A double-layer core-shell structure artificial fibrous dura mater for spinal cord injury and a preparation method thereof

Abstract

The present application belongs to the technical field of medical devices, and relates to medical consumables for treating spinal cord injury, in particular to a double-layer core-shell structure artificial fiber dura mater for spinal cord injury and a preparation method thereof. The double-layer core-shell structure artificial fiber is used to form a film structure, the double-layer core-shell structure artificial fiber is composed of an outer shell structure and an inner core structure, the material of the outer shell structure is composed of PLGA and PEG, the outer shell structure is loaded with NM, the material of the inner core structure is PLA, and the inner core structure is loaded with NT3. The double-layer core-shell structure artificial fiber dura mater provided by the present application has the advantages of reducing spinal cord edema, preventing fiber adhesion, improving the microenvironment, promoting the repair of spinal cord injury, and the like.

Description

Technical Field

[0001] This invention belongs to the field of medical device technology and relates to medical consumables for the treatment of spinal cord injury, specifically to a double-layered core-shell structure artificial fibrous dura mater for spinal cord injury and its preparation method. Background Technology

[0002] The information disclosed in this background section is intended only to enhance understanding of the overall background of the invention and is not necessarily to be construed as an admission or in any way implying that such information constitutes prior art known to those skilled in the art.

[0003] Spinal cord injury (SCI) is a severe injury to the central nervous system, characterized by high rates of disability and mortality. It can cause not only loss of motor and sensory function but also multiple organ failure leading to death. While external compression caused by fractures and dislocations can be treated surgically, simple laminectomy neglects the restraining role of the dura mater and cannot break the vicious cycle or effectively reduce intramedullary pressure. Therefore, early dural endoscopic myotomy to relieve dural restraint is a necessary means to break this vicious cycle and is key to effectively repairing spinal cord injuries.

[0004] The imbalance of the local microenvironment following spinal cord injury (SCI) triggers a cascade of inflammatory responses, which is a major reason why surgical treatment for structural reconstruction often fails to restore function. Microenvironmental imbalance is a primary cause of hindering SCI regeneration and recovery. At the tissue level, this imbalance manifests as hemorrhage and local ischemia, glial scar formation, and an imbalance between demyelination and remyelination. At the cellular level, it involves the differentiation of endogenous stem cells and microglia. At the molecular level, it includes imbalances in neurotrophic factors and their propeptides, cytokines, inflammatory factors, and chemokines. Summary of the Invention

[0005] To address the shortcomings of existing technologies, the present invention aims to provide a double-layered core-shell structured artificial fibrous dura mater for spinal cord injury and its preparation method. The double-layered core-shell structured artificial fibrous dura mater provided by the present invention has advantages such as reducing spinal cord edema, preventing fiber adhesion, improving the microenvironment, and promoting spinal cord injury repair.

[0006] To achieve the above objectives, the technical solution of the present invention is as follows:

[0007] In a first aspect, there is a double-shell artificial fibrous dura mater for spinal cord injury, which is composed of a membrane structure made of double-shell artificial fibers. The double-shell artificial fibers are composed of an outer shell structure and an inner core structure. The outer shell structure is made of polylactic-co-glycolic acid (PLGA) and polyethylene glycol (PEG), and the outer shell structure is loaded with naphthostat mesylate (NM). The inner core structure is made of polylactic acid (PLA), and the inner core structure is loaded with neurotrophic factor 3 (NT3).

[0008] According to the inventor's research, the main problems with using medication to restore function after surgical reconstruction of the reconstructed structure are as follows: (1) The blood-spinal barrier prevents drugs from entering the cerebrospinal fluid. The blood-spinal barrier can provide a stable microenvironment for the central nervous system and prevent the invasion of bacteria, viruses, etc., but it also brings great difficulties to the application of drugs. In order to achieve an effective therapeutic concentration of drugs in the cerebrospinal fluid, it is necessary to increase the dose of systemic administration so that more drugs can cross the blood-spinal barrier, which also brings a lot of side effects to patients. (2) Oral drugs are first absorbed through the digestive tract mucosa and then enter the blood circulation system. It is difficult to achieve an effective drug concentration at the site of injury. In addition, the kidneys clear the drugs, which makes the drug stay in the body for too short a time and the metabolic rate too fast. Moreover, the local injection method has the significant disadvantage that the drug cannot accumulate and be released at the site of injury.

[0009] Therefore, this invention loads drugs onto an artificial dura mater to allow the drugs to act on the affected area in situ, thereby bypassing the blood-spinal cord barrier and avoiding the systemic side effects of oral medication. Firstly, this invention selects PLA and PLGA, which have anti-adhesion properties, not only preventing fibrous adhesion but also ensuring that the thickness and mechanical properties of the prepared artificial membrane meet the requirements of the dura mater. Secondly, this invention uses PLGA and PEG as the outer shell structure materials, which have the advantages of rapid degradation and high hydrophilicity, and PLA as the inner core structure material, which has a relatively long degradation time. Loading NM and NT3 onto the outer shell structure and inner core structure respectively allows the dura mater to sequentially release NM, which has a stabilizing effect on the microenvironment, and NT3, which stimulates neurogenesis. This bypasses the blood-spinal cord barrier and avoids the systemic side effects of oral medication, thereby reducing spinal cord edema, improving the microenvironment, and promoting spinal cord injury repair.

[0010] Secondly, a method for preparing the above-mentioned double-layered core-shell structured artificial fibrous dura mater for spinal cord injury includes the following steps:

[0011] PLGA, PEG and NM were dissolved to prepare a shell solution;

[0012] PLA and NT3 were dissolved to prepare a nucleo-fluid mixture;

[0013] The shell liquid and the core liquid are coaxially spun and then cooled and dried to obtain the final product.

[0014] The beneficial effects of this invention are as follows:

[0015] 1. This invention uses PLA and PLGA as the main materials for dura mater, which not only have anti-adhesion properties, but also the thickness and mechanical properties of the dura mater prepared with them meet the requirements for use.

[0016] 2. This invention uses PLGA as the main material for the outer shell structure and introduces PEG, which makes the outer shell structure have the characteristics of fast degradation and high hydrophilicity. PLA is used as the material for the inner core structure, so that the degradation rate of the inner core structure is lower than that of the outer shell structure. Then, NM and NT3 are loaded onto the outer shell structure and the inner core structure respectively, thereby realizing the sequential release of NM and NT3. This allows the artificial dura mater to act on the affected area in situ after drug loading, bypassing the blood-spinal cord barrier and avoiding the systemic side effects of oral medication. At the same time, it reduces spinal cord edema, improves the microenvironment, and promotes spinal cord injury repair.

[0017] In summary, the double-layered core-shell structured artificial fibrous dura mater provided by this invention can realize a novel multi-intervention repair mode for temporary treatment after spinal cord injury, rapid decompression, restoration of microenvironmental stability, reduction of spinal cord edema, and prevention of fibrous adhesion, providing support for the final clinical translation and commercialization. Attached Figure Description

[0018] The accompanying drawings, which form part of this invention, are used to provide a further understanding of the invention. The illustrative embodiments of the invention and their descriptions are used to explain the invention and do not constitute an improper limitation of the invention.

[0019] Figure 1 Images of the dura mater prepared in Examples 2 and 3 of the present invention, a is Example 2, b is Example 3;

[0020] Figure 2 SEM image of the dura mater prepared in Example 1 of this invention;

[0021] Figure 3 SEM image of the dura mater prepared in Example 1 of this invention;

[0022] Figure 4 The figures show the hydrophilicity test results of the dura maters prepared in Examples 1, 4, 5 and the comparative examples of the present invention;

[0023] Figure 5 The figures show the mechanical properties of the dura mater prepared in Examples 1, 2, and 3 of this invention. The thickness of the dura mater prepared in Example 1 is 0.32 mm, the thickness of the dura mater prepared in Example 2 is 0.15 mm, and the thickness of the dura mater prepared in Example 3 is 0.53 mm.

[0024] Figure 6 The images are fluorescent staining results of the dura mater loaded with neural stem cells prepared in Example 1 of this invention, with a blank control and b being the dura mater loaded with neural stem cells.

[0025] Figure 7 This is a graph showing the results of cell proliferation experiments performed on cells seeded onto different samples in the experimental examples of this invention;

[0026] Figure 8 The results of PCR tests on specific neural markers performed on cells seeded onto different samples in the experimental examples of this invention are shown. Tuj1 (a) and MAP2 (b) are related microtubule protein markers, and GFAP (c) is an astrocyte marker.

[0027] Figure 9 The image shows the PCR test results of specific inflammatory markers performed on cells seeded onto different samples in the experimental examples of this invention, where iNOS (a) and IL-1β (b) are inflammatory microglial cell markers;

[0028] Figure 10 The images show the tissue immunofluorescence results of mice using different fibrous membranes and whether or not a fibrous membrane was used in the experimental examples of this invention. iNOS and IBA-1 are inflammatory microglial cell markers. The scale bar is 100 μm.

[0029] Figure 11 The images show the tissue immunofluorescence results of mice using different fibrous membranes and whether or not fibrous membranes were used in the experimental examples of this invention. MAP2 is a microtubule-associated protein marker, and GFAP is an astrocyte marker. The scale bar is 100 μm.

[0030] Figure 12 The figures show the results of the inclined plane test and muscle weight measurement of mice using different fiber membranes in the experimental examples of this invention. a is the inclined plane test, which can reflect the behavioral ability of mice, and b is the muscle weight, which can reflect the motor function of mice. Detailed Implementation

[0031] It should be noted that the following detailed descriptions are exemplary and intended to provide further illustration of the invention. Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains.

[0032] It should be noted that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to limit the scope of exemplary embodiments according to the invention. As used herein, the singular form is intended to include the plural form as well, unless the context clearly indicates otherwise. Furthermore, it should be understood that when the terms "comprising" and / or "including" are used in this specification, they indicate the presence of features, steps, operations, devices, components, and / or combinations thereof.

[0033] In view of the problem that surgical treatment of spinal cord injury restores structural reconstruction but has difficulty in restoring function, this invention proposes a double-layered core-shell structured artificial fibrous dura mater for spinal cord injury and its preparation method.

[0034] In a typical embodiment of the present invention, a double-shell artificial fiber dura mater for spinal cord injury is provided, which is composed of a membrane structure made of double-shell artificial fibers. The double-shell artificial fibers are composed of an outer shell structure and an inner core structure. The outer shell structure is made of PLGA and PEG and is loaded with NM. The inner core structure is made of PLA and is loaded with NT3.

[0035] In some embodiments, PEG is 5 to 20% of the total weight of PLGA and PEG.

[0036] In some embodiments, the molar ratio of lactic acid units to glycolic acid in the PLGA is 1:0.8~1.2.

[0037] In some embodiments, the weight-average molecular weight of PLGA is 880,000 to 1,170,000.

[0038] In some embodiments, the weight-average molecular weight of PLA is 100,000 to 130,000.

[0039] In some embodiments, the weight of NM is 8 to 12% of the weight of the outer shell structure.

[0040] In some embodiments, the ratio of NT3 added to the weight of the inner core structure is 0.8~1.2:0.1, μL: g.

[0041] In some embodiments, the weight ratio of the outer shell structure to the inner core structure is 3.5 to 4.5:1.

[0042] In some embodiments, the thickness is 0.1 to 0.5 mm.

[0043] Another embodiment of the present invention provides a method for preparing the above-mentioned double-layered core-shell structured artificial fibrous dura mater for spinal cord injury, comprising the following steps:

[0044] PLGA, PEG and NM were dissolved to prepare a shell solution;

[0045] PLA and NT3 were dissolved to prepare a nucleolytic solution;

[0046] The shell liquid and the core liquid are coaxially spun and then cooled and dried to obtain the final product.

[0047] In some embodiments, the solvent for the shell liquid is hexafluoroisopropanol.

[0048] In some embodiments, the total weight ratio of PLGA and PEG to the volume ratio of solvent in the shell solution is 0.8~1.2:5, g:ml.

[0049] In some embodiments, the solvent for the nucleus fluid is hexafluoroisopropanol.

[0050] In some embodiments, the weight ratio of PLA to solvent in the nuclear liquid is 0.8~1.2:8, g: ml.

[0051] In some embodiments, during coaxial spinning, the flow rate ratio of shell liquid to core liquid is 3.8 to 4.2:1.

[0052] In some embodiments, during coaxial spinning, the ratio of the inner tube diameter to the outer tube diameter of the spinning nozzle is 1:1.5~2.0. Specifically, the inner tube diameter of the spinning nozzle is 0.38~0.40 mm. Specifically, the outer tube diameter of the spinning nozzle is 0.70~0.75 mm.

[0053] In some embodiments, the cooling and drying time is 45-50 h. After freeze-drying, store at 3-5 °C for use and at -22 to -18 °C for storage.

[0054] To enable those skilled in the art to better understand the technical solution of the present invention, the technical solution of the present invention will be described in detail below with reference to specific embodiments.

[0055] Example 1

[0056] A method for preparing a double-layered core-shell structured artificial fibrous dura mater for spinal cord injury, comprising the following steps:

[0057] (1) Using PLGA (50:50, Mw = 880000-1170000) and PEG (Mn = 6000) as shell matrix, with PEG having a mass percentage of 20%, the shell matrix and hexafluoroisopropanol were mixed and dissolved at a mass / volume ratio of 1:5, g: ml. Then, NM was added at a mass ratio of 1:10, g: ml to NM and shell matrix, and the mixture was ultrasonically dispersed for 1 min to obtain the shell liquid.

[0058] (2) PLA (Mw = 100000-130000) and hexafluoroisopropanol were mixed and dissolved at a mass / volume ratio of 1:8. Then, NT3 was added at a ratio of 1 μg NT3 per 0.1g PLA and ultrasonically dispersed for 1 min to obtain the nucleolysin.

[0059] (3) Connect two independent syringes, the core liquid and the shell liquid, with a coaxial spinning nozzle. The inner tube diameter of the spinning nozzle is 0.40 mm, the flow rate is 0.5 mL / h, the outer tube diameter is 0.72 mm, the flow rate is 2 mL / h, the ambient temperature is 25℃, the ambient humidity is 65%, the distance from the spinning nozzle to the receiver is 18 mm, and the spinning time is 3 h.

[0060] (4) The sample was collected on aluminum foil, freeze-dried for 24 h, and stored at 4℃ when used and at -20℃ when stored.

[0061] Example 2

[0062] A method for preparing a double-layered core-shell structured artificial fibrous dura mater for spinal cord injury, comprising the following steps:

[0063] (1) Using PLGA (50:50, Mw = 880000-1170000) and PEG (Mn = 6000) as shell matrix, with PEG having a mass percentage of 20%, the shell matrix and hexafluoroisopropanol were mixed and dissolved at a mass / volume ratio of 1:5, g: ml. Then, NM was added at a mass ratio of 1:10, g: ml to NM and shell matrix, and the mixture was ultrasonically dispersed for 1 min to obtain the shell liquid.

[0064] (2) PLA (Mw = 100000-130000) and hexafluoroisopropanol were mixed and dissolved at a mass / volume ratio of 1:8. Then, NT3 was added at a ratio of 1 μg NT3 per 0.1g PLA and ultrasonically dispersed for 1 min to obtain the nucleolysin.

[0065] (3) Connect two independent syringes, the core liquid and the shell liquid, with a coaxial spinning nozzle. The inner tube diameter of the spinning nozzle is 0.40 mm, the flow rate is 0.5 mL / h, the outer tube diameter is 0.72 mm, the flow rate is 2 mL / h, the ambient temperature is 25℃, the ambient humidity is 65%, the distance from the spinning nozzle to the receiver is 18 mm, and the spinning time is 1 h.

[0066] (4) The sample was collected on aluminum foil, freeze-dried for 24 h, and stored at 4℃ when used and at -20℃ when stored.

[0067] Example 3

[0068] A method for preparing a double-layered core-shell structured artificial fibrous dura mater for spinal cord injury, comprising the following steps:

[0069] (1) Using PLGA (50:50, Mw = 880000-1170000) and PEG (Mn = 6000) as shell matrix, with PEG having a mass percentage of 20%, the shell matrix and hexafluoroisopropanol were mixed and dissolved at a mass / volume ratio of 1:5, g: ml. Then, NM was added at a mass ratio of 1:10, g: ml to NM and shell matrix, and the mixture was ultrasonically dispersed for 1 min to obtain the shell liquid.

[0070] (2) PLA (Mw = 100000-130000) and hexafluoroisopropanol were mixed and dissolved at a mass / volume ratio of 1:8. Then, NT3 was added at a ratio of 1 μg NT3 per 0.1g PLA and ultrasonically dispersed for 1 min to obtain the nucleolysin.

[0071] (3) Connect two independent syringes, the core liquid and the shell liquid, with a coaxial spinning nozzle. The inner tube diameter of the spinning nozzle is 0.40 mm, the flow rate is 0.5 mL / h, the outer tube diameter is 0.72 mm, the flow rate is 2 mL / h, the ambient temperature is 25℃, the ambient humidity is 65%, the distance from the spinning nozzle to the receiver is 18 mm, and the spinning time is 5 h.

[0072] (4) The sample was collected on aluminum foil, freeze-dried for 24 h, and stored at 4℃ when used and at -20℃ when stored.

[0073] Example 4

[0074] A method for preparing a double-layered core-shell structured artificial fibrous dura mater for spinal cord injury, comprising the following steps:

[0075] (1) Using PLGA (50:50, Mw = 880000-1170000) and PEG (Mn = 6000) as shell matrix, with PEG having a mass percentage of 5%, the shell matrix and hexafluoroisopropanol were mixed and dissolved at a mass / volume ratio of 1:5, g: ml. Then, NM was added at a mass ratio of 1:10, g: ml to NM and shell matrix, and the mixture was ultrasonically dispersed for 1 min to obtain the shell liquid.

[0076] (2) PLA (Mw = 100000-130000) and hexafluoroisopropanol were mixed and dissolved at a mass / volume ratio of 1:8. Then, NT3 was added at a ratio of 1 μg NT3 per 0.1g PLA and ultrasonically dispersed for 1 min to obtain the nucleolysin.

[0077] (3) Connect two independent syringes, the core liquid and the shell liquid, with a coaxial spinning nozzle. The inner tube diameter of the spinning nozzle is 0.40 mm, the flow rate is 0.5 mL / h, the outer tube diameter is 0.72 mm, the flow rate is 2 mL / h, the ambient temperature is 25℃, the ambient humidity is 65%, the distance from the spinning nozzle to the receiver is 18 mm, and the spinning time is 3 h.

[0078] (4) The sample was collected on aluminum foil, freeze-dried for 24 h, and stored at 4℃ when used and at -20℃ when stored.

[0079] Example 5

[0080] A method for preparing a double-layered core-shell structured artificial fibrous dura mater for spinal cord injury, comprising the following steps:

[0081] (1) Using PLGA (50:50, Mw = 880000-1170000) and PEG (Mn = 6000) as shell matrix, with PEG having a mass percentage of 10%, the shell matrix and hexafluoroisopropanol were mixed and dissolved at a mass / volume ratio of 1:5, g: ml. Then, NM was added at a mass ratio of 1:10, g: ml to NM and shell matrix, and the mixture was ultrasonically dispersed for 1 min to obtain the shell liquid.

[0082] (2) PLA (Mw = 100000-130000) and hexafluoroisopropanol were mixed and dissolved at a mass / volume ratio of 1:8. Then, NT3 was added at a ratio of 1 μg NT3 per 0.1g PLA and ultrasonically dispersed for 1 min to obtain the nucleolysin.

[0083] (3) Connect two independent syringes, the core liquid and the shell liquid, with a coaxial spinning nozzle. The inner tube diameter of the spinning nozzle is 0.40 mm, the flow rate is 0.5 mL / h, the outer tube diameter is 0.72 mm, the flow rate is 2 mL / h, the ambient temperature is 25℃, the ambient humidity is 65%, the distance from the spinning nozzle to the receiver is 18 mm, and the spinning time is 3 h.

[0084] (4) The sample was collected on aluminum foil, freeze-dried for 24 h, and stored at 4℃ when used and at -20℃ when stored.

[0085] Comparative Example

[0086] A method for preparing a double-layered core-shell structured artificial fibrous dura mater for spinal cord injury, comprising the following steps:

[0087] (1) Using PLGA (50:50, Mw = 880000-1170000) as the shell matrix, the shell matrix and hexafluoroisopropanol were mixed and dissolved at a mass / volume ratio of 1:5, g: ml. Then, NM was added at a mass ratio of 1:10, g: ml to NM and the shell matrix. The mixture was ultrasonically dispersed for 1 min to obtain the shell liquid.

[0088] (2) PLA (Mw = 100000-130000) and hexafluoroisopropanol were mixed and dissolved at a mass / volume ratio of 1:8. Then, NT3 was added at a ratio of 1 μg NT3 per 0.1g PLA and ultrasonically dispersed for 1 min to obtain the nucleolysin.

[0089] (3) Connect two independent syringes, the core liquid and the shell liquid, with a coaxial spinning nozzle. The inner tube diameter of the spinning nozzle is 0.40 mm, the flow rate is 0.5 mL / h, the outer tube diameter is 0.72 mm, the flow rate is 2 mL / h, the ambient temperature is 25℃, the ambient humidity is 65%, the distance from the spinning nozzle to the receiver is 18 mm, and the spinning time is 3 h.

[0090] (4) The sample was collected on aluminum foil, freeze-dried for 24 h, and stored at 4℃ when used and at -20℃ when stored.

[0091] The artificial fibrous dura mater prepared in Examples 2 and 3 of this invention is as follows: Figure 1 As shown, where, Figure 1 The thickness at circle a is 0.15mm. Figure 1 The thickness at circle b is 0.53 mm, indicating that the thickness of the artificial fiber dura mater can be controlled by the spinning time.

[0092] The microstructure of the artificial fiber dura mater prepared in Example 1 of this invention is as follows: Figures 2-3 As shown, Figure 2 The crisscrossing spinning structure is visible, and the spinning structure is smooth and porous. Figure 3 It shows a bamboo-like structure with a clearly visible core-shell structure.

[0093] Figure 4 The results indicate that the hydrophilicity of the artificial fiber dura mater can be adjusted by regulating the PEG content in the shell fluid, with the best hydrophilicity observed when the PEG content is 20%. Increasing the hydrophilicity of the artificial fiber dura mater facilitates better drug release, improves the anti-inflammatory microenvironment, and promotes spinal cord injury reduction.

[0094] The mechanical properties of the artificial fiber dura maters prepared in Examples 1-3 were tested, such as... Figure 5 As shown, the artificial fiber dura maters prepared in Examples 1 to 3 all have good mechanical properties. Among them, the artificial fiber dura mater prepared in Example 1 by spinning for 3 hours has the best mechanical properties.

[0095] Experimental example:

[0096] The artificial fibrous dura mater prepared in Example 1 was used in an in vitro experiment to induce neural differentiation of neural stem cells and to investigate the anti-inflammatory effects of BV2 cells. The specific steps are as follows:

[0097] (1) The fibrous membrane was sterilized under ultraviolet irradiation, soaked in culture medium, and then placed in an incubator at a constant temperature of 37°C for later use. The biocompatibility of the material was verified using mesenchymal stem cells. Cell proliferation experiments showed that the fibrous membrane had good biocompatibility, such as... Figures 6-7 As shown.

[0098] (2) Verification of the neurotransmitter-promoting effect of the material by inoculating neural stem cells after sterilization of the fibrous membrane. Seven days after inoculation, the cells inoculated on different samples were lysed to extract RNA, and several specific neural markers were identified by polymerase chain reaction (PCR), such as... Figure 8 As shown, the fibrous membrane can indeed effectively promote the differentiation of neural stem cells into neural cells, and its differentiation effect is far superior to that of the blank control and pure fibrous membrane.

[0099] (3) Microglia were inoculated into the sterilized fiber membrane to verify the anti-inflammatory effect of the material. One day after inoculation, the cells inoculated onto different samples were lysed to extract RNA. Polymerase chain reaction (PCR) was used to identify several specific inflammatory markers, such as... Figure 9 As shown, the fibrous membrane can indeed effectively reduce the number of inflammatory microglia.

[0100] (4) The anti-inflammatory effect of the sterilized fibrous membrane was verified by applying it to C57 mice, a spinal cord injury and contusion model. One day after treatment with the fibrous membrane, frozen sections of spinal cord tissue from mice with and without the fibrous membrane and with different fibrous membranes were prepared. Immunofluorescence was used to identify several specific inflammatory markers, such as… Figure 10 As shown, the fibrous membrane can indeed effectively reduce the number of inflammatory microglia.

[0101] (5) The sterilized fibrous membrane was used to treat C57 mice with spinal cord injury and contusion to verify its effect on promoting neural differentiation. Twenty-eight days after treatment with the fibrous membrane, frozen sections of spinal cord tissue from mice with and without the fibrous membrane, and mice with different fibrous membranes, were prepared. Immunofluorescence was used to identify several specific neural markers, such as… Figure 11 As shown, the fibrous membrane can indeed effectively promote the differentiation of neural stem cells into neural cells.

[0102] (6) The sterilized fiber membrane was used to verify its effect on promoting spinal cord injury repair in C57 mice, a model of spinal cord injury and contusion. Twenty-eight days after treatment with the fiber membrane, mice using different fiber membranes underwent an inclined plane test for functional assessment, and the gastrocnemius muscles of the mice were collected for muscle weight measurement. Figure 12 As shown, the fibrous membrane can indeed effectively promote the repair of spinal cord injuries.

[0103] The above description is merely a preferred embodiment of the present invention and is not intended to limit the invention. Various modifications and variations can be made to the present invention by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the scope of protection of the present invention.

Claims

1. A double-layered core-shell structured artificial fibrous dura mater for spinal cord injury, characterized in that, It is composed of a membrane structure made of artificial fibers with a double core-shell structure, which consists of an outer shell structure and an inner core structure. The outer shell structure is made of PLGA and PEG and is loaded with naphthostat mesylate (NM). The inner core structure is made of PLA and is loaded with neurotrophic factor 3 (NT3).

2. The double-layered core-shell artificial fibrous dura mater for spinal cord injury as described in claim 1, characterized in that, PEG is 5-20% of the total weight of PLGA and PEG.

3. The double-layered core-shell artificial fibrous dura mater for spinal cord injury as described in claim 1, characterized in that, The molar ratio of lactic acid units to glycolic acid in PLGA is 1:0.8~1.2; Alternatively, the weight-average molecular weight of PLGA is 880,000 to 117,000; Alternatively, the weight-average molecular weight of PLA is 100,000 to 130,000.

4. The double-layered core-shell artificial fibrous dura mater for spinal cord injury as described in claim 1, characterized in that, The weight of NM is 8-12% of the weight of the outer shell structure; Alternatively, the ratio of NT3 addition to the weight of the inner core structure is 0.8~1.2:0.1, μL: g; Alternatively, the weight ratio of the outer shell structure to the inner core structure is 3.5 to 4.5:

1.

5. The double-layered core-shell artificial fibrous dura mater for spinal cord injury as described in claim 1, characterized in that, The thickness is 0.1~0.5 mm.

6. A method for preparing a double-layered core-shell structured artificial fibrous dura mater for spinal cord injury as described in any one of claims 1 to 5, characterized in that, Includes the following steps: PLGA, PEG and NM were dissolved to prepare a shell solution; PLA and NT3 were dissolved to prepare a nucleolytic solution; The shell liquid and the core liquid are coaxially spun and then freeze-dried to obtain the product.

7. The method for preparing a double-layered core-shell structured artificial fibrous dura mater for spinal cord injury as described in claim 6, characterized in that, The solvent for the shell liquid is hexafluoroisopropanol; Alternatively, in the shell solution, the total weight of PLGA and PEG to the volume ratio of the solvent is 0.8~1.2:5, g: ml.

8. The method for preparing a double-layered core-shell structured artificial fibrous dura mater for spinal cord injury as described in claim 6, characterized in that, The solvent for the nuclear fluid is hexafluoroisopropanol; Alternatively, in the nuclear liquid, the weight ratio of PLA to solvent volume is 0.8~1.2:8, g: ml.

9. The method for preparing a double-layered core-shell structured artificial fibrous dura mater for spinal cord injury as described in claim 6, characterized in that, During coaxial spinning, the flow rate ratio of shell liquid to core liquid is 3.8~4.2:1; Alternatively, during coaxial spinning, the ratio of the inner tube diameter of the spinning nozzle to the outer tube diameter of the spinning nozzle is 1:1.5~2.

0.

10. The method for preparing a double-layered core-shell structured artificial fibrous dura mater for spinal cord injury as described in claim 6, characterized in that, The freeze-drying time is 45-50 hours.

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