule drug solution is 1-5 mg / mL, and the solvent used for the small molecule drug solution is water and / or dimethyl sulfoxide.
[0023] The third aspect of this application provides the application of magnetic drug-loaded hydrogel microspheres as described in the first aspect of this application or magnetic drug-loaded hydrogel microspheres prepared by the method described in the second aspect of this application in the prevention or treatment of electroactive organ damage.
[0024] According to some embodiments of the application described in this application, the electroactive organ includes the spinal cord, brain, peripheral nervous system, or heart.
[0025] The beneficial effects of this application include: the magnetic drug-loaded hydrogel microspheres described in this application are injectable and magnetically responsive, and can simultaneously load bioactive macromolecules and small molecules. When intervened by an external magnetic field and combined with the magnetic response characteristics, the anti-inflammatory small molecules and nerve growth-promoting macromolecules contained in the magnetic drug-loaded hydrogel microspheres can be released in a dual sustained manner, synergistically reducing the harm of spinal cord injury.
[0026] The magnetic drug-loaded hydrogel microspheres described in this application utilize magnetic nanoparticles that, on the one hand, can flexibly load small-molecule anti-inflammatory drugs through surface functionalization; on the other hand, they also act as magnetic response cues, synergizing with an external static magnetic field to achieve magnetic intervention in spinal cord injury tissue. The material is injected into the spinal cord injury site in an injectable form, and an external static magnetic field is applied to achieve synergistic effects with the internal magnetic nanoparticles. The static magnetic field combined with the magnetic nanoparticles promotes the proliferation and differentiation of nerve cells, and the drug-loaded magnetic nanoparticles also exhibit a significant protective effect on nerve cells under oxidative stress. The preparation method of the magnetic drug-loaded hydrogel microspheres described in this application is simple, mild, and rapid. Attached Figure Description
[0027] Figure 1 The sustained-release curve of curcumin in the magnetic drug-loaded hydrogel microspheres described in Example 1 of this application; Figure 2 The survival rate (left) and proliferation activity (right) of cells with different concentrations of functionalized magnetic nanoparticles on the magnetic drug-loaded hydrogel microspheres described in Example 1 of this application. Figure 3 This demonstrates the protective effect of the magnetic drug-loaded hydrogel microspheres described in Example 1 of this application on cells under an oxidative stress model. Figure 3 The left side shows a flow cytometry plot of apoptosis rate. Figure 3 The right side shows the cell survival rate statistics; Figure 4 This is a diagram showing the area of damage after spinal cord injury using the magnetic drug-loaded hydrogel microspheres described in Example 1 of this application. Detailed Implementation
[0028] The embodiments of the present invention are described in detail below. These embodiments are exemplary and intended to explain the present invention, and should not be construed as limiting the present invention.
[0029] In this invention, the terms "one embodiment," "some embodiments," "example," "specific example," or "some examples," etc., refer to a specific feature, structure, material, or characteristic described in connection with that embodiment or example, which is included in at least one embodiment or example of the invention. In this specification, the illustrative expressions of the above terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in one or more embodiments or examples. Moreover, without contradiction, those skilled in the art can combine and integrate the different embodiments or examples described in this specification, as well as the features of different embodiments or examples.
[0030] This application provides a magnetic drug-loaded hydrogel microsphere comprising the following raw materials in parts by weight: 1-40 parts of hydrogel crosslinking precursor, 2-30 parts of initiator, 0.1-1 parts of magnetic nanoparticles loaded with small molecule drugs, and 0-1 parts of macromolecule drugs.
[0031] The magnetic drug-loaded hydrogel microspheres described in this application are internally loaded with magnetic nanoparticles modified with the anti-inflammatory small molecule curcumin. Basic fibroblast cytokine (bFGF) was further selected as an example of a macromolecular drug, and it was in situ crosslinked into the three-dimensional network of the gel through a gelation process, thereby realizing the construction of a dual drug loading system of large and small molecules. Based on this, the material was injected into the spinal cord injury site in an injectable form.
[0032] The spinal cord possesses typical electrophysiological characteristics and exhibits close electromagnetic interactions. Secondary damage following spinal cord injury dominates neuronal apoptosis and persistent immune inflammation. Therefore, a key issue is how to construct repair materials that match the dynamic pathological process while adapting to the physiological characteristics of the spinal cord. The magnetic drug-loaded hydrogel microspheres described in this application employ a multimodal synergistic repair strategy that integrates magnetic regulatory cues and flexibly loads large / small molecule drugs via injection. Magnetic nanoparticles can, on the one hand, achieve flexible loading of small molecule anti-inflammatory drugs through surface functionalization to inhibit immune inflammation following spinal cord injury; on the other hand, they also act as magnetic cues, synergizing with external static magnetic fields to achieve magnetic intervention on injured spinal cord tissues.
[0033] In some embodiments of this application, the hydrogel crosslinking precursor includes ionic hydrogel crosslinking precursors and / or photosensitive hydrogel crosslinking precursors.
[0034] In some embodiments of this application, the ionic hydrogel crosslinking precursor includes sodium alginate.
[0035] In some embodiments of this application, the photosensitive hydrogel crosslinking precursor includes at least one of methacrylamide hyaluronic acid, methacrylamide gelatin, methacrylamide polyvinyl alcohol, methacrylamide sodium alginate, methacrylamide silk fibroin, methacrylamide chitosan, methacrylamide glucose, methacrylamide chondroitin sulfate, and methacrylamide diacrylate.
[0036] In some embodiments of this application, the initiator includes an ionic initiator and / or a photoinitiator.
[0037] In some embodiments of this application, the ion initiator includes calcium chloride.
[0038] In some embodiments of this application, the photoinitiator includes at least one of lithium phenyl (2,4,6-trimethylbenzoyl)phosphate, IRGACURE 2959, and ruthenium.
[0039] In some embodiments of this application, the magnetic nanoparticles in the magnetic nanoparticles loaded with small molecule drugs include at least one of Fe3O4, γ-Fe2O3, MnFe2O4, NaGdF4, and Gd2O3.
[0040] In some embodiments of this application, the small molecule drug in the magnetic nanoparticles loaded with small molecule drugs includes at least one of natural small molecule drugs, nonsteroidal anti-inflammatory drugs (NSAIDs), and steroidal anti-inflammatory drugs; preferably, the natural small molecule drug includes curcumin; preferably, the NSAID includes aspirin and / or indomethacin; preferably, the steroidal anti-inflammatory drug includes fluticasone and / or hydroxychloroquine.
[0041] In some embodiments of this application, the mass ratio of magnetic nanoparticles to small molecule drugs in the magnetic nanoparticles loaded with small molecule drugs is (45-55):1; for example, 45:1, 48:1, 50:1, 55:1, etc.
[0042] In some embodiments of this application, the macromolecular drug includes at least one of basic fibroblast growth factor (bFGF), epidermal growth factor (EGF), and vascular endothelial growth factor (VEGF). The macromolecular drug is in situ crosslinked into the three-dimensional network of the gel through a gelation process, thereby constructing a dual drug loading system of large and small molecules for inhibiting neuronal apoptosis after spinal cord injury. In this dual drug loading system, the small molecule drug exerts an anti-inflammatory effect, while the large molecule drug promotes regeneration.
[0043] In some embodiments of this application, a gel medium is also included; the gel medium is equivalent to a solvent for dissolving the hydrogel precursor and the initiator.
[0044] In some embodiments of this application, the gel medium includes phosphate buffer.
[0045] In some embodiments of this application, the magnetic drug-loaded hydrogel microspheres have a particle size of 60-70 μm, such as 60 μm, 63 μm, 65 μm, 68 μm, 70 μm, etc.
[0046] This application also provides a method for preparing the magnetic drug-loaded hydrogel microspheres described in the first aspect of this application, comprising the following steps: (1) The hydrogel crosslinking precursor, initiator, magnetic nanoparticles loaded with small molecule drugs and macromolecule drugs are mixed to obtain an aqueous phase; (2) Droplets were prepared by shearing the aqueous phase with the oil phase and then solidified; the oil phase was removed to obtain the magnetic drug-loaded hydrogel microspheres.
[0047] The method for preparing magnetic drug-loaded hydrogel microspheres described in this application is simple, mild, and rapid, and it constructs a multimodal synergistic repair strategy that integrates magnetic modulation cues in an injectable form and flexibly carries large / small molecule drugs.
[0048] In some embodiments of this application, in step (2), the oil phase includes at least one of paraffin oil, edible oil and surfactantin HFE7500.
[0049] In some embodiments of this application, in step (2), the volume ratio of the aqueous phase to the oil phase is 1:(4-10); for example, 1:4, 1:6, 1:8, 1:10, etc.
[0050] In some embodiments of this application, the method for preparing droplets in step (2) includes microfluidics.
[0051] In some embodiments of this application, the step of preparing magnetic nanoparticles loaded with small molecule drugs is also included; Specifically, the following steps are included: Magnetic nanoparticles and a buffer solution containing nanoparticle modifiers are mixed to obtain functionalized magnetic nanoparticle powder; the functionalized magnetic nanoparticle powder is mixed with a small molecule drug solution, filtered, washed, and dried to obtain the magnetic nanoparticles loaded with the small molecule drug.
[0052] In some embodiments of this application, the buffer solution includes a Tris-HCl buffer solution with a pH of 8.5, and the concentration of the nanoparticle modifier in the buffer solution is 1-3 mg / ml; for example, 1 mg / ml, 2 mg / ml, 3 mg / ml, etc.
[0053] In some embodiments of this application, the nanoparticle modifier includes polydopamine; polydopamine has the function of connecting magnetic nanoparticles and small molecule drugs.
[0054] In some embodiments of this application, the mass ratio of the magnetic nanoparticles to the nanoparticle modifier is (4-6):3; for example, 4:3, 5:3, 6:3, etc.
[0055] In some embodiments of this application, the concentration of the small molecule drug solution is 1-5 mg / mL, such as 1 mg / mL, 2 mg / mL, 3 mg / mL, 5 mg / mL, etc., and the solvent used for the small molecule drug solution is water and / or dimethyl sulfoxide.
[0056] In some embodiments of this application, polydopamine powder was dispersed in a 10 mM Tris-HCl buffer solution (pH=8.5); 0.1-1.0 g of Fe3O4 magnetic nanoparticles were added to the buffer solution and mechanically stirred at room temperature (25°C) for 12 h. The resulting precipitate was washed three times alternately with anhydrous ethanol and deionized water to obtain functionalized magnetic nanoparticle powder; curcumin was dissolved in a mixed solution of water and DMSO (volume ratio of 10:1) to obtain a curcumin solution with a concentration of 1-5 mg / L. 200 μL of the above curcumin solution was mixed with 5 mL of ultrapure water, and 10-50 mg of functionalized magnetic nanoparticle powder was added. The mixture was stirred at room temperature for 24 h, washed, and then freeze-dried to obtain the magnetic nanoparticles loaded with the small molecule drug.
[0057] This application also provides the application of magnetic drug-loaded hydrogel microspheres as described in the first aspect of this application or magnetic drug-loaded hydrogel microspheres prepared by the method described in the second aspect of this application in the prevention or treatment of electroactive organ damage.
[0058] The application of the magnetic drug-loaded hydrogel microspheres described in this application to drugs for the prevention or treatment of electroactive organ injury promotes the transformation of spinal cord injury repair from single neuroprotection to a precise "neuro-immune co-regulation" model, and provides new ideas for the construction of novel spinal cord injury repair materials.
[0059] In some embodiments of this application, the electroactive organ includes the spinal cord, brain, peripheral nervous system, or heart.
[0060] The technical solution of this application will be further described below with reference to specific embodiments.
[0061] Example 1 A method for preparing magnetic drug-loaded hydrogel microspheres includes the following steps: (1) Mix 0.1g Fe3O4 magnetic nanoparticles with 30ml Tris-HCl buffer containing polydopamine (where the mass concentration of polydopamine in the Tris-HCl buffer is 2g / L and the pH of the buffer is 8.5), stir at 25℃ for 12h, filter, and wash the precipitate three times alternately with anhydrous ethanol and deionized water to obtain functionalized magnetic nanoparticle powder. 1 mg of curcumin was dissolved in 1 mL of a mixture of water and DMSO (volume ratio of 10:1). 200 μL of the curcumin solution was mixed with 5 mL of ultrapure water. 10 mg of the functionalized magnetic nanoparticle powder was added to the mixture. The mixture was stirred at 25 °C for 24 h. After filtration, washing, and drying, curcumin-loaded magnetic nanoparticles were obtained. (2) Mix 10 mg of methacrylamide hyaluronic acid, 2.5 mg of phenyl (2,4,6-trimethylbenzoyl) lithium phosphate, 20 μg of curcumin-loaded magnetic nanoparticles, 300 μg of basic fibroblast growth factor and 1 mL of phosphate buffer as the aqueous phase; (3) 2% surfactant in HFE7500 is used as the oil phase. A cross-shaped microfluidic chip is introduced to control the volume ratio of the water phase and the oil phase to 1:8. During the curing process, blue light with a wavelength of 405nm is applied to the end of the droplet collection channel to achieve cross-linking of droplets to microspheres. The obtained droplets are centrifuged and washed to remove the surface oil phase, and the magnetic drug-loaded hydrogel microspheres with a particle size of 65-75 μm are obtained.
[0062] Example 2 The only difference between the preparation method of the magnetic drug-loaded hydrogel microspheres in Example 2 and Example 1 is that aspirin is used instead of curcumin in the preparation process of the magnetic drug-loaded hydrogel microspheres in Example 2, and the rest of the operation is the same as in Example 1.
[0063] Example 3 The only difference between the preparation method of the magnetic drug-loaded hydrogel microspheres in Example 3 and that in Example 1 is that vascular endothelial growth factor (VEGF) is used instead of basic fibroblast growth factor (bFGF) in the preparation process of the magnetic drug-loaded hydrogel microspheres in Example 3. The rest of the operation is the same as in Example 1.
[0064] Example 4 The only difference between the preparation method of the magnetic drug-loaded hydrogel microspheres in Example 4 and Example 1 is that methacrylated gelatin is used instead of methacrylated hyaluronic acid in the preparation process of the magnetic drug-loaded hydrogel microspheres in Example 4, and the rest of the operation is the same as in Example 1.
[0065] Example 5 The difference between the preparation method of the magnetic drug-loaded hydrogel microspheres in Example 5 and that in Example 1 is that paraffin oil-span 80 is used instead of 2% surfactant in HFE7500 in the preparation process of the magnetic drug-loaded hydrogel microspheres in Example 5, and the rest of the operation is the same as in Example 1.
[0066] Example 6 The only difference between the preparation method of the magnetic drug-loaded hydrogel microspheres in Example 6 and Example 1 is that the volume ratio of the aqueous phase to the oil phase in the preparation process of the magnetic drug-loaded hydrogel microspheres in Example 6 is 1:10, and the rest of the operation is the same as in Example 1.
[0067] Comparative Example 1 The only difference between the preparation method of the magnetic drug-loaded hydrogel microspheres in Comparative Example 1 and Example 1 is that no small molecule drugs or large molecule drugs were added during the preparation process of the magnetic drug-loaded hydrogel microspheres in Comparative Example 1.
[0068] The specific operating steps include: (1) A mixture of methacrylamide hyaluronic acid, phenyl (2,4,6-trimethylbenzoyl) lithium phosphate, magnetic nanoparticles and phosphate buffer was used as the aqueous phase; (3) 2% surfactant in HFE7500 is used as the oil phase. A cross-shaped microfluidic chip is introduced to control the volume ratio of the water phase and the oil phase to 1:8. During the curing process, blue light with a wavelength of 405nm is applied to the end of the droplet collection channel to achieve cross-linking of droplets to microspheres. The obtained droplets are centrifuged and washed to remove the surface oil phase, and the magnetic drug-loaded hydrogel microspheres with a particle size of 65-75 μm are obtained.
[0069] This application presents a study on the effects of magnetic drug-loaded hydrogel microspheres. 1. Study on the release rate of curcumin contained in the magnetic drug-loaded hydrogel microspheres described in Example 1 of this application. Quantitative analysis of drug release behavior of the magnetic drug-loaded hydrogel microspheres based on standard curve: The magnetic drug-loaded hydrogel microspheres described in Example 1 of this application were dispersed in 1 mL of PBS solution (37°C, shake incubation). A standard curve for curcumin was established using UV-Vis spectrophotometry at 37°C. Calibration was performed by measuring the absorbance values at the characteristic absorption wavelength of 429 nm of a series of serially diluted standard solutions (concentration gradients: 3.125, 6.25, 12.5, 25, 50 μmol / L). Quantitatively, 1 mL of release medium was collected at preset time points, and an equal volume of fresh medium (PBS solution) was simultaneously added to maintain a constant system. The concentration of curcumin released at each time point was calculated using the standard curve equation.
[0070] The results are as follows Figure 1 As shown.
[0071] from Figure 1 As can be seen from this, curcumin in the magnetic drug-loaded hydrogel microspheres described in Example 1 of this application is rapidly released within 5 days, and then slowly released in the later stage, and its process matches the pathological process after spinal cord injury.
[0072] 2. Study on the effect of the magnetic drug-loaded hydrogel microspheres described in Example 1 of this application Research Methods: The magnetic drug-loaded hydrogel microspheres described in Example 1 were co-cultured with PC-12 cells. Under the same culture conditions, the cell viability was marked using an AM-PI live / dead fluorescence staining kit, and the cell proliferation activity under different concentrations and time conditions was statistically analyzed. The results are as follows: Figure 2 As shown.
[0073] from Figure 2 As shown in Figure A, to assess the biocompatibility of the hydrogel microspheres, extracts of hydrogel microspheres loaded with different concentrations of MNPs were co-cultured with PC-12 cells. The AM-PI live / dead fluorescence staining kit was used to label cell viability. Figure A shows that the hydrogel microspheres loaded with 50 μg / mL MNPs maintained high compatibility with the cells, and the cell viability remained above 90% after 5 days of culture. Furthermore, to further investigate the effect of MNPs alone on cell proliferation, MNPs were directly co-cultured with cells, and the cell proliferation activity was statistically analyzed under different concentrations and time periods. The results showed that cells in all groups maintained high proliferation activity (Figure B). Notably, within the concentration range of 0-30 μg / mL, MNPs exhibited a concentration-dependent proliferative effect. Based on this dose-response relationship, the concentration for subsequent experiments was determined to be 20 μg / mL.
[0074] 3. Study on the anti-inflammatory effect of the magnetic drug-loaded hydrogel microspheres described in Example 1 of this application. PC-12 cells were cultured in DMEM medium (normal medium) containing 10% FBS as the Control group; PC-12 cells were first treated with normal culture medium for 24 h, then with culture medium containing 150 mM hydrogen peroxide tert-butanol (TBHP) for 1 h, and then replaced with normal culture medium as the TBHP group. PC-12 cells were first treated with a culture medium containing the magnetic drug-loaded hydrogel microspheres described in Example 1 for 24 h (where the concentration of the magnetic drug-loaded hydrogel microspheres was 20 μg / mL), then treated with a culture medium containing 150 mM TBHP for 1 h, and then replaced with normal culture medium, which was used as the Example 1 group (denoted as TBHP+MNPs@PDA@Cur). PC-12 cells were first treated with a culture medium containing magnetic hydrogel microspheres as described in Comparative Example 1 for 24 h (where the concentration of magnetic hydrogel microspheres was 20 μg / mL), then treated with a culture medium containing 150 mM TBHP for 1 h, and then replaced with normal culture medium as Comparative Example 1 (denoted as TBHP+MNPs).
[0075] The results are as follows Figure 3 As shown.
[0076] from Figure 3 As can be seen from the above, the decrease in cell apoptosis rate after treatment with the magnetic drug-loaded hydrogel microspheres described in this application is attributed to: ① Fe3O4MNPs have certain catalase and superoxide dismutase activities, which have a protective effect on cells under oxidative stress conditions; ② the small molecule drug curcumin also has anti-inflammatory effects.
[0077] 4. Study on the therapeutic effect of the magnetic drug-loaded hydrogel microspheres described in Example 1 of this application on spinal cord injury. Modeling and intervention of spinal cord injury in rats: The location of the T9 segment of the rat spinal cord was determined. Using this segment as the center, a surgical approach was established 15 mm above and below the longitudinal axis. The paravertebral muscles were dissected to the outer edge of the facet joints, and the facet joints were removed. A laminectomy was performed centered on this segment, exposing approximately 15 mm of spinal cord parenchyma. The rats were then fixed to the Allen impactor fixation splint. The T7 and T11 spinous processes were clamped with clamps, and the spinal cord impactor was used to confirm that the impact center was in the midline of the rat spinal cord.
[0078] After model construction, 10 μl of each of the following materials was injected into the spinal cord injury site of rats: hydrogel microspheres of magnetic nanoparticles as described in Example 1, and hydrogel microspheres of magnetic nanoparticles as described in Comparative Example 1 (injected into two groups of rats respectively); rats injected with the hydrogel microspheres of magnetic nanoparticles as described in Example 1 underwent static magnetic field intervention for 12 hours / day (denoted as SM-HA@D), one group of rats injected with the hydrogel microspheres of magnetic nanoparticles as described in Comparative Example 1 underwent static magnetic field intervention for 12 hours / day (denoted as SM-HA), and the other rat did not undergo static magnetic field intervention (denoted as M-HA). The spinal cord injury treatment results were assessed after 30 days, and histological analysis was performed. The results are as follows: Figure 4 As shown.
[0079] Note: The Sham group includes: sham surgery group; model group (SCI), magnetic nanoparticle-microsphere group (M-HA), static magnetic field combined with magnetic nanoparticle-microsphere group (SM-HA), and static magnetic field combined with magnetic nanoparticle-drug-loaded microsphere group (SM-HA@D).
[0080] from Figure 4The results show that the SCI group had the largest spinal cord injury area, with fibrosis caused by astrocyte aggregation in the cystic area. With the intervention of magnetic nanoparticles, external static magnetic field intervention, and the construction of a dual drug delivery system, the SM-HA and SM-HA@D groups showed smaller spinal cord injury areas.
[0081] Although the above embodiments have been shown and described, it is understood that the above embodiments are exemplary and should not be construed as limiting the present invention. Any changes, modifications, substitutions and variations made to the above embodiments by those skilled in the art are within the protection scope of the present invention.
Claims
1. A magnetic drug-loaded hydrogel microsphere, characterized in that, The raw materials include the following parts by weight: 1-40 parts of hydrogel crosslinking precursor, 2-30 parts of initiator, 0.1-1 parts of magnetic nanoparticles loaded with small molecule drugs, and 0-1 parts of macromolecule drugs.
2. The magnetic drug-loaded hydrogel microspheres according to claim 1, characterized in that, The hydrogel crosslinking precursor includes an ionic hydrogel crosslinking precursor and / or a photosensitive hydrogel crosslinking precursor; preferably, the ionic hydrogel crosslinking precursor includes sodium alginate; preferably, the photosensitive hydrogel crosslinking precursor includes at least one of methacrylamide hyaluronic acid, methacrylamide gelatin, methacrylamide polyvinyl alcohol, methacrylamide sodium alginate, methacrylamide silk fibroin, methacrylamide chitosan, methacrylamide glucose, methacrylamide chondroitin sulfate, and methacrylamide diacrylate. And / or, the initiator comprises an ionic initiator and / or a photoinitiator; preferably, the ionic initiator comprises calcium chloride; preferably, the photoinitiator comprises at least one of lithium phenyl (2,4,6-trimethylbenzoyl)phosphate, IRGACURE 2959 and ruthenium; And / or, the magnetic nanoparticles in the magnetic nanoparticles loaded with small molecule drugs include at least one of Fe3O4, γ-Fe2O3, MnFe2O4, NaGdF4, and Gd2O3; And / or, the small molecule drug in the magnetic nanoparticles loaded with the small molecule drug includes at least one of a natural small molecule drug, a nonsteroidal anti-inflammatory drug (NSAID), and a steroidal anti-inflammatory drug; preferably, the natural small molecule drug includes curcumin; preferably, the NSAID includes aspirin and / or indomethacin; preferably, the steroidal anti-inflammatory drug includes fluticasone and / or hydroxychloroquine; And / or, the mass ratio of magnetic nanoparticles to small molecule drugs in the magnetic nanoparticles loaded with small molecule drugs is (45-55):1; And / or, the macromolecular drug includes at least one of basic fibroblast growth factor (bFGF), epidermal growth factor (EGF), and vascular endothelial growth factor (VEGF).
3. The magnetic drug-loaded hydrogel microspheres according to claim 1, characterized in that, It also includes gel media; Preferably, the gel medium comprises phosphate buffer.
4. The magnetic drug-loaded hydrogel microspheres according to claim 1, characterized in that, The magnetic drug-loaded hydrogel microspheres have a particle size of 60-75 μm.
5. The method for preparing the magnetic drug-loaded hydrogel microspheres according to any one of claims 1-4, characterized in that, Includes the following steps: (1) The hydrogel crosslinking precursor, initiator, magnetic nanoparticles loaded with small molecule drugs and macromolecule drugs are mixed to obtain an aqueous phase; (2) Droplets were prepared by shearing the aqueous phase with the oil phase and then solidified; the oil phase was removed to obtain the magnetic drug-loaded hydrogel microspheres.
6. The method for preparing magnetic drug-loaded hydrogel microspheres according to claim 5, characterized in that, In step (2), the oil phase includes at least one of paraffin oil, edible oil, and surfactant in HFE7500; And / or, in step (2), the volume ratio of the aqueous phase to the oil phase is 1:(4-10). And / or, in step (2), the method for preparing droplets includes microfluidics.
7. The method for preparing magnetic drug-loaded hydrogel microspheres according to claim 5, characterized in that, It also includes the step of preparing magnetic nanoparticles loaded with small molecule drugs; Specifically, the following steps are included: Magnetic nanoparticles and a buffer solution containing nanoparticle modifiers are mixed to obtain functionalized magnetic nanoparticle powder; the functionalized magnetic nanoparticle powder is mixed with a small molecule drug solution, filtered, washed, and dried to obtain the magnetic nanoparticles loaded with the small molecule drug.
8. The method for preparing magnetic drug-loaded hydrogel microspheres according to claim 7, characterized in that, The buffer solution includes a Tris-HCl buffer solution with a pH of 8.5, and the concentration of the nanoparticle modifier in the buffer solution is 1-3 mg / ml. And / or, the nanoparticle modifier includes polydopamine; And / or, the mass ratio of the magnetic nanoparticles to the nanoparticle modifier is (4-6):3; And / or, the concentration of the small molecule drug solution is 1-5 mg / mL, and the solvent used for the small molecule drug solution is water and / or dimethyl sulfoxide.
9. The use of the magnetic drug-loaded hydrogel microspheres according to any one of claims 1-4 or the magnetic drug-loaded hydrogel microspheres obtained by the preparation method according to any one of claims 5-8 in the prevention or treatment of electroactive organ damage.
10. The application according to claim 9, characterized in that, The electrically active organs include the spinal cord, brain, peripheral nervous system, or heart.