Sleeve patch and preparation method thereof

Abstract

The invention discloses a rotator cuff patch and a preparation method thereof, and belongs to the technical field of repair materials. According to the preparation method provided by the invention, the bioactive material is combined, the fibers are gradually transited from random arrangement to aligned arrangement through electrostatic spinning, the difference of fiber orientation structures between the random fiber layer and the aligned fiber layer is effectively reduced, and the bonding force between the layers of the patch is greatly enhanced in cooperation with steam welding treatment; the layering phenomenon is effectively avoided; the tensile strength of the rotator cuff patch prepared by the method reaches 18 MPa, and the rotator cuff patch has excellent mechanical properties, can meet the requirements on flexibility and tear resistance during sewing, can provide enough mechanical support for the rotator cuff in the repair process, and adapts to the complex shoulder movement environment.

Description

Technical Field

[0001] The present invention belongs to the technical field of repair materials, and particularly relates to a rotator cuff patch and a preparation method thereof. Background Art

[0002] Rotator cuff injury is a common musculoskeletal disease, and surgical repair is a commonly used treatment method. However, the high retear rate after surgical repair remains a major clinical problem. Statistical data shows that the retear rate after rotator cuff tear repair is between 20% and 30%, and for massive rotator cuff tears, this probability can be as high as 40% - 50%. Such a high retear rate not only affects the long-term functional prognosis of patients, but also increases the burden on the medical system.

[0003] To solve this problem, the use of patch materials in rotator cuff repair has received increasing attention. Existing patch preparation methods include electrospinning, weaving technology, 3D printing, etc. Among various patch preparation methods, electrospinning has become a promising technology. Electrospun patches have many advantages, such as a high specific surface area, the ability to mimic the extracellular matrix structure at the nanoscale, and the potential to incorporate bioactive substances. These characteristics make electrospun patches potentially suitable for promoting cell adhesion, proliferation, and tissue ingrowth at the repair site, thereby enhancing the healing process of the rotator cuff.

[0004] However, the rotator cuff has a complex hierarchical structure. The tendon-bone interface of the rotator cuff consists of four different regions: tendon, unmineralized fibrocartilage, mineralized fibrocartilage, and bone. This complex structure is crucial for the smooth transition of mechanical properties from the soft tendon to the hard bone. Conventional electrospun patch fibers have a single arrangement and cannot mimic the complex structure of natural rotator cuff tissue, making it difficult to achieve ideal effects in terms of mechanical properties and tissue repair guidance. In existing multi-layer patches that mimic the natural tendon-bone interface structure, the bonding force between layers is insufficient, and delamination is likely to occur. This not only affects the mechanical properties and suturing of the patch, but also may lead to a decrease in the stability of the patch in the body, thereby affecting the repair effect. Summary of the Invention

[0005] In view of the above-mentioned prior art, the present invention provides a rotator cuff patch and a preparation method thereof, which solve the problems of single fiber arrangement and poor stability of existing rotator cuff patches.

[0006] To achieve the above object, the technical solution adopted by the present invention is: to provide a preparation method of a rotator cuff patch, comprising the following steps: S1: Freeze-mill the acellular matrix of porcine small intestinal submucosa to obtain a substrate powder, and then place the substrate powder in a pepsin solution for digestion for 12 - 72 h and then freeze-dry to obtain a pretreated substrate; S2: Disperse the pretreated substrate into Solvent 1 to obtain a dispersion; then dissolve the poly(lactic-co-glycolic acid) copolymer in the dispersion to obtain a spinning solution. S3: Perform electrospinning on the spinning solution to obtain a spun film. S4: Place the spun film in Solvent 2 for steam welding, and then perform vacuum drying to obtain the rotator cuff patch.

[0007] The beneficial effects of the present invention are as follows: The preparation method provided by the present invention uses a bioactive material as the raw material. Through pretreatment, electrospinning, and steam welding, a rotator cuff patch with biological activity is obtained. In this method, fibers with a gradually changing structure from random to aligned are stacked layer by layer to form a fiber film. Then, through solvent vapor, partial dissolution of the fiber surface is promoted, and after drying, a firm physical bond is formed between the fibers, effectively avoiding delamination. Moreover, the preparation steps of this method are simple, the materials are easily available, and it has excellent industrial prospects.

[0008] Based on the above technical solutions, the present invention can be further improved as follows.

[0009] Further, the concentration of the pepsin solution is 0.1 wt% - 2 wt%.

[0010] The beneficial effect of adopting the further technical solution is: Porcine small intestinal submucosa acellular matrix particles with smaller particle sizes can be obtained.

[0011] Further, Solvent 1 is hexafluoroisopropanol or a mixture of dichloromethane and hexafluoroisopropanol, and the concentration of the pretreated substrate in the dispersion is 0.1 wt% - 10 wt%.

[0012] Further, the volume ratio of dichloromethane to hexafluoroisopropanol in the mixture of dichloromethane and hexafluoroisopropanol is 1 - 5:5 - 9.

[0013] Further, the process parameters of electrospinning are positive high voltage 7 - 12 kV, negative high voltage -1 kV, injection rate 0.5 - 1.5 mm / min, collection distance 10 - 30 cm, and needle diameter 18 - 25 G.

[0014] The beneficial effect of adopting the further technical solution is: Through electrospinning technology, the fibers change from a random structure to an aligned structure, and are stacked layer by layer to form a fiber film, effectively reducing the difference in fiber orientation structure between the random fiber layer and the aligned fiber layer.

[0015] Further, Solvent 2 is dichloromethane or hexafluoroisopropanol.

[0016] Further, the reagent vapor pressure for steam welding is 10 - 80 kPa, and the time is 0.5 - 6 h.

[0017] The beneficial effects of adopting the further technical solution are as follows: Steam welding can enhance the bonding force between the layers of the rotator cuff patch, effectively avoid delamination, and ensure the stability of rotator cuff patch repair.

[0018] The present invention also provides a rotator cuff patch prepared by the preparation method of the rotator cuff patch.

[0019] The beneficial effects of the present invention are as follows: The rotator cuff patch prepared by the present invention undergoes a gradual transition, and the fibers gradually transition from random arrangement to aligned arrangement, effectively reducing the difference in fiber orientation structure between the random fiber layer and the aligned fiber layer. Combined with steam welding treatment, the bonding force between the layers of the patch is greatly enhanced, effectively avoiding delamination, prolonging the service life of the patch in the body, and ensuring the stability of the repair process; moreover, the tensile strength of the rotator cuff patch reaches 18 MPa, and it has excellent mechanical properties, which can not only meet the requirements for flexibility and tear resistance during suture, but also provide sufficient mechanical support for the rotator cuff during the repair process, adapting to the complex movement environment of the shoulder. Description of the Drawings

[0020] Figure 1 is a process flow chart; Figure 2 is a scanning electron micrograph of the starting fiber collection surface of the rotator cuff patch of Example 1; Figure 3 is a scanning electron micrograph of the final fiber collection surface of the rotator cuff patch of Example 1; Figure 4 is a scanning electron micrograph of the starting fiber collection surface of the rotator cuff patch of Comparative Example 1; Figure 5 is a scanning electron micrograph of the final fiber collection surface of the rotator cuff patch of Comparative Example 1; Figure 6 is a comparison chart of the tensile strength of the rotator cuff patches of Example 1 and Comparative Example 1; Figure 7 is a test result detection chart of the cell scratch experiment. Detailed Embodiments

[0021] The following describes in detail the specific embodiments of the present invention in conjunction with the embodiments.

[0022] Example 1 A rotator cuff patch, the process of which is as Figure 1 shown, and the preparation method includes the following steps: S1: Freeze-mill the acellular matrix of porcine small intestinal submucosa to obtain a substrate powder, and then soak the substrate powder in a 1 wt% pepsin solution for 24 h and freeze-dry it to obtain a pretreated substrate; S2: Disperse 1 g of the pretreated substrate into 40 g of hexafluoroisopropanol to obtain a dispersion; then dissolve 10 g of poly(lactic-co-glycolic acid) copolymer in the dispersion to obtain a spinning solution. S3: Transfer the electrospinning solution to a 20-ml syringe, install a needle with a diameter of 21G on the syringe, install the syringe on an electrospinning machine, set the distance between the needle and the collector to 15 cm, connect a positive high voltage to the needle end, connect a negative high voltage to the roller collector end, and cover the collector with aluminum foil to collect the spun film (spinning parameters: positive high voltage 10 kV, negative high voltage -1 kV, extrusion rate 1 mm / min, initial roller collector rotation speed 500 rmp, increase the roller rotation speed by 100 rmp every 30 min, and the final roller rotation speed is 2000 rmp). S4: Cut the spun film into 15 cm * 15 cm and place it in a 60 cm * 60 cm * 60 cm square acrylic container (the container contains dichloromethane), set the reagent vapor pressure to 50 kPa, and perform steam welding for 2 h. After steam welding, place the spun film in a 38 °C vacuum oven and dry for 24 h to obtain the rotator cuff patch.

[0023] Example 2 A rotator cuff patch, the process of which is as Figure 1 shown, and its preparation method includes the following steps: S1: Freeze-mill the acellular matrix of porcine small intestinal submucosa to obtain substrate powder, and then place the substrate powder in a 0.1 wt% pepsin solution and digest for 72 h, followed by freeze-drying to obtain the pretreated substrate. S2: Disperse 1 g of the pretreated substrate into a mixture of 100 g of dichloromethane and hexafluoroisopropanol (the volume ratio of dichloromethane to hexafluoroisopropanol is 1:9) to obtain a dispersion; then dissolve 10 g of poly(lactic-co-glycolic acid) copolymer in the dispersion to obtain a spinning solution. S3: Transfer the electrospinning solution to a 20-ml syringe, install a needle with a diameter of 18G on the syringe, install the syringe on an electrospinning machine, set the distance between the needle and the collector to 10 cm, connect a positive high voltage to the needle end, connect a negative high voltage to the roller collector end, and cover the collector with aluminum foil to collect the spun film (spinning parameters: positive high voltage 7 kV, negative high voltage -1 kV, extrusion rate 0.5 mm / min, initial roller collector rotation speed 500 rmp, increase the roller rotation speed by 100 rmp every 30 min, and the final roller rotation speed is 2000 rmp). S4: Cut the spun film into 15 cm * 15 cm and place it in a 60 cm * 60 cm * 60 cm square acrylic container (the container contains hexafluoroisopropanol), set the reagent vapor pressure to 10 kPa, and perform steam welding for 6 h. After steam welding, place the spun film in a 38 °C vacuum oven and dry for 24 h to obtain the rotator cuff patch.

[0024] Example 3 A rotator cuff patch, the process of which is as Figure 1 shown, and its preparation method includes the following steps: S1: Freeze-mill the acellular matrix of porcine small intestinal submucosa to obtain base material powder, and then place the base material powder in a 2wt% pepsin solution for digestion for 12 h and then freeze-dry to obtain a pretreated base material; S2: Take 1 g of the pretreated base material and disperse it in a mixed solution of 9 g of dichloromethane and hexafluoroisopropanol (the volume ratio of dichloromethane to hexafluoroisopropanol is 1:1) to obtain a dispersion; then dissolve 2.5 g of poly(lactic-co-glycolic acid) copolymer in the dispersion to obtain a spinning solution; S3: Transfer the electrospinning solution to a 20 ml syringe, install a needle with a diameter of 25G on the syringe, install the syringe on an electrospinning machine, the distance between the needle and the collector is 30 cm, connect a positive high voltage to the needle end, connect a negative high voltage to the roller receiver end, and cover the receiver with aluminum foil for collecting the spun film (the spinning parameters are: positive high voltage 12 kV, negative high voltage -1 kV, extrusion rate 1.5 mm / min, initial roller collector rotation speed 500 rmp, increase the roller speed by 100 rmp every 30 min, and the final roller speed is 2000 rmp); S4: Cut the spun film into 15 cm * 15 cm and place it in a 60 cm * 60 cm * 60 cm square acrylic container (there is hexafluoroisopropanol in the container), set the reagent vapor pressure to 80 kPa, and the time to 0.5 h for steam welding. After steam welding, place the spun film in a 38 °C vacuum oven for drying for 24 h to obtain the rotator cuff patch.

[0025] Comparative Example 1 S1: Freeze-mill the acellular matrix of porcine small intestinal submucosa to obtain base material powder, and then place the base material powder in a 1wt% pepsin solution for digestion for 24 h and then freeze-dry to obtain a pretreated base material; S2: Take 1 g of the pretreated base material and disperse it in 40 g of hexafluoroisopropanol to obtain a dispersion; then dissolve 10 g of poly(lactic-co-glycolic acid) copolymer in the dispersion to obtain a spinning solution; S3: Transfer the electrospinning solution into a 20 ml syringe. Install a needle with a diameter of 21G on the syringe, mount the syringe on the electrospinning machine, keep the distance between the needle and the collector at 15 cm, connect a positive high voltage to the needle end, connect a negative high voltage to the roller collector end, cover the collector with aluminum foil for collecting the spun film (the electrospinning parameters are: positive high voltage 10 kV, negative high voltage -1 kV, extrusion rate 1 mm / min, initial roller collector rotation speed 500 rmp, increase the roller speed by 100 rmp every 30 min, and the final roller speed is 2000 rmp); then place the spun film in a vacuum oven at 38 °C and dry for 24 h to obtain the rotator cuff patch.

[0026] Comparative Example 2 A rotator cuff patch, the process of which is as Figure 1 shown, and the preparation method includes the following steps: S1: Freeze-mill the acellular matrix of porcine small intestinal submucosa to obtain the substrate powder, then soak the substrate powder in a 1 wt% pepsin solution for digestion for 24 h and then freeze-dry to obtain the pretreated substrate; S2: Take 1 g of the pretreated substrate and disperse it in 40 g of hexafluoroisopropanol to obtain a dispersion; then dissolve 10 g of poly(lactic-co-glycolic acid) in the dispersion to obtain the electrospinning solution; S3: Transfer the electrospinning solution into a 20 ml syringe. Install a needle with a diameter of 21G on the syringe, mount the syringe on the electrospinning machine, keep the distance between the needle and the collector at 15 cm, connect a positive high voltage to the needle end, connect a negative high voltage to the roller collector end, cover the collector with aluminum foil for collecting the spun film (the electrospinning parameters are: positive high voltage 10 kV, negative high voltage -1 kV, extrusion rate 1 mm / min, initial roller collector rotation speed 500 rmp, increase the roller speed by 500 rmp every 30 min, and the final roller speed is 2000 rmp); S4: Cut the spun film into 15 cm * 15 cm and place it in a 60 cm * 60 cm * 60 cm square acrylic container (there is dichloromethane in the container), set the reagent vapor pressure to 50 kPa and the time to 2 h for steam welding. After steam welding, place the spun film in a vacuum oven at 38 °C and dry for 24 h to obtain the rotator cuff patch.

[0027] Comparative Example 3 A rotator cuff patch, the process of which is as Figure 1 shown, and the preparation method includes the following steps: S1: Freeze-mill the acellular matrix of porcine small intestinal submucosa to obtain the substrate powder, then soak the substrate powder in a 1 wt% pepsin solution for digestion for 24 h and then freeze-dry to obtain the pretreated substrate; S2: Disperse 1 g of the pretreated substrate into 40 g of hexafluoroisopropanol to obtain a dispersion; then dissolve 10 g of poly (lactic - co - glycolic acid) in the dispersion to obtain a spinning solution. S3: Transfer the electrospinning solution to a 20 - ml syringe. Install a needle with a diameter of 21G on the syringe, mount the syringe on an electrospinning machine. The distance between the needle and the collector is 15 cm. Connect a positive high voltage to the needle end and a negative high voltage to the roller receiver end. Cover the receiver with aluminum foil to collect the spun - fiber membrane (spinning parameters: positive high voltage 10 kV, negative high voltage - 1 kV, extrusion rate 1 mm / min, roller collector rotation speed 500 rmp, for 10 h). S4: Cut the spun - fiber membrane into 15 cm * 15 cm and place it into a 60 cm * 60 cm * 60 cm square acrylic container (the container contains dichloromethane). Set the reagent vapor pressure to 50 kPa and the time to 2 h for steam welding. After steam welding, put the spun - fiber membrane into a 38 °C vacuum oven and dry it for 24 h to obtain the rotator cuff patch.

[0028] Experimental Example 1 Microscopic analysis: Analyze the two surfaces of the rotator cuff patches prepared in Example 1 and Comparative Example 1 using a scanning electron microscope. The results are as Figures 2 - 5 shown. Figure 2 The randomly arranged fibers collected at a roller rotation speed of 500 rmp for the rotator cuff patch prepared in Example 1. The fiber surface is smooth without beads. Figure 3 The aligned fibers collected at a roller rotation speed of 2000 rmp. The fibers are highly oriented, the surface is smooth without beads, and after steam welding, bonding occurs at the fiber intersections for both randomly arranged fibers and aligned fibers. Figure 4 The randomly arranged fibers collected at a roller rotation speed of 500 rmp for the rotator cuff patch prepared in Comparative Example 1. The fiber surface is smooth without beads. Figure 5 The aligned fibers collected at a roller rotation speed of 2000 rmp. The fibers are highly oriented, the surface is smooth without beads, and there is no bonding at the fiber intersections for both randomly arranged fibers and aligned fibers.

[0029] Experimental Example 2 Tensile experiment: Make the rotator cuff patches prepared in Example 1 and Comparative Example 1 into specimens with a width of 10 mm, an effective tensile length of 20 mm, and a thickness of 0.5 mm. Use an American INSTRON 5966 universal material testing machine for tensile testing. Set the tensile rate to 10 mm / min and the temperature to 25 °C. The experimental results are as Figure 6 shown. It can be seen that for the rotator cuff patch prepared in Example 1, that is, after steam welding, the fiber tensile strength is significantly improved.

[0030] Detect Examples 2 - 3 and Comparative Examples 2 - 3 in the same way. The results are shown in Table 1.

[0031] Table 1 Performance Statistics Table

[0032] Experimental Example 3 Cell Scratch Assay: The effect of the rotator cuff patch on the migration ability of NIH / 3T3 cells was evaluated by the cell scratch assay. A blank control group (ordinary culture plate) and an experimental group (the rotator cuff patch obtained in Example 1 covering the culture plate) were set up, and each group repeated 3 independent experiments. The following are the specific steps: S1: The rotator cuff patch was soaked in 75% ethanol for 2 h and then rinsed 3 times with PBS, and then equilibrated in complete medium at 37 °C for 2 h to stabilize the surface properties; S2: Cells were seeded into the control group and the experimental group at a density of 5×10 5 cells / well, and both groups were cultured at 37 °C and 5% CO2 for 24 h until a confluent monolayer was formed.

[0033] S3: A sterile pipette tip was used to make a vertical scratch. In the experimental group, the force was controlled to avoid membrane damage. After standing for 10 min to eliminate membrane rebound, the integrity of the scratch was confirmed by microscope and then the serum-free medium was replaced; S4: Cultured at 37 °C and 5% CO2 for 24 h, and the scratch was photographed at fixed positions at 0 h, 12 h, and 24 h; S5: The image was converted into an 8-bit grayscale image using ImageJ software. After setting the threshold, the scratch area was calculated. The cell migration rate was calculated according to the following formula: Cell migration rate (%) = (Initial scratch area - Scratch area at time t) / Initial scratch area. The detection results are as Figure 7 shown.

[0034] Although the specific implementation manners of the present invention have been described in detail in combination with the embodiments, it should not be construed as a limitation on the protection scope of this patent. Within the scope described in the claims, various modifications and deformations that can be made by those skilled in the art without creative labor still fall within the protection scope of this patent.

Claims

1. A preparation method of a rotator cuff patch, characterized in that, It includes the following steps: S1: Freeze-mill the acellular matrix of porcine small intestinal submucosa to obtain a base material powder, and then place the base material powder in a pepsin solution for digestion for 12 - 72 h and then freeze-dry to obtain a pretreated base material; S2: Disperse the pretreated base material in Solvent 1 to obtain a dispersion; then dissolve the poly(lactic-co-glycolic acid) copolymer in the dispersion to obtain a spinning solution; S3: Perform electrospinning on the spinning solution to obtain a spun film; S4: Place the spun film in Solvent 2 for steam welding and then vacuum dry to obtain a rotator cuff patch.

2. The preparation method of the rotator cuff patch according to claim 1, characterized in that: The concentration of the pepsin solution is 0.1 wt% - 2 wt%.

3. The preparation method of the rotator cuff patch according to claim 1, characterized in that: The Solvent 1 is hexafluoroisopropanol or a mixture of dichloromethane and hexafluoroisopropanol, and the concentration of the pretreated base material in the dispersion is 0.1 wt% - 10 wt%.

4. The preparation method of the rotator cuff patch according to claim 3, wherein: In the mixture of dichloromethane and hexafluoroisopropanol, the volume ratio of dichloromethane to hexafluoroisopropanol is 1 - 5:5 - 9.

5. The preparation method of the rotator cuff patch according to claim 1, characterized in that: The concentration of the poly(lactic-co-glycolic acid) copolymer in the spinning solution is 10 wt% - 25 wt%.

6. The preparation method of the rotator cuff patch according to claim 1, wherein: The process parameters of the electrospinning are a positive high voltage of 7 - 12 kV, a negative high voltage of -1 kV, a pushing rate of 0.5 - 1.5 mm / min, a collection distance of 10 - 30 cm, and a needle diameter of 18 - 25 G.

7. The preparation method of the rotator cuff patch according to claim 1, wherein: The Solvent 2 is dichloromethane or hexafluoroisopropanol.

8. The preparation method of the rotator cuff patch according to claim 1, characterized in that: The reagent vapor pressure of the steam welding is 10 - 80 kPa and the time is 0.5 - 6 h.

9. A rotator cuff patch prepared by the preparation method of the rotator cuff patch according to any one of claims 1 - 8.

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