Magnetic responsive elastin-like polypeptide hydrogel and its preparation method and application

Abstract

The present invention provides a magnetically responsive elastin-like polypeptide hydrogel and a preparation method and application thereof. The preparation method of the magnetically responsive elastin-like polypeptide hydrogel comprises: (1) mixing nanoclay with water to obtain a nanoclay dispersion; (2) adding elastin-like polypeptide to the nanoclay dispersion and stirring until completely dissolved; (3) adding magnetic nanoparticles to the above system and stirring to obtain a magnetically responsive hydrogel that can be used for 3D printing, and the printed structure is solidified by a disuccinimidyl suberate solution to complete the shaping process. The preparation method is simple, and the gel system has magnetic responsiveness, printability and biocompatibility, and has good application prospects in the fields of tissue engineering and regenerative medicine.

Description

Technical Field

[0001] The present invention belongs to the field of material engineering, and in particular, relates to a magnetically responsive elastin-like polypeptide hydrogel and a preparation method and application thereof. Background Art

[0002] Hydrogel is a soft material with a network cross-linked structure and rich in water molecules. Because of its characteristics similar to the extracellular matrix, it has received more and more attention in the biomedical field. With the development of science and technology, many fields require the materials used to become "smart" to adapt to a wider range of application scenarios. Depending on whether the hydrogel can respond to external stimuli, it can be divided into traditional hydrogels and smart hydrogels. As magnetic field manipulation technology becomes more and more mature, magnetically responsive hydrogels that can respond to external magnetic fields have gradually become a highlight in the research of smart hydrogels. At the same time, combined with 3D printing technology, how to improve the printability of magnetically responsive hydrogels is particularly critical to expanding its application range. Summary of the invention

[0003] The purpose of the present invention is to provide a novel magnetically responsive elastin-like polypeptide hydrogel and a preparation method and application thereof.

[0004] The present invention is conceived as follows: to provide a 3D printed magnetically responsive elastin-like polypeptide hydrogel and a preparation method thereof. The hydrogel material has simple ingredients and is easy to prepare, and is composed of nanoclay, elastin-like polypeptide and magnetic nanoparticles. Nanoclay, as a rheological additive, can give the gel system an excellent extrusion molding effect. Since the nanoclay used is rich in lysine groups, it can generate intermolecular forces with the nanoclay to form a physically cross-linked hydrogel system. Subsequently, the magnetic nanoparticles are wrapped in the hydrogel system, and the entire system can respond under the action of an external magnetic field. The gel system has both magnetic responsiveness, printability and biocompatibility, and has good application prospects in the biomedical field.

[0005] In order to achieve the purpose of the present invention, in a first aspect, the present invention provides a method for preparing a magnetically responsive elastin-like polypeptide hydrogel, comprising the following steps:

[0006] (1) mixing nanoclay with water to obtain a nanoclay dispersion;

[0007] (2) adding the elastin-like polypeptide to the nanoclay dispersion and stirring until completely dissolved;

[0008] (3) Adding magnetic nanoparticles to the system of step (2) and stirring to obtain a magnetically responsive elastin-like polypeptide hydrogel.

[0009] In the present invention, the amino acid sequence of the elastin-like polypeptide is [(VPGVG)2VPGKG(VPGVG)2] 25 (SEQ ID NO: 1).

[0010] Furthermore, the nanoclay is of gel type, and the particle diameter of the nanoclay is 20-100 nm.

[0011] Furthermore, the concentration of the nanoclay dispersion in step (1) is 3%-15% w / v.

[0012] Furthermore, the concentration of the elastin-like polypeptide in the system of step (2) is 1% to 20% w / v.

[0013] Furthermore, the magnetic nanoparticles in step (3) are Fe3O4, with a diameter of 10-100 nm and a concentration of 2% to 25% w / v.

[0014] The above steps (1) to (3) are all carried out at room temperature.

[0015] In a second aspect, the present invention provides a magnetically responsive elastin-like polypeptide hydrogel prepared according to the method.

[0016] In a third aspect, the present invention provides any of the following applications of the magnetically responsive elastin-like polypeptide hydrogel:

[0017] 1) Used in biomaterials engineering;

[0018] 2) Used in tissue engineering;

[0019] 3) Used in medicine and environment fields;

[0020] 4) Used for 3D printing.

[0021] In a fourth aspect, the present invention provides a method for using the magnetically responsive elastin-like polypeptide hydrogel, comprising: using the magnetically responsive elastin-like polypeptide hydrogel for 3D printing, and then immersing the printed structure in a curing agent for curing and shaping.

[0022] The curing agent may be a disuccinimidyl suberate (DSS) solution, glutaraldehyde or BS3 (sulfo-DSS) and the like.

[0023] Furthermore, the concentration of the disuccinimidyl suberate solution is 5% to 20% w / v, and the printed structure is immersed in the disuccinimidyl suberate solution for 2 to 500 minutes.

[0024] Compared with existing hydrogels, the magnetically responsive elastin-like polypeptide hydrogel for 3D printing provided by the present invention has the following advantages:

[0025] (I) From the perspective of material sources, elastin-like polypeptides are polypeptides with elastin characteristics that are designed and synthesized based on the amino acid sequence of natural elastin. The hydrogels prepared from them have good mechanical properties and have therefore received widespread attention in the biomedical field.

[0026] (ii) The elastin-like protein sequence used in the present invention contains abundant lysine groups, which can produce intermolecular interactions with nanoclay to form an extrudable, self-healing polymer network, which can be used as a 3D printing material.

[0027] (iii) Thanks to the pre-mixed magnetic nanoparticles, the hydrogel can respond to an external magnetic field.

[0028] (iv) The materials used in the hydrogel have good biocompatibility. The preparation process is simple and the conditions are mild. The prepared magnetically responsive elastin-like polypeptide hydrogel has good application prospects in many fields. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] Figure 1 This is a digital photo of the 3D printed magnetically responsive elastin-like polypeptide hydrogel and its printed grid structure prepared in a preferred embodiment of the present invention.

[0030] Figure 2 The self-healing rheological results of the 3D printed magnetically responsive elastin-like polypeptide hydrogel prepared in the preferred embodiment of the present invention.

[0031] Figure 3 This is a microscope photograph of vascular endothelial cells cultured in a 3D printed magnetically responsive elastin-like polypeptide hydrogel prepared in a preferred embodiment of the present invention.

[0032] Figure 4 This is a diagram showing the live and dead results of vascular endothelial cells cultured in a 3D printed magnetically responsive elastin-like polypeptide hydrogel prepared in a preferred embodiment of the present invention. DETAILED DESCRIPTION

[0033] The present invention provides a method for preparing a 3D printed magnetically responsive hydrogel, comprising the following steps:

[0034] (1) adding nanoclay into an aqueous solution and stirring the solution thoroughly to obtain a uniform nanoclay dispersion;

[0035] (2) adding elastin-like polypeptide to the nanoclay dispersion and stirring thoroughly until completely dissolved;

[0036] (3) adding magnetic nanoparticles and stirring thoroughly to obtain a magnetically responsive elastin-like polypeptide hydrogel that can be used for 3D printing; the printed structure is immersed in a disuccinimidyl suberate solution and solidified to complete the shaping process.

[0037] In the above step (1), nanoclay is added into the aqueous solution and stirred thoroughly to obtain a uniform nanoclay dispersion.

[0038] In the above method, the nanoclay is of gel type, the diameter of the nanoclay particles is 20-100 nm, and the concentration is 5%-15% w / v.

[0039] In the above step (2), the elastin-like polypeptide is added to the nanoclay dispersion and stirred thoroughly until it is completely dissolved.

[0040] In the above method, the amino acid sequence of the elastin-like polypeptide is [(VPGVG)2VPGKG(VPGVG)2] 25 , concentration is 1% ~ 20% w / v.

[0041] In the above step (3), magnetic nanoparticles are added and stirred thoroughly to obtain a magnetically responsive hydrogel that can be used for 3D printing. The printed structure is solidified by a disuccinimidyl suberate solution to complete the shaping process.

[0042] In the above method, the diameter of the magnetic nanoparticles is 10-100 nm, and the concentration is 2% to 25% w / v.

[0043] In the above method, the concentration of the disuccinimidyl suberate solution used is 5% to 20% w / v, and the soaking time is 2 to 500 minutes.

[0044] The following examples are used to illustrate the present invention, but are not intended to limit the scope of the present invention. Unless otherwise specified, the technical means used in the examples are conventional means well known to those skilled in the art, and the raw materials used are all commercially available products.

[0045] The nanoclay used in the following examples has a particle diameter of 20-100 nm and is purchased from BYK Chemicals of Germany.

[0046] The magnetic nanoparticles were Fe3O4 with a diameter of 10-100 nm, purchased from Beijing Dekedaojin Technology Co., Ltd.

[0047] In the present invention, the preparation method of the disuccinimidyl suberate solution is as follows: DSS is first dissolved with a small amount of co-solvent DMF, and then added into water at a volume ratio of 1:10 to obtain the solution.

[0048] Example 1 Preparation of magnetically responsive elastin-like polypeptide hydrogel for 3D printing

[0049] (1) Add nanoclay to the aqueous solution and stir thoroughly to obtain a uniform nanoclay dispersion. The details are as follows:

[0050] 7 g of nanoclay was added to 100 mL of aqueous solution and stirred thoroughly until completely dissolved to obtain a uniform nanoclay dispersion.

[0051] (2) Add elastin-like polypeptide to the nanoclay dispersion and stir thoroughly until completely dissolved. The details are as follows:

[0052] Add 5 g of elastin-like polypeptide (SEQ ID NO: 1) to the prepared nanoclay dispersion and stir thoroughly until completely dissolved.

[0053] (3) Add magnetic nanoparticles and stir thoroughly to obtain a magnetically responsive hydrogel that can be used for 3D printing. The printed structure is cured with disuccinimidyl suberate to complete the shaping process. The details are as follows:

[0054] Add 5 g of magnetic nanoparticles to the above system and stir thoroughly to obtain a magnetically responsive hydrogel that can be used for 3D printing, such as Figure 1 The printed structure was immersed in a 5% w / v disuccinimidyl suberate solution for 30 min for secondary cross-linking, thus completing the shaping process of the printed structure.

[0055] Example 2 Preparation of magnetically responsive elastin-like polypeptide hydrogel for 3D printing

[0056] (1) Add nanoclay to the aqueous solution and stir thoroughly to obtain a uniform nanoclay dispersion. The details are as follows:

[0057] 10 g of nanoclay was added to 100 mL of aqueous solution and stirred thoroughly until completely dissolved to obtain a uniform nanoclay dispersion.

[0058] (2) Add elastin-like polypeptide to the nanoclay dispersion and stir thoroughly until completely dissolved. The details are as follows:

[0059] 3 g of elastin-like polypeptide (SEQ ID NO: 1) was added to the prepared nanoclay dispersion and stirred thoroughly until completely dissolved.

[0060] (3) Add magnetic nanoparticles and stir thoroughly to obtain a magnetically responsive hydrogel that can be used for 3D printing. The printed structure is cured with disuccinimidyl suberate to complete the shaping process. The details are as follows:

[0061] 10 g of magnetic nanoparticles were added to the above system and stirred thoroughly to obtain a magnetically responsive hydrogel that can be used for 3D printing. The printed structure was immersed in a 10% w / v disuccinimidyl suberate solution for 60 min for secondary crosslinking, thus completing the shaping process of the printed structure. The experimental results were similar to those of Example 1.

[0062] Example 3 Performance of 3D printed magnetically responsive elastin-like polypeptide hydrogel

[0063] 1. Rheological investigation of self-healing of 3D printed magnetically responsive elastin-like polypeptide hydrogel

[0064] The self-healing properties of the hydrogel (prepared in Example 1) were verified using a rheometer (MCR302, Anton Paar, Austria). A flat plate mold (25 mm in diameter) was used for rheological measurements. The hydrogel sample was first placed between the flat plates, and then the sample was repeatedly subjected to high strain (300%) and low strain (1%) shear tests, and the changes in the storage modulus G′ and the loss modulus G″ were recorded.

[0065] The experimental results are shown in Figure 2 ,from Figure 2 It can be seen that under high-strain shear, the network breaks and the loss modulus is greater than the storage modulus, but under low-strain conditions, the broken network can self-heal and re-exhibit the original state where the storage modulus is higher than the loss modulus. Under repeated high-low strain shear, the hydrogel maintains a consistent modulus change, showing that its internal cross-linked network has certain self-healing characteristics.

[0066] 2. 3D printed magnetically responsive elastin-like polypeptide hydrogels for culturing vascular endothelial cells

[0067] Endothelial cells (human umbilical vein endothelial cells HUVEC) were cultured in a system of RPMI 1640 basal medium + 10% fetal bovine serum (FBS) + 1% penicillin / streptomycin. The hydrogel scaffold prepared by an extrusion 3D printer was soaked in the medium overnight, and then the endothelial cells were planted on the surface of the hydrogel and cultured for 1, 3, 7, and 14 days, respectively. The cells were observed and photographed using a fluorescence microscope, and tested using a live-death test kit to obtain the cell survival rate.

[0068] Microscope photos of 3D printed magnetically responsive elastin-like polypeptide hydrogels culturing vascular endothelial cells are shown in Figure 3 ,from Figure 3 It can be seen that vascular endothelial cells successfully adhered to the hydrogel scaffold.

[0069] The results of 3D printed magnetically responsive elastin-like polypeptide hydrogels culturing vascular endothelial cells are shown in Figure 4 ,from Figure 4 It can be seen that the vascular endothelial cells maintained a survival rate of more than 80% throughout the culture process, which shows that the hydrogel has good cell compatibility.

[0070] Although the present invention has been described in detail above with general descriptions and specific embodiments, it is obvious to those skilled in the art that some modifications or improvements can be made based on the present invention. Therefore, these modifications or improvements made without departing from the spirit of the present invention all fall within the scope of protection claimed by the present invention.

Claims

1. A method for preparing a magnetically responsive elastin-like polypeptide hydrogel, characterized in that: The following steps are involved: (1) mixing nanoclay with water to obtain a nanoclay dispersion; (2) adding elastin-like polypeptide to the nanoclay dispersion and stirring until completely dissolved; (3) adding magnetic nanoparticles to the system of step (2) and stirring to obtain a magnetically responsive elastin-like polypeptide hydrogel; Wherein, the amino acid sequence of the elastin-like polypeptide is shown in SEQ ID NO: 1; The nanoclay is of gel type, and the particle diameter of the nanoclay is 20-100 nm; The concentration of the nanoclay dispersion in step (1) is 3%-15% w / v; The concentration of the elastin-like polypeptide in the system of step (2) is 1% to 20% w / v; The magnetic nanoparticles in step (3) are Fe3O4 with a diameter of 10-100 nm and a concentration of 2%-25% w / v.

2. The method according to claim 1, characterized in that The method is carried out at room temperature.

3. A magnetically responsive elastin-like polypeptide hydrogel prepared according to the method of claim 1 or 2.

4. Any of the following uses of the magnetically responsive elastin-like polypeptide hydrogel according to claim 3: 1) Used for preparing biological materials; 2) Application in the preparation of materials for use in the medical and environmental fields; 3) Used as 3D printing material.

5. The method for using the magnetically responsive elastin-like polypeptide hydrogel according to claim 3, characterized in that: include: Using the magnetically responsive elastin-like polypeptide hydrogel for 3D printing, and then immersing the printed structure in a curing agent for curing and shaping; The curing agent is disuccinimidyl suberate solution, glutaraldehyde or BS3.

6. The method according to claim 5, characterized in that The concentration of the disuccinimidyl suberate solution is 5% to 20% w / v, and the printed structure is immersed in the disuccinimidyl suberate solution for 2 to 500 minutes.

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