4d-printed bone tissue scaffolds with tunable microenvironments, methods of making, and uses thereof

Abstract

The application provides a 4D printed bone tissue scaffold with adjustable microenvironment and a preparation method and application thereof, and relates to the technical field of bone repair. The 4D printed bone tissue scaffold with adjustable microenvironment comprises a scaffold structure and a film covering the surface of the scaffold structure, and the scaffold structure is provided with holes; under the condition that the scaffold structure is deformed under stress, the holes are configured to switch between a contact state and a separation state. During the deformation of the scaffold structure under stress, the holes can switch between the contact state and the separation state, generate a triboelectric effect, and further generate a micro-current, so as to construct a physical microenvironment conducive to cell growth, and further conducive to the regeneration of damaged bone tissue; as a mechanical wave, ultrasound provides additional mechanical stimulation at the bone defect site, can make the scaffold structure deform under stress, and can also make the scaffold structure generate a piezoelectric effect and a triboelectric effect, induce the generation of a micro-current on the surface of the scaffold structure, and thus construct a physical microenvironment conducive to cell growth, and further conducive to the regeneration of damaged bone tissue.

Description

Technical Field

[0001] This invention relates to the field of bone repair technology, and more specifically, to 4D-printed bone tissue scaffolds with adjustable microenvironments, their preparation methods, and applications. Background Technology

[0002] In clinical practice, bone defects larger than the critical size of 2.5 cm caused by trauma, infection debridement, tumor resection, or congenital malformations often result in the body's inability to spontaneously repair the defect, frequently leading to severe incomplete healing. In such cases, bone grafting is the most common treatment, often using autologous or allogeneic tissue transplantation. However, autologous tissue transplantation often involves new trauma or is limited by donor site conditions, making it difficult to achieve the desired therapeutic effect. Allogeneic tissue transplantation presents challenges related to donor availability and immune rejection. Especially for larger bone defects, traditional treatment methods are no longer sufficient to meet medical needs.

[0003] The development of tissue engineering technology has provided another option for the repair and reconstruction of damaged tissues. Bioscaffolds are a key factor in tissue engineering research, serving as templates to guide the regeneration of damaged tissues. However, existing bioscaffolds lack a suitable physical microenvironment for cell growth, making it difficult to achieve the function of regenerating damaged bone tissue. Summary of the Invention

[0004] The problem addressed by this invention is how to solve the difficulty in regenerating damaged bone tissue due to the lack of a suitable physical microenvironment for cell growth in existing biological scaffolds.

[0005] To address the aforementioned issues, this invention provides a 4D-printed bone tissue scaffold with adjustable microenvironment, its preparation method, and its applications.

[0006] In a first aspect, the present invention provides a 4D-printed bone tissue scaffold with adjustable microenvironment, comprising a scaffold structure and a membrane covering the surface of the scaffold structure, wherein the scaffold structure is provided with pores.

[0007] The holes have a contact state where their edges are in contact and a separation state where their edges are not in contact. When the support structure is deformed under the stimulation of ultrasound, the holes are configured to switch between the contact state and the separation state.

[0008] Optionally, the surface of the membrane away from the support structure is provided with micropillars, which have collapsed and upright states, and are configured to switch between collapsed and upright states under ultrasonic stimulation.

[0009] Optionally, the ultrasound waves are 1-3MHz wavelength and 30mW / cm. 2 -1000 mW / cm 2 Low-intensity pulsed ultrasound.

[0010] Alternatively, the scaffold structure may include shape memory polymers, carbon nanotubes, barium titanate, and Fe3O4 nanoparticles.

[0011] Alternatively, the membrane may comprise shape memory polymers, carbon nanotubes, barium titanate, and Fe3O4 nanoparticles.

[0012] Optionally, the mass of carbon nanotubes is 2-5 wt% of the mass of shape memory polymer, the mass of barium titanate is 3-5 wt% of the mass of shape memory polymer, and the mass of Fe3O4 nanoparticles is 10-15 wt% of the mass of shape memory polymer.

[0013] Optionally, the pore size is 200-500 micrometers, and the porosity of the pores on the support structure is 25-80%.

[0014] Secondly, the present invention provides a method for preparing a 4D-printed bone tissue scaffold with an adjustable microenvironment, comprising the following steps:

[0015] Based on the bone defect, a structural model of the scaffold structure is designed, and the scaffold structure with holes is printed using fused deposition modeling or direct-write 4D printing methods.

[0016] The film is printed using a photopolymerization printing method;

[0017] A film is attached to the surface of the support structure.

[0018] Optionally, the steps of designing a structural model of the scaffold structure according to the bone defect, and printing it using fused deposition modeling or direct-write 4D printing methods to obtain a scaffold structure with holes include:

[0019] Based on CT scans and reconstruction software, a 3D printed model of a scaffold structure with holes was designed according to the shape and anatomical structure of the bone defect.

[0020] The 3D printing model is imported into a 3D printer, and the scaffold structure is prepared using fused deposition modeling or direct-write 4D printing methods.

[0021] Thirdly, the present invention provides an application of a 4D-printed bone tissue scaffold with adjustable microenvironment as described in any of the preceding claims in the field of filling bone defects and inducing osteoogenesis.

[0022] The beneficial effects of the adjustable microenvironment 4D-printed bone tissue scaffold, its preparation method, and its application of the present invention are as follows: First, the scaffold structure has pores. During the deformation process under stress, the pores can switch between a contact state and a separation state, and the distance between the pores changes. This change in distance generates a triboelectric effect, which in turn generates a microcurrent. With the generation of the microcurrent, a physical microenvironment conducive to cell growth is constructed, which in turn facilitates the regeneration of damaged bone tissue. Second, ultrasound, as a mechanical wave, provides additional mechanical stimulation at the bone defect site. This can cause the scaffold structure to undergo microdeformation under stress, and can also generate piezoelectric and triboelectric effects, inducing a microcurrent on the surface of the scaffold structure, thereby constructing a physical microenvironment conducive to cell growth, which in turn facilitates the regeneration of damaged bone tissue. Attached Figure Description

[0023] Figure 1 This is a schematic diagram of the structure of the micropillars on the membrane in a collapsed state according to an embodiment of the present invention;

[0024] Figure 2 This is a schematic diagram of the structure of the micropillars on the membrane in an upright state according to an embodiment of the present invention. Detailed Implementation

[0025] To make the above-mentioned objects, features, and advantages of the present invention more apparent and understandable, specific embodiments of the present invention will be described in detail below with reference to the accompanying drawings. Although some embodiments of the present invention are shown in the drawings, it should be understood that the present invention can be implemented in various forms and should not be construed as limited to the embodiments set forth herein. Rather, these embodiments are provided to provide a more thorough and complete understanding of the present invention. It should be understood that the accompanying drawings and embodiments of the present invention are for illustrative purposes only and are not intended to limit the scope of protection of the present invention.

[0026] Unless otherwise defined, all technical and scientific terms used in this application have the same meaning as commonly understood by one of ordinary skill in the art to which this application pertains; the terminology used in the description of this application is for the purpose of describing particular embodiments only and is not intended to limit this application.

[0027] The term "comprising" and its variations as used herein are open-ended, meaning "including but not limited to"; the term "based on" means "at least partially based on"; the term "one embodiment" means "at least one embodiment"; the term "another embodiment" means "at least one additional embodiment"; the term "some embodiments" means "at least some embodiments"; and the term "optionally" means "optional embodiments". Definitions of other terms will be given in the description below. It should be noted that the concepts of "first," "second," etc., mentioned in this invention are used to distinguish different objects, not to describe a specific order or hierarchy. Furthermore, the terms "first" and "second" are used for descriptive purposes only and should not be construed as indicating or implying relative importance or implicitly specifying the number of indicated technical features. Thus, a feature defined with "first" or "second" may explicitly or implicitly include one or more of that feature. In the description of this application, unless otherwise stated, "a plurality of" means two or more.

[0028] In related technologies, the mechanical microenvironment surrounding cells plays a crucial role in cell growth, development, and differentiation. This mechanical microenvironment includes mechanical pressure, microgravity, magnetic fields, electric fields, and extracellular matrix stiffness, all of which can influence cellular biological functions by regulating intracellular signaling pathways. For example, natural bone exhibits piezoelectric behavior under mechanical stress, with a piezoelectric constant of 0.7 pC / N–2.3 pC / N. This is closely related to the movement of asymmetric collagen molecules and ionic liquids generated by collagen fiber sliding. Piezoelectric and triboelectric biomaterials generate surface charges similar to those on the bone surface under external stress. Implanting piezoelectric bioscaffold materials at bone defect sites generates stress through physiological load, thereby producing electrical stimulation and promoting bone tissue regeneration.

[0029] Low-intensity pulsed ultrasound (LIPUS) is a non-invasive physical therapy used to accelerate bone repair and promote bone healing. As a mechanical wave, LIPUS can provide additional mechanical stimulation at bone defects, not only activating kinase pathways and upregulating osteogenic protein expression to promote osteogenesis, but also acting on piezoelectric and triboelectric biomaterials to induce piezoelectric and triboelectric effects, further promoting bone regeneration. Studies have shown that when LIPUS and piezoelectric biomaterials are applied simultaneously in bone tissue engineering, cell proliferation and osteogenic differentiation in vitro, as well as in vivo osteoogenesis in animal models, are more effective than using LIPUS or piezoelectric biomaterials alone. Therefore, introducing LIPUS as a biophysical stimulus into bone tissue engineering can not only induce microcurrents on the surfaces of piezoelectric and triboelectric materials, but also serve as a mechanical stimulus to further promote bone tissue formation. In addition, low-frequency ultrasound can drive the microstructures on the scaffold surface to oscillate back and forth, thereby constructing the biomechanical microenvironment required for cell growth.

[0030] To address the problems existing in the aforementioned related technologies, this embodiment provides a 4D-printed bone tissue scaffold with adjustable microenvironment, its preparation method, and its application.

[0031] This invention provides a 4D-printed bone tissue scaffold with adjustable microenvironment, comprising a scaffold structure and a membrane covering the surface of the scaffold structure, wherein the scaffold structure has pores.

[0032] The holes have a contact state where their edges are in contact and a separation state where their edges are not in contact. When the support structure is deformed under the stimulation of ultrasound, the holes are configured to switch between the contact state and the separation state.

[0033] In this embodiment, firstly, the scaffold structure has holes. During the deformation process under stress, the holes can switch between a contact state and a separation state, and the distance between the holes changes. This change in distance generates a triboelectric effect, accompanied by the generation of microcurrents, thereby constructing a physical microenvironment conducive to cell growth, which in turn facilitates the regeneration of damaged bone tissue. Secondly, ultrasound, as a mechanical wave, provides additional mechanical stimulation at the bone defect site, which can cause the scaffold structure to undergo microdeformation under stress. It can also generate piezoelectric and triboelectric effects on the scaffold structure, inducing microcurrents on the surface of the scaffold structure, thereby constructing a physical microenvironment conducive to cell growth, which in turn facilitates the regeneration of damaged bone tissue.

[0034] Optionally, the surface of the membrane away from the support structure is provided with micropillars, which have collapsed and upright states, and are configured to switch between collapsed and upright states under ultrasonic stimulation.

[0035] In this optional embodiment, such as Figure 1 and Figure 2 As shown, the surface of the scaffold structure is covered with a membrane containing micropillars. Under the stimulation of ultrasound, the micropillars on the surface of the membrane will deform. During the repeated process of uprighting and collapsing, the micropillars will exert mechanical stimulation on the cells, constructing a physical microenvironment conducive to cell growth, thereby inducing bone tissue regeneration.

[0036] Optionally, the ultrasound waves are 1-3MHz wavelength and 30mW / cm. 2 -1000 mW / cm 2 Low-intensity pulsed ultrasound.

[0037] In this optional embodiment, a wavelength of 1-3 MHz and 30 mW / cm 2 -1000 mW / cm 2Low-intensity pulsed ultrasound (LIPUS) is a non-invasive physical therapy used to accelerate bone repair and promote bone healing. As a mechanical wave, it provides additional mechanical stimulation at the bone defect site. It can not only activate kinase pathways and upregulate osteoblast protein expression to promote osteoogenesis, but also act on the scaffold structure of piezoelectric and triboelectric biomaterials to induce piezoelectric and triboelectric effects on the scaffold structure, thereby creating a physical microenvironment conducive to cell growth and thus facilitating the regeneration of damaged bone tissue.

[0038] Specifically, low-intensity pulsed ultrasound stimulation for 5-30 minutes allows the scaffold structure to generate microcurrents under ultrasound stimulation, and the micropillars on the membrane surface can vibrate under ultrasound stimulation, completing the collapse-to-standing conversion action.

[0039] Alternatively, the scaffold structure may include shape memory polymers, carbon nanotubes, barium titanate, and Fe3O4 nanoparticles.

[0040] In this optional embodiment, fused deposition modeling (FDM) or direct-write 4D printing methods are used, with shape memory polymers, carbon nanotubes, barium titanate, and Fe3O4 nanoparticles as the main raw materials. These are blended to create a printing filament for 4D printing. Utilizing its variable stiffness properties, the resulting bone tissue engineering scaffold structure exhibits good retention capabilities. This scaffold structure can be implanted into the body via minimally invasive surgery and unfolds under stimulation such as a magnetic field. The bone tissue scaffold can also be implanted even with uncertain bone defect shapes. Utilizing its shape memory effect, the scaffold material is in a relatively soft state above the glass transition temperature, allowing the bone tissue scaffold to self-fill irregular bone defects. Furthermore, using carbon nanotubes to fabricate triboelectric / piezoelectric scaffold structures significantly improves their output performance. This is mainly due to the excellent electrical conductivity and mechanical strength of carbon nanotubes, which helps to improve the generation and charge transport efficiency of triboelectric charges.

[0041] Specifically, the shape memory polymer is one or more of the following: shape memory polylactic acid, shape memory polyurethane, shape memory polycaprolactone, shape memory chitosan, and their complexes and derivatives.

[0042] Specifically, the particle size of barium titanate is 200-800 nm, the particle size of carbon nanotubes is 20-50 nm, and the particle size of Fe3O4 nanoparticles is 20-50 nm.

[0043] Alternatively, the membrane may comprise shape memory polymers, carbon nanotubes, barium titanate, and Fe3O4 nanoparticles.

[0044] In this optional embodiment, the membrane has high precision and is prepared using a photopolymerization printing method, which allows it to perfectly adhere to the surface of the bone tissue scaffold structure under thermal stimulation.

[0045] Optionally, the mass of carbon nanotubes is 2-5 wt% of the mass of shape memory polymer, the mass of barium titanate is 3-5 wt% of the mass of shape memory polymer, and the mass of Fe3O4 nanoparticles is 10-15 wt% of the mass of shape memory polymer.

[0046] In this optional embodiment, the raw materials in this proportion are blended to obtain a printing line for 4D printing, which has good variable stiffness properties, thereby resulting in a better ability to maintain the bone tissue engineering scaffold structure.

[0047] Optionally, the pore size is 200-500 micrometers, and the porosity of the pores on the support structure is 25-80%.

[0048] In this optional embodiment, the distance between the holes changes during the compression process of the support structure. This change in distance will generate a triboelectric effect, which in turn will generate a microcurrent.

[0049] Secondly, the present invention provides a method for preparing a 4D-printed bone tissue scaffold with an adjustable microenvironment, comprising the following steps:

[0050] Based on the bone defect, a structural model of the scaffold structure is designed, and the scaffold structure with holes is printed using fused deposition modeling or direct-write 4D printing methods.

[0051] The film is printed using a photopolymerization printing method;

[0052] A film is attached to the surface of the support structure.

[0053] In this embodiment, the 4D-printed bone tissue scaffold with adjustable microenvironment, prepared by 3D laser-based direct-write 4D printing technology, can maintain cell viability very well.

[0054] Optionally, the steps of designing a structural model of the scaffold structure according to the bone defect, and printing it using fused deposition modeling or direct-write 4D printing methods to obtain a scaffold structure with holes, specifically include:

[0055] Based on CT scans and reconstruction software, a 3D printed model of a scaffold structure with holes was designed according to the shape and anatomical structure of the bone defect.

[0056] The 3D printing model is imported into a 3D printer, and the scaffold structure is prepared using fused deposition modeling or direct-write 4D printing methods.

[0057] Thirdly, this invention provides an application of a 4D-printed bone tissue scaffold with an adjustable microenvironment as described in any of the preceding claims in the field of filling bone defects and inducing osteoogenesis.

[0058] The present invention will be further described below with reference to specific embodiments.

[0059] Example 1: The fabrication of a 4D-printed bone tissue scaffold with adjustable microenvironment includes the following steps:

[0060] 1. Prepare a 50wt% shape memory polylactic acid solution, then add 2wt% carbon nanotubes, 5wt% barium titanate, and 15wt% Fe3O4 nanoparticles (by mass of shape memory polylactic acid) to the solution, mix thoroughly to obtain solution A.

[0061] 2. Place the solution A obtained in step 1 in a petri dish and evaporate it completely to obtain a blocky solid, then crush it.

[0062] Third, use the pulverized material from step two to prepare filaments for 4D printing, and use 4D printing technology to prepare the support structure.

[0063] Fourth, take the pulverized material from step two and use photopolymerization printing technology to prepare a thin film with micropillars. Utilize the variable stiffness characteristics of the thin film to perfectly attach it to the surface of the scaffold structure to obtain a 4D printed bone tissue scaffold with an adjustable microenvironment.

[0064] While the present invention has been disclosed above, its scope of protection is not limited thereto. Those skilled in the art can make various changes and modifications without departing from the spirit and scope of the present invention, and all such changes and modifications will fall within the scope of protection of the present invention.

Claims

1. A 4D-printed bone tissue scaffold with adjustable microenvironment, characterized in that, It includes a support structure and a membrane covering the surface of the support structure, wherein the support structure has holes; The holes have a contact state where their opposite edges are in contact and a separation state where their opposite edges are not in contact. When the support structure is deformed under the stimulation of ultrasound, the holes are configured to switch between the contact state and the separation state, and the distance between the holes changes. This change in distance will generate a triboelectric effect, which in turn will generate a microcurrent. The membrane has micropillars on its surface away from the scaffold structure. These micropillars have both collapsed and upright states and are configured to switch between these states under ultrasonic stimulation. During repeated switching between the upright and collapsed states, the micropillars apply mechanical stimulation to the cells. The ultrasonic waves, acting as mechanical waves, provide additional mechanical stimulation at the bone defect site, causing the scaffold structure to deform under stress and generating piezoelectric and triboelectric effects. This induces current on the surface of the scaffold structure, thereby constructing a physical microenvironment conducive to cell growth and ultimately promoting the regeneration of damaged bone tissue. The scaffold structure comprises shape memory polymer, carbon nanotubes, barium titanate, and Fe3O4 nanoparticles. The membrane comprises shape memory polymer, carbon nanotubes, barium titanate, and Fe3O4 nanoparticles, wherein the mass of the carbon nanotubes is 2-5 wt% of the mass of the shape memory polymer, the mass of the barium titanate is 3-5 wt% of the mass of the shape memory polymer, and the mass of the Fe3O4 nanoparticles is 10-15 wt% of the mass of the shape memory polymer. The pore diameter is 200-500 micrometers, and the porosity of the pores in the support structure is 25-80%. The ultrasonic wave has a wavelength of 1-3MHz and a power of 30mW / cm. 2 -1000 mW / cm 2 Low-intensity pulsed ultrasound.

2. A method for preparing a 4D-printed bone tissue scaffold with adjustable microenvironment, characterized in that, The preparation of the 4D-printed bone tissue scaffold with adjustable microenvironment as described in claim 1 includes the following steps: Based on the bone defect, a structural model of the scaffold structure is designed, and the scaffold structure with holes is obtained by printing it using fused deposition modeling or direct-write 4D printing methods. The film is printed using a photopolymerization printing method; A film is attached to the surface of the support structure.

3. The method for preparing a 4D-printed bone tissue scaffold with adjustable microenvironment according to claim 2, characterized in that, The step of designing a structural model of the scaffold structure based on the bone defect, and printing it using fused deposition modeling or direct-write 4D printing methods to obtain the scaffold structure with holes includes: Based on CT scans and reconstruction software, a 3D printed model of the scaffold structure with holes is designed according to the shape and anatomical structure of the bone defect. The 3D printing model is imported into a 3D printer, and the scaffold structure is prepared using fused deposition modeling or direct-write 4D printing methods.

4. The application of a 4D-printed bone tissue scaffold with adjustable microenvironment as described in claim 1 in the field of filling bone defects to induce osteoogenesis.

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