CN111317860A

CN111317860A - Film-coated biological ceramic artificial bone and preparation method thereof

Info

Publication number: CN111317860A
Application number: CN202010131299.XA
Authority: CN
Inventors: 曾庆丰; 魏静; 益明星
Original assignee: Xi'an Particle Cloud Biotechnology Co ltd
Current assignee: Xi'an Particle Cloud Biotechnology Co ltd
Priority date: 2020-02-28
Filing date: 2020-02-28
Publication date: 2020-06-23

Abstract

The invention provides a film-coated biological ceramic artificial bone and a preparation method thereof, wherein the film-coated biological ceramic artificial bone comprises a biological ceramic artificial bone and a film coated on the outer surface of the biological ceramic artificial bone; the raw material for preparing the coating comprises silk fibroin. The artificial bone is covered with the membrane, so that the cell planting efficiency and the adhesion capability of the artificial bone can be effectively improved, meanwhile, the bone nonunion phenomenon caused by periosteum defect is avoided, and the osteogenesis repair rate is effectively improved. On the other hand, the coating is prepared from silk fibroin, the silk fibroin has no toxic or side effect on human bodies and can be hydrolyzed and degraded by protease, the degradation product has no toxic or side effect on tissues, and the silk fibroin can be taken out without secondary operation after being used, so that the pain of patients is reduced.

Description

Film-coated biological ceramic artificial bone and preparation method thereof

Technical Field

The invention belongs to the technical field of medical treatment, and relates to a film-coated biological ceramic artificial bone and a preparation method thereof.

Background

At present, the treatment means of bone defects mainly comprise autologous bone transplantation, allogeneic or xenogeneic bone transplantation and fibular bone transplantation with vascular pedicles. However, the above methods all have certain limitations: the autologous bone transplantation is mainly suitable for defects below 5cm and has limited sources; allogeneic or xenogeneic bone transplantation is also difficult to widely accept due to rejection and infection factors; the fibula transplantation with vascular pedicle is suitable for treating large bone defects, but has higher requirements on the source, the operation of operators and the postoperative management.

The fracture healing integrates the synergistic effects of inflammatory cells, stem cells, osteoblasts, osteoclasts, growth factors and the like, in the healing process, the periosteum plays a vital role, the strength of the membrane is maintained by an outer fibrous layer in a double-layer structure, a highly vascularized inner forming layer provides rich mesenchymal progenitor cells and transforming growth factors, the complete coverage of the periosteum effectively promotes the regeneration of bones at the fracture ends, and when bone defects are caused by severe injury, the integrity of the periosteum is usually destroyed, so that fracture repair obstacles are caused, and bone nonunion is caused.

Generally, the repair and reconstruction after bone defect mainly regenerates bone through bone conduction and bone induction, and is a process that new bone tissues gradually crawl to replace absorbed necrotic and organized tissues and perform reconstruction and healing. However, the biological environment favorable for the formation of new bones is required to exist in the autologous bone transplantation, the allogeneic bone transplantation and the fibular transplantation repair with the vascular pedicle.

The bone regeneration membrane is guided to maintain good biological environment at the bone defect part through the barrier function of the bone regeneration membrane, and further the fibrous tissue is prevented from growing in. In addition, the bone regeneration membrane is guided to promote the adhesion and proliferation of osteogenic precursor cells through the support effect of the bone regeneration membrane, so that the fracture healing is accelerated, the bone regeneration membrane can be used for large-section bone defects, is beneficial to repairing irregular bone defects, is not limited by material supply and demand, and provides a new method for treating the bone defects.

The guided bone regeneration membrane can wrap the bone defect part, provides a relatively stable environment for bone formation, effectively prevents the embedding of fibrous soft tissue at the defect part, thereby guiding the bone regeneration at the bone defect part in the stable environment, has good biocompatibility and a nanofiber structure, can induce the adhesion and proliferation of mesenchymal stem cells and osteoprogenitor cells and the differentiation of the mesenchymal stem cells and osteoprogenitor cells, and can ensure the infiltration of cell nutrition and the growth of blood vessels.

At present, the materials used for the guided bone regeneration membrane which is widely used mainly include polytetrafluoroethylene, titanium alloy, polyacetal, and the like. The material has the characteristics of biological inertia, strong stability and no reaction with the body, but because the material can not be absorbed in the human body, the material needs to be taken out by a secondary operation, thereby increasing the pain of a patient and the treatment time. Therefore, the guided bone regeneration membrane which has the characteristics of good biocompatibility, tissue and cell affinity, degradability, capability of directly participating in tissue repair and the like is in urgent need of development.

Disclosure of Invention

Aiming at the problems in the prior art, the invention provides a coated biological ceramic artificial bone and a preparation method thereof, wherein the coating can maintain a good biological environment at a bone defect part, is degradable, can be absorbed by a human body and does not need to be taken out by a secondary operation.

The invention is realized by the following technical scheme:

a film-coated biological ceramic artificial bone comprises a biological ceramic artificial bone and a film coated on the outer surface of the biological ceramic artificial bone; the raw material for preparing the coating comprises silk fibroin.

Preferably, the raw material for preparing the coating film also comprises a biological macromolecular substance, and the biological macromolecular substance is chitosan, polylactic acid, gelatin, hyaluronic acid, polycaprolactone, polylactic acid-glycolic acid or collagen.

Further, the mass ratio of the silk fibroin to the biomacromolecule substance is 9: (3-4).

Preferably, the raw material for preparing the coating also comprises an anticoagulant.

Preferably, the thickness of the coating is 1.0mm to 1.6 mm.

Preferably, the bioceramic powder used for preparing the bioceramic artificial bone is one or more of hydroxyapatite, β -tricalcium phosphate, calcium carbonate, calcium silicate and calcium sulfate.

The preparation method of the film-coated biological ceramic artificial bone comprises the following steps:

step 1, preparing a biological ceramic artificial bone;

and 2, coating the raw materials for preparing the coating film on the outer surface of the bioceramic artificial bone by an electrostatic spinning method.

Preferably, in the step 1, the bioceramic artificial bone is prepared by adopting a silk-free 3D printing method.

Further, step 1 comprises:

step 1.1, uniformly mixing biological ceramic powder and a binder to obtain slurry, and defoaming to obtain uniform printing slurry; obtaining an artificial bone three-dimensional model according to CT, MRI or X-ray of a bone defect part of a patient;

and step 1.2, printing the printing slurry by using a 3D biological ceramic printer according to the three-dimensional model of the artificial bone to obtain the biological ceramic artificial bone, and performing freeze-drying treatment on the printed biological ceramic artificial bone.

Preferably, step 2 comprises:

step 2.1, dissolving the raw materials for preparing the coating film in a solvent, stirring and preparing into an electrostatic spinning mixed solution;

step 2.2, placing the biological ceramic artificial bone on a receiving device of an electrostatic spinning machine, and then carrying out electrostatic spinning film covering by adopting an electrostatic spinning mixed solution;

and 2.3, drying the biological ceramic artificial bone coated with the film.

Compared with the prior art, the invention has the following beneficial technical effects:

when the membrane-covered biological ceramic artificial bone is used for repairing bone defects, on one hand, the membrane has the function of guiding a bone regeneration membrane, can maintain a relatively stable biological environment at the bone defect, can effectively prevent fibrous soft tissue at the bone defect from being embedded, can ensure the sufficiency of time and space required by membrane-below osteogenesis, ensures that the growth of the bone is not interfered by epithelial connective tissue before the bone is generated, guides the bone regeneration at the bone defect in the stable environment, strengthens the bone healing, and effectively avoids the repair obstacle caused by the defect of the bone membrane; the bioceramic artificial bone has osteoinductivity and osteoconductivity, the cell planting efficiency and the adhesion capacity of the artificial bone can be effectively improved by covering the artificial bone with a film, meanwhile, the phenomenon of bone nonunion caused by periosteal defect is avoided, and the osteogenic repair rate is effectively improved. On the other hand, the coating is prepared from silk fibroin, the silk fibroin has no toxic action on a human body, the cell adhesion performance is good, the biocompatibility is excellent, more importantly, the silk fibroin can be hydrolyzed and degraded by protease, the degradation product has no toxic or side effect on tissues, the silk fibroin can be taken out without secondary operation after being used, the pain of a patient is reduced, and the silk fibroin film has remarkable beneficial effects. The degradation and absorption time of the material and the influence of the degradation products on osteogenesis are the key problems of the degradable material in treating bone defects, the optimal degradation time of the material should be in the range of 6 weeks to several months, and the degradation products should be non-toxic and not hinder new bone formation, and the conditions are met by the silk fibroin.

Furthermore, biological macromolecular substances are added in the raw materials of the coating, so that the coating is more favorable for forming the film. When chitosan is used, the product also has the biological functions of stopping bleeding, diminishing inflammation, sterilizing, promoting wound healing and regulating body's immunity.

Furthermore, the film can be loaded with drugs, and the film-coated biological ceramic artificial bone loaded with the drugs has excellent anti-inflammatory, analgesic and anticoagulant properties, and can effectively avoid complications such as infection, thrombus and the like in the bone repair process, especially the lower limb bone repair process.

According to the invention, the biological ceramic artificial bone is coated with the coating by adopting an electrostatic spinning technology, and the nano fibers in the coating have the characteristics of high specific surface area, good dissolution rate, small size and high porosity, so that the adhesion, migration and proliferation of cells are facilitated, the medicine can be rapidly infiltrated when the medicine is loaded, and meanwhile, the recrystallization of the medicine is limited due to the rapid volatilization of the solvent.

Furthermore, the 3D printing technology is adopted to regulate and control the aperture, porosity and geometric appearance of the artificial bone, and the individualized and accurate treatment of the bone trauma patient is really realized.

Drawings

FIG. 1 is a flow chart of the preparation of an electrospun coated bioceramic artificial bone according to example 1;

FIG. 2 is a flow chart of the preparation of the bio-ceramic artificial bone coated by electrostatic spinning in example 2;

figure 3 is a schematic representation of silk fibroin;

fig. 4 is an SEM image (5000 ×) of the silk fibroin/chitosan electrospun fiber prepared in example 1.

Detailed Description

The present invention will now be described in further detail with reference to specific examples, which are intended to be illustrative, but not limiting, of the invention.

The film-coated biological ceramic artificial bone comprises a biological ceramic artificial bone and a film coated on the outer surface of the biological ceramic artificial bone.

The biological ceramic artificial bone is prepared from biological ceramic powder and a binder, wherein the biological ceramic powder is one or a mixture of more of Hydroxyapatite (HA), β -tricalcium phosphate (β -TCP), calcium carbonate, calcium silicate and calcium sulfate, the hydroxyapatite and the β -tricalcium phosphate are used as bioactive ceramics, are similar to minerals in natural bone matrix in structure and composition, are good bionic materials, have good biocompatibility, bioactivity, bone inductivity and bone conductivity, and are widely applied to bone repair materials.

The binder is preferably polyvinyl alcohol or collagen. Polyvinyl alcohol belongs to water-soluble polymer, and can be dissolved in water together with silk fibroin to form a mixed system.

The preparation raw material of the coating comprises silk fibroin, and also can comprise biological macromolecular substances, wherein the biological macromolecular substances are chitosan, polylactic acid, gelatin, hyaluronic acid, polycaprolactone, polylactic acid-glycolic acid or collagen, and other medicines, such as aspirin, warfarin, dipyridamole and other anticoagulant medicines can be added.

The silk fibroin is a natural high molecular protein which is originated from silk and has high purity, easy purification and separation, no toxic and harmful effects on human bodies, good cell adhesion performance, excellent biocompatibility, capability of being hydrolyzed and degraded by protease and no toxic or side effect on tissues of degradation products.

The chitosan is obtained by deacetylating chitin widely existing in nature, and has a chemical name of polyglucosamine (1-4) -2-amino-B-D glucose. The chitosan is non-toxic, non-irritant and good in biocompatibility, has a structure similar to that of an extracellular matrix of a human body, and can be degraded by lysozyme in a living body to generate natural metabolites so as to be completely absorbed. Meanwhile, chitosan also has the biological functions of stopping bleeding, diminishing inflammation, sterilizing, promoting wound healing and regulating the immunity of organisms.

In the postoperative recovery period of a bone defect patient, the blood circulation of limbs becomes slow due to limb braking, particularly lower limb braking, so that the venous blood flow rate becomes slow, thrombus is easily formed, and the life is threatened. Generally, patients need to take anticoagulant drugs such as aspirin enteric-coated tablets, warfarin, dipyridamole and the like orally after operation.

Aspirin has strong antipyretic and analgesic effects, is applied to rheumatism, toothache, arthralgia, cold, fever and the like, and is the most widely applied antipyretic analgesic and anti-inflammatory drug. The mechanism of action of aspirin anticoagulation is that it prevents thromboxane A2(TXA2) from generating by preventing platelet epoxidase from converting arachidonic acid into intermediate of prostaglandin (TXA2 can promote platelet aggregation), and further prevents platelet aggregation, so that it is not easy to release blood coagulation factors, and has anticoagulation effect.

The membrane-covered biological ceramic artificial bone has the double functions of drug carrier and bone defect repair, and can effectively avoid infection and thrombosis complications while repairing bone injury.

The preparation method of the film-coated biological ceramic artificial bone combines a filament-free 3D printing method and an electrostatic spinning method.

The preparation method comprises the following steps:

(1) preparing printing slurry, namely uniformly mixing biological ceramic powder and a binder to obtain slurry, defoaming to obtain uniform printing slurry, and then filling the printing slurry into a charging barrel, wherein the biological ceramic powder comprises hydroxyapatite and β -tricalcium phosphate;

(2) artificial bone three-dimensional model: the method comprises the steps of obtaining CT or MRI or X-ray of a bone defect part of a patient, processing data by using three-dimensional reconstruction software, obtaining a host bone prototype model, and designing an artificial bone three-dimensional model STL file suitable for a bone defect anatomical structure by using three-dimensional modeling software.

(3) Silkless 3D printing: the artificial bone is printed by a 3D biological ceramic printer (Xian Point cloud Biotechnology Co., Ltd.). Fixing the charging barrel filled with printing slurry in the step (1) on a 3D biological ceramic printer, loading the STL file of the artificial bone three-dimensional model designed in the step (2) into point cloud printing software (PCLAb), setting printing parameters, and carrying out silk-free 3D printing to obtain the biological ceramic artificial bone.

(4) Freeze-drying: and freeze-drying the printed bioceramic artificial bone by adopting a freeze-drying technology.

(5) Preparing silk fibroin: firstly, silk is processed by 0.1mol/L Na₂CO₃Boiling the solution, and placing in 75 deg.C CaCl₂/H₂O/C₂H₅Dissolving the OH ternary solvent (the molar ratio is 1:8:2) to prepare a fibroin-calcium chloride solution, filtering the fibroin-calcium chloride solution, putting the filtered fibroin-calcium chloride solution into a dialysis bag with the cut-off molecular weight of 12000 for dialysis, and finally freeze-drying the dialyzed solution to obtain the silk fibroin.

(6) Preparing an electrostatic spinning solution: dissolving silk fibroin or a mixture of silk fibroin and biomacromolecule substances and/or other medicines in a solvent, and magnetically stirring at normal temperature to prepare a uniform electrostatic spinning mixed solution.

(7) Film covering: and (3) placing the freeze-dried bioceramic artificial bone on a receiving device of an electrostatic spinning machine, and then carrying out electrostatic spinning and film coating.

(8) Drying, packaging and sterilizing: and drying, packaging and sterilizing the coated bioceramic artificial bone to obtain a finished product.

The electrostatic spinning film-coated biological ceramic artificial bone can be prepared by combining the two methods. The film-coated biological ceramic artificial bone not only has good bone conduction, bone induction and biocompatibility, but also has the effects of diminishing inflammation, easing pain, resisting blood coagulation and promoting wound healing.

Example 1: see FIG. 1

(1) Preparing an adhesive: preparing 8g of polyvinyl alcohol and 92g of water for injection into a polyvinyl alcohol aqueous solution with the mass fraction of 8%, placing the polyvinyl alcohol aqueous solution in a solvent bottle with a cover, heating and swelling the polyvinyl alcohol aqueous solution in a water bath at 60 ℃ for 4 hours, and then stirring the polyvinyl alcohol aqueous solution in a magnetic stirrer at 95 ℃ at the rotating speed of 160r/min for 2 hours to completely dissolve the polyvinyl alcohol aqueous solution to form a uniform solution;

(2) preparing printing slurry, namely mixing hydroxyapatite and β -tricalcium phosphate in a mass ratio of 3:2 to form biological ceramic powder, then mixing the biological ceramic powder and an adhesive in a mass ratio of 1:1.05, placing the mixture in a homogenizer at a rotating speed of 1800r/min for mixing for 8min to form uniform printing slurry, and then filling the printing slurry into a charging barrel for later use.

(3) Artificial bone three-dimensional model: acquiring CT or MRI or X-ray of a bone defect part of a patient, processing data by using three-dimensional reconstruction software, acquiring a host bone prototype model, and designing an artificial bone three-dimensional model STL file suitable for a bone defect anatomical structure by using three-dimensional modeling software.

(4) Printing: the artificial bone is printed by a 3D biological ceramic printer (Xian Point cloud Biotechnology Co., Ltd.). Fixing the charging barrel filled with printing slurry in the step (2) on a 3D biological ceramic printer, loading the STL file of the artificial bone three-dimensional model designed in the step (3) into point cloud printing software (PCLAb), and setting printing process parameters as follows: the printing speed is 10mm/s, the printing layer thickness is 150 micrometers, the average pore diameter is 450 micrometers, the printing slurry is uniformly extruded at a constant speed through a spiral propeller, the workbench performs synthetic motion along the x-y axis, the printing head moves along the z axis, and the printing is sequentially performed layer by layer, so that the printing of the bioceramic artificial bone is finally completed.

(5) And (3) freeze drying: placing the bioceramic artificial bone formed in the step (4) in a freeze drying box for freeze drying treatment for 24 hours;

(6) preparing silk fibroin:

a) placing silk in 0.1mol/L Na₂CO₃Boiling the solution for 2h, then washing with deionized water for three times, and airing at room temperature in a ventilated place to obtain the refined silk.

b) Placing the obtained refined silk in 75 deg.C CaCl at bath ratio of 1:18₂/H₂O/C₂H₅Dissolving the OH ternary solvent (the molar ratio is 1:8:2) for 1.5h to obtain a fibroin-calcium chloride solution. Then the fibroin-calcium chloride solution is filtered and put into a dialysis bag with cut-off molecular weight of 12000 for dialysis for 3d, and deionized water is changed every two hours.

c) Freeze drying the dialyzed solution in a freeze dryer for 2d to obtain spongy silk fibroin as shown in figure 3.

(7) Preparing an electrostatic spinning solution: dissolving 9g of silk fibroin and 4g of chitosan in 50g of formic acid solution, and magnetically stirring for 3 hours at normal temperature to prepare a uniform electrostatic spinning mixed solution.

(8) Electrostatic spinning and film covering: injecting the prepared electrostatic spinning mixed solution into a 20mL syringe, fixing the syringe on a micro-injection pump, adjusting the distance between a needle (the caliber of the needle is 0.7mm) and a receiving aluminum foil to be 18cm, connecting an anode with the needle, connecting a cathode with the receiving aluminum foil, and fixing the biological ceramic artificial bone subjected to freeze-drying treatment on the receiving aluminum foil. Setting the flow rate of the micro-injection pump to be 0.5mL/h, adjusting the voltage of a high-voltage direct-current power supply to be 22kv, forming the nano-fibers by the electrostatic spinning solution under the action of a high electric field, and spinning for 6 h. When spinning is carried out for 3 hours, the upper position and the lower position of the artificial bone of the biological ceramics are turned, so that the surface of the artificial bone of the biological ceramics can be evenly coated with an electrostatic spinning film of 1.0mm to 1.6 mm.

(9) Drying and sterilizing: and (3) drying the biological ceramic artificial bone coated with the electrostatic spinning film in a vacuum oven at 30 ℃ for 24h, packaging and carrying out irradiation sterilization treatment to obtain a finished product.

Example 2: see FIG. 2

(1) Preparing an adhesive: preparing 10g of polyvinyl alcohol and 90g of water for injection into a polyvinyl alcohol aqueous solution with the mass fraction of 10%, placing the polyvinyl alcohol aqueous solution in a solvent bottle with a cover, heating and swelling the polyvinyl alcohol aqueous solution in a water bath at 60 ℃ for 4 hours, and then stirring the polyvinyl alcohol aqueous solution in a magnetic stirrer at 95 ℃ at the rotating speed of 160r/min for 2 hours to completely dissolve the polyvinyl alcohol aqueous solution to form a uniform solution;

(2) preparing printing paste, namely mixing β -tricalcium phosphate and an adhesive in a mass ratio of 1:0.8, placing the mixture in a homogenizer at a rotating speed of 1800r/min for mixing for 8min to form uniform printing paste, and then filling the printing paste into a charging barrel.

(4) Printing: the artificial bone is printed by a 3D biological ceramic printer (Xian Point cloud Biotechnology Co., Ltd.). Fixing the charging barrel filled with printing slurry in the step (2) on a 3D biological ceramic printer, loading the STL file of the artificial bone three-dimensional model designed in the step (3) into point cloud printing software (PCLAb), and setting printing process parameters as follows: the printing speed is 10mm/s, the printing layer thickness is 150 micrometers, the average pore diameter is 450 micrometers, the slurry is uniformly extruded at a constant speed through a spiral propeller, the workbench performs synthetic motion along the x-y axis, the printing head moves along the z axis, and the printing is sequentially performed layer by layer, so that the printing of the bioceramic artificial bone is finally completed.

(6) preparing silk fibroin:

(7) Preparing an electrostatic spinning solution: dissolving 9g of silk fibroin, 3g of polylactic acid and 1g of aspirin in 30g of mixed solution of trifluoroacetic acid and dichloromethane (volume ratio of 7:3), and magnetically stirring for 3 hours at normal temperature to prepare uniform electrostatic spinning mixed solution.

(8) Electrostatic spinning and film covering: injecting the prepared electrostatic spinning mixed solution into a 20mL syringe, fixing the syringe on a micro-injection pump, adjusting the distance between a needle (the caliber of the needle is 0.7mm) and a receiving aluminum foil to be 18cm, connecting an anode with the needle, connecting a cathode with the receiving aluminum foil, and fixing the biological ceramic artificial bone subjected to freeze-drying treatment on the receiving aluminum foil. Setting the flow rate of the micro-injection pump to be 0.5mL/h, adjusting the voltage of a high-voltage direct-current power supply to be 22kv, forming the nano-fibers by the spinning solution under the action of a high electric field, and spinning for 6 h. When spinning is carried out for 3 hours, the upper position and the lower position of the artificial bone of the biological ceramics are turned, so that the surface of the artificial bone of the biological ceramics can be evenly coated with an electrostatic spinning film of 1.0mm to 1.6 mm.

Example 3

(1) Preparing an adhesive: dissolving 3g of collagen in 47g of 0.05mol/L acetic acid solution, placing on a magnetic stirrer, and stirring for 90min at the rotating speed of 160r/min to obtain a binder;

(2) preparing printing slurry, namely mixing β -tricalcium phosphate, calcium silicate and calcium carbonate in a mass ratio of 2:1:1 to form biological ceramic powder, then mixing the biological ceramic powder and an adhesive in a mass ratio of 1:1.2, placing the mixture in a homogenizer at a rotating speed of 1800r/min for mixing for 8min to form uniform printing slurry, and then filling the printing slurry into a charging barrel.

(5) And (3) freeze drying: placing the formed biological ceramic artificial bone in the step (4) in a freeze drying box for freeze drying treatment for 24 hours;

(6) preparing silk fibroin:

(7) Preparing an electrostatic spinning solution: 6g of polylactic glycolic acid, 3g of silk fibroin and 3g of collagen were dissolved in 57g of hexafluoroisopropanol solution, and the solution was magnetically stirred at room temperature for 3 hours to prepare a uniform electrospinning mixed solution.

Example 4

(2) preparing printing slurry, namely mixing calcium sulfate and β -tricalcium phosphate in a mass ratio of 1:2 to form biological ceramic powder, mixing the biological ceramic powder and an adhesive in a mass ratio of 1:1.05, placing the mixture in a homogenizer at a rotating speed of 1800r/min for mixing for 8min to form uniform printing slurry, and then filling the printing slurry into a charging barrel.

(4) Printing: the artificial bone is printed by a 3D biological ceramic printer (Xian Point cloud Biotechnology Co., Ltd.). Firstly, fixing the charging barrel (2) filled with printing slurry on a 3D biological ceramic printer, then loading the STL file of the artificial bone three-dimensional model (4) into point cloud printing software (PCLAb), and setting the printing process parameters as follows: the printing speed is 10mm/s, the printing layer thickness is 150 micrometers, the average pore diameter is 450 micrometers, the slurry is uniformly extruded at a constant speed through a spiral propeller, the workbench performs synthetic motion along the x-y axis, the printing head moves along the z axis, and the printing is sequentially performed layer by layer, so that the printing of the bioceramic artificial bone is finally completed.

(6) preparing silk fibroin:

(7) Preparing an electrostatic spinning solution: dissolving 9g of silk fibroin and 3g of polylactic acid in a mixed solution of trifluoroacetic acid and dichloromethane (volume ratio of 7:3), and magnetically stirring for 3 hours at normal temperature to prepare a uniform electrostatic spinning mixed solution.

Example 5

(2) preparing printing slurry, namely mixing calcium silicate and β -tricalcium phosphate in a mass ratio of 1:2 to form biological ceramic powder, then mixing the biological ceramic powder and an adhesive in a mass ratio of 1:1.05, placing the mixture in a homogenizer at a rotating speed of 1800r/min for mixing for 8min to form uniform printing slurry, and then filling the printing slurry into a charging barrel.

(6) preparing an electrostatic spinning solution: 5g of polycaprolactone and 6g of gelatin are dissolved in 48g of hexafluoroisopropanol, 2.6g of glacial acetic acid is added at the same time, and then the mixture is magnetically stirred for 3 hours at normal temperature to prepare a uniform electrostatic spinning mixed solution.

(7) Electrostatic spinning and film covering: injecting the prepared electrostatic spinning mixed solution into a 20mL syringe, fixing the syringe on a micro-injection pump, adjusting the distance between a needle (the caliber of the needle is 0.7mm) and a receiving aluminum foil to be 18cm, connecting an anode with the needle, connecting a cathode with the receiving aluminum foil, and fixing the biological ceramic artificial bone subjected to freeze-drying treatment on the receiving aluminum foil. Setting the flow rate of the micro-injection pump to be 0.5mL/h, adjusting the voltage of a high-voltage direct-current power supply to be 22kv, forming the nano-fibers by the spinning solution under the action of a high electric field, and spinning for 6 h. When spinning is carried out for 3 hours, the upper position and the lower position of the artificial bone of the biological ceramics are turned, so that the surface of the artificial bone of the biological ceramics can be evenly coated with an electrostatic spinning film of 1.0mm to 1.6 mm.

(8) Drying and sterilizing: and (3) drying the biological ceramic artificial bone coated with the electrostatic spinning film in a vacuum oven at 30 ℃ for 24h, packaging and carrying out irradiation sterilization treatment to obtain a finished product.

Example 6

(6) preparing an electrostatic spinning solution: 7g of hyaluronic acid and 4g of chitosan were dissolved in 50g of formic acid solution. Then stirring for 3h under magnetic force at normal temperature to prepare uniform electrostatic spinning mixed solution.

Fig. 4 is an SEM image of the silk fibroin/chitosan electrospun fiber prepared in example 1, and it can be seen that the silk fibroin/chitosan can be used to prepare nano-sized fibers, and the prepared nano-sized fibers have uniform thickness, continuous fibers, and smooth surfaces.

The silk fibroin/chitosan nanofiber membrane prepared in example 1 had a hemolysis rate of 3.6%, and had a certain anticoagulation property.

Example 2 the hemolysis rate of the polylactic acid/silk fibroin nanofiber membrane added with the anti-thrombotic powder aspirin is 0.6%, and the higher the mass fraction of aspirin is, the lower the hemolysis rate is, which shows that the addition of aspirin can effectively improve the hemolysis performance of the polylactic acid/silk fibroin nanofiber membrane.

Experiments prove that the planting efficiency of the rabbit bone marrow stromal stem cells of the traditional bioceramic artificial bone (namely, the uncoated bioceramic artificial bone) is 79.2%, and the planting efficiency of the rabbit bone marrow stromal stem cells of the coated bioceramic artificial bone in the example 1 can be improved to 98.7%. Compared with the traditional bioceramic artificial bone, the coated bioceramic artificial bone has the advantages that the planting efficiency of rabbit bone marrow matrix stem cells is obviously improved, and the cell adhesion capability is also obviously enhanced, so that the coated bioceramic artificial bone product has important significance in improving the repair rate of human bone defects.

Example 2A coated bioceramic artificial bone containing aspirin is prepared by embedding aspirin powder in fiber, gradually releasing degraded aspirin powder along with the swelling of fiber, and implanting the bone into human body for 72h with cumulative release rate of above 10%, and has high clinical application value in relieving pain, diminishing inflammation and resisting blood coagulation.

Claims

1. A film-coated biological ceramic artificial bone is characterized by comprising a biological ceramic artificial bone and a film coated on the outer surface of the biological ceramic artificial bone; the raw material for preparing the coating comprises silk fibroin.

2. The coated bioceramic artificial bone according to claim 1, wherein the raw material for preparing the coating further comprises a biomacromolecule substance, and the biomacromolecule substance is chitosan, polylactic acid, gelatin, hyaluronic acid, polycaprolactone, polylactic acid-glycolic acid or collagen.

3. The coated bioceramic artificial bone according to claim 2, wherein the mass ratio of silk fibroin to biomacromolecule substance is 9: (3-4).

4. The coated bioceramic artificial bone according to claim 1, wherein the raw material for preparing the coating further comprises an anticoagulant drug.

5. The coated bioceramic artificial bone according to claim 1, wherein the coating has a thickness of 1.0-1.6 mm.

6. The coated bioceramic artificial bone according to claim 1, wherein the bioceramic powder used for preparing the bioceramic artificial bone is a mixture of one or more of hydroxyapatite, β -tricalcium phosphate, calcium carbonate, calcium silicate and calcium sulfate.

7. The method for preparing the film-coated bioceramic artificial bone according to any one of claims 1-6, comprising the following steps:

step 1, preparing a biological ceramic artificial bone;

8. The method for preparing the membrane-coated bioceramic artificial bone according to claim 7, wherein in the step 1, the bioceramic artificial bone is prepared by adopting a silk-free 3D printing method.

9. The method for preparing the film-coated bioceramic artificial bone according to claim 8, wherein the step 1 comprises:

10. The method for preparing the film-coated bioceramic artificial bone according to claim 7, wherein the step 2 comprises:

and 2.3, drying the biological ceramic artificial bone coated with the film.