CN112870439A - Nano fiber bone tissue engineering scaffold with core-shell-series crystal structure and preparation method thereof - Google Patents
Nano fiber bone tissue engineering scaffold with core-shell-series crystal structure and preparation method thereof Download PDFInfo
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- CN112870439A CN112870439A CN202110186329.1A CN202110186329A CN112870439A CN 112870439 A CN112870439 A CN 112870439A CN 202110186329 A CN202110186329 A CN 202110186329A CN 112870439 A CN112870439 A CN 112870439A
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- 239000011258 core-shell material Substances 0.000 claims abstract description 26
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- 229940010747 sodium hyaluronate Drugs 0.000 claims abstract description 14
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Abstract
The core-shell structure realizes the functions of loading and slow release of drugs, and the cluster crystal structure formed by the nano wafers grown in situ on the surface of the nano fibers improves the hydrophilicity and the cell compatibility of the nano fiber membrane with the core-shell structure. The method comprises the following steps: (1) taking ammonia sugar and sodium hyaluronate hydrosol as water phases; (2) dichloromethane, Span80 and the sol are stirred and mixed at high speed to obtain water-in-oil emulsion; (3) dissolving polycaprolactone and N, N-dimethylformamide in the emulsion, and performing electrostatic spinning to obtain a nanofiber membrane with a core-shell structure; (4) soaking the nanofiber membrane with the core-shell structure in a polycaprolactone dilute solution to obtain the nanofiber bone tissue engineering scaffold with the core-shell-string crystal structure. The nanofiber bone tissue engineering scaffold with the core-shell-tandem structure has certain application value in the biomedical fields of drug loading, slow release, bone repair and the like.
Description
Technical Field
The invention relates to a nanofiber bone tissue engineering scaffold with a core-shell-tandem crystal structure and a preparation method thereof, belongs to the field of biomedical engineering bone repair scaffolds, and has wide application value in the aspects of effectively controlling drug slow release and bone repair.
Background
Bone is a complex organ with the functions of hematopoiesis, storage of important minerals, preservation of important organs and promotion of exercise. Natural bone is composed of approximately 70wt% inorganic nanocrystals (such as hydroxyapatite) and 30wt% organic matrix (such as collagen nanofibers). Due to the insufficient number of blood vessels in bone tissue, with a high mineral content, bone healing is very difficult in cases of massive tissue loss. Although human bone has the ability to heal and regenerate, this ability is ineffective for injuries caused by large bone defects, fractures, traffic accidents, and bone tumor resection. The treatment is mainly carried out by adopting autologous bone transplantation and xenogenic bone transplantation means in clinic, but the application of the technologies is limited by the problems of limited donor sources, complicated operation, immune reaction and the like, and a new thought is provided for solving the problem by tissue engineering. Therefore, the construction of a bone repair scaffold capable of rapidly inducing osteogenesis and highly matching the shape of a defect is receiving much attention.
The bone tissue engineering scaffold should be designed to have proper porosity, biodegradability and specific functions related to tissue regeneration, such as specific shape and size. The electrostatic spinning technology is a commonly used technology for preparing a tissue engineering scaffold, and the produced fibers can effectively imitate the shape of the natural extracellular matrix of bone tissues. The fiber produced by electrostatic spinning has very high specific surface area, and the pore diameter can be adjusted from several microns to tens of microns after modification. Although the electrostatic spinning technology has a wide application prospect in bionic reconstruction, materials for preparing electrostatic spinning still need to be carefully selected. The composition of the material can affect the mechanical properties, plasticity, biocompatibility and degradability of the stent, thereby further affecting the repair efficiency. In addition, the morphology, hydrophilicity, and surface activity of the scaffold have some effect on cell adhesion, migration, and proliferation.
Polycaprolactone is a semi-crystalline polymer with good biocompatibility and degradability, and the product produced after degradation is non-toxic. However, polycaprolactone is highly hydrophobic, poorly attached to cells, and lack of biological activity limits its further use. Glucosamine can induce osteoblast differentiation and inhibit osteoclast differentiation, thereby increasing bone matrix deposition, reducing bone resorption, and finally promoting bone formation.
Disclosure of Invention
The invention provides a nanofiber bone tissue engineering scaffold with a core-shell-series crystal structure and a preparation method thereof, wherein the nanofiber bone tissue engineering scaffold has good mechanical properties and biocompatibility, can be applied to tissue engineering bone repair and has a drug slow-release behavior.
In order to achieve the purpose, the invention provides the following technical scheme:
the nanofiber bone tissue engineering scaffold with the core-shell-tandem structure is characterized in that: the inner core of the fiber is sodium hyaluronate loaded with glucosamine, and the outer shell is biodegradable polycaprolactone.
The preparation method of the nanofiber bone tissue engineering scaffold with the core-shell-tandem crystal structure comprises the following specific steps: and (2) stirring the sodium hyaluronate (glucosamine) hydrosol and dichloromethane at a high speed to obtain a water-in-oil type emulsion, adding polycaprolactone to prepare a spinning solution with a certain concentration, and performing electrostatic spinning to obtain the core-shell structured nanofiber membrane. And soaking the obtained nanofiber membrane with the core-shell structure in a polycaprolactone dilute solution, and drying in vacuum to obtain the nanofiber bone tissue engineering scaffold with the core-shell-crystal structure.
In order to solve the problem of poor hydrophilicity of the core-shell structure nanofiber membrane, the invention adopts a solution crystallization method to enable the nanofiber of the core-shell structure to induce homogeneous polycaprolactone to form an interface crystallization structure so as to improve the hydrophilicity of the core-shell structure nanofiber membrane.
Preferably, the sodium hyaluronate is derived from streptococcus equi.
Preferably, the molecular weight of the polycaprolactone is 80000.
Preferably, the glucosamine is D (+) -glucosamine hydrochloride.
Preferably, the high-speed stirring time is 3min, and the rotating speed is 30000 r/min.
Preferably, the diluted polycaprolactone solution is diluted by glacial acetic acid and deionized water.
Preferably, the soaking time is 5min, 10min, 15min or 20 min.
The most preferred soaking time is 15 min.
Preferably, the vacuum drying time is 24 h.
Preferably, the mechanical property, hydrophilicity and hydrophobicity and cell adhesion of the core-shell-crystal structure nanofiber bone tissue engineering scaffold are adjusted by adjusting and controlling the soaking time of the core-shell structure nanofiber membrane.
Compared with the prior art, the invention has the beneficial effects that: the nanofiber bone tissue engineering scaffold with the core-shell-tandem structure, which is prepared by the invention, has good fiber morphology, good biocompatibility and degradability, and can slowly release glucosamine. The crystal-string structure can improve the hydrophilicity of the core-shell structure nanofiber membrane, can better simulate the microstructure of collagen fibers in extracellular matrix, has the capability of guiding bone regeneration, has obvious effect, and can be used for repairing bone tissue defects.
Drawings
Fig. 1 is an SEM image of the core-shell structured nanofiber membrane in example 1.
Fig. 2 is a diameter distribution diagram of the core-shell structured nanofiber membrane in example 1.
FIG. 3 is an SEM image of a core-shell-tandem structure nanofiber bone tissue engineering scaffold soaked for 5min in example 2.
FIG. 4 is an SEM image of a core-shell-tandem structure nanofiber bone tissue engineering scaffold soaked for 10min in example 2.
FIG. 5 is an SEM image of a core-shell-tandem structure nanofiber bone tissue engineering scaffold soaked for 15min in example 2.
FIG. 6 is an SEM image of a core-shell-tandem structure nanofiber bone tissue engineering scaffold soaked for 20min in example 2.
FIG. 7 shows the hydrophilic and hydrophobic properties of the nanofiber bone tissue engineering scaffold with core-shell-tandem structure in example 2.
FIG. 8 is an SEM image of a core-shell structured nanofiber membrane loaded with an aminosugar in example 3.
FIG. 9 is a distribution diagram of the diameter of the core-shell structured nanofiber membrane loaded with aminosugars in example 3.
FIG. 10 is a graph of cell proliferation on different nanofiber scaffold materials.
Detailed Description
In order to make the technical solutions and advantages of the present invention clearer, the present invention will be further described with reference to specific embodiments. Obviously. The described embodiments are only a few embodiments of the present invention, not all embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Example 1
Dissolving 12mg of sodium hyaluronate in 988 microliters of deionized water, and uniformly mixing to obtain 1.2wt% sodium hyaluronate hydrosol. A mixture containing 3.03ml of methylene chloride and 0.01g of span-80 was mixed with the sodium hyaluronate sol and stirred at high speed to obtain a uniform water-in-oil (W/O) emulsion. Dissolving 0.5g of polycaprolactone and 2.11ml of N, N-dimethylformamide in the emulsion to obtain an electrostatic spinning solution, sucking the spinning solution into an injector for electrostatic spinning, wherein the spinning voltage is 18kv, the distance between a spinning nozzle and a receiving plate is 15cm, the propelling speed of a propelling pump is 0.001mm/s, and preparing the nanofiber membrane MS with a core-shell structure.
FIG. 1 and FIG. 2 are scanning electron microscope pictures of the scaffold, which can see the smooth and continuous nanofiber structure, and the diameter of the nanofiber is 127 +/-33 nm by ImageJ software analysis.
Example 2
Dissolving 0.2g of polycaprolactone in 20ml of mixed solvent of glacial acetic acid and deionized water, and magnetically stirring at 60 ℃, wherein the volume ratio of the glacial acetic acid to the deionized water is 3: 1. And cooling the solution to room temperature after full dissolution, and soaking the core-shell structured nanofiber membrane obtained in the example 1 in a polycaprolactone dilute solution for 5min, 10min, 15min and 20min respectively. And then vacuum drying is carried out for 24h, and residual solvent is removed, so as to obtain the nanofiber bone tissue engineering scaffolds SK5, SK10, SK15 and SK20 with the core-shell-series crystal structure.
Fig. 3 to 6 are scanning electron microscope images of the core-shell-tandem structure nanofiber bone tissue engineering scaffold with different soaking times, and it is determined that the tandem structure on the surface of the core-shell structure nanofiber can improve the hydrophilicity of the core-shell structure nanofiber membrane, and the core-shell-tandem structure nanofiber bone tissue engineering scaffold SK15 has the most significant effect (shown in fig. 7).
Example 3
Dissolving 12mg of sodium hyaluronate in 988 microliters of deionized water, and uniformly mixing to obtain 1.2wt% sodium hyaluronate hydrosol. Then, 10 μ l of glucosamine (100 μ g/ml) was mixed into 50 μ l of hyaluronic acid hydrosol. A mixture containing 3.03ml of methylene chloride and 0.01g of span-80 was mixed with the above sol and stirred at a high speed to obtain a uniform water-in-oil (W/O) type emulsion. Dissolving 0.5g of polycaprolactone and 2.11ml of N, N-dimethylformamide in the emulsion to obtain an electrostatic spinning solution, sucking the spinning solution into an injector for electrostatic spinning, wherein the spinning voltage is 18kv, the distance between a spinning nozzle and a receiving plate is 15cm, the propelling speed of a propelling pump is 0.001mm/s, and preparing the nano-fiber membrane MS-GLU (shown in figures 8 and 9) with the core-shell structure carrying glucosamine.
Example 4
Dissolving 0.2g of polycaprolactone in 20ml of mixed solvent of glacial acetic acid and deionized water, and magnetically stirring at 60 ℃, wherein the volume ratio of the glacial acetic acid to the deionized water is 3: 1. And after the solution is fully dissolved, cooling the solution to room temperature, and soaking the nano-fiber membrane loaded with the glucosamine core-shell structure obtained in the embodiment 3 in the diluted polycaprolactone solution for 15 min. And then vacuum drying is carried out for 24h, and residual solvent is removed, so that the nanofiber bone tissue engineering scaffold SKMS-GLU with the serial crystal-core shell structure loaded with glucosamine is obtained.
Example 5
The nanofiber scaffolds in the above examples were prepared into a disc shape with a diameter of 1cm and placed in a 24-well plate, and after co-culturing with cells for 1, 4, and 7 days, 50 μ l of CCK-8 solution was added to each well, and the blank control group without the nanofiber scaffold was used. After incubation in the cell incubator for 0.5 hour, absorbance was measured at 450 nm. Sample SK15 is designated as SKMS.
The results are shown in fig. 10, the samples of examples 2, 3 and 4 are favorable for cell proliferation, and the sample of example 4 has more significant proliferation effect.
Claims (6)
1. The nanofiber bone tissue engineering scaffold with the core-shell-tandem structure is characterized in that: the inner core of the fiber is sodium hyaluronate loaded with glucosamine, and the outer shell is biodegradable polycaprolactone.
2. The nanofiber bone tissue engineering scaffold with core-shell-tandem structure according to claim 1, wherein: and adopting a solution crystallization method to induce the homogeneous polycaprolactone to form an interface crystal structure by the nano-fiber with the core-shell structure.
3. The nanofiber bone tissue engineering scaffold with the core-shell-series crystal structure according to claim 2, wherein the mechanical property, hydrophilicity and hydrophobicity and cell adhesion of the nanofiber bone tissue engineering scaffold with the core-shell-series crystal structure are adjusted by adjusting and controlling the soaking time of the nanofiber membrane with the core-shell structure.
4. The nanofiber bone tissue engineering scaffold with core-shell-tandem structure according to claim 1 or 2, wherein: the preparation method comprises the following steps:
(1) dissolving sodium hyaluronate in deionized water, uniformly mixing to prepare 1.2wt% of sodium hyaluronate hydrosol, and mixing 10 mul of glucosamine with the concentration of 100 mug/ml into 50 mul of sodium hyaluronate hydrosol;
(2) mixing a mixed solution containing 3.03ml of dichloromethane and 0.01g of span-80 with the sodium hyaluronate sol, and stirring at a high speed to obtain a uniform water-in-oil type emulsion;
(3) dissolving 0.5g of polycaprolactone and 2.11ml of N, N-dimethylformamide in the emulsion to obtain an electrostatic spinning solution, and obtaining a nanofiber membrane with a core-shell structure by using an electrostatic spinning technology;
(4) dissolving polycaprolactone in a mixed solvent of glacial acetic acid and deionized water, and magnetically stirring at 60 ℃; and after the nano fiber membrane is fully dissolved, cooling the solution to room temperature, soaking the nano fiber membrane with the core-shell structure in a diluted polycaprolactone solution, carrying out vacuum drying for 24 hours, and removing the residual solvent to obtain the nano fiber bone tissue engineering scaffold with the core-shell-series crystal structure.
5. The nanofiber bone tissue engineering scaffold with the core-shell-crystal-string structure according to claim 4, wherein the electrostatic spinning process parameters are as follows: the voltage was 18kv, the distance between the spinneret and the receiving plate was 15cm, and the advancing speed of the advancing pump was 0.001 mm/s.
6. The nanofiber bone tissue engineering scaffold with the core-shell-tandem crystal structure according to claim 4, wherein the mass fraction of polycaprolactone in the polycaprolactone dilute solution is 1%, the volume ratio of glacial acetic acid to deionized water is 3:1, and the molecular weight of polycaprolactone is 80000.
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN113730658A (en) * | 2021-09-22 | 2021-12-03 | 太原理工大学 | Bionic bone cartilage integrated repair support and preparation method thereof |
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| CN113832611A (en) * | 2021-10-27 | 2021-12-24 | 中国药科大学 | Phase-change coaxial nanofiber membrane for epidermal desensitization treatment and preparation method and application thereof |
| CN113832611B (en) * | 2021-10-27 | 2023-02-24 | 中国药科大学 | Phase-change coaxial nanofiber membrane for epidermal desensitization treatment and preparation method and application thereof |
| CN114225116A (en) * | 2022-01-25 | 2022-03-25 | 奥精医疗科技股份有限公司 | Artificial periosteum capable of slowly releasing hyaluronic acid and growth factors and preparation method thereof |
| CN115300669A (en) * | 2022-08-24 | 2022-11-08 | 上海睿植康医疗科技有限公司 | Fiber membrane and preparation method and application thereof |
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