Method for preparing degradable magnesium metal mechanical enhanced piezoelectric composite bone-guided regeneration membrane

Abstract

The invention provides a method for preparing a degradable magnesium metal mechanical enhanced piezoelectric composite bone-guided regeneration membrane. A magnesium foil or a magnesium net (which is cut into a magnesium net with small holes through laser cutting) or a magnesium support (which is cut through laser cutting) is firstly subjected to heat treatment by HF (hydrogen fluoride), and a layer of MgF4 coating is obtained on the surface of the magnesium foil or the magnesium net or the magnesium support. Then preparing an electrostatic spinning membrane with a piezoelectric effect on the surface of the sample; preparing a piezoelectric spinning fiber membrane with fibers arranged in parallel on one side close to the fiber tissue so as to shield the ingrowth of the fiber tissue; a disordered and loose piezoelectric spinning fiber membrane is prepared on the side close to the bone tissue by adopting an electrostatic spinning technology. The electrostatic spinning membrane provided by the invention has piezoelectric property, and when the piezoelectric spinning membrane is subjected to physiological mechanical stress (such as muscle contraction and chewing pressure), weak charges can be generated to actively regulate and control physiological activities of osteoblasts and provide repair power for bone regeneration.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of electrospinning and membrane layer assembly, and particularly relates to a method for preparing a piezoelectric composite bone guide regeneration membrane mechanically reinforced by degradable magnesium metal. BACKGROUND

[0002] The key to the clinical application of guided bone regeneration (GBR) technology lies in the use of barrier membranes. The function of the membrane is to isolate the rapidly proliferating epithelium and connective tissue, to build a relatively isolated space environment for bone tissue regeneration, so as to maximize the proliferation and differentiation of cells with osteogenic potential, and ultimately promote the formation of new bone in the bone defect area.

[0003] The ideal GBR barrier membrane needs to meet the following core requirements: ① Excellent tissue compatibility: support the repair process of bone or soft tissue defects. ② Reliable osteogenic space maintenance ability: provide and stabilize sufficient space volume to ensure the smooth growth of new bone. ③ Selective permeable barrier function: effectively block the invasion of epithelial connective tissue into the bone defect area, while allowing oxygen, blood and bioactive factors to penetrate. ④ Good clinical operability: easy to operate, with appropriate shaping ability and mechanical strength. ⑤ Promote healing of biological activity.

[0004] The currently commonly used GBR membrane materials can be divided into three categories according to the source: artificial synthetic polymer materials: such as polytetrafluoroethylene (PTFE) and aliphatic polyester (such as PLA, PGA, PCL). Natural source polymer materials: including collagen membrane (the structural feature is dense on the outside and porous on the inside), chitosan membrane (often loaded with hydroxyapatite HA microspheres) and alginate membrane (good barrier performance, but insufficient mechanical strength). Metal materials: such as titanium and its alloy membrane (relying on high stiffness to maintain space). According to the retention time in vivo, these materials can be divided into: absorbable membrane: advantages: avoid secondary surgery, reduce surgical complexity, patient complications and membrane exposure risk. Limitations: low mechanical strength, easy to collapse, affecting bone growth space; often need to cooperate with grafting materials; the duration of the barrier effect is not easy to accurately control; membrane micro-motion may interfere with the stability of the grafting material and the integrity of the blood clot. Non-absorbable membrane: advantages: outstanding mechanical stability, good biocompatibility, appropriate strength and plasticity. Limitations: increased risk of membrane exposure, which may lead to infection; need for secondary surgery; usually need to be fixed with membrane nails. Collagen membrane is a widely used absorbable membrane in clinical practice, known for its excellent biocompatibility, low immunogenicity and tissue integration ability, and does not need to be removed. However, its high cost, rapid degradation rate, poor plasticity, soft texture and insufficient space maintenance resulting in collapse and displacement cannot be ignored. Even when used in combination with bone grafting materials, there is still the disadvantage of difficulty in accurately controlling the osteogenic position.

[0005] Synthetic absorbable membranes such as polylactic acid (PLA) and polyglycolide (PGA) can cause local acidity to rise during degradation, which adversely affects tissue regeneration. Such membranes can also induce mild to severe inflammatory reactions, attracting lymphocyte and multinucleated giant cell infiltration. Inflammation superimposed on local pH decline can even damage newly formed bone tissue. Non-absorbable polytetrafluoroethylene (PTFE) membranes have excellent mechanical stability and space maintenance ability, but their tissue affinity is poor, and there is a high risk of membrane exposure and infection, which also requires secondary surgery for removal. Titanium mesh membranes perform well in maintaining space and reducing bacterial adhesion, but still face problems such as membrane exposure, soft tissue isolation, and the need for secondary removal. In addition, compared with non-absorbable membranes, absorbable membranes with better tissue integration can bring better soft tissue response. Therefore, the development of GBR membranes based on hard absorbable materials is considered a potential way to solve the many shortcomings of existing materials. Existing collagen membranes are mainly used for small-range bone defects (such as single-tooth site repair), while large-range or complex bone defects (such as multiple tooth loss, postoperative jaw tumor) rely on non-degradable high-strength support membranes (such as titanium mesh or titanium reinforced membrane), which have complications and secondary surgery problems.

[0006] Magnesium, as a biodegradable metal, exhibits good mechanical properties, controllable degradation characteristics, and biocompatibility, and has been applied in cardiovascular stents, tracheal stents, orthopedic fixation screws, craniofacial fixation systems, and bone repair plates. The mechanical stability of magnesium implants is closer to natural bone and can be completely absorbed in the body. In the human body environment, the final products of degradable magnesium are magnesium ions (Mg 2+ ), hydroxide ions (OH - ) and hydrogen gas (H2). Among them, magnesium ions can promote the activation and differentiation of osteoblasts by activating the Pi3K / Akt signaling pathway; hydroxide ions help to increase the local microenvironment pH, thereby reducing the risk of inflammation and infection; hydrogen gas production may lead to gas cavity formation, but this phenomenon usually only occurs during the metal phase corrosion stage, and the gas cavity can be spontaneously absorbed, and has no obvious negative impact on bone formation. Currently, the research of degradable magnesium metal membranes is still in its early stages. For example, Frank Witte's team developed a non-porous GBR membrane of pure magnesium (99.95%) with a thickness of about 140 μm and a matching fixation screw, but still faces problems such as too fast degradation speed and stress corrosion.

[0007] Electrospinning is an effective method to prepare polymer nanofiber membranes (ordered or disordered). The resulting nanofiber membranes have high specific surface area and high porosity, which are conducive to cell adhesion and proliferation, and are widely used in drug controlled release, wound dressing and tissue engineering. This technology provides a new idea for the preparation of GBR membranes. By adjusting the solution system, spinning parameters (such as voltage, time, solution advancing rate, temperature, humidity) and equipment improvement, the fiber structure, size, mechanical and biological properties can be accurately controlled to meet different needs. Numerous studies have confirmed the feasibility of using electrospinning technology to prepare GBR membranes. However, its limitation is that the commonly used spinning raw materials are mostly high molecular polymers, and the acidic by-products generated by their degradation may stimulate the surrounding tissue and even cause inflammation. In addition, the polymer GBR membranes prepared by electrospinning method are usually soft in mechanical properties and easy to collapse, which is difficult to provide sufficient mechanical support to the bone defect area. The present invention aims at the situation that the non-degradable titanium mesh is often used for clinical large-scale bone defects and complex bone defects, which leads to high clinical complications and the need for secondary surgery to remove, and develops a GBR barrier membrane with mechanical support, degradable function, piezoelectric effect and easy molding. SUMMARY

[0008] In order to overcome the problems of the prior art, the present application provides a method for preparing a degradable bone guided regeneration membrane by preparing an asymmetric piezoelectric spinning membrane on both sides of a magnesium foil / magnesium mesh / magnesium support.

[0009] A method for preparing a degradable bone guided regeneration membrane by preparing an asymmetric piezoelectric membrane layer on both sides of a magnesium foil / magnesium mesh / magnesium support, comprising the following steps:

[0010] Step S1: laser cutting magnesium metal sheet: using a laser cutting machine to cut small holes of different shapes on magnesium metal sheets of different thicknesses to obtain magnesium meshes with different porosities; using a laser cutting machine to cut magnesium metal sheets to obtain magnesium supports of different shapes;

[0011] Step S2: treating the magnesium foil / magnesium mesh / magnesium support obtained in step S1;

[0012] Step S3: electrospinning the magnesium foil / magnesium mesh / magnesium support treated in step S2 to prepare a membrane layer, thereby obtaining a degradable bone guided regeneration membrane.

[0013] Preferably, in the step S1 of the preparation method of the present application, the magnesium metal sheet is a magnesium alloy or a pure magnesium metal, the magnesium alloy is a magnesium-zinc alloy or a magnesium-zinc-calcium alloy; the purity of the pure magnesium metal is >99.9%, and the thickness is 50-500 μm; the small holes of different shapes are round holes or triangular holes; the porosity is 10%-90%.

[0014] Preferably, the treatment process of step S2 of the preparation method of the present application is hydrofluoric acid treatment, specifically: placing the magnesium mesh / magnesium stent into a hydrofluoric acid solution for 30 minutes to 12 hours to obtain a surface coating containing MgF4.

[0015] Preferably, the concentration of the hydrofluoric acid solution in the preparation method of the present application is 0.5wt% to 5wt%, and the soaking time is 30 minutes to 12 hours.

[0016] Preferably, the specific process of step S3 of the preparation method of the present application is as follows:

[0017] GBR membrane with dense ordered electrospun membrane on one side of the magnesium mesh / magnesium stent and loose electrospun membrane on the other side: place the magnesium mesh / magnesium stent treated by step S2 above on the drum receiving plate of the electrospinning machine to prepare a dense parallel ordered nanofiber membrane, with the following spinning parameters: voltage: 10-50kV, temperature: 15-35℃, humidity: 10%-30%, time: 3-12 hours, and the rotation speed of the drum is 1500-4000rpm.

[0018] Place the above obtained sample on the flat receiving plate of the electrospinning machine, and use the electrospinning machine to prepare a loose unordered nanofiber membrane on the other side. The electrospinning parameters used are: voltage: 10-50kV, temperature: 15-35℃, humidity: 10%-30%, time: 3-12 hours.

[0019] Preferably, the preparation method of the present application uses a degradable polymer solution or a mixture of two organic solvents in the process of leaching and electrospinning.

[0020] The concentration of the degradable polymer solution is 1%-40%, including a mixture of poly(vinylidene fluoride-trifluoroethylene) copolymer (PVDF-TrFE) and polycaprolactone (PCL) or poly(lactic acid) (PLA), a mixture of silk fibroin (SF) and polycaprolactone (PCL), and a poly-L-lactic acid solution (PLLA); the mixing ratio of the mixture of two organic solvents is 1:9 to 9:1, and the organic solvent is hexafluoroisopropanol.

[0021] The application provides a method for preparing a piezoelectric composite bone guided regeneration (GBR) film with mechanical enhancement of degradable magnesium metal, i.e. using magnesium metal as a support, preparing a degradable polymer film with piezoelectric effect in different morphological structures on both sides of the magnesium metal by electrospinning technology, combining the advantages of degradable magnesium metal and electrospun nanofiber film, so that the composite film has degradable performance, piezoelectric effect, good mechanical properties and plasticity, and biocompatibility, one side of the composite film can promote fibrous tissue regeneration and promote wound healing, and the other side can promote bone tissue regeneration, so as to meet the performance requirements of the GBR film in clinical application. Specifically, ① magnesium metal sheet (including: magnesium alloy sheet or high-purity magnesium sheet (thickness: 50-500 μm), which is cut into a magnesium mesh with small holes or a magnesium support with a specific structure by laser cutting, ② the magnesium sheet or magnesium mesh or magnesium support is first subjected to heat treatment by HF, and a MgF4 coating is formed on the surface, ③ then, the electrospun film is prepared on the surface of the magnesium foil or magnesium mesh or magnesium support with the MgF4 coating: a dense polymer film layer with piezoelectric effect is prepared on the surface of the sample by a hot pressing method on the side close to the fibrous tissue, or a dense parallel piezoelectric nanofiber film is prepared by an electrospinning method, and the designed structure and piezoelectric effect can shield the ingrowth of fibrous tissue and tissue healing. The piezoelectric nanofiber film is prepared on the side close to the bone tissue by an electrospinning technology, and the designed structure and piezoelectric effect can promote the bone.

[0022] Compared with the prior art, the preparation method has the following advantages:

[0023] 1. Good mechanical properties and plasticity: the commonly used electrospun polymer film is relatively soft and has poor mechanical properties. In the application, the GBR film has a sandwich structure, the middle layer is a magnesium foil, a magnesium metal mesh or a magnesium metal support, which serves as a mechanical support and endows the GBR film with good mechanical properties and plasticity, so that the clinician can perform free hand shaping according to different bone defect shapes, and the clinical operation is more convenient.

[0024] 2. Reducing suture tension: the thickness of the sandwich GBR can be adjusted (less than 2 mm), the GBR film is relatively thin, and after the gum is sutured, the suture tension of the soft tissue can be reduced, the gum tissue is effectively prevented from cracking at the suture site, the GBR film is exposed, and bacterial contamination of the GBR film is avoided.

[0025] 3. Avoiding soft tissue invasion: the film layer close to the gum on one side of the GBR film in the application is a dense film layer, which avoids the invasion of the soft tissue into the film layer and the ingrowth into the bone defect space.

[0026] 4. Piezoelectric effect promotes osteogenesis: The electrospun membrane in the application has piezoelectric properties. When the membrane is subjected to physiological mechanical stress (such as muscle contraction, chewing pressure), it will generate weak electric charge. This electric signal can activate the calcium ion channel in the osteoblasts, MAPK signal pathway, and up-regulate the expression of bone morphogenetic protein (BMP-2), Runx2 and other osteogenesis-related genes.

[0027] 5. Piezoelectric effect promotes angiogenesis and nutrient transport: The electric field generated by the piezoelectric properties of the electrospun membrane in the application can induce the migration and tubular structure formation of vascular endothelial cells, accelerate the growth of new blood vessels into the bone defect area, and provide oxygen and nutrients for bone regeneration.

[0028] 6. Antibacterial properties of piezoelectric effect: The piezoelectric effect of the electrospun membrane in the application endows the GBR membrane with antibacterial properties.

[0029] 7. Adjustable structure: The structure size of the membrane layer (such as thickness, membrane structure on both sides, porosity of the fiber membrane layer, etc.) can be adjusted according to the needs, and the GBR membrane can be adjusted according to the actual application scenario in clinical practice.

[0030] 8. Low cost: The electrospinning equipment used in the application can be assembled by oneself, and the cost is low.

[0031] 9. High efficiency: The entire preparation process can be controlled within 24 hours. BRIEF DESCRIPTION OF DRAWINGS

[0032] Figure 1 The figure is a flowchart of the preparation method of the application.

[0033] Figure 2 The figure is a schematic diagram of the magnesium mesh structure obtained by laser cutting.

[0034] Figure 3 The figure is a physical diagram of the piezoelectric electrospun fiber composite GBR membrane reinforced by magnesium mechanics.

[0035] Figure 4 The figure is a micro-SEM morphology diagram of PCL / SF piezoelectric electrospun membrane, wherein a is a micro-SEM morphology diagram magnified by 4000 times, and b is a micro-SEM morphology diagram magnified by 10000 times.

[0036] Figure 5 The figure is an XRD spectrum of PCL / SF piezoelectric electrospun membrane (after alcohol treatment, the 2θ scanning range of PCL / SF electrospun membrane is 5°-40°, and the β-sheet structure peak value of silk fibroin is about 20.4°). Figure 6The piezoelectric properties of PCL / SF piezoelectric spun film (the piezoelectric properties of PCL / SF spun film after alcohol treatment: at impact frequencies of 1Hz and 6Hz, the output voltage of the spun film is between -0.5V and 0.5V within 35s). Detailed Implementation

[0037] The technical solution of the present invention will now be described in detail with reference to the accompanying drawings:

[0038] like Figure 1 As shown, a method for preparing a biodegradable magnesium metal mechanically reinforced piezoelectric composite bone-guided regeneration membrane includes the following steps:

[0039] Step S1: Using a laser cutting machine, small holes of different shapes (round holes, triangular holes, etc.) are cut on magnesium metal sheets of different thicknesses (magnesium alloys: such as magnesium-zinc alloys, magnesium-zinc-calcium alloys; pure magnesium metal (purity: >99.9%) thickness: 50μm-500μm) to obtain magnesium meshes with different porosities (10%-90%). Figure 2 (As shown). Magnesium metal sheets were cut using a laser cutting machine to obtain magnesium supports of different shapes.

[0040] Step S2: Treat the magnesium mesh / magnesium support obtained in step S1 with hydrofluoric acid: Immerse the magnesium metal / magnesium mesh / magnesium support in a hydrofluoric acid solution (0.5wt%-5wt%) for 30 minutes to 12 hours to obtain a surface coating containing MgF4.

[0041] Step S3: Electrospinning is performed on the magnesium foil / magnesium mesh / magnesium scaffold treated in step S2 to prepare a piezoelectric film layer, thereby obtaining a biodegradable bone-guided regeneration membrane.

[0042] Preparation of GBR membranes with one side being a dense, ordered piezoelectric electrospun membrane and the other side being a loose piezoelectric electrospun membrane: Magnesium foil / magnesium mesh / magnesium scaffold with a surface coated with MgF4 and a polymer coating, as described above, was placed on the receiving plate of an electrospinning machine to prepare a dense, parallel-aligned, ordered nanofiber membrane (the resulting nanofiber morphology is shown in the figure). Figure 4Fig. 1 shows the micro-SEM morphology of the electrospun nanofiber film, wherein a is a micro-SEM morphology at 4000 times magnification, and b is a micro-SEM morphology at 10000 times magnification. The concentration of the degradable polymer solution is 1%-40%, including a mixed solution of poly(vinylidene fluoride-trifluoroethylene) copolymer (PVDF-TrFE) and polycaprolactone (PCL) or polylactic acid (PLA), a mixed solution of silk fibroin (SF) and polycaprolactone (PCL), and a poly-L-lactic acid solution (PLLA); the mixing ratio of the mixed solution of the two organic solvents is 1:9 to 9:1, and the organic solvent is hexafluoroisopropanol. The organic solvent is hexafluoroisopropanol (HFIP). The spinning parameters are as follows: voltage: 10-50 kV, temperature: 15-35°C, humidity: 10%-30%, time: 3-12 hours, and the rotating speed of the drum is 1500-4000 rpm. The sample obtained above is placed on the flat receiving plate of the electrospinning machine, and a loose and disordered piezoelectric nanofiber film is prepared on the other side of the electrospinning machine (the concentration of the degradable polymer solution is 1%-40%, including a mixed solution of poly(vinylidene fluoride-trifluoroethylene) copolymer (PVDF-TrFE) and polycaprolactone (PCL) or polylactic acid (PLA), a mixed solution of silk fibroin (SF) and polycaprolactone (PCL), and a poly-L-lactic acid solution (PLLA); the mixing ratio of the mixed solution of the two organic solvents is 1:9 to 9:1, and the organic solvent is hexafluoroisopropanol. The organic solvent is hexafluoroisopropanol (HFIP). The electrospinning parameters used are as follows: voltage: 10-50 kV, temperature: 15-35°C, humidity: 10%-30%, and time: 3-12 hours). The piezoelectric nanofiber film is combined with the magnesium stent to form a composite electrospun film. Figure 6 The film layer has piezoelectric properties.

[0043] Figure 3 Fig. 3 shows the composite electrospun film, in which the magnesium stent is covered with the electrospun piezoelectric nanofiber film on both sides.

[0044] Figure 5 Fig. 5 shows the XRD pattern of the PCL / SF electrospun film after alcohol treatment, in which there is a clear silk fibroin β-sheet structure peak at 20.4°.

[0045] Compared with the prior art, the degradable GBR film prepared by the preparation method of the present application has the following advantages:

[0046] 1. Mechanical properties: The commonly used degradable GBR film is soft and has poor mechanical properties, which is easy to cause collapse. Although the non-degradable GBR film has good mechanical properties, it needs to be removed by secondary surgery. The present application uses degradable magnesium as the middle layer and degradable piezoelectric spinning film on both sides, so that the GBR film has degradability and also has certain mechanical support.

[0047] 2. Multifunctionality: The electrospinning technique is used to prepare a loose piezoelectric nanofiber membrane on the end close to the bone tissue, and a dense parallel piezoelectric nanofiber membrane on the end close to the soft tissue. The asymmetric structure endows the GBR membrane with multifunctionality.

[0048] 3. Plasticity: The currently commonly used titanium metal GBR membrane needs to be shaped with the aid of tools to conform to the shape of the defect site due to its high hardness. The prepared degradable magnesium polymer composite can be shaped by hand, satisfying the free hand operation method of clinicians and being more convenient.

[0049] 4. The prepared electrospun nanofiber membrane with piezoelectric effect generates weak electric charges when subjected to physiological mechanical stress (such as muscle contraction and chewing pressure), has the functions of promoting bone formation, promoting angiogenesis and nutrient transport, and antibacterial performance.

[0050] Example One

[0051] Laser-cut high-purity magnesium mesh: High-purity magnesium foil (purity: 99.98%), cut by a laser cutting machine, pattern: equilateral triangle with a side length of 2 mm, magnesium mesh with a porosity of 30-90%, and a high-purity magnesium mesh with a thickness of 0.16 mm obtained by chemical polishing.

[0052] The magnesium mesh is immersed in a 10-30% HF solution at 20°C for 6 hours. Then the residual HF solution is rinsed clean with deionized water. The obtained magnesium mesh coated with MgF4 is electrospun: placed on the drum receiving plate of the electrospinning machine to prepare a dense parallel piezoelectric nanofiber membrane. The spinning solution is a mixed solution of PVDF-TrFE and PCL (concentration: 8 wt%, ratio: 7:3, solvent: hexafluoroisopropanol, solution flow rate: 0.4 ml / min, spinning time: 6 hours, drum rotation speed: 4000 rpm, positive and negative electrode distance: 30 cm).

[0053] The obtained sample is turned over and placed on the flat plate receiving plate of the electrospinning machine to prepare a loose piezoelectric nanofiber membrane. The spinning solution is a mixed solution of PVDF-TrFE and PCL (concentration: 8 wt%, ratio: 7:3, solvent: hexafluoroisopropanol, solution flow rate: 0.4 ml / min, spinning time: 6 hours, positive and negative electrode distance: 30 cm).

[0054] Example Two

[0055] Laser-cut high-purity magnesium mesh: High-purity magnesium foil (purity: 99.98%), cut by a laser cutting machine, pattern: equilateral triangle with a side length of 2 mm, magnesium mesh with a porosity of 30-90%, and a high-purity magnesium mesh with a thickness of 0.16 mm obtained by chemical polishing.

[0056] The magnesium mesh was immersed in 10-30% HF solution at 20 °C for 6 hours. The residual HF solution was then washed away with deionized water. The obtained magnesium mesh with MgF4coating was electrospun to prepare a dense and parallel-aligned piezoelectric nanofiber membrane on the drum receiving plate of the electrospinning machine. The spinning solution was a mixed solution of PCL and silk fibroin (concentration: 10 wt%, ratio: 7:3, solvent: hexafluoroisopropanol, solution flow rate: 0.4 ml / h, spinning time: 6 hours, drum rotation speed: 4000 rpm, positive and negative electrode distance: 30 cm).

[0057] The obtained sample was turned over and placed on the flat plate receiving plate of the electrospinning machine to prepare a disordered and loose piezoelectric nanofiber membrane. The spinning solution was a mixed solution of PVDF-TrFE and PCL (concentration: 10 wt%, ratio: 7:3, solvent: hexafluoroisopropanol, solution flow rate: 0.4 ml / h, spinning time: 6 hours, positive and negative electrode distance: 30 cm).

[0058] Example Three

[0059] High-purity magnesium foil (purity > 99.9%, thickness 0.16 mm) was taken and laser-cut to obtain a porous magnesium mesh with a porosity of about 30-90% (e.g., equilateral triangular hole structure with a side length of 2 mm). After chemical polishing treatment, the magnesium mesh was immersed in 10-30% HF solution at 20 °C for 6 hours, and then washed with deionized water to obtain a magnesium mesh with a MgF4coating on the surface.

[0060] The obtained magnesium mesh with MgF4coating was electrospun to prepare a dense and parallel-aligned piezoelectric nanofiber membrane on the drum receiving plate of the electrospinning machine. The spinning solution was a mixed solution of PCL and silk fibroin (concentration: 8 wt%, ratio: 7:3, solvent: hexafluoroisopropanol, solution flow rate: 0.4 ml / h, spinning time: 6 hours, voltage: 30 kv, drum rotation speed: 4000 rpm, positive and negative electrode distance: 25 cm, needle gauge: 21G).

[0061] The obtained sample was turned over and placed on the flat plate receiving plate of the electrospinning machine to prepare a disordered and loose piezoelectric nanofiber membrane. The spinning solution was a mixed solution of PCL and silk fibroin (SF) (concentration: 8 wt%, ratio: 7:3, solvent: hexafluoroisopropanol, solution flow rate: 0.4 ml / h, spinning time: 6 hours, voltage: 30 kv, positive and negative electrode distance: 25 cm, needle gauge: 21G).

[0062] The composite piezoelectric nanofiber membrane was immersed in 75% alcohol for 0.5-1 h, and then dried at room temperature, finally obtaining a stable magnesium mesh / nanofiber composite membrane in a sandwich structure: the magnesium mesh is in the middle, one side is a layer of parallel dense PCL / SF nanofiber, and the other side is a layer of disordered loose PCL / SF nanofiber.

[0063] Example Four

[0064] Take high-purity magnesium foil (purity ≥ 99.9%, thickness 0.16 mm), and laser cut to obtain a porous magnesium mesh with a porosity of about 30-90% (for example, an equilateral triangular hole structure with a side length of 2 mm). After chemical polishing treatment, the magnesium mesh is placed in a 10-30% HF solution and soaked at 20°C for 6 h. After taking out, it is washed with deionized water to obtain a magnesium mesh with a MgF4 coating on the surface.

[0065] The obtained magnesium mesh with MgF4 coating is electrospun: place it on the drum receiving plate of the electrospinning machine to prepare a dense parallel piezoelectric nanofiber membrane. The spinning solution is a mixed solution of PCL and silk fibroin (concentration: 10wt%, ratio: 7:3, solvent: hexafluoroisopropanol, solution flow rate: 0.4ml / h, spinning time: 6 hours, voltage: 30kV, drum speed: 4000rpm, positive and negative electrode distance: 25cm, needle size: 21G).

[0066] The obtained sample is turned over and placed on the flat plate receiving plate of the electrospinning machine to prepare a disordered loose piezoelectric nanofiber membrane. The spinning solution is a mixed solution of PCL and silk fibroin (SF) (concentration: 10wt%, ratio: 7:3, solvent: hexafluoroisopropanol, solution flow rate: 0.4ml / h, spinning time: 6 hours, voltage: 30kV, positive and negative electrode distance: 25cm, needle size: 21G).

[0067] The composite piezoelectric nanofiber membrane was immersed in 75% alcohol for 0.5-1 h, and then dried at room temperature, finally obtaining a stable magnesium mesh / nanofiber composite membrane in a sandwich structure: the magnesium mesh is in the middle, one side is a layer of parallel dense PCL / SF nanofiber, and the other side is a layer of disordered loose PCL / SF nanofiber.

[0068] The application provides a method for preparing a degradable magnesium metal mechanically reinforced piezoelectric composite bone guide regeneration film. A magnesium foil or a magnesium mesh (a magnesium mesh with small holes cut by a laser) or a magnesium support (cut by a laser) is first subjected to heat treatment by using HF, so that a MgF4 coating layer is obtained on the surface. Then, an electrospun film with a piezoelectric effect is prepared on the surface of the sample: a piezoelectric electrospun fiber film with parallel arranged fibers is prepared on the side close to the fibrous tissue, so as to shield the ingrowth of the fibrous tissue. An unordered loose piezoelectric electrospun fiber film is prepared on the side close to the bone tissue by using an electrospinning technology. The electrospun film in the application has piezoelectric performance. When the piezoelectric electrospun film is subjected to physiological mechanical stress (such as muscle contraction and mastication pressure), a weak electric charge is generated, active regulation of the physiological activity of osteoblasts is realized, and a repair driving force for bone regeneration is provided.

Claims

1. A method for preparing a biodegradable magnesium metal mechanically reinforced piezoelectric composite bone-guided regeneration membrane, characterized in that... Includes the following steps: Step S1: Laser cutting of magnesium foil: Using a laser cutting machine, small holes of different shapes are cut on magnesium metal sheets of different thicknesses to obtain magnesium meshes with different porosities; using a laser cutting machine, magnesium foil is cut to obtain magnesium supports of different shapes. Step S2: Process the magnesium mesh / magnesium support obtained in step S1; Step S3: Electrospinning is performed on the magnesium mesh / magnesium scaffold treated in step S2 to prepare a piezoelectric fiber membrane, thereby obtaining a biodegradable bone-guided regeneration membrane.

2. The method for preparing a biodegradable magnesium metal mechanically reinforced piezoelectric composite bone-guided regeneration membrane according to claim 1, characterized in that... In step S1 above, the magnesium foil is a magnesium alloy or pure magnesium metal, wherein the magnesium alloy is a magnesium-zinc alloy or a magnesium-zinc-calcium alloy; the purity of the pure magnesium metal is >99.9%, and the thickness is 50μm-500μm; the small holes of different shapes are round holes or triangular holes; the porosity is 10%-90%.

3. The method for preparing a biodegradable magnesium metal mechanically enhanced piezoelectric composite bone-guided regeneration membrane according to claim 1, characterized in that... The above step S2 process is as follows: hydrofluoric acid treatment, specifically: immersing the magnesium mesh / magnesium support in a hydrofluoric acid solution for 30 minutes to 12 hours to obtain a surface coating containing MgF4.

4. The method for preparing a biodegradable magnesium metal mechanically reinforced piezoelectric composite bone-guided regeneration membrane according to claim 3, characterized in that... The concentration of the above hydrofluoric acid solution is 0.5wt%-5wt%, and the soaking time is 30 minutes-3 hours.

5. The method for preparing a biodegradable magnesium metal mechanically reinforced piezoelectric composite bone-guided regeneration membrane according to claim 1, characterized in that... The above step S2 process is as follows: polymer solution immersion treatment, specifically: immersing magnesium foil / magnesium mesh / magnesium support in a 1%-40% biodegradable polymer solution or a mixture of two organic solvents, then lifting the magnesium foil / magnesium mesh / magnesium support so that the solution covers the entire surface of the magnesium metal sheet, the metal part of the magnesium foil / magnesium mesh / magnesium support is wrapped, but the pore part is not covered; obtaining a dense polymer film layer of different thicknesses covering the magnesium metal; then placing it in a fume hood at room temperature to wait for the solvent of the polymer coating to completely evaporate, or placing it in an oven at 30 degrees-100 degrees to dry the polymer coating.

6. The method for preparing a biodegradable magnesium metal mechanically reinforced piezoelectric composite bone-guided regeneration membrane according to claim 5, characterized in that... The number of extractions is 1-10; the thickness of the dense polymer film is 10μm-50μm.

7. The method for preparing a biodegradable magnesium metal mechanically reinforced piezoelectric composite bone-guided regeneration membrane according to claim 1, characterized in that... The specific process of step S3 is as follows: A GBR membrane with parallel piezoelectric fibers is prepared on one side of a magnesium foil / magnesium mesh / magnesium support, and a disordered and loose electrospun membrane is prepared on the other side: The magnesium foil / magnesium mesh / magnesium support treated in step S2 is placed on the flat receiving plate of an electrospinning machine, and a loose and disordered nano-piezoelectric fiber membrane is prepared on the other side using an electrospinning machine. The electrospinning parameters used are: voltage: 10-50kV, temperature: 15-35℃, humidity: 10%-30%, time: 3-12 hours.

8. The method for preparing a biodegradable magnesium metal mechanically reinforced piezoelectric composite bone-guided regeneration membrane according to claim 1, characterized in that... The specific process of step S3 is as follows: A dense, ordered piezoelectric fiber spun membrane is prepared on one side of a magnesium mesh / magnesium support, and a loose electrospun membrane is prepared on the other side to form a GBR membrane: The magnesium mesh / magnesium support treated in step S2 is placed on the high-speed roller receiving plate of an electrospinning machine to prepare a dense, parallel, ordered nanofiber membrane. Spinning parameters: voltage: 10-50kV, temperature: 15-35℃, humidity: 10%-30%, time: 3-12 hours, roller speed: 1500-4000rpm.

9. The method for preparing a biodegradable magnesium metal mechanically enhanced piezoelectric composite bone-guided regeneration membrane according to claim 5, 7, or 8, characterized in that... Electrospinning processes all utilize biodegradable polymer solutions with piezoelectric effects or mixtures of two organic solvents. The concentration of the polymer solution is 1%-40%, including a mixture of poly(vinylidene fluoride-trifluoroethylene) copolymer with polycaprolactone or polylactic acid, a mixture of silk fibroin and polycaprolactone, and a poly-L-lactic acid solution or a mixture of poly-L-lactic acid with collagen or gelatin. The mixing ratio of the two organic solvents is 1:9 to 9:1, and the organic solvent is hexafluoroisopropanol.

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