CN108992212B

CN108992212B - Bone-cartilage integrated repair scaffold and preparation method thereof

Info

Publication number: CN108992212B
Application number: CN201810879249.2A
Authority: CN
Inventors: 张婧; 邓坤学; 袁玉宇
Original assignee: Medprin Regenerative Medical Technologies Co Ltd
Current assignee: Medprin Regenerative Medical Technologies Co Ltd
Priority date: 2018-08-03
Filing date: 2018-08-03
Publication date: 2020-11-24
Anticipated expiration: 2038-08-03
Also published as: CN108992212A

Abstract

The invention provides a bone-cartilage integrated repair scaffold and a preparation method thereof. The bone-cartilage integrated repair scaffold comprises: a bone repair layer; a cartilage repair layer; the bone repair layer is connected with the cartilage repair layer, wherein the bone repair layer comprises a biodegradable synthetic polymer compound and a powder component with osteogenic activity, and the cartilage repair layer comprises a biodegradable synthetic polymer compound and a powder component with chondrogenic activity. The bone-cartilage integrated repair scaffold can specifically stimulate defect parts aiming at cartilage and subchondral bone, and promote tissue regeneration. Furthermore, the cartilage repair layer of the bone-cartilage integrated repair bracket contains chondrocytes, so that the repair rate of cartilage parts can be increased, and the repair quality of cartilage can be improved.

Description

Bone-cartilage integrated repair scaffold and preparation method thereof

Technical Field

The invention relates to a bone-cartilage integrated repair scaffold and a preparation method thereof, belonging to the field of medical implant materials.

Background

Joint cartilage degradation or pathological changes caused by trauma, disease, or external loads that are often borne by long-term joint movement are clinically common conditions. The inside of the articular cartilage has no blood supply, no nerve and no immune response, and almost has no self-healing capability, so once suffering from diseases or injuries, the articular cartilage gradually deteriorates, finally leads to movement dysfunction and suffers from joint diseases, the life quality is greatly influenced, and even the artificial joint needs to be replaced for patients with the later-stage articular cartilage injury. Research shows that cartilage and subchondral bone are integrated, the former plays a load role, the latter plays a mechanical supporting and impact force absorbing role, and the two are interdependent and inseparable.

Cartilage repair has made great progress, but still has a great distance from functional repair. Cartilage repair techniques aim to reduce pain and try to repair the original function of the tissue, making it not only morphologically similar to natural transparent tissue, but also keeping its composition and mechanical properties close to those of natural tissue in a long-term repair effect.

The current clinical techniqueMainly comprises the following steps: cartilage planing, intra-articular cavity cleaning and lavage, micro-fracture, autologous/allogenic cartilage transplantation, and autologous/allogenic chondrocyte transplantation. Typically damage less than 2cm²The method mainly cleans joint fragments, is suitable for the elderly patients with low requirements, cannot recover the mobility as soon as possible after the operation, belongs to conservative treatment, but can relieve the pain as soon as possible after the operation and basically recovers the normal life. The basic principle of the micro-fracture operation is to drill a hole with the size of about 0.5 mm-1 mm on the surface of the articular cartilage, so that the mesenchymal stem cells in the marrow cavity can permeate into the defect part to promote the tissue repair. In a short period, the repair effect is good, and the tissue can recover normal mechanical properties, but long-term observation shows that the tissue can not completely form a similar transparent cartilage tissue and fibrosis can occur (cited document [1 ]]Reference [2 ]])。

When articular cartilage is damaged in a full-thickness way, even if stem cells in subchondral bone marrow migrate to a defect part, fibrocartilage-like repair can be generated, but cartilage tissue formed by the repair has far different morphological and functional characteristics from normal articular cartilage, in fact, the damaged joint accelerates the ossification of the joint, and the degradation rate of extracellular matrix is greater than the synthesis of new matrix, thereby causing arthritis. Therefore, cartilage regeneration should not be limited to cartilage repair, and attention should be paid to the effect of subchondral bone on cartilage repair in order to achieve functional repair.

At present, the traditional technology is adopted for treating, the repaired tissue does not have the long-term biological performance of the natural cartilage-like tissue, and joint degeneration can occur within a period of time. The traditional treatment techniques can only meet the needs of patients in a short term, but not only are the traditional treatment techniques expensive, but also inflammation, rejection reaction and the like occur, and the long-term treatment effect is still controversial. For defects exceeding 3cm³The size can be repaired by adopting autologous bone/cartilage transplantation, the defect part can be filled with fresh and healthy autologous tissues, and the tissue repair is facilitated, but the method is not always sufficient in donor and weak in interface combination to generate rejection reaction.

In recent years, research reports that the micro-fracture is combined with bioactive scaffold materials, stem cells are enriched through the biological materials, and the repair of hyaline cartilage tissues is promoted by combining the micro-fracture. Erggelet et al adopts a PGA and hyaluronic acid compounded stent to implant at the defect part, and combines with microfracture, so as to promote the penetration and migration of bone marrow stem cells, obviously enhance II-type collagen secretion, and finally achieve the aim of repairing the full-thickness cartilage defect in sheep (cited document [3 ]). However, the method is only the scaffold repair, has low overall repair rate, is only directed at the middle-stage cartilage defect, and does not relate to the cartilage and subchondral bone double regions.

Cited documents:

[1]Mithoefer K.,Mcadams T.,Williams R.J.,et al.Clinical Efficacy of the Microfracture Technique for Articular Cartilage Repair in the Knee An Evidence-Based Systematic Analysis[J].The American journal of sports medicine,2009,37(10):2053-63.

[2]Saris D.B.,Vanlauwe J.,Victor J.,et al.Treatment of symptomatic cartilage defects of the knee characterized chondrocyte implantation results in better clinical outcome at 36months in a randomized trial compared to microfracture[J].The American journal of sports medicine,2009,37(1suppl):10S-9S.

[3]Erggelet C.,Endres M.,Neumann K.,et al.Formation of cartilage repair tissue in articular cartilage defects pretreated with microfracture and covered with cell‐free polymer‐based implants[J].Journal of Orthopaedic Research,2009,27(10):1353-60.

disclosure of Invention

Problems to be solved by the invention

In view of the technical problems of the bone-cartilage integrated repair scaffold in the prior art, for example: the ability to resume activity as soon as possible; the similar transparent cartilage tissue can not be formed after long-term use; the technical problem of fibrosis is easy to occur; the biological performance of the natural cartilage-like tissue is not long-lasting, and joint degeneration can occur within a period of time; donor deficiency, interface binding weakness and rejection reaction. The invention firstly provides a bone-cartilage integrated repair support, which can simultaneously realize cartilage repair and subchondral bone repair in the full-layer defect of cartilage, can pointedly stimulate the defect part and promote tissue regeneration, is beneficial to tissue repair, does not generate rejection reaction, does not generate the problems of fibrosis and the like.

Furthermore, the invention also provides a preparation method of the bone-cartilage integrated repair scaffold, which has the advantages of easily obtained raw materials and simple preparation method.

Means for solving the problems

The invention provides a bone-cartilage integrated repair scaffold, which comprises:

a bone repair layer;

a cartilage repair layer;

the bone repair layer is connected with the cartilage repair layer, wherein,

the material of the bone repair layer comprises biodegradable synthetic macromolecular compound and powder component with osteogenic activity,

the cartilage repair layer is made of biodegradable synthetic high molecular compound and powder component with cartilage forming activity.

The bone-cartilage integrated repair scaffold comprises a biodegradable synthetic polymer compound, wherein the biodegradable synthetic polymer compound comprises one or a combination of more than two of polycaprolactone, polylactic acid, polyglycolic acid and polylactic acid-glycolic acid copolymer.

The bone-cartilage integrated repair scaffold comprises a powder component with osteogenic activity, wherein the powder component with osteogenic activity comprises one or more of an inorganic component with osteogenic activity, polymer microspheres with osteogenic activity and an animal-derived bone matrix.

The bone-cartilage integrated repair scaffold according to the present invention, wherein the polymer microspheres having osteogenic activity comprise growth factors promoting osteoblast proliferation; preferably, the content of the growth factor is 0.1 ng/g-50 ng/g based on the mass of the polymer microsphere with osteogenic activity.

The bone-cartilage integrated repair scaffold comprises a powder component with chondrogenic activity, wherein the powder component with chondrogenic activity comprises animal cartilage matrix and/or high-molecular microspheres with chondrogenic activity.

The integrated bone-cartilage repair scaffold according to the present invention, wherein the polymer microspheres having chondrogenic activity comprise growth factors that promote chondrocyte proliferation; preferably, the content of the growth factor is 0.1 ng/g-50 ng/g based on the mass of the polymer microsphere with chondrogenic activity.

The bone-cartilage integrated repair scaffold according to the present invention further comprises chondrocytes in the cartilage repair layer.

The bone-cartilage integrated repairing bracket comprises a bone repairing layer, a cartilage repairing layer and a fixing layer, wherein the bone repairing layer and the cartilage repairing layer comprise fiber bundles; preferably, the fiber bundle comprises one or more than two fiber filaments, and the average diameter of each fiber filament is between 50 and 1000 microns; preferably, the surface of the fiber filament has a concave structure.

The bone-cartilage integrated repair scaffold is characterized in that the powder component with osteogenic activity exists on the surface and/or inside the fiber filaments of the bone repair layer; the powder component with chondrogenic activity exists on the surface and/or inside the fiber silk of the cartilage repair layer;

preferably, the particle size of the powder component having osteogenic activity and the powder component having chondrogenic activity is 1nm to 500 μm.

The bone-cartilage integrated repair scaffold at least partially has a plurality of pore structures, and the average pore diameter of the pore structures is between 50 and 1000 microns; the porosity of the bone-cartilage integrated repair scaffold is between 50% and 95%; preferably, at least part of the pore structure is formed by overlapping the fiber bundles and penetrates through the bone repair layer and the cartilage repair layer; and/or, at least part of the pore structure is formed inside the fiber filament.

The bone-cartilage integrated repairing scaffold is characterized in that the compressive modulus of the bone repairing layer is 0.1-15 MPa; the compression modulus of the cartilage repair layer is 0.1 MPa-15 MPa.

The invention also provides a preparation method of the bone-cartilage integrated repairing scaffold, which comprises the step of preparing the bone-cartilage integrated repairing scaffold by using a 3D printing technology; the bone-cartilage integrated repair scaffold comprises:

a bone repair layer;

a cartilage repair layer;

the bone repair layer is connected with the cartilage repair layer, wherein,

the material of the bone repair layer comprises a biodegradable synthetic high molecular compound and a powder component with osteogenesis activity, and the material of the cartilage repair layer comprises a biodegradable synthetic high molecular compound and a powder component with chondrogenesis activity.

The preparation method of the bone-cartilage integrated repair scaffold comprises the following steps:

dissolving a biodegradable synthetic high molecular compound in a first solvent to serve as a matrix solution, wherein preferably, the content of the synthetic high molecular compound in the matrix solution is 0.05 g/mL-1 g/mL;

dividing the matrix solution into two parts, and respectively dispersing a powder component with osteogenic activity and a powder component with chondrogenic activity into the two parts of the matrix solution to obtain osteogenic component slurry and chondrogenic component slurry; preferably, the adding amount of the powder component with osteogenic activity in the osteogenic component slurry is 0.01 g/mL-1 g/mL, and the adding amount of the powder component with chondrogenic activity in the chondrogenic component slurry is 0.01 g/mL-1 g/mL;

and respectively printing the osteogenic component slurry and the chondrogenic component slurry by using a 3D printing technology to form a bone repair layer and a cartilage repair layer.

The production method according to the present invention, wherein the first solvent includes one or a combination of two or more of dioxane, acetic acid, hexafluoroisopropanol, and dimethyl sulfoxide.

The manufacturing method according to the present invention, wherein the 3D printing technique includes: respectively printing and extruding the osteogenic component slurry and the chondrogenic component slurry to a low-temperature printing receiving platform, and then solidifying and forming, wherein preferably, the temperature of the low-temperature printing receiving platform is-30 ℃ to 15 ℃.

The manufacturing method according to the present invention, wherein the 3D printing technique includes: respectively printing and extruding the osteogenic component slurry and the chondrogenic component slurry into a second solvent for precipitation molding, wherein

The second solvent is a poor solvent for synthesizing a high molecular compound, and the second solvent comprises water and/or ethanol.

The preparation method according to the present invention further comprises a post-treatment step of sterilizing and/or pouring hydrogel containing chondrocytes into the cartilage repair layer, wherein

The hydrogel containing chondrocytes is prepared by extracting chondrocytes, expanding the cells in the hydrogel so that the content of the chondrocytes is between 1,000,000 and 50,000,000 per mL,

preferably, the hydrogel is derived from at least one or a combination of two or more of gelatin, collagen, sodium alginate, agarose and fibrinogen.

ADVANTAGEOUS EFFECTS OF INVENTION

The bone-cartilage integrated repair bracket can pointedly stimulate the defect part and simultaneously give consideration to cartilage repair and subchondral bone repair; the degradable synthetic polymer material is selected to ensure that the bone repair layer and the cartilage repair layer have higher compression elastic modulus, so that the bone-cartilage integrated scaffold is not easy to deform after being implanted.

The bone-cartilage integrated repair scaffold has the pore diameter and porosity suitable for tissue adhesion growth, and the pore structure is favorable for promoting the release of active components of bone and cartilage in the bone-cartilage integrated repair scaffold and promoting the regeneration of tissues.

Furthermore, the cartilage repair layer of the bone-cartilage integrated repair bracket can be perfused with chondrocytes, so that the repair rate of cartilage parts can be increased, and the repair quality of cartilage can be improved.

Furthermore, the preparation method of the bone-cartilage integrated repair scaffold can keep the activity of each component in the bone-cartilage integrated repair scaffold, has higher safety and lower cost, is easy to form, and is suitable for industrial large-scale production.

Drawings

Fig. 1 is a perspective view illustrating a bone-cartilage integrated repair scaffold according to an embodiment of the present invention.

Fig. 2 shows an electron microscope image of the integrated bone-cartilage repair scaffold prepared according to one embodiment of the present invention.

Fig. 3 is a partially enlarged view of an electron microscope of the integrated bone-cartilage repair scaffold according to the embodiment of the present invention.

FIG. 4 is a graph showing immunohistochemical staining of cartilage extracellular matrix type II collagen at cartilage sites 1 month after the integrated bone-cartilage repair scaffold of example 1 was implanted subcutaneously in nude mice.

Fig. 5 is a graph showing aliskiren blue staining of acidic glycosaminoglycan at a cartilage site 1 month after the implantation of the integrated bone-cartilage repair scaffold of example 1 subcutaneously in nude mice.

Description of the reference numerals

1: a bone repair layer; 2: a cartilage repair layer; 3: a fiber bundle.

Detailed Description

Various exemplary embodiments, features and aspects of the invention will be described in detail below. The word "exemplary" is used exclusively herein to mean "serving as an example, embodiment, or illustration. Any embodiment described herein as "exemplary" is not necessarily to be construed as preferred or advantageous over other embodiments.

Furthermore, in the following detailed description, numerous specific details are set forth in order to provide a better understanding of the present invention. It will be understood by those skilled in the art that the present invention may be practiced without some of these specific details. In other instances, methods, means, devices and steps which are well known to those skilled in the art have not been described in detail so as not to obscure the invention.

First embodiment

In a first embodiment of the present invention, a bone-cartilage integrated repair scaffold is provided. The bone-cartilage integrated repair scaffold comprises: the bone repair layer 1 and the cartilage repair layer 2 are connected; in the present invention, the bone repair layer 1 and the cartilage repair layer 2 may be directly contacted, or may be connected through some other possible active layer.

In the present invention, the material of the bone repair layer 1 includes a biodegradable synthetic polymer compound and a powder component having osteogenic activity, and the material of the cartilage repair layer 2 includes a biodegradable synthetic polymer compound and a powder component having chondrogenic activity.

The length of the bone repair layer 1 (for example, the thickness of the bone repair layer 1) in the direction perpendicular to the bone repair layer 1 (the z direction of the bone repair layer 1) is 5mm to 15mm, for example, 7mm to 12 mm. Preferably, the compressive modulus of the bone repair layer 1 is in the range of 0.1 to 15MPa, may be in the range of 0.5 to 10MPa, and may be in the range of 1 to 5 MPa.

The length of the cartilage repair layer 2 (for example, the thickness of the cartilage repair layer 2) in the direction perpendicular to the cartilage repair layer 2 (the z direction of the cartilage repair layer 2) is 1mm to 5mm, for example, 2mm to 4 mm. Preferably, the cartilage repair layer 2 has a compressive modulus in the range of 0.1 to 15MPa, 0.5 to 10MPa, or 1 to 5 MPa.

The invention relates to a method for testing compression modulus, which adopts a universal mechanical testing machine. Placing a bone-cartilage integrated restoration bracket or a bone restoration layer or a cartilage restoration layer with a certain shape, for example, the upper surface and the lower surface of the bone-cartilage integrated restoration bracket are close to be parallel between two pressing plates of a universal mechanical testing machine, setting the compression rate of the pressing plates to be 0.5mm/min, and finishing compression after the bone-cartilage integrated restoration bracket is cracked to obtain a stress-strain curve. And selecting a linear region at the front section of the curve, and calculating to obtain the compression modulus of the stent.

Generally, cartilage and subchondral bone are a whole, the former plays a role in load, the latter plays a role in mechanical support and impact absorption, and the two are interdependent and inseparable. Cartilage regeneration should not be limited to cartilage repair, and to achieve functional repair, attention should be paid to the effects of subchondral bone on cartilage repair.

The material of the bone repair layer 1 of the bone-cartilage integrated repair bracket comprises a biodegradable synthetic polymer compound and a powder component with osteogenesis activity, and the material of the cartilage repair layer 2 comprises a biodegradable synthetic polymer compound and a powder component with chondrogenesis activity, so that the defect part can be stimulated in a targeted manner, and cartilage repair and subchondral bone repair are both considered; the bone repair layer 1 and the cartilage repair layer 2 both have higher compressive elastic modulus by selecting biodegradable synthetic polymer materials, so that the bone-cartilage integrated repair scaffold is not easy to deform after being implanted.

In addition, in the present invention, the bone-cartilage integrated repair scaffold may further include other layer structures, and the other layer structures included in the scaffold are not particularly limited, and may achieve the functions of the present invention. Specifically, the method comprises the following steps:

synthesis of Polymer Compound

The biodegradable synthetic high molecular compound has good biocompatibility, and the mechanical property and the degradation speed of the material can be adjusted and controlled by adjusting and increasing the molecular weight and selecting different polymerization modes and forming means. Preferably, the biodegradable synthetic polymer compound may include one or a combination of two or more of polycaprolactone, polylactic acid, polyglycolic acid, and polylactic acid-glycolic acid copolymer, and the like.

In the present invention, the biodegradable synthetic polymer compound used for the bone repair layer 1 and the biodegradable synthetic polymer compound used for the cartilage repair layer 2 may be the same or different.

Powder component with osteogenic activity

In the invention, the powder component with osteogenesis activity comprises one or more of inorganic matter component with osteogenesis activity, high molecular microspheres with osteogenesis activity and animal-derived bone matrix.

The inorganic substance component having osteogenic activity of the present invention may be various inorganic substances having osteogenic activity commonly used in the art. For example, it may be one or a combination of two or more of hydroxyapatite, tricalcium phosphate, tetracalcium phosphate, octacalcium phosphate, calcium oxide, calcium silicate, calcium sulfate, calcium carbonate, strontium phosphate, sodium phosphate, magnesium oxide, silicon oxide, zinc phosphate, zinc oxide, bioglass, and metal magnesium powder.

The polymer microsphere with osteogenic activity is biodegradable polymer microsphere containing growth factor for promoting bone cell proliferation. Among them, the growth factor that promotes the proliferation of the osteocyte can be various growth factors that promote the proliferation and differentiation of the osteocyte commonly used in the art. In the polymer microsphere with osteogenic activity, the adding amount of the growth factor for promoting the proliferation of the osteocyte is 0.1 ng/g-50 ng/g based on the mass of the polymer microsphere with osteogenic activity, for example: 0.5ng/g, 1ng/g, 10ng/g, 20ng/g, 30ng/g, 40ng/g, etc. In the present invention, the proliferation and differentiation of osteocytes can be rapidly promoted despite the use of such a low content of growth factors that promote the proliferation of osteocytes, ranging from 0.1ng/g to 50 ng/g.

Specifically, the growth factor that promotes the proliferation of bone cells according to the present invention may be one or a combination of two or more of the bone morphogenetic protein family (BMP), the fibroblast growth factor family (FGF), the insulin-like growth factor family (IGF), the platelet-derived growth factor (PDGF), the transforming growth factor- β family (TGF- β), and the like. In the polymer microsphere with chondrogenic activity, the degradable polymer microsphere matrix can be derived from one or more of gelatin, collagen, sodium alginate, chitosan, hyaluronic acid, polylactic acid, polycaprolactone, polyglycolic acid, polylactic acid-glycolic acid copolymer, polyurethane and the like. The degradable polymer microsphere matrix can be used as a carrier of growth factors for promoting osteogenesis.

In the present invention, the animal-derived bone matrix may be a demineralized bone matrix, a non-demineralized bone matrix, a defatted and deproteinized bone powder having no immunogenicity, or the like.

Preferably, the particle size of the powder component having osteogenic activity of the present invention is between 1nm and 500. mu.m, preferably between 50nm and 200. mu.m, and may be between 10nm and 400. mu.m, or may be between 100nm and 100. mu.m, etc.

Powder composition with chondrogenic activity

The powder component with chondrogenic activity comprises animal cartilage matrix and/or high molecular microspheres with chondrogenic activity.

The animal cartilage matrix of the present invention includes defatted deproteinized animal cartilage powder without immunogenicity.

The polymer microsphere with chondrogenic activity is biodegradable polymer microsphere containing growth factors for promoting chondrocyte proliferation. Among them, the growth factor that promotes the proliferation of chondrocytes may be various growth factors that are commonly used in the art to promote the proliferation and differentiation of chondrocytes. The content of the growth factor for promoting the proliferation of the chondrocytes is 0.1 ng/g-50 ng/g based on the mass of the polymer microsphere with chondrogenic activity, for example: 0.5ng/g, 1ng/g, 10ng/g, 20ng/g, 30ng/g, 40ng/g, etc. In the present invention, the proliferation and differentiation of chondrocytes can be rapidly promoted despite the use of such a low content of growth factors that promote the proliferation of chondrocytes, ranging from 0.1ng/g to 50 ng/g.

Specifically, the growth factor that promotes the proliferation of chondrocytes according to the present invention may be one or a combination of two or more of the bone morphogenetic protein family (BMP), transforming growth factor- β family (TGF- β), platelet-derived growth factor (PDGF), insulin-like growth factor family (IGF), Wingless family (Wnt), Hedgehog family, and the like. In the polymer microsphere with chondrogenic activity, the degradable polymer microsphere matrix can be derived from one or more of gelatin, collagen, sodium alginate, chitosan, hyaluronic acid, polylactic acid, polycaprolactone, polyglycolic acid, polylactic acid-glycolic acid copolymer, polyurethane and the like. The degradable polymer microsphere matrix can be used as a carrier of growth factors for promoting chondrogenesis.

Preferably, the particle size of the powder component having chondrogenic activity according to the invention is between 1nm and 500. mu.m, preferably between 50nm and 200. mu.m; may be 10nm to 400 μm, or may be 100nm to 100 μm.

Chondrocytes

The cartilage repair layer 2 of the present invention preferably further contains chondrocytes. The chondrocytes may be healthy hyaline chondrocytes extracted from an affected part of a patient, which are mixed into a sterile hydrogel and expanded in vitro in a hydrogel three-dimensional environment. And (3) pouring hydrogel containing a proper amount of hyaline chondrocytes into the cartilage repair layer 2 of the bone-cartilage integrated repair scaffold to obtain the cartilage repair layer 2 containing chondrocytes. Preferably, the hydrogel contains hyaline chondrocytes in an amount of 1,000,000 to 50,000,000 per mL. The cartilage cells of the invention can accelerate the repair rate of cartilage parts and improve the quality of cartilage repair.

Fiber bundle

In the invention, the bone repair layer 1 and the cartilage repair layer 2 contain fiber bundles 3. Preferably, the fiber bundle 3 comprises one or more than two fiber filaments, and the average diameter of each fiber filament is between 50 μm and 1000 μm, for example: may be 100 to 800 μm, may be 200 to 600 μm, or the like. More than two fiber filaments in the fiber bundle 3 can be tightly combined together. The material of the fiber yarn is derived from the biodegradable synthetic polymer compound.

As shown in fig. 2 and 3, it can be seen from the electron microscope images that the powder component having osteogenic activity exists on the surface and/or inside of the fiber filaments of the bone repair layer 1, and the powder component having chondrogenic activity exists on the surface and/or inside of the fiber filaments of the cartilage repair layer 2. Preferably, the powder component with osteogenic activity is uniformly distributed on the surface and/or inside of the fiber filaments of the bone repair layer 1, and the powder component with chondrogenic activity is uniformly distributed on the surface and/or inside of the fiber filaments of the cartilage repair layer 2. In addition, as shown in fig. 2 and 3, the surface of the fiber filament may further have a concave structure, and the concave structure may be formed by drying the bone-cartilage integrated repair scaffold to remove a solvent during the preparation process.

The fiber yarn has good biocompatibility, can better promote the repair of human subchondral bone, and does not produce adverse effect.

Pore structure

The bone-cartilage integrated repair scaffold at least partially has a plurality of pore structures, and the average pore diameter of the pore structures is between 50 and 1000 μm, such as: may be 100 to 800 μm, may be 200 to 600 μm, and the like, and is preferably 50 to 400 μm; the porosity of the bone-cartilage integrated repair scaffold is between 50% and 95%, for example: may be 51% to 95%, may be 60% to 85%, may be 70% to 80%, and the like.

Preferably, at least part of the pore structure is formed by overlapping the fiber bundles 3 and penetrates through the bone repair layer 1 and the cartilage repair layer 2. The pore structure may be formed by overlapping the fiber bundles 3 with each other during the preparation process. For example: when the bone-cartilage integrated repair scaffold is prepared by using a 3D printing process, the fiber yarns are formed in the printing process and are made into the fiber bundles 3, and the fiber bundles 3 are mutually overlapped in the printing process. The angle between the two sets of mutually overlapping fiber bundles 3 may be more than 0 ° to 90 °, for example, 10 ° to 80 °, 20 ° to 60 °, 30 ° to 50 °, and preferably 70 ° to 90 °.

Preferably, in the present invention, at least a part of the pore structure may also be present inside the fiber filaments. For example: may be formed by drying the bone-cartilage integrated repair scaffold to remove the solvent during the preparation process.

The porosity detection method of the invention can be according to the method commonly used in the prior art, and specifically can be as follows: the porosity of the sample to be tested is calculated according to the following formula,

in the formula: m is the sample mass (g);

v is the sample volume (cm)³)；

ρ s is the skeleton density (g/cm) of the sample material³) Or referred to as true density.

Second embodiment

The second embodiment of the present invention provides a method for preparing a bone-cartilage integrated repair scaffold, comprising the steps of preparing the bone-cartilage integrated repair scaffold by using a 3D printing technique; the integrated bone-cartilage repair scaffold is the integrated bone-cartilage repair scaffold in the first embodiment. The preparation method of the invention can keep the activity of each component in the bone-cartilage integrated repair scaffold, has higher safety and lower cost, is easy to form and is suitable for industrial large-scale production. Specifically, the preparation method of the present invention can be performed according to the following steps:

dissolving a biodegradable synthetic high molecular compound in a first solvent to be used as a matrix solution;

dividing the matrix solution into two parts, and respectively dispersing a powder component with osteogenic activity and a powder component with chondrogenic activity into the two parts of the matrix solution to obtain osteogenic component slurry and chondrogenic component slurry;

and respectively printing the osteogenic component slurry and the chondrogenic component slurry by using the 3D printing technology to form a bone repair layer 1 and a cartilage repair layer 2.

Generally, when printing, the bone component slurry can be printed firstly, and then the cartilage component slurry can be printed; or the paste can be printed into cartilage component paste firstly and then into bone component paste; it is also possible to print separately and to connect the bone repair layer 1 and the cartilage repair layer 2 by means of an optional active layer. The printing sequence is not particularly limited, and the prepared product can realize the functions of the invention.

The 3D printing technology belongs to one of the rapid forming technologies, and is a technology for constructing an object by using a bondable material such as powdered metal or plastic and the like in a layer-by-layer stacking and accumulating mode on the basis of a digital model file. For example, the printer may be instructed to print layer by modeling it with Computer Aided Design (CAD) or computer animation modeling software and "partitioning" the created three-dimensional model into layer-by-layer sections. Specifically, a printer (e.g., FDM 3D printer) reads cross-sectional information from a document, prints the cross-sectional information layer by layer using a liquid, powder, or sheet material, and bonds the layers in various ways to produce a solid body.

In the invention, the biodegradable synthetic high molecular compound also has good biocompatibility, and the mechanical property and the degradation speed of the material can be adjusted and controlled by adjusting and increasing the molecular weight and selecting different polymerization modes and forming means. Preferably, the biodegradable synthetic polymer compound may include one or a combination of two or more of polycaprolactone, polylactic acid, polyglycolic acid, and polylactic acid-glycolic acid copolymer.

In the invention, the first solvent can be a solvent with a higher freezing point, and the freezing point can be-10-20 ℃ generally. The first solvent is used as a good solvent of the biodegradable synthetic polymer compound, so that the biodegradable synthetic polymer compound can be well dissolved in the first solvent. For example: the first solvent of the present invention may be one or a combination of two or more of dioxane, acetic acid, hexafluoroisopropanol, dimethyl sulfoxide, and the like. In the present invention, the first solvent is not limited to the above-listed first solvents, and may be any of the first solvents in the prior art that can achieve the present invention. In the present invention, the content of the biodegradable synthetic high molecular compound in the first solvent is 0.05g/mL to 1g/mL, for example: may be 0.1 g/mL-0.8 g/mL or 0.3 g/mL-0.6 g/mL.

The powder component having osteogenic activity and the powder component having chondrogenic activity in the present embodiment are the powder component having osteogenic activity and the powder component having chondrogenic activity in the first embodiment. In the osteogenic component slurry, the addition amount of the powder component with osteogenic activity is 0.01 g/mL-1 g/mL, for example: may be 0.05 g/mL-0.8 g/mL or 0.1 g/mL-0.5 g/mL; the amount of the powder component having a chondrogenic activity to be added to the chondrogenic component slurry may be 0.01g/mL to 1g/mL, for example, 0.05g/mL to 0.8g/mL, or 0.1g/mL to 0.5 g/mL.

In a specific implementation manner of this embodiment, the 3D printing technology may include: and printing the osteogenic component slurry and the chondrogenic component slurry by using a low-temperature deposition technology. In the present invention, the low temperature deposition technique includes: and respectively printing and extruding the osteogenic component slurry and the chondrogenic component slurry to a printing receiving platform, and then solidifying and molding.

In the specific printing process, under the condition that the temperature of the printing receiving platform and the printing cavity is lower than the freezing point, the osteogenic component slurry and the chondrogenic component slurry are immediately solidified and molded after being extruded. Generally, the temperature of the print receiving platform and the print chamber can be between-30 ℃ and 15 ℃, and the freezing point of the first solvent is generally higher than the temperature of the print receiving platform and the print chamber. And further freeze-drying the molded bone-cartilage integrated repair scaffold, then soaking and cleaning the dried bone-cartilage integrated repair scaffold in ethanol, removing residual organic solvent and drying.

In another specific embodiment of this embodiment, the 3D printing technique includes: and printing the bone repair layer 1 and/or the cartilage repair layer 2 by adopting a process phase separation technology. In the present invention, the process phase separation technique comprises: respectively printing and extruding the osteogenic component slurry and the chondrogenic component slurry into a second solvent for precipitation molding, wherein the second solvent is a poor solvent of a biodegradable synthetic high molecular compound.

In a specific printing process, a biodegradable synthetic high molecular compound is dissolved in a first solvent, and a second solvent is contained in a printing receiving platform, wherein the first solvent and the second solvent can be mixed and dissolved. Because the second solvent is a poor solvent of the biodegradable synthetic high molecular compound, when the printing ink is extruded into the second solvent, the first solvent and the second solvent can be quickly mixed and dissolved, and the biodegradable synthetic high molecular compound in the first solvent is separated out in a phase separation manner, so that the bone-cartilage integrated repairing scaffold is formed. And further, the printed and molded bone-cartilage integrated repair scaffold can be continuously soaked in ethanol and/or water, and the residual organic solvent is removed and dried.

Preferably, the second solvent of the present invention comprises water and/or ethanol. In the present invention, the second solvent is not limited to the above two solvents, and may be any second solvent capable of implementing the present invention in the prior art.

In addition, when performing the process phase separation technique to print the osteogenic and chondrogenic component slurries, the first solvent may be any feasible solvent, such as: it may be dioxane, acetic acid, chloroform, hexafluoroisopropanol, dimethyl sulfoxide, etc.

Preferably, the preparation process further comprises a post-treatment step, such as: the prepared bone-cartilage integrated restoration scaffold can be further subjected to gamma-ray irradiation sterilization and then packaged to obtain a finished product bone-cartilage integrated restoration scaffold. The following steps are repeated: the hydrogel containing chondrocytes, which is prepared by extracting chondrocytes and expanding the cells in the hydrogel so that the content of the chondrocytes is between 1,000,000 and 50,000,000/mL, may be perfused into the cartilage repair layer 2 in the integrated bone-cartilage repair scaffold. Preferably, the hydrogel is derived from at least one or a combination of two or more of gelatin, collagen, sodium alginate, agarose and fibrinogen.

In addition, a strippable film can be arranged on one side of the bone-cartilage integrated repairing scaffold prepared by the invention, which is close to the bone repairing layer 1 and/or the cartilage repairing layer 2. The strippable film can be prepared from any material, and the performance of the bone-cartilage integrated repair scaffold is not changed.

Third embodiment

A third embodiment of the present invention provides an integrated bone-cartilage repair scaffold according to the first embodiment of the present invention and an integrated bone-cartilage repair scaffold obtained by the method for preparing an integrated bone-cartilage repair scaffold according to the second embodiment of the present invention, and applications of the integrated bone-cartilage repair scaffold in cartilage defect repair products. The cartilage defect repair article of the present invention may be directed to the repair of articular cartilage defects, and more preferably, to the repair of full thickness cartilage defects of the knee.

Generally, the quality of the repaired tissue is evaluated by the morphology of the new tissue, and the natural articular cartilage is hyaline cartilage and is composed of chondrocytes and ECM. Cells contained in the tissue of the native articular cartilage should be simple chondrocytes, and the secreted ECM should include type II collagen and glycosaminoglycan. The cartilage repair layer 2 of the existing repair scaffold mainly comprises fibrocartilage, which is different from natural articular cartilage. The invention considers both cartilage repair and subchondral bone repair in the full-layer defect of the cartilage, and the cartilage repair layer 2 contains active ingredients for promoting the proliferation of chondrocytes, so that the new tissue can be hyaline cartilage, and in addition, the cartilage repair layer 2 can also contain hyaline chondrocytes, thereby further accelerating the repair rate of cartilage parts and improving the quality of cartilage repair.

Examples

Embodiments of the present invention will be described in detail below with reference to examples, but those skilled in the art will appreciate that the following examples are only illustrative of the present invention and should not be construed as limiting the scope of the present invention. The examples, in which specific conditions are not specified, were conducted under conventional conditions or conditions recommended by the manufacturer. The reagents or instruments used are not indicated by the manufacturer, and are all conventional products commercially available.

Example 1

In the embodiment, polycaprolactone is selected as a biodegradable synthetic high molecular compound, dioxane is used as a first solvent, and a low-temperature deposition printing technology is adopted for printing so as to prepare the bone-cartilage integrated repair scaffold. The specific implementation steps are as follows:

(1) dissolving polycaprolactone into dioxane, wherein the content of the polycaprolactone is 1 g/mL. And dividing the dissolved solution into two parts to obtain a first matrix solution and a second matrix solution. Hydroxyapatite with the granularity of 10-100 mu m is selected as a powder component with osteogenic activity, and the powder component with osteogenic activity is uniformly dispersed into the first matrix solution according to the addition amount of 0.05g/mL to form osteogenic component slurry.

(2) Selecting gelatin microspheres containing transforming growth factor (TGF-beta 1) and having the particle size of 10-100 μm as a powder component with chondrogenic activity, wherein the content of the transforming growth factor (TGF-beta 1) is 0.5ng/g based on the mass of the gelatin microspheres with chondrogenic activity. The powder component with chondrogenic activity is uniformly dispersed in the second matrix solution according to the addition amount of 0.3g/mL to form chondrogenic component slurry.

(3) Establishing a cylindrical scaffold model by adopting modeling software, sequentially printing a bone repair layer and a cartilage repair layer at the temperature of-10 ℃, wherein the average diameter of fibers formed in the printing process is 200 mu m, and obtaining the bone-cartilage integrated repair scaffold I. The bone-cartilage integrated repairing scaffold I is subjected to freeze drying, and then the freeze-dried bone-cartilage integrated repairing scaffold I is soaked in ethanol for 3d to remove residual solvent, and then is subjected to air drying treatment. And finally, carrying out gamma-ray irradiation sterilization on the dried bone-cartilage integrated repair scaffold I.

(4) The hyaline chondrocytes were extracted and expanded in agarose gel. Then, the hydrogel containing the hyaline chondrocytes is poured into the cartilage repair layer of the integrated bone-cartilage repair scaffold I. In the hydrogel containing hyaline chondrocytes, the content of the hyaline chondrocytes is 5,000,000 cells/mL.

Example 2

In the embodiment, polylactic acid is used as a biodegradable synthetic polymer compound, dioxane is used as a first solvent, and a low-temperature deposition printing technology is adopted for printing to prepare the bone-cartilage integrated repair scaffold. The specific implementation steps are as follows:

(1) the polylactic acid is dissolved in dioxane, and the content of the polylactic acid is 0.5 g/mL. And dividing the dissolved solution into two parts to obtain a first matrix solution and a second matrix solution. Tricalcium phosphate with the granularity of 100 nm-10 mu m is selected as a powder component with osteogenic activity, and the powder component with osteogenic activity is uniformly dispersed into the first matrix solution according to the addition amount of 0.3g/mL to form osteogenic component slurry.

(2) Selecting sodium alginate microspheres containing platelet-derived growth factors (PDGF) and having the particle size of 100 nm-10 μm as powder components with chondrogenic activity, wherein the content of the platelet-derived growth factors (PDGF) is 10ng/g based on the mass of the sodium alginate microspheres with chondrogenic activity. The powder component with chondrogenic activity is uniformly dispersed in the second matrix solution according to the addition amount of 0.7g/mL to form chondrogenic component slurry.

(3) Establishing a cylindrical scaffold model by adopting modeling software, sequentially printing a bone repair layer and a cartilage repair layer at the temperature of-20 ℃, and obtaining a bone-cartilage integrated repair scaffold II, wherein the average diameter of fibers formed in the printing process is 250 mu m. And (3) freeze-drying the bone-cartilage integrated repair scaffold II, then soaking the freeze-dried bone-cartilage integrated repair scaffold II in absolute ethyl alcohol for 3d to remove residual solvent, and then carrying out air drying treatment. And finally, carrying out gamma-ray irradiation sterilization on the dried bone-cartilage integrated repair bracket II.

(4) The hyaline chondrocytes were extracted and expanded in agarose gel. Then, the hydrogel containing the hyaline chondrocytes is poured into the cartilage repair layer of the integrated bone-cartilage repair scaffold II. In the hydrogel containing hyaline chondrocytes, the content of the hyaline chondrocytes is 1,000,000 cells/mL.

Example 3

In the embodiment, the combination of polycaprolactone and polylactic acid-glycolic acid copolymer is used as a biodegradable synthetic high molecular compound, hexafluoroisopropanol is used as a first solvent, and the process split-phase printing technology is adopted for printing to prepare the bone-cartilage integrated repair scaffold. The specific implementation steps are as follows:

(1) dissolving polycaprolactone and polylactic acid-glycolic acid copolymer in hexafluoroisopropanol, wherein the total content of the polycaprolactone and the polylactic acid-glycolic acid copolymer is 0.7g/mL, and the mass ratio of the polycaprolactone to the polylactic acid-glycolic acid copolymer is 1: 1. And dividing the dissolved solution into two parts to obtain a first matrix solution and a second matrix solution. The non-immunogenic defatted and deproteinized bone powder with the granularity of 100 nm-500 nm is selected as a powder component with osteogenic activity, and the powder component with osteogenic activity is uniformly dispersed into a first matrix solution according to the addition amount of 0.5g/mL to form osteogenic component slurry.

(2) Selecting the non-immunogenic defatted deproteinized cartilage powder with the granularity of 100 nm-10 mu m as the powder component with chondrogenic activity, and uniformly dispersing the powder component with chondrogenic activity into a second matrix solution according to the addition amount of 0.5g/mL to form chondrogenic component slurry.

(3) And establishing a cylindrical bracket model by adopting modeling software, and sequentially printing a bone repair layer and a cartilage repair layer. And adding a second solvent absolute ethyl alcohol into a receiving groove of the printing receiving platform, extruding the printing ink into the absolute ethyl alcohol, rapidly dissolving hexafluoroisopropanol into the absolute ethyl alcohol, forming fibers with the average diameter of 300 mu m in the printing process, and performing phase separation and forming on polycaprolactone and polylactic acid-glycolic acid copolymer to obtain the bone-cartilage integrated repair scaffold III. Soaking the bone-cartilage integrated repair scaffold III in absolute ethyl alcohol for 3d to remove residual solvent, and then performing air drying treatment. And finally, carrying out gamma-ray irradiation sterilization on the air-dried bone-cartilage integrated repair bracket III.

(4) The hyaline chondrocytes were extracted and expanded in agarose gel. Then, the hydrogel containing the hyaline chondrocytes is poured into the cartilage repair layer of the integrated bone-cartilage repair scaffold III. The content of hyaline chondrocytes in the hydrogel containing hyaline chondrocytes was 20,000,000 cells/mL.

Example 4

In the embodiment, polycaprolactone is selected as a biodegradable synthetic high molecular compound, acetic acid is selected as a first solvent, and a process split-phase printing technology is adopted for printing so as to prepare the bone-cartilage integrated repair scaffold. The specific implementation steps are as follows:

(1) dissolving polycaprolactone in acetic acid, wherein the content of polycaprolactone is 0.5 g/mL. And dividing the dissolved solution into two parts to obtain a first matrix solution and a second matrix solution. Bioglass with the granularity of 100-200 mu m is selected as a powder component with osteogenic activity, and the powder component with osteogenic activity is uniformly dispersed into the first matrix solution according to the addition amount of 0.3g/mL to form osteogenic component slurry.

(2) Selecting the non-immunogenic defatted deproteinized cartilage powder with the granularity of 100 nm-10 mu m as a powder component with chondrogenic activity, and uniformly dispersing the powder component with chondrogenic activity into a second matrix solution according to the addition amount of 0.2g/mL to form chondrogenic component slurry.

(3) And establishing a cylindrical bracket model by adopting modeling software, and sequentially printing a bone repair layer and a cartilage repair layer. And adding a second solvent deionized water into the receiving groove of the printing receiving platform, extruding printing ink into the deionized water, rapidly dissolving acetic acid into the deionized water, forming fibers with the average diameter of 300 mu m in the printing process, and forming after phase separation of polycaprolactone to obtain the bone-cartilage integrated repair scaffold IV. And soaking the bone-cartilage integrated restoration support IV in deionized water for 3d to remove residual solvent, then carrying out air drying treatment, and finally carrying out gamma-ray irradiation sterilization on the air-dried bone-cartilage integrated restoration support IV.

(4) The hyaline chondrocytes were extracted and expanded in agarose gel. The hydrogel containing hyaline chondrocytes was then infused into the cartilage repair layer of the integrated bone-cartilage repair scaffold IV. The content of hyaline chondrocytes in the hyaline chondrocyte-containing hydrogel is 50,000,000 cells/mL.

Performance testing

The average pore size, bone repair layer thickness, cartilage repair layer thickness, porosity and compressive modulus test data for the integrated bone-cartilage repair scaffolds I-IV of examples 1-4 above are set forth in table 1 below.

TABLE 1

As can be seen from Table 1, the scaffolds I-IV for integrated bone-cartilage repair of examples 1-4 of the present application not only have porosity and pore size suitable for tissue adhesion growth, but also facilitate the release of the osteogenic active ingredient and/or the chondrogenic active ingredient in the scaffold into the interstitial space of the tissue to promote tissue repair; the compression modulus is large, the deformation resistance of the bone-cartilage integrated repair scaffold after being implanted into a human body is strong, and better support performance is provided before new autologous cartilage grows out.

Animal experiments

The integrated bone-cartilage repair scaffold of example 3 was implanted under the skin of nude mice, removed 1 month after the operation, and the cartilage site was sectioned and analyzed for immunohistochemical staining of cartilage matrix type II collagen and staining of acid glycosaminoglycan with aliskiren blue. As shown in particular in fig. 4 and 5.

As can be seen from FIGS. 4 and 5, the cells secreted a certain amount of the cartilage matrix type II collagen and acidic glycosaminoglycan after the cell-containing cartilage layer scaffold was implanted subcutaneously in nude mice for 1 month. Indicating that the scaffold has good chondrogenic activity.

Further, the bone-cartilage one-body repair scaffold of examples 1-2 and 4 of the present invention was implanted under the skin of nude mice, was taken out 1 month after the operation, and was identical to fig. 4 and 5 after sectioning the cell-containing site, and then subjecting the section to type II collagen immunohistochemical staining and acid glycosaminoglycan azulene staining.

The above description is only for the specific embodiments of the present invention, but the scope of the present invention is not limited thereto, and any person skilled in the art can easily conceive of the changes or substitutions within the technical scope of the present invention, and all the changes or substitutions should be covered within the scope of the present invention. Therefore, the protection scope of the present invention shall be subject to the protection scope of the appended claims.

Claims

1. A bone-cartilage integrated repair scaffold, comprising:

a bone repair layer;

a cartilage repair layer;

the bone repair layer is connected with the cartilage repair layer, wherein,

the material of the bone repair layer comprises a biodegradable synthetic high molecular compound and a powder component with osteogenesis activity;

the bone repair layer and the cartilage repair layer comprise fiber bundles, and the fiber bundles comprise one or more than two fiber yarns; the bone-cartilage integrated repair scaffold at least partially has a plurality of pore structures, at least part of the pore structures are formed by mutual overlapping of the fiber bundles and penetrate through the bone repair layer and the cartilage repair layer, and the fiber yarns are made of biodegradable synthetic high molecular compounds;

the powder component with osteogenic activity exists in the inner part of the fiber silk of the bone repair layer, or on the surface and the inner part; the powder component with chondrogenic activity is present inside, or on the surface and inside, the filaments of the cartilage repair layer.

2. The integrated bone-cartilage repair scaffold according to claim 1, wherein said biodegradable synthetic polymer compound comprises one or a combination of two or more of polycaprolactone, polylactic acid, polyglycolic acid and polylactic acid-glycolic acid copolymer.

3. The integrated bone-cartilage repair scaffold according to claim 1 or 2, wherein the powder component having osteogenic activity comprises one or a combination of two or more of an inorganic component having osteogenic activity, polymeric microspheres having osteogenic activity and an animal-derived bone matrix.

4. The integrated bone-cartilage repair scaffold according to claim 3, wherein said polymer microspheres with osteogenic activity comprise growth factors that promote osteoblast proliferation.

5. The integrated bone-cartilage repair scaffold according to claim 4, wherein the growth factor is contained in an amount of 0.1ng/g to 50ng/g based on the mass of the polymer microspheres having osteogenic activity.

6. The integrated bone-cartilage repair scaffold according to claim 1 or 2, wherein the powder component having chondrogenic activity comprises cartilage matrix of animal origin and/or polymeric microspheres having chondrogenic activity.

7. The integrated bone-cartilage repair scaffold according to claim 6, wherein said polymer microspheres with chondrogenic activity comprise growth factors that promote chondrocyte proliferation.

8. The integrated bone-cartilage repair scaffold according to claim 7, wherein the growth factor is contained in an amount of 0.1ng/g to 50ng/g based on the mass of the polymer microspheres having chondrogenic activity.

9. The integrated bone-cartilage repair scaffold according to claim 1 or 2, wherein the cartilage repair layer further comprises chondrocytes.

10. The integrated bone-cartilage repair scaffold according to claim 1 or 2, wherein the average diameter of the individual fiber filaments is between 50 μm and 1000 μm; the surface of the fiber filaments is provided with a concave structure.

11. The integrated bone-cartilage repair scaffold according to claim 1 or 2, wherein the particle size of the powder component having osteogenic activity and the powder component having chondrogenic activity is 1nm to 500 μm.

12. The integrated bone-cartilage repair scaffold according to claim 1 or 2, wherein the average pore diameter of the pore structure is between 50 μm and 1000 μm; the porosity of the bone-cartilage integrated repair scaffold is 50% -95%.

13. The integrated bone-cartilage repair scaffold according to claim 12, wherein at least part of the pore structure is formed inside the fiber filaments.

14. The integrated bone-cartilage repair scaffold according to claim 1, wherein the compressive modulus of the bone repair layer is 0.1MPa to 15 MPa; the compressive modulus of the cartilage repair layer is 0.1 MPa-15 MPa.

15. A preparation method of a bone-cartilage integrated repair scaffold is characterized by comprising the step of preparing the bone-cartilage integrated repair scaffold by using a 3D printing technology; the bone-cartilage integrated repair scaffold comprises:

a bone repair layer;

a cartilage repair layer;

the bone repair layer is connected with the cartilage repair layer, wherein,

the cartilage repair layer is made of biodegradable synthetic high molecular compound and powder components with chondrogenic activity;

16. The method for preparing the integrated bone-cartilage repair scaffold according to claim 15, comprising the steps of:

17. The method for preparing a scaffold for integrated bone-cartilage repair according to claim 16, wherein the content of the synthetic polymer compound in the matrix solution is 0.05g/mL to 1 g/mL.

18. The method for preparing the bone-cartilage integrated repair scaffold according to claim 16, wherein the adding amount of the powder component having osteogenic activity in the slurry of osteogenic components is 0.01g/mL to 1g/mL, and the adding amount of the powder component having chondrogenic activity in the slurry of osteogenic components is 0.01g/mL to 1 g/mL.

19. The method according to claim 16, wherein the first solvent comprises one or a combination of two or more of dioxane, acetic acid, hexafluoroisopropanol, and dimethyl sulfoxide.

20. The method for preparing according to any one of claims 16 to 19, wherein the 3D printing technique comprises: and respectively printing and extruding the osteogenic component slurry and the chondrogenic component slurry to a printing receiving platform, and then solidifying and molding.

21. The method of claim 20, wherein the print receiving platform has a temperature of-30 ℃ to 15 ℃.

22. The method for preparing according to any one of claims 16 to 19, wherein the 3D printing technique comprises: respectively printing and extruding the osteogenic component slurry and the chondrogenic component slurry into a second solvent for precipitation molding, wherein

23. The method for preparing according to any one of claims 16 to 19, further comprising a post-treatment step of sterilization, and/or perfusion of a hydrogel containing chondrocytes to the chondral repair layer, wherein

The preparation method of the hydrogel containing the chondrocytes comprises the steps of extracting the chondrocytes, and amplifying the cells in the hydrogel, so that the content of the chondrocytes is 1,000,000-50,000,000/mL.

24. The method according to claim 23, wherein the hydrogel is derived from at least one or a combination of two or more of gelatin, collagen, sodium alginate, agarose, and fibrinogen.