CN112386744B - Drug sustained-release hydrogel semi-embedded composite stent and preparation method thereof - Google Patents
Drug sustained-release hydrogel semi-embedded composite stent and preparation method thereof Download PDFInfo
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- A61L2430/00—Materials or treatment for tissue regeneration
- A61L2430/06—Materials or treatment for tissue regeneration for cartilage reconstruction, e.g. meniscus
Abstract
The invention relates to the technical field of biomedical materials, in particular to a drug sustained-release hydrogel semi-embedded composite stent and a preparation method thereof, wherein the preparation method comprises the following steps: (1) preparing an alendronate sodium deposition type hydroxyapatite scaffold; (2) preparing a pre-crosslinked drug sustained-release hydrogel solution; (3) sealing the holes at the lower layer of the hydroxyapatite bracket by adopting gelatin; (4) injecting the pre-crosslinked drug sustained-release hydrogel solution in the step (2) on the upper layer of the hydroxyapatite scaffold, and carrying out photo-crosslinking; (5) and (3) cleaning the hydrogel semi-embedded composite stent, and dissolving and removing gelatin sealing the holes at the lower layer of the stent to obtain the drug sustained-release hydrogel semi-embedded composite stent. The preparation method has the advantages of convenient operation and control, high production efficiency, stable product quality and contribution to industrial production, and the prepared composite scaffold has the advantages of bionic structure, mechanical matching, high biological activity and capability of promoting the directional and regional differentiation of stem cells.
Description
Technical Field
The invention relates to the technical field of biomedical materials, in particular to a drug sustained-release hydrogel semi-embedded composite stent and a preparation method thereof.
Background
The structurally complex bone joint is composed of articular cartilage, calcified cartilage and subchondral bone. Tissues of three structures are mutually affected, and damage or lesion of any one structure can influence the exertion of biological functions of the other two structures, so that the later development of the overall damage of the bone joint is likely to be caused. Therefore, in the treatment of osteoarticular diseases, simultaneous repair of cartilage and subchondral bone is required to achieve radical cure. And because of the chemical composition of cartilage and subchondral bone, the mechanical properties and cell differentiation types have great difference, thereby forming a great problem of bone joint defect repair. Osteoarticular damage seriously afflicts human health as a common disease and has a very limited self-repairing ability as a terminal differentiation product, so many studies have been conducted around the repair of osteoarticular damage.
Currently, clinical therapies (including drug therapy, surgical transplantation, etc.) are limited in their application and efficacy due to side effects, immune rejection, etc. With the rapid development of tissue engineering, the implantation of biological scaffolds as an effective tissue repair means in biomedical engineering has obvious advantages, but at the same time, there are still many difficulties to overcome. Because of the complex structure of the bone joint, the different functional areas have a specific distribution of components and cell types. The single-component biological scaffold has the limitation of the single-component biological scaffold, and can only be used for repairing a single tissue, so that the two-phase and multi-phase scaffolds are adopted to repair the bone and cartilage structures of the bone joints, and the important significance is achieved.
Disclosure of Invention
In order to overcome the defects of the prior art, the invention aims to provide the preparation method of the drug sustained-release hydrogel semi-embedded composite scaffold for repairing the osteochondral full-thickness injury, the preparation method is convenient to operate and control, high in production efficiency, stable in product quality and beneficial to industrial production, and the prepared drug sustained-release hydrogel semi-embedded composite scaffold has the advantages of bionic structure, mechanical matching and high bioactivity and can promote the directional zoning and differentiation of stem cells.
In order to achieve the purpose, the invention adopts the following technical scheme: a preparation method of a drug sustained-release hydrogel semi-embedded composite stent comprises the following steps:
(1) preparing an alendronate sodium (ALN) deposition type hydroxyapatite scaffold;
(2) preparing a pre-crosslinked drug sustained-release hydrogel solution;
(3) sealing the holes at the lower layer of the hydroxyapatite bracket by adopting gelatin;
(4) injecting the pre-crosslinked drug sustained-release hydrogel solution in the step (2) on the upper layer of the hydroxyapatite scaffold in the step (3) by means of a mold to perform photocrosslinking;
(5) cleaning the hydrogel semi-embedded composite stent formed in the step (4), dissolving and removing gelatin sealing the holes at the lower layer of the stent to obtain the drug sustained-release hydrogel semi-embedded composite stent
Further, in the step (1), the plasma treatment atmosphere is oxygen or air, and the treatment time is 1-15 min.
The semi-embedded composite scaffold of the drug slow-release hydrogel prepared by combining the biological hydrogel and the 3D-printed hydroxyapatite scaffold is a bionic double-layer osteochondral scaffold, can be used for repairing osteochondral full-layer injuries, has a bionic structure and components of an osteochondral composite layer at a bone joint, supports the growth of stem cells, can regulate the differentiation of the stem cells by combined drug molecules, is stably combined and has a transition structure, has good advantages when being used for synchronously repairing a bone injury layer and a cartilage injury layer, and has a great clinical application value; the preparation method has the advantages of simple process, convenient operation and control, high production efficiency and stable product quality, and is beneficial to industrial production.
Further, in the step (1), the preparation method of the alendronate sodium deposition-type hydroxyapatite scaffold comprises the following steps:
a1, obtaining a hydroxyapatite three-dimensional scaffold with 60-100% of scaffold pores by a 3D printing technology;
a2, soaking the hydroxyapatite three-dimensional scaffold prepared in the step A1 in an alendronate sodium solution to obtain the alendronate sodium deposition type hydroxyapatite scaffold, wherein the concentration of the alendronate sodium solution is 2-10 mM.
The drug sustained-release hydrogel semi-embedded composite scaffold takes a hydroxyapatite scaffold as a framework, and the alendronate sodium (ALN) is deposited on the hydroxyapatite scaffold, so that the alendronate sodium and the hydroxyapatite have strong affinity, the biocompatibility of the composite scaffold is greatly improved, the composite scaffold can be well combined with bones, the bone conduction effect and the bone induction effect of the composite scaffold can also effectively promote the growth of the bones, the alendronate sodium can effectively resist osteoporosis, the bone absorption resisting effect is realized by inhibiting the activity of osteoclasts, the bone absorption resisting effect is strong, and the bone mineralization inhibiting effect is not existed. Among hydroxyapatite scaffolds, Hydroxyapatite (HA) is an important component of bone tissue, HAs excellent biocompatibility, and HAs osteoinductivity and osteoconductivity. The sodium alendronate has bisphosphonic acid groups, can be combined with Ca2+ rich in hydroxyapatite scaffold, and has the effects of promoting osteogenesis and inhibiting osteoclast.
Further, in the step (2), the preparation method of the pre-crosslinked drug sustained-release hydrogel solution comprises the following steps:
b1, chemically grafting carbon-carbon double bonds to hyaluronic acid molecules to prepare modified hyaluronic acid; synthesizing carbon-carbon double bond grafted modified beta-cyclodextrin molecules;
b2, mixing the modified beta-cyclodextrin molecule prepared in the step B1 and the hydrophobic drug for promoting the chondrogenic differentiation of the stem cells in an aqueous solution in a ratio of 1.1-1.3: 1, stirring for 23-25 h to obtain drug-coated composite molecules;
b3, carrying out stirring and dissolving on the drug-loaded composite molecule prepared in the step B2 and the modified hyaluronic acid prepared in the step B1 in an aqueous solution, and then adding a biological initiator to prepare a pre-crosslinked drug sustained-release hydrogel solution.
Furthermore, in the pre-crosslinked drug sustained-release hydrogel solution, the concentration of the drug-loaded composite molecules is 2-6wt%, and the concentration of the modified hyaluronic acid is 2-4 wt%. Further, the hydrophobic drug for promoting the chondrogenic differentiation of stem cells is a chondrogenic differentiation factor (KGN). The concentration of the hydrophobic drug of the stem cell chondrogenic differentiation promoting factor is 0.1-1 mu M. Chondrogenic differentiation factor KGN: binding to fibrillar proteins disrupts their interaction with the transcription factor core binding factor B subunit (CBF β), and through the recombinant CBF β -Runx1 transcription program, promotes the selective differentiation of pluripotent mesenchymal stem cells into chondrocytes, thereby stimulating cartilage repair.
Further, in the step B1, the step of chemically grafting carbon-carbon double bonds to the hyaluronic acid molecule is: first, 3.5-4.5g of sodium hyaluronate is dissolved in 180-220mL of DPBS solution, 16-18mL of methacrylic anhydride is added dropwise at the temperature of 3-5 ℃ at the speed of 0.8-1.2 s/drop while stirring vigorously, and the reaction is carried out overnight at room temperature. Precipitating white floccule with ethanol, dissolving in deionized water (140-160 mL), dialyzing with dialysis bag (12-14 kDa) for one week, and freeze-drying to obtain hyaluronic acid as main component of articular cartilage and synovial fluid, which is suitable for repairing articular cartilage. Hyaluronic acid and glycoprotein contained in synovial fluid impart lubricity and viscoelasticity to the synovial fluid, and contribute to reduction of friction between soft tissues and between cartilage.
Further, in the step B1, the step of chemically grafting carbon-carbon double bonds to the β -cyclodextrin molecule (β -CD) is: firstly, beta-CD is recrystallized in deionized water (88-92 ℃), and vacuum drying is carried out for 11-13h at 58-62 ℃ to obtain the recrystallized beta-CD. Dissolving 9-11g of recrystallized beta-CD in 45-55mL of N, N-dimethylformamide, sequentially adding 18-22 mu L of stannous octoate and 2.10-2.18g of acrylic acid-2-isocyano ethyl ester under the protection of nitrogen environment, and reacting at room temperature for 0.8-1.2 h. Then, the reaction temperature is adjusted to 38-42 ℃ to continue the reaction for 3.5-4.5 h. After the reaction was completed, the reaction solution was repeatedly recrystallized from acetone. Finally, the precipitate is dried in vacuum at 34-36 ℃ for 46-50h, and the product is stored at low temperature. Modified cyclodextrin: the molecular cavity can promote the loading and slow release of lipophilic drugs; the double bonds grafted on the cyclodextrin molecules can be moderately chemically crosslinked with the modified hyaluronic acid, so that the strength of the hydrogel is enhanced. Too high hydrogel strength can lead to decreased survival of the internal cells.
Further, in the step B3, the biological initiator is at least one of biological initiators I2959 and LAP, and the addition amount of the biological initiator in the aqueous solution is 0.4-0.6 wt%.
The invention prepares the pre-crosslinked drug sustained-release hydrogel solution by matching the cartilage differentiation promoting factor with the modified beta-cyclodextrin molecule and the modified hyaluronic acid, so that the composite scaffold has the drug sustained-release function, and can induce mesenchymal stem cells to differentiate towards cartilage cells and promote cartilage injury repair; the stent is implanted into an animal body, the drug release time is long, and no thrombus or serious inflammatory reaction occurs at the stent. The modified hyaluronic acid is prepared from hyaluronic acid with the average molecular weight of 200-400 w.
Further, the step (3) is specifically as follows: immersing the lower layer holes of the hydroxyapatite scaffold obtained in the step (1) by using gelatin solution at the temperature of 48-52 ℃, immersing the hydroxyapatite scaffold at the height of 7/12-3/4 by using the gelatin solution, and sealing the holes at the lower layer of the hydroxyapatite scaffold by using gelatin as solid jelly at the temperature of 2-8 ℃, wherein the concentration of the gelatin solution is 13-17 wt%.
Further, in the step (4), the pre-crosslinked drug sustained-release hydrogel solution in the step (2) is injected on the upper layer of the hydroxyapatite scaffold in the step (3) by means of a mold, and is crosslinked for 9-11min under ultraviolet light, wherein the ultraviolet intensity is 5.6-15.0 mW/cm2。
Further, in the step (4), the pre-crosslinked hydrogel solution permeates into the porous hydroxyapatite scaffold with the height of 1/4-5/12.
Further, in the step (5), the hydrogel semi-embedded composite stent is cleaned by water at 37-43 ℃, and gelatin for sealing the lower-layer holes of the stent is removed.
The invention also provides a drug sustained-release hydrogel semi-embedded composite stent, which is prepared by the preparation method of the drug sustained-release hydrogel semi-embedded composite stent.
Furthermore, the drug slow-release hydrogel semi-embedded composite scaffold comprises drug slow-release hydrogel positioned on the upper layer and a hydroxyapatite three-dimensional scaffold positioned on the lower layer.
Further, the drug sustained-release hydrogel comprises the following components:
2-6wt% of modified cyclodextrin
2-4wt% of modified hyaluronic acid
0.1-1 μ M of chondrogenic differentiation factor (KGN).
The porosity of the hydroxyapatite three-dimensional scaffold is 60-100%, and the load concentration of the sodium alendronate is 1-10 mu M.
The other raw materials are aqueous solution, and the aqueous solution is sterile water or cell culture medium. If cells are to be coated, the remaining raw material is a cell complete medium. Adding a photoinitiator to the aqueous solution: 0.04-0.06wt% of I2959 (UV initiated) or LAP (blue initiated).
The invention has the beneficial effects that: the drug slow-release hydrogel semi-embedded composite scaffold for repairing osteochondral full-layer damage, provided by the invention, has a bionic structure and components of an osteochondral composite layer at a bone joint, supports the growth of stem cells, can regulate the differentiation of the stem cells by combined drug molecules, is stably combined and has a transition structure, has an excellent technical effect when being used for synchronously repairing the osteochondral damaged layer and the cartilage damaged layer, and has a great clinical application value; the preparation method has the advantages of convenient operation and control, high production efficiency and stable product quality, and is beneficial to industrial production.
Detailed Description
In order to facilitate understanding of those skilled in the art, the present invention will be further described with reference to the following examples and drawings, which are not intended to limit the present invention.
Example 1
In this embodiment, a method for preparing a drug sustained-release hydrogel semi-embedded composite stent includes the following steps:
(1) preparing an alendronate sodium (ALN) deposition type hydroxyapatite scaffold;
(2) preparing a pre-crosslinked drug sustained-release hydrogel solution;
(3) sealing the holes at the lower layer of the hydroxyapatite bracket by adopting gelatin;
(4) injecting the pre-crosslinked drug sustained-release hydrogel solution in the step (2) on the upper layer of the hydroxyapatite scaffold in the step (3) by means of a mould, and crosslinking under ultraviolet light;
(5) and (4) cleaning the hydrogel semi-embedded composite stent formed in the step (4), and dissolving and removing gelatin sealing the holes at the lower layer of the stent to obtain the drug sustained-release hydrogel semi-embedded composite stent.
Further, in the step (1), the preparation method of the alendronate sodium deposition-type hydroxyapatite scaffold comprises the following steps:
a1, obtaining a hydroxyapatite three-dimensional scaffold with 63% of scaffold pores by a 3D printing technology;
a2, soaking the hydroxyapatite three-dimensional scaffold prepared in the step A1 in an alendronate sodium solution to obtain the alendronate sodium deposition type hydroxyapatite scaffold, wherein the concentration of the alendronate sodium solution is 2 mu M.
Specifically, the alendronate sodium deposition type hydroxyapatite scaffold is prepared as follows: hydroxyapatite powder and a dispersant PAA-NH4 (solid content 1.5%) were dissolved in water/glycerol (w/w, 7/3) in parts by weight, using NH3·H2Adjusting the pH value to 9.00 to prepare a mixture, wherein the adding concentration of the hydroxyapatite powder is 49wt%, and the adding amount of a dispersing agent PAA-NH4 is 1.5% of the mass of the hydroxyapatite powder; and (3) putting the mixture into a ball milling pot, grinding for 12h, and then adding hydroxypropyl methyl cellulose and n-octanol into the ball milling pot, wherein the addition amount of the hydroxypropyl methyl cellulose is 3 wt%, so as to obtain slurry with proper viscosity. A cylindrical hydroxyapatite three-dimensional scaffold (the radius is 5 mm multiplied by the height is 3 mm) with 63% of scaffold pores is obtained through a 3D printing technology, the printing distance is 300 mu m, and the printed scaffold is sintered for 3 hours at a high temperature of 1130 ℃. Soaking the obtained scaffold in 2 mM alendronate sodium (ALN) solutionObtaining the ALN deposition type hydroxyapatite bracket.
Further, in the step (2), the preparation method of the pre-crosslinked drug sustained-release hydrogel solution comprises the following steps:
b1, chemically grafting carbon-carbon double bonds to hyaluronic acid molecules to prepare modified hyaluronic acid; synthesizing carbon-carbon double bond grafted modified beta-cyclodextrin molecules;
b2, mixing the modified beta-cyclodextrin molecule prepared in the step B1 and hydrophobic drug for promoting stem cell chondrogenic differentiation in an aqueous solution in a ratio of 1.2: 1, stirring for 24 hours to obtain drug-coated composite molecules;
b3, carrying out stirring and dissolving on the drug-loaded composite molecule prepared in the step B2 and the modified hyaluronic acid prepared in the step B1 in an aqueous solution, and then adding a biological initiator to prepare a pre-crosslinked drug sustained-release hydrogel solution.
Furthermore, in the pre-crosslinked drug sustained-release hydrogel solution, the concentration of the drug-loaded composite molecules is 4wt%, and the concentration of the modified hyaluronic acid is 2 wt%.
Further, in the step B1, the step of chemically grafting carbon-carbon double bonds to the hyaluronic acid molecule is: first, 4g of sodium hyaluronate was dissolved in 200mL of DPBS solution, 17mL of methacrylic anhydride was added dropwise at 4 ℃ at a rate of 1 s/drop while vigorously stirring, and the reaction was carried out overnight at room temperature. Precipitating white floccule with ethanol, dissolving in deionized water (150 mL), dialyzing with dialysis bag (12-14 kDa) for one week, and freeze drying
Further, in the step B1, the step of chemically grafting carbon-carbon double bonds to the β -cyclodextrin molecule (β -CD) is: beta-CD is first recrystallized in deionized water (90 ℃) and dried in vacuum at 60 ℃ for 12h to obtain the recrystallized beta-CD. Dissolving 10g of recrystallized beta-CD in 50mL of N, N-dimethylformamide, sequentially adding 20 mu L of stannous octoate and 2.14g of 2-isocyanoethyl acrylate under the protection of nitrogen environment, and reacting at room temperature for 1 h. Subsequently, the reaction temperature was adjusted to 40 ℃ and the reaction was continued for 4 hours. After the reaction was completed, the reaction solution was repeatedly recrystallized from acetone. Finally, the precipitate was dried under vacuum at 35 ℃ for 2 days and the product was stored at low temperature.
Further, in the step B2, the hydrophobic drug for promoting chondrogenic differentiation of stem cells is a chondrogenic differentiation factor (KGN). The concentration of the hydrophobic drug that promotes chondrogenic differentiation of stem cells was 0.5. mu.M.
Further, in the step B3, the photoinitiator is a photoinitiator I2959, and the addition amount of the photoinitiator in the aqueous solution is 0.5 wt%.
Further, the step (3) is specifically as follows: pouring a gelatin solution at the temperature of 50 ℃ into a flat-bottom disc, immersing the lower layer holes of the hydroxyapatite bracket obtained in the step (1), immersing the gelatin solution into 2/3 of the height of the hydroxyapatite bracket, and sealing the holes at the lower layer of the hydroxyapatite bracket by using gelatin which is solid jelly after 30min at the temperature of 4 ℃, wherein the concentration of the gelatin solution is 15 wt%.
Further, in the step (4), a PDMA mold is sleeved on the top of the hydroxyapatite bracket and is attached to the gelatin at the bottom, a pre-crosslinked drug sustained-release hydrogel solution is injected into the PDMA mold, and crosslinking is performed for 10 min under ultraviolet light with the ultraviolet intensity of 5.6mW/cm2。
Further, in the step (5), the hydrogel semi-embedded composite scaffold is cleaned by water at 40 ℃, and gelatin for sealing the lower-layer holes of the scaffold is removed.
The embodiment also provides a drug sustained-release hydrogel semi-embedded composite stent, which is prepared by the preparation method of the drug sustained-release hydrogel semi-embedded composite stent. Fig. 1 is a simple hydroxyapatite three-dimensional scaffold. Fig. 2 is a diagram illustrating an example of the alendronate sodium-deposited hydroxyapatite scaffold according to the present embodiment. Fig. 3 is an EDS spectrum of the alendronate sodium deposition-type hydroxyapatite scaffold of the present example. Fig. 4 is a scanning electron microscope image of the alendronate sodium deposition-type hydroxyapatite scaffold of the present embodiment. Fig. 5 is another scanning electron microscope image of the alendronate sodium deposition-type hydroxyapatite scaffold of the present embodiment.
Example 2
The present embodiment is different from embodiment 1 in that: allenThe hydroxyapatite scaffold of the sodium phosphonate deposition type was prepared as follows: the preparation method comprises the following steps: hydroxyapatite powder and dispersant PAA-NH4 (solid content dissolved in water/glycerol (w/w, 7/3) in parts by weight, combined with NH3·H2Adjusting the pH value to 9.50 to prepare a mixture, wherein the adding concentration of the hydroxyapatite powder is 45wt%, and the adding amount of a dispersing agent PAA-NH4 is 1.5% of the mass of the hydroxyapatite powder; and (3) putting the mixture into a ball milling pot, grinding for 12h, and then adding hydroxypropyl methyl cellulose and n-octanol into the ball milling pot to obtain slurry with proper viscosity. A cylindrical hydroxyapatite three-dimensional scaffold with 60% of scaffold pores is obtained through a 3D printing technology, and the printed scaffold is sintered for 3 hours at a high temperature of 1110 ℃. The bracket is printed by adopting a 30-degree oblique angle, the radius of the bracket is 6 mm, the height of the bracket is 3 mm, the bracket size of the printing needle is 250 micrometers, and the distance between the printing needle and the bracket is 300 micrometers. The obtained scaffold was immersed in alendronate sodium (ALN) solution (1. mu.M) to obtain ALN-deposited hydroxyapatite scaffold. The porosity of the hydroxyapatite scaffold was 50%.
The rest of this embodiment is the same as embodiment 1, and is not described herein again.
Example 3
The present embodiment is different from embodiment 1 in that:
in the step (1), the preparation method of the alendronate sodium deposition type hydroxyapatite scaffold comprises the following steps: soaking the hydroxyapatite three-dimensional scaffold in an alendronate sodium (ALN) solution (the concentration of the alendronate sodium solution is 10 mu M), and taking out the hydroxyapatite three-dimensional scaffold for 12 hours to obtain the hydroxyapatite scaffold deposited with the alendronate sodium in low density, wherein the alendronate sodium deposit is in a fine needle shape by the observation of a scanning electron microscope.
The rest of this embodiment is the same as embodiment 1, and is not described herein again.
Example 4
The present embodiment is different from embodiment 3 in that:
in the step (1), the preparation method of the alendronate sodium deposition type hydroxyapatite scaffold comprises the following steps: soaking the hydroxyapatite three-dimensional scaffold in an alendronate sodium (ALN) solution (the concentration of the alendronate sodium solution is 10 mu M), and taking out the hydroxyapatite three-dimensional scaffold for 12 hours to obtain the hydroxyapatite scaffold deposited with the alendronate sodium in low density, wherein the alendronate sodium deposit is in a petal shape by the observation of a scanning electron microscope. The scanning electron micrograph of the alendronate sodium deposition-type hydroxyapatite scaffold of the present example is shown in fig. 4.
The rest of this embodiment is the same as embodiment 3, and is not described herein again.
Example 5
The present embodiment is different from embodiment 1 in that:
in the step (2), the preparation method of the pre-crosslinked drug sustained-release hydrogel solution comprises the following steps:
b1, chemically grafting carbon-carbon double bonds to hyaluronic acid molecules to prepare modified hyaluronic acid; synthesizing carbon-carbon double bond grafted modified beta-cyclodextrin molecules;
b2, mixing the modified beta-cyclodextrin molecule prepared in the step B1 and hydrophobic drug for promoting stem cell chondrogenic differentiation in an aqueous solution in a ratio of 1.2: 1, stirring for 24 hours to obtain drug-coated composite molecules;
b3, carrying out stirring and dissolving on the drug-loaded composite molecule prepared in the step B2 and the modified hyaluronic acid prepared in the step B1 in an aqueous solution, and then adding a biological initiator to prepare a pre-crosslinked drug sustained-release hydrogel solution.
Furthermore, in the pre-crosslinked drug sustained-release hydrogel solution, the concentration of the drug-loaded composite molecules is 4wt%, and the concentration of the modified hyaluronic acid is 2 wt%.
Further, in the step B3, the photoinitiator is a photoinitiator LAP, and the addition amount of the photoinitiator in the aqueous solution is 0.25 wt%.
Further, in the step (4), a PDMA mold is sleeved on the top of the hydroxyapatite support and is attached to the gelatin at the bottom, a pre-crosslinked drug sustained-release hydrogel solution is injected into the PDMA mold, and crosslinking is performed for 15min under blue light.
The rest of this embodiment is the same as embodiment 1, and is not described herein again.
The invention provides a drug slow-release hydrogel semi-embedded composite scaffold for repairing osteochondral full-layer damage, which has a bionic structure and components of an osteochondral composite layer at a bone joint, supports the growth of stem cells, can regulate the differentiation of the stem cells by combined drug molecules, is stably combined and has a transition structure, has good advantages when used for synchronously repairing a bone damage layer and a cartilage damage layer, and has great clinical application value; the preparation method has the advantages of convenient operation and control, high production efficiency and stable product quality, and is beneficial to industrial production.
All the technical features in the embodiment can be freely combined according to actual needs.
The above embodiments are preferred implementations of the present invention, and the present invention can be implemented in other ways without departing from the spirit of the present invention.
Claims (8)
1. A preparation method of a drug sustained-release hydrogel semi-embedded composite stent is characterized by comprising the following steps: the method comprises the following steps:
(1) preparing an alendronate sodium deposition type hydroxyapatite scaffold;
(2) preparing a pre-crosslinked drug sustained-release hydrogel solution;
(3) sealing the holes at the lower layer of the hydroxyapatite bracket by adopting gelatin;
(4) injecting the pre-crosslinked drug sustained-release hydrogel solution in the step (2) on the upper layer of the hydroxyapatite scaffold in the step (3) to perform photocrosslinking;
(5) cleaning the hydrogel semi-embedded composite stent formed in the step (4), and dissolving and removing gelatin sealing the holes at the lower layer of the stent to obtain the drug sustained-release hydrogel semi-embedded composite stent;
in the step (1), the preparation method of the alendronate sodium deposition type hydroxyapatite scaffold comprises the following steps:
a1, obtaining a hydroxyapatite three-dimensional scaffold by a 3D printing technology;
a2, soaking the hydroxyapatite three-dimensional scaffold prepared in the step A1 in an alendronate sodium solution to obtain an alendronate sodium deposition type hydroxyapatite scaffold, wherein the concentration of the alendronate sodium solution is 1-10 mu M;
in the step (2), the preparation method of the pre-crosslinked drug sustained-release hydrogel solution comprises the following steps:
b1, chemically grafting carbon-carbon double bonds to hyaluronic acid molecules to prepare modified hyaluronic acid; synthesizing carbon-carbon double bond grafted modified beta-cyclodextrin molecules;
b2, mixing the modified beta-cyclodextrin molecule prepared in the step B1 and the hydrophobic drug for promoting the chondrogenic differentiation of the stem cells in an aqueous solution in a ratio of 1.1-1.3: 1, stirring for 23-25 h to obtain drug-coated composite molecules;
b3, mixing and stirring the drug-loaded composite molecules prepared in the step B2 and the modified hyaluronic acid prepared in the step B1 in an aqueous solution for dissolving, and then adding a biological initiator to prepare a pre-crosslinked drug sustained-release hydrogel solution.
2. The preparation method of the drug sustained-release hydrogel semi-embedded composite stent according to claim 1, which is characterized in that: in the pre-crosslinked drug sustained-release hydrogel solution, the concentration of drug-coated composite molecules is 2-6wt%, and the concentration of modified hyaluronic acid is 2-4 wt%.
3. The preparation method of the drug sustained-release hydrogel semi-embedded composite stent according to claim 1, which is characterized in that: in the step B3, the biological initiator is at least one of biological initiators I2959 and LAP, and the addition amount of the biological initiator in the aqueous solution is 0.4-0.6 wt%.
4. The preparation method of the drug sustained-release hydrogel semi-embedded composite stent according to claim 1, which is characterized in that: the step (3) is specifically as follows: immersing the lower layer holes of the hydroxyapatite scaffold obtained in the step (1) by using gelatin solution at the temperature of 48-52 ℃, immersing the hydroxyapatite scaffold at the height of 7/12-3/4 by using the gelatin solution, and sealing the holes at the lower layer of the hydroxyapatite scaffold by using gelatin as solid jelly at the temperature of 2-8 ℃, wherein the concentration of the gelatin solution is 13-17 wt%.
5. The preparation method of the drug sustained-release hydrogel semi-embedded composite stent according to claim 1, which is characterized in that: in the step (4), the pre-crosslinked drug slow-release hydrogel solution in the step (2) is injected on the upper layer of the hydroxyapatite scaffold in the step (3) by means of a mold, and is crosslinked for 9-11min under ultraviolet light, wherein the ultraviolet intensity is 5.6-15.0 mW/cm2。
6. The preparation method of the drug sustained-release hydrogel semi-embedded composite stent according to claim 1, which is characterized in that: in the step (4), the pre-crosslinked hydrogel solution permeates into the porous hydroxyapatite scaffold with the height of 1/4-5/12.
7. The preparation method of the drug sustained-release hydrogel semi-embedded composite stent according to claim 1, which is characterized in that: in the step (5), the hydrogel semi-embedded composite bracket is cleaned by water at 37-43 ℃, and gelatin sealing the lower-layer holes of the bracket is removed.
8. A drug sustained-release hydrogel semi-embedded composite stent is characterized in that: the drug sustained-release hydrogel semi-embedded composite stent is prepared by the preparation method of the drug sustained-release hydrogel semi-embedded composite stent of any one of claims 1 to 7.
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| CA2866483A1 (en) * | 2011-12-29 | 2013-07-04 | "Nextgen" Company Limited | A biocomposite for regeneration of injured tissue and organs, a kit for making the biocomposite, a method of making the biocomposite and a method of treating inquiries |
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| CA2866483A1 (en) * | 2011-12-29 | 2013-07-04 | "Nextgen" Company Limited | A biocomposite for regeneration of injured tissue and organs, a kit for making the biocomposite, a method of making the biocomposite and a method of treating inquiries |
| CN104587531A (en) * | 2014-12-25 | 2015-05-06 | 南京臻泉医药科技有限公司 | Preparation method for gel scaffold for repairing articular cartilage injuries |
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