A manganese-polydopamine-based organic-inorganic hybrid gel material for promoting oral oropharyngeal fistula repair and its preparation method

Abstract

The present invention discloses a manganese-polydopamine-based organic-inorganic hybrid gel material for promoting the repair of oral oropharyngeal fistulas and a preparation method thereof, the method comprising the following steps: 1) preparing manganese dioxide-coated manganese-polydopamine-based mesoporous nanocarriers by a micelle template method; 2) loading small molecule drugs and growth factors into the manganese-polydopamine-based mesoporous nanocarriers; 3) preparing an antibacterial polyethylene glycol-based gel matrix material; 4) blending and compounding the drug-loaded nanoparticles with the gel matrix to form an organic-inorganic composite gel material. The manganese-polydopamine-based organic-inorganic composite gel material provided by the present invention has significant peroxide scavenging ability, can effectively load a variety of drugs and release them in a controllable manner in response to a peroxidative microenvironment. The material is particularly suitable for the treatment of through-hole oral fistula-type wounds, can improve the wound's peroxidation and inflammatory microenvironment, achieve rapid repair of inflammatory oral oropharyngeal fistulas, and has good application prospects.

Description

Technical Field

[0001] The present invention relates to the field of novel oral biomaterials, and more particularly to a manganese-polydopamine-based organic-inorganic hybrid gel material for promoting the repair of oral oropharyngeal fistulas and a preparation method thereof. Background Art

[0002] Oropharyngeal fistulas are complex, penetrating soft tissue defects in the oropharynx caused by congenital malformations, major head and neck trauma, severe oral inflammation, post-operative tumor surgery, and radiotherapy, with an incidence rate as high as 65%. Due to the oropharynx's variable biological environment, complex anatomical structure, constant tissue movement, and continuous infiltration of saliva and food, once a fistula forms in the oropharynx, the wound surface often overflows with saliva and food, leading to recurrent bacterial and fungal inflammation and prolonged healing, significantly impacting the patient's quality of life and delaying subsequent treatment. In some patients, the spread of inflammation can lead to death. This is a malignant disease with a high morbidity, low cure rate, and significant impact. Therefore, oropharyngeal fistulas have been a problem that has plagued the field of surgical medicine for nearly a century.

[0003] However, patents targeting fistulas (particularly oral fistulas) are primarily focused on medical devices, such as drainage tubes and stents for preventing oral fistulas after surgery, as well as surgical sutures or staplers for physically closing fistula wounds. Only one invention patent uses a photosensitive hydrogel to prevent fistulas at anastomotic sites after colorectal cancer surgery. Research articles on oral fistulas also focus primarily on basic pathology, prevention, and surgical treatment. Studies addressing the closure and repair of existing oral oropharyngeal fistulas, which present complex physiological environments and severe inflammation, are scarce. Currently, clinical treatment for oral oropharyngeal fistulas relies primarily on conventional conservative interventions, primarily iodoform or Dakin's solution packing, and negative pressure wound therapy (NPWT). These conventional methods are prone to bacterial accumulation, persistent inflammation, and delayed wound healing. Furthermore, NPWT requires patient restriction of activity and may sometimes require nasogastric tube placement or gastrectomy, further complicating treatment. If treatment fails, surgical intervention, such as resection of adjacent tissue, is necessary. These methods are complex, expensive, and lengthy, often resulting in poor patient tolerance. Other clinical treatments for oral oropharyngeal fistulas include drug coating, imported collagen plugging, or complex flap transplantation, which are accompanied by problems such as poor mechanical strength of the filler, weak tissue adhesion, low biocompatibility, difficulty in degradation, high price, and large surgical trauma. At the same time, the lengthy treatment process causes secondary injuries such as malnutrition and respiratory obstruction, requiring patients to use radical methods such as gastric tube insertion, gastrostomy, and tracheotomy to ensure eating and breathing, causing additional pain to patients. Therefore, it is urgent to develop a bioactive material that can efficiently block and repair oral oropharyngeal fistulas to achieve fast, convenient, painless, and low-cost treatment of difficult-to-heal fistulas.

[0004] Among existing new biomaterials, polymer hydrogels have shown great application potential in wound closure, wound repair, and soft tissue regeneration. However, the drug burst release effect of hydrogels loaded through physical adsorption or entrapment is significant, making it difficult to maintain long-term therapeutic effects and simultaneously load multiple drugs with very different physical and chemical properties. Immobilization through covalent binding has high requirements for the types of gel and drug molecules, complex preparation, and single drug release kinetics, which can only achieve single-stage release and cannot match the multi-stage characteristics of complex wound repair. It is difficult to meet the needs of the variable pathological environment and complex treatment process of fistula diseases. The excellent drug delivery function of granular nanodrug carriers is widely used in the construction of nanodrugs. Their morphology, size and structure are highly adjustable, which can meet the needs of simultaneous loading of multiple drugs and large loading capacity. However, the uncontrolled mechanical properties and low plasticity of nanodrug carriers limit their application in wound closure.

[0005] The combination of traditional inorganic nanoparticles and polymer hydrogel materials has the following main problems: (1) The properties of inorganic nanoparticles and polymer hydrogel materials are quite different, resulting in poor material compatibility and interfacial interaction; (2) The cross-linking process of hydrogels during photocuring or thermal gelation may destroy the stability of nanoparticles; (3) The addition of inorganic nanomaterials leads to a decrease in the mechanical properties of hydrogels, making it difficult to support the movement requirements of wounds.

[0006] Therefore, it is urgent to combine the advantages of traditional inorganic nanoparticles and polymer hydrogels and overcome the above difficulties to propose a composite biomaterial that has multiple biological functions such as wound sealing, antibacterial and anti-inflammatory, and tissue regeneration. Summary of the Invention

[0007] The purpose of the present invention is to provide a manganese-polydopamine-based organic-inorganic hybrid gel material that promotes the repair of oral oropharyngeal fistulas and a preparation method thereof, thereby solving the problem in the prior art that refractory oral oropharyngeal fistulas are difficult to treat due to the complex wound microenvironment and the long and complicated process.

[0008] In order to solve the above problems, the present invention adopts the following technical solutions:

[0009] According to a first aspect of the present invention, a method for preparing a manganese-polydopamine-based organic-inorganic hybrid gel material for promoting oral oropharyngeal fistula repair is provided, comprising the following steps: S1: preparing manganese dioxide-coated manganese-polydopamine-based mesoporous nanocarriers MnPDA@MnO2 using a micelle template method; S2: loading small molecule drugs and growth factor-type macromolecular drugs into the manganese dioxide-coated manganese-polydopamine-based mesoporous nanocarriers MnPDA@MnO2 to obtain drug-loaded nanoparticles MnPDA@MnO2; S3: dissolving amino-terminated polyethylene glycol in water, adding a small amount of paraformaldehyde to the solution, and stirring to prepare a polyethylene glycol-based gel matrix material; S4: physically blending the drug-loaded nanoparticles MnPDA@MnO2 with the polyethylene glycol-based gel matrix, volatilizing and concentrating, and pressing to form a manganese-polydopamine-based organic-inorganic hybrid gel material.

[0010] Preferably, step S1 includes: S11: dissolving block copolymer surfactants Pluronic® F-127 and Pluronic® P-123 and a pore-enlarging agent 1,3,5-trimethylbenzene in water, and stirring to form a composite micelle emulsion; S12: using the composite micelle as a template, dissolving manganese chloride and dopamine molecules in the composite micelle emulsion, adding trishydroxymethylaminoethane to catalyze the polymerization reaction of dopamine, and then removing the micelle template to obtain manganese-polydopamine-based mesoporous nanoparticles; S13: dispersing the manganese-polydopamine-based mesoporous nanoparticles in water, reacting with potassium permanganate, and obtaining a manganese dioxide-coated manganese-polydopamine-based mesoporous nanocarrier MnPDA@MnO2 with a mesoporous structure inside.

[0011] Preferably, step S2 includes: S21: dispersing the mesoporous nanocarrier MnPDA@MnO2 in water, adding concentrated hydrochloric acid and sonicating to obtain activated MnPDA@MnO2; S22: dispersing the activated MnPDA@MnO2 in ethanol, reacting with (3-aminopropyl)triethoxysilane, and performing surface grafting to change the surface charge; S23: dispersing the grafted MnPDA@MnO2 nanoparticles in ethanol, adding small molecule drugs at a certain mass ratio, stirring and incubating, to obtain nanoparticles MnPDA@MnO2 loaded with small molecule drugs; S24: dispersing the nanoparticles MnPDA@MnO2 loaded with small molecule drugs again in phosphate buffer, adding growth factor-type macromolecular drugs at a certain mass ratio, stirring and incubating, and obtaining drug-loaded nanoparticles MnPDA@MnO2 after centrifugation.

[0012] Preferably, in step S11, the mass ratio of Pluronic® F-127 to Pluronic® P-123 is 1:3-3:1. It should be understood that the use of surfactants and the appropriate ratio are key to the synthesis of mesoporous structures. In a preferred embodiment, the mass ratio of the two is 3:1.

[0013] Preferably, in step S12, the amount of tris(hydroxymethylaminoethane) added may be 1 mg / mL-4 mg / mL. In a preferred embodiment, the amount of tris(hydroxymethylaminoethane) added is 3 mg / mL.

[0014] Preferably, in step S12, the mass ratio of manganese chloride tetrahydrate to dopamine molecules is 1.8:6-1:6. In a preferred embodiment, the mass ratio of manganese chloride tetrahydrate to dopamine molecules is 1:6. Exceeding this preferred range may result in a polymerization rate that is too slow or too fast, affecting the mesoporous structure of the support material.

[0015] Preferably, the extraction in step S12 is ultrasonically cleaned several times using N,N-dimethylformamide as a template remover, and then repeatedly cleaned and centrifuged several times using ethanol and water as cleaning agents. In a preferred embodiment, ultrasonically cleaned three times using N,N-dimethylformamide as a template remover, and then repeatedly cleaned and centrifuged three times using ethanol and deionized water as cleaning agents.

[0016] Preferably, in step S13, the mass ratio of potassium permanganate to manganese-polydopamine-based mesoporous nanoparticles can be 10:1-2.5:1. In a preferred embodiment, the mass ratio of potassium permanganate to manganese-polydopamine-based mesoporous nanoparticles is 5:1. Exceeding this preferred range, too low a mass ratio will result in uneven coating of the manganese dioxide shell, affecting the subsequent activity of catalase-like enzymes. Too high a mass ratio will result in clogging of the mesoporous structure of the mesoporous manganese dopamine, affecting the subsequent activity of superoxide dismutase-like enzymes.

[0017] Preferably, the small molecule drug in step S23 may be quercetin, 5-fluorouracil, dexamethasone or curcumin. In a preferred embodiment, quercetin is used as the small molecule drug.

[0018] Preferably, the growth factor macromolecular drug in step S24 can be basic fibroblast growth factor, epidermal growth factor or transforming growth factor-α. In a preferred embodiment, basic fibroblast growth factor is used as the loaded growth factor.

[0019] Preferably, the molecular weight of the amino-terminated polyethylene glycol in step S3 is 1, 2, 5, 10, or 20 kDa. In a preferred embodiment, the molecular weight of the amino-terminated polyethylene glycol is 2 kDa. It should be understood that the lower the molecular weight of the amino-terminated polyethylene glycol used, the better the antibacterial properties of the hydrogel. However, the gel prepared with a molecular weight of 1 kDa has poor formability. The most preferred molecular weight is 2 kDa.

[0020] Preferably, in step S4, the concentration of the drug-loaded MnPDA@MnO2 nanoparticles in the gel matrix is 0.50%-2.58%. In a preferred embodiment, the concentration of the drug-loaded MnPDA@MnO2 nanoparticles in the gel matrix is 0.50%. Exceeding this preferred range, too low a concentration can reduce the anti-inflammatory and repair properties of the composite material, while too high a concentration can lead to decreased mechanical properties of the gel.

[0021] Preferably, the water used in the preparation method is deionized water.

[0022] According to a preferred embodiment of the present invention, the preparation method of the manganese-polydopamine-based organic-inorganic hybrid gel material comprises the following steps: 1) dissolving polyoxypropylene-polyoxyethylene copolymer and pore-expanding agent 1,3,5-trimethylbenzene in water and stirring to form a uniform composite micelle emulsion; 2) using the composite micelles synthesized in step 1) as a template, dissolving manganese chloride tetrahydrate and dopamine molecules in the composite micelle solution formed in step 1), adding trishydroxymethylaminoethane to catalyze the polymerization reaction of dopamine, and cross-linking it with manganese ions to form manganese-polydopamine replica micelle aggregates. The mesoporous manganese-polydopamine nanoparticles are obtained by dispersing the mesoporous manganese-polydopamine nanoparticles obtained in step 2) in water, reacting with potassium permanganate and stirring to obtain manganese dioxide-coated manganese-polydopamine mesoporous nanocarriers MnPDA@MnO2 with a mesoporous structure inside the particles and a manganese dioxide shell on the outside; 4) the mesoporous nanocarriers MnPDA@MnO2 obtained in step 3) are dispersed in water, concentrated hydrochloric acid is added and ultrasonicated to obtain activated MnPDA@MnO2. A@MnO2; 5) dispersing the activated MnPDA@MnO2 in ethanol, reacting with (3-aminopropyl)triethoxysilane and stirring to perform surface grafting to change the surface charge; 6) dispersing the grafted nanoparticles obtained in step 5) in ethanol, adding small molecule drugs at a mass ratio of 4:5, stirring and incubating for several hours to obtain nanoparticles MnPDA@MnO2 loaded with small molecule drugs; 7) dispersing the drug-loaded nanoparticles obtained in step 6) in phosphate buffer again, adding growth factor macromolecular drugs at a mass ratio of 1:1, and incubating The mixture was stirred for several hours and centrifuged to obtain drug-loaded nanoparticles MnPDA@MnO2; 8) amino-terminated polyethylene glycol was dissolved in water, a small amount of paraformaldehyde was added to the solution, and the solution was stirred at 70°C for 1 hour to obtain a polyethylene glycol-based hydrogel matrix having a dynamic covalent aminal cross-linked network; 9) a certain amount of drug-loaded nanoparticles MnPDA@MnO2 prepared in step 7) was physically blended with the polyethylene glycol hydrogel matrix prepared in step 8), and the obtained mixture was evaporated and concentrated, and a film was pressed in a mold to form a manganese-polydopamine-based organic-inorganic hybrid gel material.

[0023] According to a second aspect of the present invention, there is provided a manganese-polydopamine-based organic-inorganic hybrid gel material for promoting oral oropharyngeal fistula repair prepared according to the above preparation method.

[0024] The polyethylene glycol-based gel matrix synthesized in the present invention is simple to synthesize, can be repeatedly reshaped, and can gradually degrade and release formaldehyde in response to a peroxide solution environment, thus exhibiting antibacterial properties. In a preferred embodiment, the manganese-polydopamine-based organic-inorganic hybrid gel material synthesized in accordance with the present invention exhibits significant bactericidal effects against Staphylococcus aureus and Escherichia coli.

[0025] According to a third aspect of the present invention, there is also provided a use of a manganese-polydopamine-based organic-inorganic hybrid gel material in the preparation of a product for repairing inflammatory oral fistula wounds.

[0026] The main point of the invention is that such a manganese-polydopamine-based porous carrier material is synthesized for the first time. The inorganic components and the organic components can be connected by coordination and hydrogen bonding. Good compatibility can be achieved by simple blending without obvious stratification. The blending and shaping process conditions are mild and will not affect the nanomaterials and the loads. The gel material and the inorganic nanomaterial jointly achieve multi-level responsiveness. It has good adhesion and does not fall off under movement conditions.

[0027] The manganese-polydopamine-based mesoporous nanocarrier (MnPDA) nanoparticles synthesized according to the present invention exhibit regular spherical shapes ranging from approximately 160 nm to 200 nm, with clearly visible irregular open pores. The mesoporous MnPDA exhibits distinct worm-like open pores with a pore size distribution of approximately 17 nm. The resulting manganese dioxide-coated MnPDA@MnO2 nanocarrier retains its spherical morphology, with a large number of dendritic shells covering the spheres and pores. After coating with manganese dioxide, the particle size significantly increases to 250 nm, while the pore size decreases to 11 nm.

[0028] The manganese dioxide-coated manganese-polydopamine-based mesoporous nanoparticles (MnPDA@MnO2) synthesized according to the present invention possess open, worm-like pores. These nanoparticles can be loaded with both small-molecule and macromolecular drugs. In a preferred embodiment, they can be loaded with the small-molecule drug quercetin, with a loading rate exceeding 20%. They can also be loaded with basic fibroblast growth factor, with a loading rate exceeding 15%.

[0029] The present invention provides a manganese dioxide-coated manganese-polydopamine-based mesoporous nanocarrier. The porous manganese-polydopamine-based mesoporous nanocarrier is a first-of-its-kind synthetic material that can simultaneously possess the functions of a manganese-polydopamine skeleton and a guest drug loading function, thereby achieving the purpose of integrating multiple functions. Currently, no similar technology has been reported.

[0030] The present invention provides a manganese dioxide-coated manganese-polydopamine-based mesoporous nanocarrier. The manganese dioxide-coated manganese-polydopamine-based mesoporous nanocarrier MnPDA@MnO2 has a manganese-polydopamine skeleton component and a manganese dioxide coating layer with obvious catalase activity (CAT enzyme) and superoxide dismutase (SOD enzyme) activity. It should be understood that it can effectively decompose peroxides, reverse the peroxidative microenvironment at the wound, and alleviate the inflammatory response at the wound.

[0031] The present invention provides a manganese dioxide-coated manganese-polydopamine-based mesoporous nanoparticle carrier. The manganese-polydopamine coordination network of the MnPDA@MnO2 mesoporous nanoparticles is capable of cleaving in the presence of hydrogen peroxide. It should be understood that the carrier has the ability to release the loaded drug in response to hydrogen peroxide. In a preferred embodiment, the MnPDA@MnO2 mesoporous nanoparticles slowly cleave and release the loaded drug in a 10mM hydrogen peroxide solution.

[0032] The manganese-polydopamine-based organic-inorganic hybrid gel material for promoting oral oropharyngeal fistula repair provided by the present invention has the following beneficial effects compared with the prior art:

[0033] The manganese dioxide-coated manganese-polydopamine mesoporous drug carrier prepared according to the present invention features uniform, small particle size, and excellent dispersibility. It also exhibits catalytic activity in the CAT and SOD cascades, enabling significant scavenging of peroxides and superoxide species from the wound microenvironment, reversing the inflammatory microenvironment and promoting wound repair. Its worm-like open-pore mesoporous structure allows for loading a wide range of small molecule drugs and growth factor-based drugs. Furthermore, the material is peroxide-responsive, cleaving in response to the peroxidative microenvironment at the wound site, releasing the loaded drugs and achieving diverse biological functions. Cellular experiments demonstrated that the manganese dioxide-coated manganese-polydopamine mesoporous drug carrier, loaded with quercetin and basic fibroblast growth factor, significantly scavenged ROS levels in oxidatively stressed HUVEC cells, promoted polarization of RAW264.7 macrophages toward an anti-inflammatory phenotype, and alleviated both cellular oxidative stress and inflammation.

[0034] 2) The present invention uses manganese dioxide-coated manganese-polydopamine mesoporous drug carriers and polyethylene glycol-based gel matrix to composite a manganese-polydopamine-based organic-inorganic hybrid gel material, and achieves the treatment of oral fistula-like wounds through the synergistic effect of the two. Only by using a simple blending method, the uniform distribution of nanoparticles in the gel matrix can be achieved. The resulting hybrid gel material has good plasticity, adhesion and self-healing properties, and integrates multiple properties such as antibacterial, antioxidant, anti-inflammatory and tissue growth promotion. In vitro cell and antibacterial experiments have shown that the material has good cell compatibility and antibacterial effects; animal experiments have shown that the hybrid gel material can not only effectively promote the repair of inflammatory oral fistula wounds in New Zealand rabbits, significantly shortening the repair time compared to the control group, but also has good biocompatibility and can effectively reduce the toxic and side effects produced during the treatment process.

[0035] In summary, the manganese-polydopamine-based organic-inorganic hybrid gel material for oral oropharyngeal fistula repair provided by the present invention has significant peroxide scavenging capabilities, can effectively load multiple drugs, and controllably releases them in response to a peroxidative microenvironment. This material is particularly suitable for treating penetrating oral fistula-type wounds, improving the wound's peroxidative and inflammatory microenvironment and enabling rapid repair of inflammatory oral oropharyngeal fistulas. It holds great promise for the treatment of oral oropharyngeal fistulas. BRIEF DESCRIPTION OF THE DRAWINGS

[0036] Figure 1 A flow chart of the synthesis of manganese dioxide-coated manganese-polydopamine-based mesoporous nanoparticles MnPDA@MnO2 according to the present invention;

[0037] Figure 2 This is a flow chart for the synthesis of a manganese-polydopamine-based organic-inorganic hybrid gel material for promoting oral oropharyngeal fistula repair according to the present invention;

[0038] Figure 3 This is a transmission electron microscopy image of the manganese-polydopamine mesoporous carrier MnPDA prepared according to Example 1 of the present invention;

[0039] Figure 4 This is a scanning electron micrograph of the manganese-polydopamine mesoporous carrier MnPDA prepared according to Example 1 of the present invention;

[0040] Figure 5 This is the nitrogen adsorption spectrum of the manganese-polydopamine mesoporous carrier MnPDA prepared in Example 1 of the present invention;

[0041] Figure 6 This is a transmission electron microscopy image of the manganese dioxide-coated manganese-polydopamine mesoporous carrier MnPDA@MnO2 prepared according to Example 1 of the present invention;

[0042] Figure 7This is a scanning electron micrograph of the manganese dioxide-coated manganese-polydopamine mesoporous carrier MnPDA@MnO2 prepared according to Example 1 of the present invention;

[0043] Figure 8 This is the nitrogen adsorption spectrum of the manganese dioxide-coated manganese-polydopamine mesoporous carrier MnPDA@MnO2 prepared in Example 1 of the present invention;

[0044] Figure 9 This is a digital photo of the manganese-polydopamine-based organic-inorganic hybrid gel material prepared in Example 1 of the present invention attached to a rubber glove accompanied by finger movement;

[0045] Figure 10 (a) SEM photograph and (b) energy dispersion spectrum element distribution diagram of the manganese-polydopamine-based organic-inorganic hybrid gel material prepared in Example 1 of the present invention;

[0046] Figure 11 Graph showing the antibacterial effects of the pure gel matrix without drug-loaded nanoparticles and the manganese-polydopamine-based organic-inorganic hybrid gel material on Staphylococcus aureus and Escherichia coli plated according to Example 1 of the present invention;

[0047] Figure 12 These are digital photographs of wound healing on days 0-12 of inflammatory oral fistula wounds in New Zealand rabbits treated with the pure gel matrix without drug-loaded nanoparticles prepared in Example 1 of the present invention and the manganese-polydopamine-based organic-inorganic hybrid gel material;

[0048] Figure 13 This is a transmission electron microscopy image of the manganese-polydopamine mesoporous carrier MnPDA prepared according to Example 2 of the present invention;

[0049] Figure 14 This is a transmission electron microscopy image of the manganese-polydopamine mesoporous carrier MnPDA prepared according to Example 3 of the present invention;

[0050] Figure 15 This is a transmission electron microscopy image of the manganese dioxide-coated manganese-polydopamine mesoporous carrier MnPDA@MnO2 prepared according to Example 4 of the present invention;

[0051] Figure 16 This is a transmission electron microscopy image of the manganese dioxide-coated manganese-polydopamine mesoporous carrier MnPDA@MnO2 prepared according to Example 5 of the present invention;

[0052] Figure 17 (a) Scanning electron microscope photograph of the manganese-polydopamine-based organic-inorganic hybrid gel material prepared in Example 6 of the present invention and (b) the corresponding energy dispersion spectrum element distribution diagram. DETAILED DESCRIPTION

[0053] The following is a preferred embodiment of the present invention with reference to the accompanying drawings to further illustrate the present invention. It should be understood that the following embodiments are only used to illustrate the present invention and are not intended to limit the scope of the present invention.

[0054] According to the present invention, a method for preparing a manganese-polydopamine-based organic-inorganic hybrid gel material for promoting oral oropharyngeal fistula repair is provided, the method comprising the steps of: 1) preparing manganese dioxide-coated manganese-polydopamine-based mesoporous nanocarriers by a micelle template method, the synthesis process of which is as follows: Figure 1 As shown in the figure; 2) loading small molecule drugs and growth factors into manganese dioxide-coated manganese-polydopamine-based mesoporous nanoparticles; 3) preparing polyethylene glycol-based gel matrix materials; 4) blending the drug-loaded manganese dioxide-coated manganese-polydopamine-based mesoporous nanoparticles with the polyethylene glycol gel matrix to form an organic-inorganic composite gel material. The overall design process of this preparation method is shown in the figure. Figure 2 shown.

[0055] Example 1

[0056] 1.1 Preparation of a manganese-polydopamine-based organic-inorganic hybrid gel material for oral oropharyngeal fistula repair

[0057] The following steps are involved:

[0058] (1) Take 4 mL of anhydrous ethanol and 6 mL of deionized water and mix them in a beaker. Add 75 mg of Pluronic® F-127 and 25 mg of Pluronic® P-123 to the above solution and stir to dissolve. Then adjust the speed to 800 rpm and add 0.4 mL of 1,3,5-trimethylbenzene and stir to form an emulsion. After resuming the speed, add 15 mg of dopamine and 2.5 mg of manganese chloride tetrahydrate and stir to dissolve. Finally, add 30 mg of tris(hydroxymethyl)aminoethane) and continue the reaction under magnetic stirring for 3 h. Collect the product by high-speed centrifugation for 15 minutes (13,000 rpm) and wash it three times with ethanol and water to remove the residual reactants. The obtained product is dissolved in N,N-dimethylformamide and ultrasonically washed three times to remove the micelle template. After washing it three times with ethanol and water, the product manganese-polydopamine mesoporous nanocarrier MnPDA is obtained.

[0059] like Figure 3 and Figure 4 As shown, the manganese-polydopamine-based mesoporous nanoparticles MnPDA nanoparticles are regular spheres with a size of about 160nm-200nm, and their irregular open pores are clearly visible. According to the nitrogen adsorption spectrum (such as Figure 5 Calculations show that the pore size distribution is approximately around 17 nm, and nitrogen adsorption data show that the mesoporous MnPDA exhibits obvious worm-like open pores.

[0060] (2) Dissolve 10 mg of MnPDA prepared in step (1) in 4 mL of deionized water, then add 50 mg of potassium permanganate powder and stir. React for 1 h. Collect the product by high-speed centrifugation for 15 min (13,000 rpm) and wash it three times with ethanol and deionized water to remove the residual reactants to obtain the product, manganese dioxide-coated manganese-polydopamine mesoporous nanocarrier MnPDA@MnO2.

[0061] like Figure 6 , Figure 7 As shown in the figure, the manganese dioxide-coated manganese-polydopamine-based mesoporous nanocarrier MnPDA@MnO2 still maintains its spherical morphology, and the surface of the sphere and the pore surface are covered with a large number of dendritic shells. After coating with manganese dioxide, its particle size increases significantly to 250nm. According to the nitrogen adsorption spectrum (such as Figure 8 Calculation shows that the pore size is reduced to 11 nm.

[0062] (3) Disperse 50 mg of the manganese dioxide-coated manganese-polydopamine mesoporous nanocarriers (MnPDA@MnO2) prepared in step (2) in 10 mL of deionized water. After thorough dissolution, add 826 μL of concentrated hydrochloric acid (12 M) and activate by sonication for 15 min. Collect the product by high-speed centrifugation for 15 min (13,000 rpm) and wash it three times with ethanol and water to remove residual reactants.

[0063] (4) The MnPDA@MnO2 prepared in step (3) was redispersed in anhydrous ethanol (10 mL). 1 mL of the product solution was taken out and diluted to 15 mL with anhydrous ethanol. After stirring evenly, APTES (60 μL) was added and reacted at 80 °C for 4 h. The product was collected by high-speed centrifugation for 15 min (13,000 rpm) and washed alternately with ethanol and water several times to remove residual reactants.

[0064] (5) Take 8 mg of the MnPDA@MnO2 powder prepared in step (4) and 10 mg of quercetin and evenly disperse them in 25 mL of anhydrous ethanol. After stirring for 24 hours, high-speed centrifugation for 15 minutes (13000 rpm) was performed to collect the product to obtain drug-loaded MnPDA@MnO2 nanoparticles loaded with quercetin drug.

[0065] (6) Take 10 mg of the nanoparticles obtained in step (5) and 10 mg of basic fibroblast growth factor and disperse them in 10 mL of phosphate buffer solution with pH = 7.4. After stirring for 24 hours, the product was collected by high-speed centrifugation for 15 minutes (13000 rpm) to obtain drug-loaded nanoparticles MnPDA@MnO2 further loaded with basic fibroblast growth factor.

[0066] (7) 1 g of amino-terminated polyethylene glycol (Mw = 2 kDa) (CAS No.: 24991-53-5, Shanghai Yien Chemical Technology Co., Ltd., Model: R011666-5 g) was added to ultrapure water (10 mL) and stirred. 30 mg of paraformaldehyde was then added to the mixture, and the solution was stirred at 70 °C for 1 h to obtain a polyethylene glycol-based hydrogel matrix with a dynamic covalent aminal cross-linked network.

[0067] (8) The hydrogel matrix prepared in step (7) was physically mixed with 0.0214 g of the drug-loaded nanoparticles MnPDA@MnO2 obtained in step (6), and stirred with magnetic stirring for 10 minutes. The resulting mixture was then evaporated and concentrated to a water content of 76%, and pressed into a mold to form 4.28 g of a manganese-polydopamine-based organic-inorganic hybrid gel material for promoting oral oropharyngeal fistula repair.

[0068] like Figure 9 As shown in Figure 2, the gel material has good adhesion and does not fall off under sports conditions. Figure 10 As shown in (a), the low-magnification SEM image shows that it has good porosity and no obvious inorganic particle agglomeration phenomenon; the element distribution diagram of the scanning image shows that the yellow manganese element is evenly distributed in the matrix, indicating that the inorganic nanomaterial MnPDA@MnO2 has good dispersion in the hydrogel matrix. Figure 10 As shown in (b), in the manganese-polydopamine-based organic-inorganic hybrid gel material prepared according to the present invention, the inorganic components and the organic components can be connected by coordination and hydrogen bonding. Good compatibility can be achieved by simply blending without obvious stratification. The blending and shaping process conditions are mild and will not affect the nanomaterials and the loads. The gel material and the inorganic nanomaterials jointly achieve multi-level responsiveness.

[0069] 1.2 Antibacterial performance testing

[0070] The antibacterial properties of the hydrogels and composite gels obtained in steps (7) and (8) of 1.1 were tested using the plate coating method. Staphylococcus aureus (S. aureus) was selected as a representative of Gram-positive bacteria, and Escherichia coli (E. coli) was selected as a representative of Gram-negative bacteria. The bacterial suspension and the test sample solution were thoroughly mixed and incubated in a constant temperature shaker (37°C, 200 rpm) for 9 hours. The absorbance at a wavelength of 600 nm was measured (blank group: bacterial suspension + PBS buffer; sample group: bacterial suspension + test sample solution).

[0071] On the agar plate of antibacterial experiment ( Figure 11 ) It can be seen intuitively that compared with the blank group, both the pure gel group and the composite gel group showed better antibacterial effects, and the incorporation of nanomaterials did not affect the performance of the hydrogel.

[0072] 1.3 Wound healing experiment using New Zealand white male rabbits as model animals

[0073] Thirty-nine male New Zealand White rabbits, approximately 16 weeks old (2.8-3.5 kg), were used as model animals. The left cheek was shaved and disinfected, and a 1.5 cm × 1.5 cm circular incision was made for each rabbit. The anatomical incision line was positioned midway between the anterior margin of the submandibular muscle and the posterior margin of the canine tooth to minimize excessive intraoperative bleeding. A through-cutaneous defect (from skin to mucosa) was created after incision. Staphylococcus aureus was applied to the incision site, and subsequent repair experiments were initiated when purulent discharge from the wound developed.

[0074] The wound sites of each rabbit were photographed and then treated in three different ways: (1) blank group (control group); (2) pure gel group; and (3) composite gel group. The materials were changed every 2 days after surgery, while the blank group underwent simple wound cleaning. In all cases, the rabbits' recovery, body weight, and food intake were closely monitored after surgery. Three rabbits in each group were euthanized on the 4th, 8th, and 12th days after surgery. Before euthanasia, the wound recovery was observed and photographed, and bacterial culture was performed to analyze the differences between the groups. Blood samples were then collected for analysis. After euthanasia, tissue specimens were collected for analysis.

[0075] In the wound healing experiments at different time periods (4th, 8th and 12th days) ( Figure 12 ), the wound healing speed of the composite gel group was significantly faster, and even basically achieved complete healing on the 12th day, and the wound closure rate reached about 95%, which was significantly higher than that of the blank group.

[0076] Example 2

[0077] The implementation method and basic formula were the same as those in step (1) of Example 1, except that 50 mg of Pluronic® F-127 and 50 mg of Pluronic® P-123 were used as templates to similarly form manganese-polydopamine mesoporous nanocarriers MnPDA having a mesoporous structure.

[0078] Figure 13 This is a transmission electron microscope photograph of Example 2. The obtained MnPDA nanoparticles are regular spherical in shape with a size of about 160 nm to 200 nm, and irregular open pores are clearly visible.

[0079] Example 3

[0080] The implementation method and basic formula were the same as those in step (1) of Example 1, except that 25 mg of Pluronic® F-127 and 75 mg of Pluronic® P-123 were used as templates to similarly form manganese-polydopamine mesoporous nanocarriers MnPDA having a mesoporous structure.

[0081] Figure 14 This is a transmission electron microscope photograph of Example 3. The obtained MnPDA nanoparticles are regular spherical in shape with a size of about 160 nm to 200 nm, and irregular open pores are clearly visible.

[0082] Example 4

[0083] The implementation method and basic formula are the same as those in step (2) of Example 1, except that the amount of potassium permanganate is changed to 100 mg, and manganese dioxide-coated manganese-polydopamine mesoporous nanocarrier MnPDA@MnO2 is also formed.

[0084] Figure 15 This is a transmission electron microscope photograph of Example 4. The particle size of the obtained MnPDA@MnO2 nanoparticles is increased to above 250nm, and there are many dendritic shells covering the spherical surface and the pore surface.

[0085] Example 5

[0086] The implementation method and basic formula are the same as those in step (2) of Example 1, except that the amount of potassium permanganate is changed to 25 mg, and manganese dioxide-coated manganese-polydopamine mesoporous nanocarriers MnPDA@MnO2 are also formed.

[0087] Figure 16 This is a transmission electron microscope photograph of Example 5. The particle size of the obtained MnPDA@MnO2 nanoparticles is increased to more than 200 nm, and the dendritic shell covering the spherical surface and the pore surface is relatively small.

[0088] Example 6

[0089] The implementation method and basic formula were the same as those in step (8) of Example 1, except that the doping amount of MnPDA@MnO2 nanoparticles was changed to 0.1128 g, and 4.37 g of manganese-polydopamine-based organic-inorganic hybrid gel material for promoting oral oropharyngeal fistula repair was obtained.

[0090] Figure 17 The scanning electron microscope photograph of Example 6 and the corresponding energy dispersion spectrum element distribution diagram show that the obtained composite gel has good porosity and no obvious particle agglomeration phenomenon. The yellow manganese element is evenly distributed in the matrix, indicating that the inorganic nanomaterial MnPDA@MnO2 has good dispersibility in the hydrogel matrix.

[0091] The above description is merely a preferred embodiment of the present invention and is not intended to limit the scope of the present invention. Various modifications are possible. Any simple, equivalent changes and modifications made in accordance with the claims and description of the present invention are within the scope of protection of the patent claims. Anything not fully described in this invention is conventional technology.

Claims

1. A method for preparing a manganese-polydopamine-based organic-inorganic hybrid gel material for promoting the repair of oral oropharyngeal fistula, characterized in that: The following steps are involved: S1: Preparation of MnO2-coated manganese-polydopamine-based mesoporous nanocarriers MnPDA@MnO2 by micelle template method; S2: loading small molecule drugs and growth factor-type macromolecular drugs into the manganese dioxide-coated manganese-polydopamine-based mesoporous nanocarrier MnPDA@MnO2 to obtain drug-loaded nanoparticles MnPDA@MnO2; S3: dissolving amino-terminated polyethylene glycol in water, adding paraformaldehyde to the solution, and stirring to prepare a polyethylene glycol-based gel matrix material; S4: Physically blending the drug-loaded nanoparticles MnPDA@MnO2 with the polyethylene glycol-based gel matrix material, volatilizing and concentrating, and pressing the film to form a manganese-polydopamine-based organic-inorganic hybrid gel material.

2. The preparation method according to claim 1, characterized in that Step S1 includes: S11: Dissolve the block copolymer surfactants Pluronic® F-127 and Pluronic® P-123 and the pore-enlarging agent 1,3,5-trimethylbenzene in water and stir to form a composite micellar emulsion; S12: using the composite micelle as a template, dissolving manganese chloride and dopamine molecules in the composite micelle emulsion, adding tris(hydroxymethylaminoethane) to catalyze the polymerization reaction of dopamine, and then removing the micelle template to obtain manganese-polydopamine-based mesoporous nanoparticles; S13: dispersing the manganese-polydopamine-based mesoporous nanoparticles in water and reacting with potassium permanganate to obtain a manganese dioxide-coated manganese-polydopamine-based mesoporous nanocarrier MnPDA@MnO2 with a mesoporous structure inside.

3. The preparation method according to claim 1, characterized in that Step S2 includes: S21: dispersing the mesoporous nanocarrier MnPDA@MnO2 in water, adding concentrated hydrochloric acid and performing sonication to obtain activated MnPDA@MnO2; S22: The activated MnPDA@MnO2 was dispersed in ethanol and reacted with (3-aminopropyl)triethoxysilane to perform surface grafting to change the surface charge; S23: dispersing the grafted MnPDA@MnO2 nanoparticles in ethanol, adding a small molecule drug at a certain mass ratio, stirring and incubating, and obtaining nanoparticles MnPDA@MnO2 loaded with small molecule drugs; S24: The MnPDA@MnO2 nanoparticles loaded with small molecule drugs are dispersed again in phosphate buffer, and growth factor-type macromolecular drugs are added at a certain mass ratio, stirred and co-incubated, and the drug-loaded nanoparticles MnPDA@MnO2 are obtained after centrifugation.

4. The preparation method according to claim 2, characterized in that In step S11, the mass ratio of Pluronic® F-127 to Pluronic® P-123 is 1:3-3:

1.

5. The preparation method according to claim 2, characterized in that In step S12, the amount of tris(hydroxymethyl)aminoethane added is 1 mg / mL-4 mg / mL, and the mass ratio of manganese chloride tetrahydrate to dopamine molecules is 1.8:6-1:6; the micelle template is removed by extraction, and ultrasonic cleaning is performed several times using N,N-dimethylformamide as a template remover, and then repeated cleaning and centrifugation are performed several times using ethanol and water as cleaning agents.

6. The preparation method according to claim 2, characterized in that In the step S13, the mass ratio of potassium permanganate to manganese-polydopamine-based mesoporous nanoparticles is 10:1-2.5:

1.

7. The preparation method according to claim 3, characterized in that In step S23, the small molecule drug is quercetin, 5-fluorouracil, dexamethasone or curcumin; the feeding mass ratio of MnPDA@MnO2 nanoparticles to small molecule drugs is 1:1-1:2; in step S24, the growth factor-type macromolecular drug is basic fibroblast growth factor, epidermal growth factor or transforming growth factor-α; the feeding mass ratio of nanoparticles MnPDA@MnO2 to growth factor-type macromolecular drugs is 1:1-1:

2.

8. The preparation method according to claim 1, characterized in that In step S4, the mass concentration of the drug-loaded nanoparticles MnPDA@MnO2 in the gel matrix is 0.50%-2.58%.

9. A manganese-polydopamine-based organic-inorganic hybrid gel material for promoting the repair of oral oropharyngeal fistula obtained by the preparation method according to any one of claims 1 to 8.

10. Use of the manganese-polydopamine-based organic-inorganic hybrid gel material for promoting oral oropharyngeal fistula repair according to claim 9 in the preparation of a product for repairing oral oropharyngeal fistula.

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