Three-stage amplification piezoelectric bioengineering patch for repairing airway injury

Abstract

The invention relates to the field of bioengineering, in particular to a three-stage amplification piezoelectric bioengineering patch for repairing airway injury and a preparation method of the three-stage amplification piezoelectric bioengineering patch. The novel three-stage amplification piezoelectric bioengineering patch capable of being used for repairing the airway injury is constructed, so that the airway epithelium injury can be stably repaired, and the regeneration of the injury part can be accelerated. Aiming at a complex environment in an airway, the patch can resist bacteria and infection, can be easily and firmly attached to the inner surface of the trachea to provide a viscous environment, and is combined with piezoelectric reaction to reconstruct a damaged environment and guide an organism to repair a damaged epithelium.

Description

Technical Field

[0001] The invention relates to the field of bioengineering, in particular to a three-stage amplification piezoelectric bioengineering patch for repairing airway injuries. Background Art

[0002] The luminal surface of the human trachea is covered with pseudostratified ciliated columnar epithelial mucosa, which constitutes an important barrier in the human respiratory portal and plays a vital role in the respiratory system. It is not only a portal that directly contacts the outside world, but also the first line of defense against harmful substances from the outside world, and against the influence of various acid-base, pathogens and other injury-causing factors. However, the tracheal epithelial barrier is susceptible to mechanical damage during clinical operations such as endotracheal intubation, tracheotomy, and lung transplantation, resulting in a decrease in local defense capabilities and causing serious complications. In a complex airway environment, infection and inflammatory reactions in the trachea not only aggravate epithelial damage, but may also lead to tracheal softening, sputum embolism, fibrosis, and even fatal fistulas and stenosis. Therefore, how to effectively repair and reconstruct damaged airway epithelium has become a major challenge in clinical treatment.

[0003] There are currently many schemes to design various devices to promote the repair of airway epithelium, including drugs, stem cell transplantation and other methods. The use of drugs such as antibiotics and glucocorticoids can reduce local epithelial damage, but the efficacy is not certain when facing severe epithelial damage. Although stem cell transplantation has shown great application prospects in some clinical studies, its safety and ethical issues still need to be studied and discussed. In addition, these methods are susceptible to factors such as infection in a complex airway environment, and the efficacy is not stable. In recent years, with the development of the field of biomaterials, the strategy of biomaterial patches has brought new hope. By covering the damaged area with biomaterial patches, the damaged environment is reconstructed, local regeneration is promoted, and the repair of epithelial structure and reconstruction of function are achieved. In particular, with the deepening of the understanding of the endogenous electric field of organisms, bioelectric materials are increasingly being used in tissue repair.

[0004] With the discovery of endogenous electric fields and cell transmembrane potentials in organisms, the potential of electrical stimulation in injury repair and functional reconstruction has become increasingly prominent. Further practice has found that biological electric fields can affect a variety of life activities such as cell migration, adhesion, proliferation, and differentiation, revealing that simulated bioelectric stimulation plays a good role in promoting biological cell growth and differentiation and tissue regeneration and repair. As for damage to organ tissues in the body, bioengineering scaffolds based on piezoelectric materials can produce potential changes by relying on tiny deformations caused by squeezing and stretching of materials by the organism without an external power supply, simulating physiological electrical stimulation to guide the body to repair and regenerate tissues, showing great application prospects. However, the use of electrical stimulation as a method of intervention in regeneration is still rare in the field of tracheal epithelial injury, and solutions without external power supply are rarely used. Therefore, it is still necessary to develop an electrical intervention solution that does not require an external power supply. Summary of the invention

[0005] The purpose of the present invention is to solve the limitations of existing treatment methods in the repair of airway epithelial damage, especially in terms of stability and promoting regeneration, and to provide a new three-stage amplification piezoelectric bioengineering patch that can be used for airway injury repair and its preparation method.

[0006] The first aspect of the present invention provides a method for preparing a three-stage amplification piezoelectric bioengineering patch for airway injury repair, comprising the following steps:

[0007] (A) Synthesis of barium titanate nanoparticles: Tetrabutyl titanate and barium nitrate are mixed in ethanol in an appropriate ratio (the molar ratio of the three substances is 1:1:10-1:1:20); under stirring conditions, gradually heated to 60-80°C to perform hydrolysis and polymerization reactions to form a sol; continued heating until gelation to form a solid gel; calcined the gel at 600-800°C for 2-4 hours to obtain barium titanate nanoparticles;

[0008] (B) Preparation of nanoclay flakes: Clay mineral (montmorillonite or kaolin) and deionized water are mixed at a mass ratio of 1:10-1:50, and treated using ultrasonic exfoliation technology for 3-6 hours; nanoclay flakes are collected by centrifugation;

[0009] (C) Synthesis of silver nanowires: Mix a silver nitrate solution and a reducing agent (sodium citrate) solution, add an appropriate amount of polyvinyl pyrrolidone (PVP) as a stabilizer (the molar ratio of the three substances is 1:2:0.1-1:2:0.5); stir at room temperature for 4-8 hours to reduce the silver ions to silver nanowires; collect the silver nanowires by centrifugation, and wash with deionized water to remove excess reducing agent and stabilizer;

[0010] (D) Preparation of polyacrylic acid substrate: polyacrylic acid monomer, initiator (potassium persulfate) and an appropriate amount of deionized water are mixed in a volume ratio of 10:1:100-10:1:200; polymerization reaction is carried out at 60-70° C. for 3-6 hours to form a polyacrylic acid substrate;

[0011] (E) Assembling gel: mixing barium titanate nanoparticles, nanoclay sheets and silver nanowires in a mass ratio of 1:1:1-1:2:1 to form a composite; uniformly dispersing the composite in a polyacrylic acid substrate to form a mixed solution; pouring the mixed solution into a mold to control the thickness to be 2-3 mm;

[0012] (F) Gel molding: Place the mold in a thermostat, set the temperature to 40-50°C, and keep it for 24 hours to solidify the gel; remove the gel from the mold and dry it until it reaches the required hardness and mechanical strength. The required hardness is 60-80 Shore hardness, and the mechanical strength is usually 10-30MPa.

[0013] The second aspect of the present invention provides a three-stage amplification piezoelectric bioengineering patch for airway injury repair, which is prepared by the preparation method described above.

[0014] The three-stage amplification piezoelectric bioengineering patch of the present invention is composed of four components, including barium titanate nanoparticles, clay nanosheets, silver nanowires, and polyacrylic acid. The polyacrylic acid is used as a base material to load other components. The clay nanosheets form a cage structure that wraps the barium titanate nanoparticles and are connected by widely deployed zinc oxide nanowires ( Figure 1 ).

[0015] These four components work together to achieve the generation of piezoelectric effect and its multi-stage amplification. They not only have antibacterial and adhesion functions, but also constitute a multifunctional composite biological patch. On the tracheal wall, the patch is subjected to forces due to the movement of the trachea and the action of airflow. These forces are evenly distributed to the clay nanosheet cages throughout the patch through silver nanowires. The clay nanosheet cages under stress vibrate synchronously to form resonance, thereby strengthening the mechanical stimulation. This resonance effect amplifies the action on the patch step by step and eventually transmits it to the barium titanate nanoparticles in the cage. As the core piezoelectric particles, barium titanate nanoparticles convert mechanical stimulation into electric fields to form the first-stage discharge. The barium titanate particles are wrapped by clay nanosheets, and the generated electric field is amplified to form a stronger electrical stimulation, which constitutes the second-stage amplification. Silver nanowires act like wires to ensure that the electric field is evenly distributed throughout the patch, enhancing the promotion effect of the nanosheet electric field on surrounding cells and tissues, and achieving the third-stage amplified electrical stimulation. In addition to this three-stage amplification mechanism, the patch also has the function of promoting cell adhesion and regeneration, as well as improving the antibacterial properties of the microenvironment ( Figure 2 ).

[0016] The third aspect of the present invention provides a use of the three-stage amplification piezoelectric bioengineering patch as described above in the preparation of a product for repairing airway injuries.

[0017] Furthermore, the product for repairing airway damage is a product that repairs airway epithelial damage and accelerates the regeneration of the damaged area.

[0018] Furthermore, the product for repairing airway damage is a product for improving tracheal stenosis.

[0019] When the present invention is used, the damaged part in the trachea is first found, and the patch of the present invention is directly attached to the damaged part under the bronchoscope. The present invention has good adhesion and can be well attached to the inner surface of the trachea without being fixed by suturing.

[0020] The advantages of the present invention are:

[0021] The present invention constructs a new type of three-stage amplification piezoelectric bioengineering patch that can be used for airway injury repair, which can not only stably repair airway epithelial damage, but also accelerate the regeneration of the damaged part. In view of the complex environment in the airway, this patch can resist bacteria and infection, and easily and firmly fit in the trachea to avoid secondary damage caused by fixing the patch. At the same time, the surface of the patch also provides a sticky environment, combined with piezoelectric reaction, to reconstruct the damaged environment and guide the body to repair the damaged epithelium. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 .The main components of the patch.

[0023] Figure 2 .Synergy between the main components of the patch.

[0024] Figure 3 .Electron microscope images of the main components of the patch.

[0025] Figure 4 .Overall view of the composite patch.

[0026] Figure 5 .Adhesion test results of the patch.

[0027] Figure 6 .Test of the adhesion of the patch to the main organs of mice. DETAILED DESCRIPTION

[0028] The specific implementation methods provided by the present invention are described in detail below in conjunction with examples.

[0029] Example 1: Preparation method of the three-stage amplification piezoelectric bioengineering patch of the present invention

[0030] The three-stage amplification piezoelectric bioengineering patch of the present invention is composed of four components, including barium titanate nanoparticles, clay nanosheets, silver nanowires, and polyacrylic acid. The polyacrylic acid is used as a base material to load other components. The clay nanosheets form a cage structure that wraps the barium titanate nanoparticles and are connected by widely deployed zinc oxide nanowires ( Figure 1 ).

[0031] 1. Raw material preparation: Barium titanate nanoparticles: prepare tetrabutyl titanate and barium salt (barium nitrate). Nanoclay flakes: select montmorillonite or kaolin. Silver nanowires: prepare silver salt and sodium citrate as reducing agent. Polyacrylic acid: prepare polyacrylic acid monomer and potassium persulfate as initiator.

[0032] 2. Synthesis of barium titanate nanoparticles: Tetrabutyl titanate and barium nitrate are mixed in ethanol in proportion (the molar ratio of the three substances is 1:1:10); under stirring conditions, gradually heat to 60°C to carry out hydrolysis and polymerization reactions to form a sol; continue heating until gelation to form a solid gel; calcine the gel at 600°C for 2 hours to obtain barium titanate nanoparticles.

[0033] 3. Preparation of nanoclay flakes: Clay minerals (montmorillonite or kaolin) were mixed in 10 times the mass of deionized water and treated with ultrasonic stripping technology for 3 hours; nanoclay flakes were collected by centrifugal separation.

[0034] 4. Synthesis of silver nanowires: Mix a silver nitrate solution and a reducing agent (sodium citrate) solution, add an appropriate amount of polyvinyl pyrrolidone (PVP) as a stabilizer (the molar ratio of the three is 1:2:0.1); stir at room temperature for 4 hours to reduce the silver ions to silver nanowires; collect the silver nanowires by centrifugation and wash with deionized water to remove excess reducing agent and stabilizer.

[0035] 5. Preparation of polyacrylic acid substrate: Mix polyacrylic acid monomer, initiator (potassium persulfate) and an appropriate amount of deionized water in a volume ratio of 10:1:200; perform polymerization reaction at 60° C. for 3 hours to form a polyacrylic acid substrate.

[0036] 6. Assembling gel: Mix barium titanate nanoparticles, nanoclay sheets and silver nanowires in a mass ratio of 1:2:1 to form a composite; evenly disperse the composite in a polyacrylic acid base to form a mixed solution; pour the mixed solution into a mold and control the thickness to be 2 mm.

[0037] 7. Gel molding: Place the mold in a constant temperature box, set the temperature to 40-50°C, and maintain it for 24 hours to solidify the gel; remove the gel from the mold and dry it until it reaches a Shore hardness of 60 and a mechanical strength of 10MPa.

[0038] Embodiment 2:

[0039] 1. Basic characteristics of the patch

[0040] The key components of the patch were successfully synthesized by the method of Example 1, and Figure 3 These components are further integrated to form a complete patch structure, as shown in Figure 4 shown.

[0041] 2. Patch Adhesion

[0042] In order to evaluate the adhesion performance of the patch, the present invention compares it with common objects ( Figure 5 Adhesion tests were performed on the main organs of mice. Figure 6 As shown in the figure, the test results show that the patch can firmly adhere to a variety of tissue surfaces. This feature not only proves its excellent adhesion, but also helps promote cell adhesion and tissue regeneration.

[0043] 3. Antibacterial properties of the patch

[0044] In order to evaluate the antibacterial properties of the patch, the present invention selected Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli) for antibacterial experiments. The antibacterial efficiency was evaluated by measuring the diameter of the antibacterial ring, and a control group was set up. As shown in Table 1, the results showed that the patch showed significant antibacterial effects on both bacteria, indicating that it has good antibacterial properties and can effectively resist the complex microbial environment in the airway.

[0045] Table 1 Antibacterial ring test of the patch

[0046]

[0047] 4. The effect of the patch on epithelial repair in vivo

[0048] In order to explore the repair effect of the patch in the epithelial injury model, the present invention conducted experiments using New Zealand rabbits. First, an epithelial injury model was established: the trachea was exposed and incised after anesthesia, and the epithelium of the predetermined area was scraped off with a disposable brush, followed by suturing and continuous monitoring. The experiment was divided into three groups: a natural control group (no treatment), a blank control group (only epithelial scraping), and an experimental group (epithelial scraping and covering with a patch). The state of the animals was closely observed after surgery, and they were killed after 28 days, and the trachea was removed for observation. The microenvironment of the epithelial defect was detected by immunofluorescence staining and histochemical staining to evaluate the regeneration and repair effect. As shown in Table 2, the results show that the intervention of the patch significantly promoted the re-covering of the epithelium, thereby enhancing the regeneration of the tissue and the reconstruction of function.

[0049] Table 2 Evaluation of the effect of the patch on epithelial regeneration

[0050]

[0051] 5. The patch regulates the microenvironment of tracheal injury and improves epithelial migration in vivo

[0052] In order to further explore the effect of the patch on the regulation of the microenvironment of tracheal injury and the improvement of epithelial migration, the present invention is based on the epithelial injury model of New Zealand rabbits. The experiment was divided into two groups: a blank group (epithelial injury only, no patch covering) and a patch group (epithelial injury and patch covering). After the operation, the state of each group of animals was closely observed, and they were killed after 28 days, and the trachea was removed for observation. By measuring the relevant parameters of tracheal stenosis (D1, D2, D3, D4), the stenosis rate (S) was calculated to evaluate the situation of tracheal stenosis. Definition S (stenosis rate) = [1-(D1*D2) / (D3*D4)]*100%. As shown in Table 3, the results showed that the average stenosis rate before airway stent implantation was 63.80%, while the average stenosis rate after 30 days of implantation was reduced to 55.50%, indicating that the implantation of the patch significantly improved the situation of tracheal stenosis.

[0053] Table 3 Evaluation of the effect of the patch on airway stenosis in rabbits

[0054]

[0055] The preferred embodiments of the present invention have been specifically described above, but the present invention is not limited to the embodiments. Those skilled in the art may make various equivalent modifications or substitutions without violating the spirit of the present invention. These equivalent modifications or substitutions are all included in the scope defined by the claims of this application.

Claims

1. A method for preparing a three-stage amplification piezoelectric bioengineering patch for airway injury repair, characterized in that: The following steps are involved: (A) Synthesis of barium titanate nanoparticles: Tetrabutyl titanate and barium nitrate are mixed in ethanol in a molar ratio of 1:1:10 to 1:1:20; the mixture is gradually heated to 60 to 80°C under stirring to perform hydrolysis and polymerization to form a sol; the mixture is further heated to gelation to form a solid gel; the gel is calcined at 600 to 800°C for 2 to 4 hours to obtain barium titanate nanoparticles; (B) Preparation of nanoclay flakes: Clay mineral and deionized water are mixed in a mass ratio of 1:10-1:50, and treated by ultrasonic stripping technology for 3-6 hours; nanoclay flakes are collected by centrifugation; (C) Synthesis of silver nanowires: a silver nitrate solution and a reducing agent solution are mixed, and polyvinyl pyrrolidone is added as a stabilizer, wherein the molar ratio of the three is 1:2:0.1-1:2:0.5; the mixture is stirred at room temperature for 4-8 hours to reduce the silver ions to form silver nanowires; the silver nanowires are collected by centrifugation, and washed with deionized water to remove excess reducing agent and stabilizer; (D) Preparation of polyacrylic acid substrate: mixing polyacrylic acid monomer, initiator and an appropriate amount of deionized water in a volume ratio of 10:1:100-10:1:200; performing polymerization reaction at 60-70° C. for 3-6 hours to form a polyacrylic acid substrate; (E) Assembling gel: mixing barium titanate nanoparticles, nanoclay sheets and silver nanowires in a mass ratio of 1:1:1-1:2:1 to form a composite; uniformly dispersing the composite in a polyacrylic acid substrate to form a mixed solution; pouring the mixed solution into a mold to control the thickness to be 2-3 mm; (F) Gel molding: placing the mold in a constant temperature box, setting the temperature to 40-50° C., and maintaining it for 24 hours to solidify the gel; taking out the gel from the mold and drying it to obtain the three-stage amplification piezoelectric bioengineering patch.

2. The method for preparing the three-stage amplification piezoelectric bioengineering patch according to claim 1, characterized in that: The clay mineral in step B is montmorillonite or kaolin.

3. The method for preparing the three-stage amplification piezoelectric bioengineering patch according to claim 1, characterized in that: In step C, the reducing agent is sodium citrate.

4. The method for preparing the three-stage amplification piezoelectric bioengineering patch according to claim 1, characterized in that: The initiator in step D is potassium persulfate.

5. The method for preparing the three-stage amplification piezoelectric bioengineering patch according to claim 1, characterized in that: In step F, the three-stage amplification piezoelectric bioengineering patch is obtained by drying until the Shore hardness is 60-80 and the mechanical strength is 10-30 MPa.

6. A three-stage amplification piezoelectric bioengineering patch for airway injury repair, characterized in that: The method is prepared according to any one of claims 1 to 5.

7. The three-stage amplification piezoelectric bioengineering patch according to claim 6, characterized in that: The three-stage amplification piezoelectric bioengineering patch is composed of four components, including barium titanate nanoparticles, clay nanosheets, silver nanowires, and polyacrylic acid. Polyacrylic acid is used as a base material to load other components. The clay nanosheets form a cage structure that wraps the barium titanate nanoparticles and are connected by widely deployed zinc oxide nanowires.

8. Use of the three-stage amplification piezoelectric bioengineering patch as claimed in claim 6 in preparing products for repairing airway injuries.

9. The use of the three-stage amplification piezoelectric bioengineering patch according to claim 8 in preparing a product for airway injury repair, characterized in that: The product for repairing airway injury is a product for repairing airway epithelial injury and accelerating the regeneration of the injured part.

10. The use of the three-stage amplification piezoelectric bioengineering patch according to claim 8 in preparing a product for airway injury repair, characterized in that: The product for repairing airway damage is a product for improving tracheal stenosis.

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