Patent Application
20230263942
This application claims the priority benefit of Taiwanese application serial no. 111106537, filed on Feb. 23, 2022. The entirety of the above-mentioned patent application is hereby incorporated by reference herein and made a part of this specification.
The disclosure relates to a technology of a double network hydrogel, and in particular to a highly compressible shape memory double network hydrogel, use and preparation method thereof, and an intervertebral disk scaffold.
Hydrogel is a material with water as the dispersion medium. A part of hydrophobic groups and hydrophilic residues are introduced into the water-soluble polymer with a network cross-linked structure. The hydrophilic residues combine with the water molecules to link the water molecules inside the network, while the hydrophobic residues expand when exposed to water. Traditionally prepared hydrogels are often composed of a single polymer network structure or a double network structure that is cross-linked by covalent or non-covalent bonds.
The traditional natural polymer hydrogel has low mechanical properties, and if it is used as a biomedical material for hard tissue replacement, it will be easily damaged by extrusion after being implanted into the body wear-bearing site. In addition, some of the current implant surgeries often result in large wound defect, which may influence the recovery rate and cause severe pain.
The disclosure provides a highly compressible shape memory double network hydrogel, capable of significantly improving the mechanical strength and compressibility of natural polymer hydrogels.
The disclosure also provides an application of a highly compressible shape memory double network hydrogel for an intervertebral disk scaffold.
The disclosure also provides a method of using a highly compressible shape memory double network hydrogel, capable of being used for the implantation of the intervertebral disk scaffold.
The disclosure further provides a preparation method of a highly compressible shape memory double network hydrogel, capable of producing a double network hydrogel with high compressibility and shape memory effect.
The highly compressible shape memory double network hydrogel of the disclosure includes a first network and a second network interpenetrating with each other. The first network is chemically crosslinked cellulose obtained by chemical crosslinking, and the chemical crosslinking is accomplished by formation of ether groups in the cellulose. The second network is a physically crosslinked alginate obtained by physical crosslinking, and the physical crosslinking is accomplished by reaction of the alginate with divalent metal ions. In a preparation process of the highly compressible shape memory double network hydrogel, the cellulose and the alginate are mixed first, the chemical crosslinking is then performed to obtain the first network, followed by the physical crosslinking to obtain the second network.
In one embodiment of the disclosure, the concentration of the cellulose is 1 to 10 wt. %.
In one embodiment of the disclosure, the double network hydrogel may further include a cross-linking agent for the chemical crosslinking. The concentration of the cross-linking agent is 5 to 10 wt. %.
In one embodiment of the disclosure, the cross-linking agent comprises epichlorohydrin (ECH), poly(ethylene glycol) diglycidyl ether or diglycidyl ether.
In one embodiment of the disclosure, the divalent metal ions are calcium ions, copper ions, ferrous ions, manganese ions, magnesium ions, strontium ions or zinc ions.
In one embodiment of the disclosure, the concentration of the calcium ions (Ca2+) is 1 to 10 wt. %, and the concentration of the alginate is 0.5 to 5 wt. %.
In one embodiment of the disclosure, the double network hydrogel may further include a chelating agent.
The intervertebral disk scaffold of the disclosure includes the highly compressible shape memory double network hydrogel.
The method of using the highly compressible shape memory double network hydrogel of the disclosure includes the followings. The highly compressible shape memory double network hydrogel is placed into a mold, and then a chelating agent is added to the highly compressible shape memory double network hydrogel.
The preparation method for a highly compressible shape memory double network hydrogel of the disclosure includes the followings. Cellulose and an alginate are mixed to obtain a mixture. A chemical crosslinking is performed on the cellulose in the mixture to form a hydrogel structure. A physical crosslinking is performed after the chemical crosslinking, so that the alginate in the hydrogel structure reacts with divalent metal ions to form a double network hydrogel.
In another embodiment of the disclosure, a cross-linking agent used in the chemical crosslinking comprises epichlorohydrin (ECH), poly(ethylene glycol) diglycidyl ether or diglycidyl ether.
In another embodiment of the disclosure, the divalent metal ions are calcium ions, copper ions, ferrous ions, manganese ions, magnesium ions, strontium ions or zinc ions.
In another embodiment of the disclosure, before the physical crosslinking is performed, the preparation method further includes shaping the hydrogel structure.
In another embodiment of the disclosure, a chelating agent may also be added to the double network hydrogel after the physical crosslinking, and then the hydrogel structure is shaped, and a solution containing the divalent metal ions is added to the shaped double network hydrogel to achieve a shape-fixed effect.
In another embodiment of the disclosure, the method of mixing the cellulose and the alginate includes adding alginate powder to a cellulose solution.
In another embodiment of the disclosure, before that the cellulose and the alginate are mixed, the preparation method further includes repeatedly freezing and thawing the cellulose solution three to five times.
In all embodiments of the disclosure, the chelating agent includes ethylenediaminetetraacetic acid (EDTA), nitrilotriacetic acid, or hydroxyethylethylenediaminetriacetic acid (HEDTA).
Based on the above, the disclosure uses different cross-linking mechanisms between two natural polymers to achieve the effect of improving the mechanical properties and shape memory effect of materials. Compared with traditional single network hydrogels, the double network hydrogels of the disclosure have stronger mechanical properties and may rebound under high compressive stress, thus having high compressibility. Compared with double network structures that are cross-linked entirely by covalent bonds, the double network hydrogel of the disclosure is suitable for intervertebral disk scaffolds subjected to higher compressive stress because its physical crosslinking part may absorb and dissipate energy by breaking bonds and then re-form new bonds, which may protect the internal structure of the polymer from damage and disintegration of the material.
To make the aforementioned more comprehensible, several accompanied with drawings are described in detail as follows.
The accompanying drawings are included to provide a further understanding of the disclosure, and are incorporated in and constitute a part of this specification. The drawings illustrate exemplary embodiments of the disclosure and, together with the description, serve to explain the principles of the disclosure.
The following provides many different embodiments for implementing different features of the disclosure. However, these embodiments are merely exemplary, and are not intended to limit the scope and application of the disclosure. Furthermore, for the sake of clarity, the relative dimensions (e.g., length, spacing, etc.) and relative positions of the compositions or structures may be reduced or enlarged.
Referring to
Referring to
Then, referring to
Next, referring to
If high strength and high compressibility are required, a chelating agent such as ethylenediaminetetraacetic acid (EDTA) may be added to the double network hydrogel 204 after physical crosslinking instead of shaping before the step in
Since the crosslinking sequence of the double network hydrogel 204 is to mix the cellulose 106 and the alginate 110 first, and to perform the physical crosslinking after the chemical crosslinking, the resulting product has high compressibility and also has shape memory effect. The following mechanical performance tests are used to verify effectiveness.
Ingredient:
The ingredients are stored at room temperature.
First, 0.8 g of NaOH and 0.4 g of urea were added to 10 mL of deionized water to prepare a solution of 4 wt. % urea/8 wt. % NaOH.
0.4 g (the concentration of 4 wt. %) of cellulose was added to the solution and mixed for 15 minutes to obtain a cellulose solution.
The cellulose solution is stored at −80° C. for 24 hours, and then the cellulose solution is repeatedly frozen and thawed for three to five times. This step is to allow the cellulose and a cross-linking agent to react at room temperature. If the number of freezing/thawing is less than three times, the reaction between the cellulose and the cross-linking agent needs to be performed at 60° C. in order to form the gel. However, if the number of freezing/thawing is greater than five, the reactivity of the hydroxyl group on the cellulose side chain is reduced, making the cellulose solution unable to be gelled.
Next, 0.2 g (the concentration of 2 wt. %) of alginate powder was added to 10 mL of the cellulose solution, and stirred for 30 minutes to obtain a mixture.
0.8 mL of epichlorohydrin (ECH)(the concentration of 8 wt. %) was added to the mixture and stirred for 15 minutes at room temperature to become a hydrogel, and then the hydrogel was stored in a specific mold.
Then, the hydrogel was immersed in a 4 wt. % calcium chloride solution (calcium ions concentration of 4 wt. %) for two hours to obtain a double network hydrogel.
The steps of Experimental Example 1 were followed, but no alginate powder was added, nor was the hydrogel immersed in calcium phosphate. Therefore, what was obtained was a single network hydrogel after chemical crosslinking of the cellulose.
The ingredients of Experimental Example 1 were used, but the preparation process was as follows.
First, 0.8 g of NaOH and 0.4 g of urea were added to 10 mL of deionized water to prepare a solution of 4 wt. % urea/8 wt. % NaOH.
0.4 g (the concentration of 4 wt. %) of cellulose was added to the solution and mixed for 15 minutes to obtain a cellulose solution.
The cellulose solution is stored at −80° C. for 24 hours, and then the cellulose solution is repeatedly frozen and thawed for three to five times. This step is to allow the cellulose and the cross-linking agent to react at room temperature. If the number of freezing/thawing is less than three times, the reaction between the cellulose and the cross-linking agent needs to be performed at 60 degrees in order to form the gel. However, if the number of freeze/thaw is greater than five, the reactivity of the hydroxyl group on the cellulose side chain is reduced, making the cellulose solution unable to be gelled.
Next, 0.8 mL of ECH was added to the cellulose solution and stirred for 15 minutes at room temperature, and then stored in a specific mold for 24 hours to become a cellulose hydrogel.
0.4 g (the concentration of 4 wt. %) of alginate powder was added to 10 mL of the deionized water to form an alginate solution.
Then, the cellulose hydrogel was immersed in the alginate solution and stirred for about 24 hours.
After that, the hydrogel was immersed in a calcium chloride solution with a concentration of 4 wt. % for two hours to obtain a double network hydrogel.
<Mechanical Strength>
Samples of Experimental Example 1, Comparative Example 1, and Comparative Example 2 were subjected to stress-strain curves, and the results are shown in
It can be seen from
<Cyclic Compression Test>
The sample of Experimental Example 1 was subjected to a cyclic compression test, and the results are shown in
It can be seen from
With the above characteristics verified by experiments, the highly compressible shape memory double network hydrogel of the disclosure may be applied to human tissues or organs that need to withstand strong compressive stress or load, such as intervertebral discs (cervical vertebrae, lumbar vertebrae, etc.) and articular cartilage.
When highly compressed, the double network hydrogel has good resilience and is therefore particularly suitable for use as a tissue graft that requires repetitive compression, such as an intervertebral disk scaffold. Thus, an intervertebral disk scaffold according to a third embodiment of the disclosure includes the highly compressible shape memory double network hydrogel. In addition, the highly compressible shape memory double network hydrogel of the disclosure also has shape memory properties that facilitate the surgical implantation process and reduce the difficulty, thus simplifying the complicated surgical procedures and reducing risks, pain and complications.
For example, in order to facilitate implantation into a small space (e.g. the space between cervical vertebrae), the highly compressible shape memory double network hydrogel of the disclosure may be shaped into a small sheet, which is then used by first placing it into an accommodation space (e.g. between cervical vertebrae and lumbar vertebrae) and then adding a chelating agent to the highly compressible shape memory double network hydrogel to expand the sheet back to its original shape, in which the chelating agent is, for example, EDTA. In another embodiment, the highly compressible shape memory double network hydrogel of the disclosure may also be placed directly into the mold without being shaped, and no chelating agent is added to retain better strength and compressibility.
The following biological experiments were conducted to demonstrate the efficacy of the highly compressible shape memory double network hydrogel for the intervertebral disk scaffold.
<Magnetic Resonance Imaging (MRI) Test>
The following table shows the details of each group of MRI test subjects.
In the table, DC represents a control group in which the intervertebral discs were removed, and no material was implanted; DN represents an experimental group in which the intervertebral discs were replaced with the double network hydrogel of Experimental Example 1; PDN and GPDN represent experimental groups in which the intervertebral discs were replaced with hydrogels carrying growth factors or different therapeutic factors, respectively.
Then, a period of time after the double network hydrogel was implanted in the caudal vertebrae of the rats, T2-MRI was used to directly observe the morphology and water content of the implanted device. The results were shown in
As can be seen from
<Immunostaining (IHC) Test>
Immunostaining was used to observe whether new intervertebral disc tissue was generated in the caudal vertebrae of the rat. Target proteins are Vimentin (green) and FOXF1 (Forkhead Box protein F1, red), both of which are specific proteins commonly found in healthy intervertebral discs, so this method may be used to determine whether the implanted double network hydrogel has a therapeutic effect on intervertebral disc regeneration and repair.
For more effective analysis, the red and green areas in
As can be seen from
To sum up, the disclosure uses natural polymers as ingredients, and through a specific crosslinking sequence, the prepared double network hydrogel has high compressibility and shape memory function. The double network hydrogel of the disclosure is mainly composed of chemical crosslinking and physical crosslinking. Chemical crosslinking forms a hard segment to stabilize the hydrogel, which is responsible for controlling the permanent shape, while physical crosslinking forms a soft segment, which is reversible and determines the temporary shape of the hydrogel. Therefore, the double network hydrogel of the disclosure can be applied zto intervertebral disk scaffolds that are subject to higher compressive stress.
It will be apparent to those skilled in the art that various modifications and variations can be made to the disclosed without departing from the scope or spirit of the disclosure. In view of the foregoing, it is intended that the disclosure covers modifications and variations provided that they fall within the scope of the following claims and their equivalents.
| Number | Date | Country | Kind |
|---|---|---|---|
| 111106537 | Feb 2022 | TW | national |
