CN115444614A - Nanofiber yarn filling type nerve conduit and preparation method and application thereof - Google Patents
Nanofiber yarn filling type nerve conduit and preparation method and application thereof Download PDFInfo
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- CN115444614A CN115444614A CN202211121415.5A CN202211121415A CN115444614A CN 115444614 A CN115444614 A CN 115444614A CN 202211121415 A CN202211121415 A CN 202211121415A CN 115444614 A CN115444614 A CN 115444614A
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Abstract
The invention relates to the technical field of biomedical materials, in particular to a nanofiber yarn filled type nerve conduit and a preparation method and application thereof.
Description
Technical Field
The invention relates to the technical field of biomedical materials, in particular to a nanofiber yarn filled nerve conduit and a preparation method and application thereof.
Background
The damage of peripheral nervous system has great influence on daily life, with the progress of science and technology, the treatment method is also developed from the nerve suture method to the tissue engineering and gene engineering treatment method, the existing gold standard for repairing longer gaps of peripheral nerve damage is still autograft, and the method is safe and effective but has the problems of limited donor source, secondary damage to donor, insufficient donor nerve matching degree and the like. The tubular structure of the synthetic nerve graft nerve conduit is used as a bridge for connecting the near end and the far end of the nerve, can provide a good regeneration environment, promotes the growth of the regenerated nerve fiber along the conduit and connects the broken nerve, protects the broken nerve from being influenced by mechanical force, can effectively reduce the loss of various growth factors and nutrient substances in the conduit, and prevents the formation of scar tissues caused by the invasion of fibroblasts.
The ideal catheter has good biocompatibility, cell adhesion and no toxic reaction, and is beneficial to cell adhesion, growth and proliferation; certain mechanical properties, namely certain supporting capacity and certain elastic properties; the high permeability and porosity are beneficial to providing nutrition support and constructing a good regeneration environment in the pipe; the processing and molding are easy, the length of the prepared diameter is controllable, and the preparation diameter is easier to adjust according to the performance; the biodegradability can be regulated and controlled, and degradation products can not cause harm to human bodies.
The specific surface area of the tube wall of the currently prepared hollow nerve conduit is small, so that the proliferation and adhesion of cells are not facilitated, and the inside of the conduit does not have a guide factor simulating a natural nerve structure, so that the axon growth disorder is caused, and the nerve regeneration is hindered. In order to improve the repair capability of the nerve conduit, the interior of the conduit is filled with physical guide factors to simulate the internal structure of the nerve to a greater extent. The amorphous filling can only macroscopically regulate and control the growth of the guided and regenerated nerve and can not guide the directional growth of axon, and the filling of the ordered directional guide structure can effectively improve the efficiency of repairing the nerve injury.
CN104739473A and a nanofiber yarn nerve conduit and a preparation method thereof, the nerve conduit is composed of a core layer and a shell layer, wherein the core layer is nanofiber yarn, and the shell layer is a tubular nanofiber membrane; the preparation method comprises the following steps: preparing nanofiber yarns by using an electrostatic spinning technology, wherein nanofibers in the yarns are arranged in parallel along the axial direction of the yarns; and arranging the nanofiber yarns in parallel to coat a nanofiber membrane to form the nerve conduit. The nerve conduit prepared by the patent has good biocompatibility, can promote adhesion, spreading and proliferation of nerve cells, wherein the parallel nanofibers on the surfaces of the nanofiber yarns are beneficial to oriented growth of the nerve cells, and the conduits with different sizes can be customized according to actual conditions to meet the requirement of peripheral nerve repair. But there are substantially no gaps between the yarns, which is not conducive to cell and tissue entry.
Disclosure of Invention
Aiming at the defects in the prior art, the nanofiber yarn filling type nerve conduit, the preparation method and the application thereof are provided, the nerve conduit with long-distance nerve injury can be repaired, the adhesion, the growth and the proliferation of cells can be guided in a directional mode, the accurate connection of the stumps is facilitated, and the mismatching rate is reduced.
In order to solve the technical problems, the technical scheme adopted by the invention is that the nanofiber yarn filled type nerve conduit comprises an external nerve conduit and a nanofiber yarn bundle filled in the external nerve conduit, wherein the external nerve conduit is of a hollow structure, the nanofiber yarn bundle is formed by arranging highly oriented nanofiber yarns, and the nanofiber yarn filled type nerve conduit is of a three-dimensional nanofiber structure.
The diameter of the nanofiber yarn filled nerve conduit is 1-50mm.
The nanofiber yarn filled nerve conduit has the diameter of 50-500 mu m.
The nanofiber yarn filled nerve conduit comprises the following steps:
(1) Obtaining a nanofiber membrane through electrostatic spinning, placing a cylindrical die with a required size on the prepared nanofiber membrane, winding the nanofiber membrane into a hollow nerve conduit by using the die, wetting the nerve conduit with alcohol, removing the nanofiber conduit, freezing the nerve conduit at-18-23 ℃ overnight or at-75-82 ℃ for 1-6 hours, taking out, and freeze-drying for 1-7 days to obtain the nerve conduit;
(2) The electrostatic spinning is directly received on the roller, the nanofiber membrane obtained on the roller is wound into yarn along the axial direction of the roller, and the yarn is cut and taken down;
(3) And (3) finishing the nanofiber yarns obtained in the step (2) into bundles and filling the bundles into the nanofiber catheters obtained in the step (1), so as to obtain the nanofiber catheters.
In the nanofiber yarn filled nerve conduit, the electrostatic spinning process parameters in the step (1) are as follows: the voltage is 5-100 kV, the receiving distance is 5-30 cm, and the spinning speed is 0.1-5 mL/h.
In the nanofiber yarn filled nerve conduit, the electrostatic spinning in the step (2) has the following process parameters: voltage 5-100 kV, receiving distance 5-30 cm, spinning speed: 0.1-5 mL/h.
In the step (1) and the step (2), silk Fibroin (SF), gelatin, polylactic acid (PLA), polycaprolactone (PCL), polydioxanone (PPDO), polylactic acid-Polycaprolactone (PLCL), glycolide-lactide copolymer (PLGA) or a plurality of them are used as raw materials, or medicines, bioactive factors and the like are further added, and electrostatic spinning is performed.
The application of the nanofiber yarn filling type nerve conduit is used for nerve conduit stents, in particular to the defect repair of large-segment peripheral nerves and the regeneration and reconstruction of nerve functions.
The nanofiber yarn filled type nerve conduit and the preparation method and application thereof have the beneficial effects that the ultramicro structure of a multi-stage nerve trunk is highly simulated, the 3D structure is filled in the external nerve conduit, the physical contact guiding effect on nerve regeneration is exerted, the natural nerve multi-stage ultramicro structure is highly simulated, and the nanofiber yarn bundle has a higher specific surface area, so that the adhesion, the oriented growth and the proliferation of cells and the maintenance of cell phenotype can be effectively promoted. The hollow nerve conduit can provide a favorable microenvironment for nerve regeneration, and scar tissues outside the conduit are prevented from entering the body.
The nerve conduit can directionally guide the adhesion, growth and proliferation of cells, is favorable for accurately connecting stumps and reduces the mismatching rate.
The nanofiber structure of nerve conduits mimics the nanostructure of extracellular matrix, and the micro-and nanostructures implanted in the conduits can effectively modulate injury-induced biological responses, including inflammation and scar formation. The nanofiber yarn bundle highly simulates a multistage ultramicro structure of natural nerves from nerve fibers to nerve bundles and then to nerve trunks, and provides an excellent intravascular environment for nerve regeneration. The nanofiber yarn has high specific surface area to volume ratio, porosity and higher cell permeability, and effectively promotes the adhesion, oriented growth and proliferation of Schwann cells and the maintenance of cell phenotype. The increased number of nanofibers is more favorable for improved chemical absorption and overall flexibility. Therefore, filling a highly oriented nanofiber yarn bundle in a nerve conduit will more efficiently repair peripheral nerve long distance injuries.
The nano fiber yarn bundle in the tube can play a role of directional contact guidance, guide the directional regeneration of the nerve axons at the proximal and distal broken ends, and cells can adhere, grow directionally, proliferate and maintain the cell phenotype along the nano fiber yarn. In the process of the growth of the regenerated nerve, the nanofiber yarn is gradually degraded to provide a growth space for the regenerated nerve.
The hollow nerve conduit provides a relatively closed internal environment for nerve regeneration, protects the new nerve from the influence of external mechanical force, can effectively reduce the loss of various growth factors and nutrient substances in the conduit, prevents the invasion of fibroblasts from causing the generation of neuroma and scars, ensures that the nerve regeneration is not hindered, can be degraded along with the damaged nerve after the damaged nerve is well recovered, and can be absorbed by human body without causing damage to the health of the human body.
The stent material of the invention is made of one or more natural or synthetic polymer materials with good biodegradability and biocompatibility. The material of the outer tube of the nerve conduit is selected from the material with better mechanical property, which is enough to support the regeneration of nerves. The degradation speed can be regulated and controlled.
The invention mainly researches the structure of the bracket, optimizes the design of the nerve conduit, prepares the nerve conduit bracket with excellent performance, explores the application prospect of the nerve conduit bracket in nerve repair and provides a new idea for repairing the defect of the nerve.
The invention skillfully utilizes the electrostatic spinning technology, and the prepared product has excellent physical and chemical properties, good biocompatibility, biodegradability, a highly bionic nerve bundle membrane structure and a highly oriented nanofiber yarn bundle filling oriented nerve conduit stent structure.
The yarn in the external nerve conduit has a larger space, so that enough space is provided for the entering of cells and tissues, and the entering of the cells and the tissues is more facilitated.
Because the oriented nanofiber yarn filling type nerve conduit is applied to the field of nerve tissue engineering and is filled to a nerve defect part to guide nerve regeneration, the structure and the performance of the nerve conduit can influence the growth of nerve cells and the regeneration of nerve tissues, and further influence the recovery of the function of regenerated nerves. The nerve conduit takes the nanofiber yarn bundle as an internal filling structure, the nanofiber yarn bundle is highly bionic in the nerve bundle structure, and the highly oriented structure can have a good regulation and control effect on the growth of nerve cells and can better guide nerve regeneration. The method has good effect on repairing long-distance damaged nerves, particularly repairing nerve defects with a long distance exceeding 10 cm.
Drawings
FIG. 1 is a process flow diagram of the present invention;
FIG. 2 is a schematic cross-sectional view of the present invention;
FIG. 3 is a cross-sectional SEM image of PLLA neuro-catheter-PLLA (6 ten thousand) nanofiber yarn bundle made in example 2;
FIG. 4 is an SEM image of PLLA (30 ten thousand) nanofiber yarns loaded with different drug contents in PLLA nerve conduits prepared in example 3;
FIG. 5 is a graph showing the tensile mechanical properties of the nanofiber yarn bundle-filled neurovascular catheters obtained in examples 2 to 4.
Detailed Description
The invention is described in detail below with reference to the figures and the embodiments.
Example 1
The nanofiber yarn filled type nerve conduit comprises an external nerve conduit and a nanofiber yarn bundle filled in the external nerve conduit, wherein the external nerve conduit is of a hollow structure, the nanofiber yarn bundle is formed by arranging highly oriented nanofiber yarns, and the nanofiber yarn filled type nerve conduit is of a three-dimensional nanofiber structure.
The diameter of the nanofiber yarn filling type nerve conduit is 1-50mm. The highly oriented nanofiber yarn has a diameter of 50-500 μm.
The nanofiber yarn filled nerve conduit comprises the following steps:
(1) Obtaining a nanofiber membrane through electrostatic spinning, placing a cylindrical die with a required size on the prepared nanofiber membrane, winding the nanofiber membrane into a hollow nerve conduit by using the die, wetting the hollow nerve conduit by using 10-100% alcohol, taking the nanofiber conduit down, freezing the nanofiber conduit at-18-23 ℃ overnight or at-75-82 ℃ for 1-6 hours, taking out, and performing freeze drying for 1-7 days to obtain the nerve conduit;
(2) The electrostatic spinning is directly received on a roller, the nanofiber membrane obtained on the roller is wound into yarn along the axial direction of the roller, and the yarn is cut and taken down;
(3) And (3) finishing the nanofiber yarns obtained in the step (2) into bundles and filling the bundles into the nanofiber catheters obtained in the step (1), so as to obtain the nanofiber catheters.
The electrostatic spinning process parameters in the step (1) are as follows: the voltage is 5-100 kV, the receiving distance is 5-30 cm, and the spinning speed is 0.1-5 mL/h.
The electrostatic spinning process parameters in the step (2) are as follows: voltage 5-100 kV, receiving distance 5-30 cm, spinning speed: 0.1-5 mL/h.
In the steps (1) and (2), one or more of Silk Fibroin (SF), gelatin, polylactic acid (PLA), polycaprolactone (PCL), polydioxanone (PPDO), polylactic acid-Polycaprolactone (PLCL) and glycolide-lactide copolymer (PLGA) are used as raw materials, or medicines, bioactive factors and the like are further added, and electrostatic spinning is carried out.
The application of the nanofiber yarn filled nerve conduit is used for nerve conduit stents, in particular to defect repair of large-segment peripheral nerves and regeneration and reconstruction of nerve functions.
Example 2:
the same parts of this embodiment as those of embodiment 1 are not described again, and the differences are:
the nanofiber yarn filled nerve conduit comprises the following steps:
(1) Dissolving 1g of PLLA with the molecular weight of 30 ten thousand in 10mL of hexafluoroisopropanol solution, continuously dissolving until a uniform solution is obtained, carrying out electrostatic spinning to obtain a nanofiber membrane, and winding the nanofiber membrane into an external nerve conduit through a die. Spinning conditions are as follows: the voltage is 12kV, the receiving distance is 20cm, and the spinning speed is 0.8mL/h. A nerve conduit of 2.5mm in diameter and 18mm in length was obtained.
(2) 3g of PLLA with the molecular weight of 6 ten thousand is dissolved in 10mL of hexafluoroisopropanol solution, the solution is continuously dissolved until a uniform solution is obtained, electrostatic spinning is carried out to directly send the solution to a receiving roller, and a nanofiber membrane received on the roller is wound into nanofiber yarns. Spinning conditions are as follows: voltage 12kV, reception distance 18cm, spinning speed: 1mL/h.
(3) And (3) arranging the nanofiber yarns obtained in the step (2) into bundles and filling the bundles into the nerve conduit obtained in the step (1), so as to obtain the nerve conduit filled with the oriented nanofiber yarn bundles with the inner diameter of 2.5 mm.
Example 3:
the same parts of this embodiment as embodiment 1 are not described again, but the differences are:
the nanofiber yarn filled nerve conduit comprises the following steps:
(1) Dissolving 1g of PLLA with the molecular weight of 30 ten thousand in 10ml of hexafluoroisopropanol solution, continuously dissolving until a uniform solution is obtained, carrying out electrostatic spinning to obtain a nanofiber membrane, and winding the nanofiber membrane into an external nerve conduit through a die. Spinning conditions are as follows: the voltage is 12kV, the receiving distance is 20cm, and the spinning speed is 1.2ml/h. A nerve conduit of 2.5mm in diameter and 10mm in length was obtained.
(2) Ultrasonically dispersing 0.1g or 0.5g of curcumin in 10ml of hexafluoroisopropanol solution, dissolving 3g of PLLA with the molecular weight of 6 ten thousand in the hexafluoroisopropanol solution containing the curcumin, carrying out electrostatic spinning to directly reach a receiving roller, and winding the nanofiber membrane received on the roller into a nanofiber-containing yarn. Spinning conditions are as follows: voltage 14kV, reception distance 18cm, spinning speed: 1mL/h.
(3) And (3) arranging the nanofiber yarns obtained in the step (2) into bundles and filling the bundles into the nerve conduit obtained in the step (1), so as to obtain the nerve conduit filled with the oriented nanofiber yarn bundles with the inner diameter of 2.5 mm.
As shown in fig. 4, the addition of curcumin reduced the diameter of the nanofiber yarn and nanofibers in the yarn that were finally obtained. The nano-fibers in all yarns are in a highly oriented structure, and the structure of the nano-fibers in peripheral nerve tissues can be well simulated.
Example 4:
the same parts of this embodiment as those of embodiment 1 are not described again, and the differences are:
the nanofiber yarn filled nerve conduit comprises the following steps:
(1) Dissolving 1g of PCL in 10mL of hexafluoroisopropanol solution, continuously dissolving until a uniform solution is obtained, carrying out electrostatic spinning to obtain a nanofiber membrane, and winding the nanofiber membrane into an external nerve conduit through a die. Spinning conditions are as follows: the pressure is 13kV, the receiving distance is 17cm, and the spinning speed is 0.1mL/h. A nerve conduit of 2.5mm in diameter and 50mm in length was obtained.
(2) Dissolving 1g PCL/SF in 10mL hexafluoroisopropanol solution, continuously dissolving until uniform solution is obtained, carrying out electrostatic spinning directly to a receiving roller, and winding the nanofiber membrane received on the roller into nanofiber yarn. Spinning conditions are as follows: voltage 13kV, receiving distance 17cm, spinning speed: 0.9mL/h.
(3) And (3) arranging the nanofiber yarns obtained in the step (2) into bundles and filling the bundles into the nerve conduit obtained in the step (1), so as to obtain the nerve conduit filled with the oriented nanofiber yarn bundles with the inner diameter of 2.5 mm.
It is to be understood that the above description is not intended to limit the present invention, and the present invention is not limited to the above examples, and that various changes, modifications, additions and substitutions which may be made by one of ordinary skill in the art within the spirit and scope of the present invention are also within the scope of the present invention.
Claims (8)
1. A nanofiber yarn filling type nerve conduit is characterized in that: the external nerve conduit is of a hollow structure, the nanofiber yarn bundle is formed by arranging highly oriented nanofiber yarns, and the nanofiber yarn filled type nerve conduit is of a three-dimensional nanofiber structure.
2. The nanofiber yarn-packed nerve conduit as claimed in claim 1, wherein the diameter thereof is 1-50mm.
3. The nanofiber yarn-filled nerve conduit as claimed in claim 2, wherein the diameter of the highly oriented nanofiber yarn is 50-500 μm.
4. The nanofiber yarn-filled nerve conduit as claimed in any one of claims 1 to 3, wherein: the method comprises the following steps:
(1) Obtaining a nanofiber membrane through electrostatic spinning, placing a cylindrical die with a required size on the prepared nanofiber membrane, winding the nanofiber membrane into a hollow nerve conduit by using the die, wetting the nerve conduit with alcohol, removing the nanofiber conduit, freezing the nerve conduit at-18-23 ℃ overnight or at-75-82 ℃ for 1-6 hours, taking out, and freeze-drying for 1-7 days to obtain the nerve conduit;
(2) The electrostatic spinning is directly received on the roller, the nanofiber membrane obtained on the roller is wound into yarn along the axial direction of the roller, and the yarn is cut and taken down;
(3) And (3) finishing the nanofiber yarns obtained in the step (2) into bundles and filling the bundles into the nanofiber conduits prepared in the step (1).
5. The nanofiber yarn filled nerve conduit according to claim 4, wherein the electrostatic spinning in the step (1) comprises the following process parameters: the voltage is 5-100 kV, the receiving distance is 5-30 cm, and the spinning speed is 0.1-5 mL/h.
6. The nanofiber yarn filled nerve conduit as claimed in claim 5, wherein the electrostatic spinning in step (2) has the following technological parameters: voltage 5-100 kV, receiving distance 5-30 cm, spinning speed: 0.1-5 mL/h.
7. The nanofiber yarn filled nerve conduit of claim 6, wherein in the steps (1) and (2), one or more of Silk Fibroin (SF), gelatin, polylactic acid (PLA), polycaprolactone (PCL), polydioxanone (PPDO), polylactic acid-Polycaprolactone (PLCL), and poly (lactide-co-lactide) (PLGA) are used as raw materials, or drugs, bioactive factors and the like are further added to carry out electrostatic spinning.
8. The application of the nanofiber yarn filling type nerve conduit is characterized in that: the nerve conduit stent is used for nerve conduit stents, particularly for defect repair of large-segment peripheral nerves and regeneration and reconstruction of nerve functions.
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