CN108310458B - Method for preparing tissue engineering scaffold material by adopting degradable synthetic polymer material - Google Patents

Method for preparing tissue engineering scaffold material by adopting degradable synthetic polymer material Download PDF

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CN108310458B
CN108310458B CN201810234110.2A CN201810234110A CN108310458B CN 108310458 B CN108310458 B CN 108310458B CN 201810234110 A CN201810234110 A CN 201810234110A CN 108310458 B CN108310458 B CN 108310458B
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奚桢浩
张嘉鹏
岑莲
赵玲
陆尔奕
崔立宇
杨盛兵
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Suzhou Mairui regenerative medicine Technology Co.,Ltd.
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    • A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
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Abstract

The invention discloses a method for preparing a tissue engineering scaffold material by adopting a degradable synthetic polymer material, which comprises the following steps: (1) forming a high molecular polymer sample by a forming technology, and then placing the high molecular polymer sample in a high-pressure container; (2) displacing gas within the high pressure vessel with a supercritical fluid; (3) heating and pressurizing the high-pressure container to saturate the high-molecular polymer sample in a high-temperature high-pressure supercritical fluid atmosphere, so that the high-molecular polymer sample is melted under the condition that the temperature is higher than the critical temperature or critical pressure of the supercritical fluid, and polymer crystallization is eliminated; (4) then cooling to the foaming temperature; (5) and (5) rapidly relieving pressure. The invention melts the crystal region of the high molecular polymer through high temperature, eliminates the polymer crystallization, solves the foaming problem of linear straight chain high crystallinity biological high molecular polymer, and finally obtains the tissue engineering scaffold material with controllable structure.

Description

Method for preparing tissue engineering scaffold material by adopting degradable synthetic polymer material
Technical Field
The invention relates to a method for preparing a tissue engineering scaffold material by adopting a degradable synthetic polymer material.
Background
In the prior art, the tissue engineering scaffold material refers to a material which can be combined with tissue living cells, can be implanted into different tissues of a living body, and can be used for replacing functions of the tissues. In order to proliferate and differentiate seed cells, it is necessary to provide a cell scaffold composed of a biomaterial, which corresponds to an artificial extracellular matrix. The tissue engineering scaffold material comprises: bone, cartilage, blood vessels, nerves, skin and artificial organs, such as liver, spleen, kidney, bladder, etc.
At present, tissue engineering scaffold materials are classified according to source as: (1) natural degradable high molecular material, natural biodegradable high molecular material refers to high molecular degradable material extracted from animal and plant tissues, such as collagen (collagen), which is the tissue component of natural bone; chitosan (chitosan), a derivative of chitin; gelatin (gelatin), agar, dextran, and hyaluronic acid. The material is characterized in that the degradation product is easy to be absorbed by organisms, but the strength and the processing performance are poor, and the degradation speed cannot be adjusted. (2) Naturally degradable inorganic materials, such as coral, are bones of natural animals, 99% of which are calcium phosphates; as another example, Coral Hydroxyapatite (CHA). They all have the porous structure of natural coral, have good porosity, can be well adhered to target cells without influencing proliferation, differentiation and osteogenesis, and are good bone tissue engineering materials. (3) The degradable synthetic polymer material can be degraded, and the commonly used degradable synthetic polymer materials include polylactic acid (poly (lactic acid), PLA), polyglycolic acid (poly (glycolic acid)), PGA, Polycaprolactone (PCL), polyether, polycarbonate, and the like. The degradation product of the material can be metabolized and eliminated in vivo, and has no harm to organism and good plasticity. Of these, PLA and PGA are most widely used in tissue engineering. (4) Synthetic degradable inorganic materials, mainly Calcium Phosphate Cement (CPC), Hydroxyapatite (HA), tricalcium phosphate (TCP), bioactive ceramics such as bioactive glass ceramic (BCG), extracellular matrix ceramic materials, and the like are commonly used. (5) The composite material comprises the following components: by combining the characteristics of different materials, certain specific functions are achieved by combining the advantages of each other. Such as polylactic acid-hydroxyapatite. Among them, the 3 rd material, namely the degradable synthetic polymer material, has made a better development. At present, the preparation method of degradable synthetic polymer materials mainly adopts a melt spinning or electrospinning mode. However, this spinning method has the following problems: (1) the prepared fiber scaffold has no compression resistance; (2) the screw shearing in the melt spinning process may cause material degradation, thereby reducing the mechanical properties of the finished product; (3) for polymers with low viscoelasticity, such as polyglycolide and the like, the inorganic filler modified spinning can not be prepared; (4) electrospinning uses organic solvents to prepare polymer solutions, which may affect cell activity.
On the other hand, the aggregation structure of the polymer material has a difference between crystalline and amorphous states, and the aggregation structure directly affects the processability of the material and the microstructure of the product. Many high molecular polymers are linear and straight-chain structures, which result in high crystallinity in the solid state, but low viscoelasticity in the molten state; thus, the use of solid state foam results in foams with too small a pore size, low porosity, and poor connectivity, which are not suitable for tissue engineering applications. In the process of melt foaming, such as injection molding or extrusion foaming, the shearing of the screw can cause the breakage of linear molecular chains of the screw, and foam pores of the foaming material collapse, which is not suitable for the application of tissue engineering.
Therefore, the method has positive practical significance on how to solve the foaming problem of the high-molecular polymer with high crystallinity and how to prepare the tissue engineering scaffold material with a controllable structure.
Disclosure of Invention
The invention aims to provide a method for preparing a tissue engineering scaffold material by adopting a degradable synthetic polymer material.
In order to achieve the purpose of the invention, the technical scheme adopted by the invention is as follows: a method for preparing tissue engineering scaffold material by adopting degradable synthetic polymer material comprises the following steps:
(1) forming a high molecular polymer sample by a forming technology, and then placing the high molecular polymer sample in a high-pressure container;
(2) displacing gas within the high pressure vessel with a supercritical fluid;
(3) heating and pressurizing the high-pressure container to saturate the high-molecular polymer sample in a high-temperature high-pressure supercritical fluid atmosphere for at least 1 minute, so that the high-molecular polymer sample is melted under the condition that the temperature is higher than the critical temperature or critical pressure of the supercritical fluid, and polymer crystallization is eliminated;
(4) then cooling to the foaming temperature for at least 1 minute;
(5) quickly decompressing to obtain the tissue engineering scaffold material;
in the step (1), the polymer sample is selected from one of the following 3:
(a) a single component high molecular weight polymer; (b) a copolymer of two or more kinds of high molecular polymers; (c) a blend of two or more high molecular polymers; wherein, the high molecular polymer is: polyglycolide with the crystallinity of 46-52 percent, polylactide glycolide with the crystallinity of 40-47 percent, polylactide with the crystallinity of 49-51 percent, polycaprolactone with the crystallinity of 65-70 percent, low-density medical grade polyethylene with the crystallinity of 55-65 percent, high-density medical grade polyethylene with the crystallinity of 80-90 percent, medical grade polyethylene terephthalate with the crystallinity of 40-60 percent, medical grade polytetrafluoroethylene with the crystallinity of 55-80 percent, medical grade polyamide PA6 with the crystallinity of 49-51 percent or medical grade polyamide PA66 with the crystallinity of 40-60 percent.
In the technical scheme, the high-pressure vessel in the step (1) is a high-temperature high-pressure reaction kettle.
In the technical scheme, the forming technology in the step (1) is one or more selected from extrusion mixing, compression molding, crushing and co-melting and electrostatic spinning.
In the above technical solution, the supercritical fluid in the step (2) is selected from CO2、N2、H2O、CH4、CH3Cl、CH2Cl2、CH3OH、C5H10、CH3CH2OH, CFC-11, HCFC-14LB, HCFC-1141B, HCFC-22, HFC-245FA, FC-365MFC and HFC-134A, HFC-152A, HFE.
In the above technical solution, in the step (3), the high-pressure vessel is heated and pressurized, so that the high molecular polymer sample is saturated in the high-temperature high-pressure supercritical fluid atmosphere, and the saturation time is 0.5 to 40 hours. Wherein, the high temperature is 1-50 ℃ higher than the melting point of the high molecular polymer, preferably 5-20 ℃; the high pressure is 5-40MPa, preferably 15-30 MP; the saturation time is preferably 1 to 10 hours, ensuring that the polymer is crystalline and amorphous.
In the above technical solution, in the step (4), the foaming temperature is between the crystallization temperature and the melting temperature of the high molecular polymer sample.
In the above technical scheme, in the step (5), the pressure relief rate of the rapid pressure relief is controlled to be 10-1000 MPa/s.
Preferably, the pressure relief rate of the rapid pressure relief is controlled to be 30-200 MPa/s. The existing pressure relief equipment can be adopted for quick pressure relief, and the pressure relief can be carried out according to a set pressure-time curve; the conventional pressure relief device mainly comprises a pressure sensor, a pressure-digital signal conversion device and a digital signal recording device.
In the technical scheme, the tissue engineering scaffold material is a porous scaffold structure, the porosity of the porous scaffold material is higher than 60%, the connectivity of the porous scaffold material is higher than 80%, and the pore size distribution of the porous scaffold material is 1-800 micrometers. Preferably, the porosity is higher than 70%, and the connectivity is higher than 85%; the pore size distribution is preferably 5 to 500. mu.m, more preferably 5 to 50 μm, and still more preferably 15 to 30 μm.
The main working principle of the invention is as follows: using supercritical fluid as physical foaming agent, using high-pressure kettle to intermittently foam; on the basis, the crystal region of the high molecular polymer is melted through high temperature, the crystallization of the polymer is eliminated, the saturation temperature is reduced, the foaming temperature is controlled to be between the crystallization temperature and the melting temperature of the polymer, and the viscoelasticity of the polymer melt is improved; thereby solving the foaming problem of linear straight-chain high-crystallinity biopolymer polymers and finally obtaining the tissue engineering scaffold material with controllable structure.
The invention also provides the tissue engineering scaffold material prepared by the preparation method. The porosity is higher than 60%, the connectivity is higher than 80%, and the pore size distribution is 1-800 microns. Preferably, the porosity is higher than 70%, and the connectivity is higher than 85%; the pore size distribution is preferably 5 to 500. mu.m, more preferably 5 to 50 μm, and still more preferably 15 to 30 μm.
Due to the application of the technical scheme, compared with the prior art, the invention has the following advantages:
1. the invention adopts the high-pressure kettle and an intermittent foaming mode, thereby avoiding the technical problems that linear molecular chains are broken and foaming material foam holes are collapsed due to screw shearing;
2. according to the invention, supercritical fluid is used as a physical foaming agent, and an autoclave is used for intermittent foaming, on the basis, the crystal region of a high molecular polymer is melted through high temperature, the crystallization of the polymer is eliminated, the saturation temperature is reduced, the foaming temperature is controlled to be between the crystallization temperature and the melting temperature of the polymer, and the viscoelasticity of the polymer melt is improved, so that the foaming problem of a linear high-crystallinity biopolymer polymer is solved, the tissue engineering scaffold material with a controllable structure is finally obtained, and a remarkable effect is obtained;
3. the preparation method is simple and easy to implement, very environment-friendly and suitable for large-scale production.
Drawings
FIG. 1 is a graph of pressure drop for example 1 of the present invention.
FIG. 2 is an SEM image of the internal cross section of a foamed product in example 1 of the present invention.
FIG. 3 is a graph of pressure drop for example 2 of the present invention.
FIG. 4 is an SEM image of the internal cross section of a foamed product in example 2 of the present invention.
Fig. 5 is a graph of pressure drop for example 3 of the present invention.
FIG. 6 is an SEM image of the internal cross section of a foamed product in example 3 of the present invention.
FIG. 7 is a graph of pressure drop for example 4 of the present invention.
FIG. 8 is an SEM image of the internal cross section of a foamed product in example 4 of the present invention.
Detailed Description
The invention is further described below with reference to the following examples:
example 1
A method for preparing tissue engineering scaffold material by adopting degradable synthetic polymer material comprises the following steps:
(1) the used raw material is polyglycolide with the crystallinity of 48 percent, and the raw material is prepared into sheet materials by a hot pressing method and then is placed in a high-temperature high-pressure reaction kettle;
(2) using supercritical carbon dioxide (scCO)2) Replacing gas in the high-temperature high-pressure reaction kettle;
(3) putting the high-temperature high-pressure reaction kettle into an oil bath kettle at 250 ℃, pressurizing to 25MPa, and saturating for 1 hour;
(4) rapidly cooling the oil bath to 220 ℃, and saturating the sheet stock in the reaction kettle for 1 hour;
(5) releasing pressure according to a set pressure drop curve to obtain the tissue engineering scaffold material; the pressure drop curve for the pressure relief is shown in figure 1.
The SEM image of the internal section of the foamed product is shown in FIG. 2. The foamed sample was prepared with an average pore size of 100 microns, a porosity of 82%, and a connectivity of 90%.
Example 2
A method for preparing tissue engineering scaffold material by adopting degradable synthetic polymer material comprises the following steps:
(1) the used raw material is polyglycolide with the crystallinity of 50 percent, and the sheet material is prepared by a hot pressing method and then is placed in a high-temperature high-pressure reaction kettle;
(2) using supercritical carbon dioxide (scCO)2) Replacing gas in the high-temperature high-pressure reaction kettle;
(3) putting the high-temperature high-pressure reaction kettle into an oil bath kettle at 250 ℃, pressurizing to 30MPa, and saturating for 1 hour;
(4) rapidly cooling the oil bath to 220 ℃, and saturating the sheet stock in the reaction kettle for 1 hour;
(5) releasing pressure according to a set pressure drop curve to obtain the tissue engineering scaffold material; the pressure drop curve for the pressure relief is shown in figure 3.
The SEM image of the internal cross section of the foamed product is shown in fig. 4. The foamed sample was prepared with an average pore size of 40 microns, a porosity of 65%, and a connectivity of 80%.
Example 3
A method for preparing tissue engineering scaffold material by adopting degradable synthetic polymer material comprises the following steps:
(1) the used raw material is Polycaprolactone (PCL) powder with the crystallinity of 67 percent, a sheet material is prepared by a hot-pressing method, and then the sheet material is placed in a high-temperature high-pressure reaction kettle;
(2) using supercritical carbon dioxide (scCO)2) Replacing gas in the high-temperature high-pressure reaction kettle;
(3) putting the high-temperature high-pressure reaction kettle into a water bath kettle at the temperature of 90 ℃, pressurizing to 30MPa, and saturating for 1 hour;
(4) rapidly cooling the oil bath to 30 ℃, and saturating the flakes in the reaction kettle for 1 hour;
(5) releasing pressure according to a set pressure drop curve to obtain the tissue engineering scaffold material; the pressure drop curve for the pressure relief is shown in figure 5.
The SEM image of the internal cross-section of the foamed product is shown in fig. 6. The foamed sample was prepared with an average pore size of 80 microns, a porosity of 80%, and a connectivity of 97%.
Example 4
A method for preparing tissue engineering scaffold material by adopting degradable synthetic polymer material comprises the following steps:
(1) the used raw material is Polylactide (PLA) powder with the crystallinity of 50 percent, and the material is prepared into sheet material by a hot-pressing method and then is placed in a high-temperature high-pressure reaction kettle;
(2) using supercritical carbon dioxide (scCO)2) Replacing gas in the high-temperature high-pressure reaction kettle;
(3) putting the high-temperature high-pressure reaction kettle into a water bath kettle at 200 ℃, pressurizing to 10MPa, and saturating for 1 hour;
(4) rapidly cooling the oil bath to 70 ℃, and saturating the sheet stock in the reaction kettle for 1 hour;
(5) releasing pressure according to a set pressure drop curve to obtain the tissue engineering scaffold material; the pressure drop curve for the pressure relief is shown in figure 7.
The SEM image of the internal cross section of the foamed product is shown in fig. 8. The foamed sample was prepared with an average pore size of 112 microns, a porosity of 75%, and a connectivity of 93%.
Comparative example 1
The method for preparing the degradable synthetic polymer material by adopting a melt spinning method is approximately as follows: (1) preparing a PCL spinning melt (melting fiber-forming polymer chips or preparing a melt by continuous polymerization); (2) extruding the melt through a spinneret orifice to form melt trickle; (3) cooling and solidifying the melt trickle to form nascent fiber; (4) oiling and winding the nascent fiber. Melt spinning is divided into direct spinning and chip spinning.
Comparative example 2
The method for preparing the degradable synthetic polymer material by adopting the electrospinning method is approximately as follows: (1) filling a PLA polymer solution in a spraying device; (2) under the action of an external electric field, the polymer liquid drops are kept at the nozzle under the action of surface tension; (3) increasing the electric field intensity, and collecting the attenuated jet flow, namely the finally formed fiber; (4) collecting and forming the non-woven fabric.
The tissue engineering scaffold materials prepared in the above examples and comparative examples were subjected to performance tests, and the results are shown in the following table:
Figure DEST_PATH_IMAGE002
as can be seen from the above table: (1) the prepared foam has better compression resistance; (2) no screw shearing is needed in the foaming process, so that the material degradation is not caused; (2) filler modified low viscoelasticity polymer products can be prepared; (3) the foaming process has no organic solvent and no toxicity.
The previous description of the disclosed embodiments is provided to enable any person skilled in the art to make or use the present invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the generic principles defined herein may be applied to other embodiments without departing from the spirit or scope of the invention. Thus, the present invention is not intended to be limited to the embodiments shown herein but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims (8)

1. A method for preparing tissue engineering scaffold material by adopting degradable synthetic polymer material is characterized by comprising the following steps:
(1) forming a high molecular polymer sample by a forming technology, and then placing the high molecular polymer sample in a high-pressure container;
(2) displacing gas within the high pressure vessel with a supercritical fluid;
(3) heating and pressurizing the high-pressure container to saturate the high-molecular polymer sample in a high-temperature high-pressure supercritical fluid atmosphere for at least 1 minute, so that the high-molecular polymer sample is melted under the condition that the temperature is higher than the critical temperature or critical pressure of the supercritical fluid, and polymer crystallization is eliminated;
(4) then cooling to the foaming temperature for at least 1 minute;
(5) quickly decompressing to obtain the tissue engineering scaffold material;
in the step (1), the polymer sample is selected from one of the following 3:
(a) a single component high molecular weight polymer; (b) a copolymer of two or more kinds of high molecular polymers; (c) a blend of two or more high molecular polymers; wherein, the high molecular polymer is: polyglycolide with the crystallinity of 46-52 percent, polylactide glycolide with the crystallinity of 40-47 percent, polylactide with the crystallinity of 49-51 percent, polycaprolactone with the crystallinity of 65-70 percent, low-density medical grade polyethylene with the crystallinity of 55-65 percent, high-density medical grade polyethylene with the crystallinity of 80-90 percent, medical grade polyethylene terephthalate with the crystallinity of 40-60 percent, medical grade polytetrafluoroethylene with the crystallinity of 55-80 percent, medical grade polyamide PA6 with the crystallinity of 49-51 percent or medical grade polyamide PA66 with the crystallinity of 40-60 percent;
in the step (4), the foaming temperature is between the crystallization temperature and the melting temperature zone of the high molecular polymer sample;
the tissue engineering scaffold material is a porous scaffold structure, the porosity of the porous scaffold material is higher than 60%, the connectivity of the porous scaffold material is higher than 80%, and the pore size distribution of the porous scaffold material is 1-800 micrometers.
2. The method of claim 1, wherein: the high-pressure vessel in the step (1) is a high-temperature high-pressure reaction kettle.
3. The method of claim 1, wherein: the forming technology in the step (1) is one or more selected from extrusion mixing, compression molding, crushing and co-melting and electrostatic spinning.
4. The method of claim 1, wherein: the supercritical fluid in the step (2) is selected from CO2、N2、CH4、CH3Cl、CH2Cl2、CH3OH、C5H10、CH3CH2OH, CFC-11, HCFC-14LB, HCFC-22, HFC-245FA, HFC-134A, HFC-152A, HFE.
5. The method of claim 1, wherein: in the step (3), the high-pressure container is heated and pressurized, so that the high-molecular polymer sample is saturated in the high-temperature high-pressure supercritical fluid atmosphere for 0.5-40 hours.
6. The method of claim 1, wherein: in the step (5), the pressure relief rate of the rapid pressure relief is controlled to be 10-1000 MPa/s.
7. The method of claim 6, wherein: the pressure relief rate of the rapid pressure relief is controlled to be 30-200 MPa/s.
8. The tissue engineering scaffold material prepared by the preparation method according to any one of claims 1 to 7.
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CN102501348A (en) * 2011-09-28 2012-06-20 华南理工大学 Preparation method of high polymer material scaffolds for tissue engineering
CN103819690A (en) * 2014-02-18 2014-05-28 上海交通大学 Process using degradation eliminating method for preparation of high-crystallinity polylactic acid

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CN102501348A (en) * 2011-09-28 2012-06-20 华南理工大学 Preparation method of high polymer material scaffolds for tissue engineering
CN103819690A (en) * 2014-02-18 2014-05-28 上海交通大学 Process using degradation eliminating method for preparation of high-crystallinity polylactic acid

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