WO2014090035A1 - β-硅酸二钙多孔生物陶瓷支架及其制备方法和应用 - Google Patents

β-硅酸二钙多孔生物陶瓷支架及其制备方法和应用 Download PDF

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WO2014090035A1
WO2014090035A1 PCT/CN2013/085130 CN2013085130W WO2014090035A1 WO 2014090035 A1 WO2014090035 A1 WO 2014090035A1 CN 2013085130 W CN2013085130 W CN 2013085130W WO 2014090035 A1 WO2014090035 A1 WO 2014090035A1
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dicalcium
porous
bioceramic
silicate
stent
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PCT/CN2013/085130
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English (en)
French (fr)
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刘海蓉
戴瑶
刘斌斌
周征
夏磊磊
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湖南大学
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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
    • A61L27/02—Inorganic materials
    • A61L27/10—Ceramics or glasses
    • 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
    • A61L27/50—Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
    • A61L27/56—Porous materials, e.g. foams or sponges
    • C—CHEMISTRY; METALLURGY
    • C04—CEMENTS; CONCRETE; ARTIFICIAL STONE; CERAMICS; REFRACTORIES
    • C04B—LIME, MAGNESIA; SLAG; CEMENTS; COMPOSITIONS THEREOF, e.g. MORTARS, CONCRETE OR LIKE BUILDING MATERIALS; ARTIFICIAL STONE; CERAMICS; REFRACTORIES; TREATMENT OF NATURAL STONE
    • C04B35/00—Shaped ceramic products characterised by their composition; Ceramics compositions; Processing powders of inorganic compounds preparatory to the manufacturing of ceramic products
    • C04B35/01—Shaped ceramic products characterised by their composition; Ceramics compositions; Processing powders of inorganic compounds preparatory to the manufacturing of ceramic products based on oxide ceramics
    • C04B35/16—Shaped ceramic products characterised by their composition; Ceramics compositions; Processing powders of inorganic compounds preparatory to the manufacturing of ceramic products based on oxide ceramics based on silicates other than clay
    • C04B35/22—Shaped ceramic products characterised by their composition; Ceramics compositions; Processing powders of inorganic compounds preparatory to the manufacturing of ceramic products based on oxide ceramics based on silicates other than clay rich in calcium oxide, e.g. wollastonite
    • C—CHEMISTRY; METALLURGY
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    • C04B35/00—Shaped ceramic products characterised by their composition; Ceramics compositions; Processing powders of inorganic compounds preparatory to the manufacturing of ceramic products
    • C04B35/622—Forming processes; Processing powders of inorganic compounds preparatory to the manufacturing of ceramic products
    • C04B35/626—Preparing or treating the powders individually or as batches ; preparing or treating macroscopic reinforcing agents for ceramic products, e.g. fibres; mechanical aspects section B
    • C04B35/62605—Treating the starting powders individually or as mixtures
    • C04B35/6261—Milling
    • C—CHEMISTRY; METALLURGY
    • C04—CEMENTS; CONCRETE; ARTIFICIAL STONE; CERAMICS; REFRACTORIES
    • C04B—LIME, MAGNESIA; SLAG; CEMENTS; COMPOSITIONS THEREOF, e.g. MORTARS, CONCRETE OR LIKE BUILDING MATERIALS; ARTIFICIAL STONE; CERAMICS; REFRACTORIES; TREATMENT OF NATURAL STONE
    • C04B35/00—Shaped ceramic products characterised by their composition; Ceramics compositions; Processing powders of inorganic compounds preparatory to the manufacturing of ceramic products
    • C04B35/622—Forming processes; Processing powders of inorganic compounds preparatory to the manufacturing of ceramic products
    • C04B35/64—Burning or sintering processes
    • C—CHEMISTRY; METALLURGY
    • C04—CEMENTS; CONCRETE; ARTIFICIAL STONE; CERAMICS; REFRACTORIES
    • C04B—LIME, MAGNESIA; SLAG; CEMENTS; COMPOSITIONS THEREOF, e.g. MORTARS, CONCRETE OR LIKE BUILDING MATERIALS; ARTIFICIAL STONE; CERAMICS; REFRACTORIES; TREATMENT OF NATURAL STONE
    • C04B38/00—Porous mortars, concrete, artificial stone or ceramic ware; Preparation thereof
    • C04B38/06—Porous mortars, concrete, artificial stone or ceramic ware; Preparation thereof by burning-out added substances by burning natural expanding materials or by sublimating or melting out added substances
    • C—CHEMISTRY; METALLURGY
    • C04—CEMENTS; CONCRETE; ARTIFICIAL STONE; CERAMICS; REFRACTORIES
    • C04B—LIME, MAGNESIA; SLAG; CEMENTS; COMPOSITIONS THEREOF, e.g. MORTARS, CONCRETE OR LIKE BUILDING MATERIALS; ARTIFICIAL STONE; CERAMICS; REFRACTORIES; TREATMENT OF NATURAL STONE
    • C04B38/00—Porous mortars, concrete, artificial stone or ceramic ware; Preparation thereof
    • C04B38/06—Porous mortars, concrete, artificial stone or ceramic ware; Preparation thereof by burning-out added substances by burning natural expanding materials or by sublimating or melting out added substances
    • C04B38/0615—Porous mortars, concrete, artificial stone or ceramic ware; Preparation thereof by burning-out added substances by burning natural expanding materials or by sublimating or melting out added substances the burned-out substance being a monolitic element having approximately the same dimensions as the final article, e.g. a porous polyurethane sheet or a prepreg obtained by bonding together resin particles
    • 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
    • A61L2400/00—Materials characterised by their function or physical properties
    • A61L2400/08—Methods for forming porous structures using a negative form which is filled and then removed by pyrolysis or dissolution
    • 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
    • A61L2430/00—Materials or treatment for tissue regeneration
    • A61L2430/02—Materials or treatment for tissue regeneration for reconstruction of bones; weight-bearing implants
    • C—CHEMISTRY; METALLURGY
    • C04—CEMENTS; CONCRETE; ARTIFICIAL STONE; CERAMICS; REFRACTORIES
    • C04B—LIME, MAGNESIA; SLAG; CEMENTS; COMPOSITIONS THEREOF, e.g. MORTARS, CONCRETE OR LIKE BUILDING MATERIALS; ARTIFICIAL STONE; CERAMICS; REFRACTORIES; TREATMENT OF NATURAL STONE
    • C04B2111/00—Mortars, concrete or artificial stone or mixtures to prepare them, characterised by specific function, property or use
    • C04B2111/00474—Uses not provided for elsewhere in C04B2111/00
    • C04B2111/00836—Uses not provided for elsewhere in C04B2111/00 for medical or dental applications
    • C—CHEMISTRY; METALLURGY
    • C04—CEMENTS; CONCRETE; ARTIFICIAL STONE; CERAMICS; REFRACTORIES
    • C04B—LIME, MAGNESIA; SLAG; CEMENTS; COMPOSITIONS THEREOF, e.g. MORTARS, CONCRETE OR LIKE BUILDING MATERIALS; ARTIFICIAL STONE; CERAMICS; REFRACTORIES; TREATMENT OF NATURAL STONE
    • C04B2235/00—Aspects relating to ceramic starting mixtures or sintered ceramic products
    • C04B2235/02—Composition of constituents of the starting material or of secondary phases of the final product
    • C04B2235/30—Constituents and secondary phases not being of a fibrous nature
    • C04B2235/44—Metal salt constituents or additives chosen for the nature of the anions, e.g. hydrides or acetylacetonate
    • C04B2235/442—Carbonates
    • C—CHEMISTRY; METALLURGY
    • C04—CEMENTS; CONCRETE; ARTIFICIAL STONE; CERAMICS; REFRACTORIES
    • C04B—LIME, MAGNESIA; SLAG; CEMENTS; COMPOSITIONS THEREOF, e.g. MORTARS, CONCRETE OR LIKE BUILDING MATERIALS; ARTIFICIAL STONE; CERAMICS; REFRACTORIES; TREATMENT OF NATURAL STONE
    • C04B2235/00—Aspects relating to ceramic starting mixtures or sintered ceramic products
    • C04B2235/02—Composition of constituents of the starting material or of secondary phases of the final product
    • C04B2235/30—Constituents and secondary phases not being of a fibrous nature
    • C04B2235/44—Metal salt constituents or additives chosen for the nature of the anions, e.g. hydrides or acetylacetonate
    • C04B2235/447—Phosphates or phosphites, e.g. orthophosphate, hypophosphite
    • C—CHEMISTRY; METALLURGY
    • C04—CEMENTS; CONCRETE; ARTIFICIAL STONE; CERAMICS; REFRACTORIES
    • C04B—LIME, MAGNESIA; SLAG; CEMENTS; COMPOSITIONS THEREOF, e.g. MORTARS, CONCRETE OR LIKE BUILDING MATERIALS; ARTIFICIAL STONE; CERAMICS; REFRACTORIES; TREATMENT OF NATURAL STONE
    • C04B2235/00—Aspects relating to ceramic starting mixtures or sintered ceramic products
    • C04B2235/65—Aspects relating to heat treatments of ceramic bodies such as green ceramics or pre-sintered ceramics, e.g. burning, sintering or melting processes
    • C04B2235/656—Aspects relating to heat treatments of ceramic bodies such as green ceramics or pre-sintered ceramics, e.g. burning, sintering or melting processes characterised by specific heating conditions during heat treatment
    • C04B2235/6562—Heating rate
    • C—CHEMISTRY; METALLURGY
    • C04—CEMENTS; CONCRETE; ARTIFICIAL STONE; CERAMICS; REFRACTORIES
    • C04B—LIME, MAGNESIA; SLAG; CEMENTS; COMPOSITIONS THEREOF, e.g. MORTARS, CONCRETE OR LIKE BUILDING MATERIALS; ARTIFICIAL STONE; CERAMICS; REFRACTORIES; TREATMENT OF NATURAL STONE
    • C04B2235/00—Aspects relating to ceramic starting mixtures or sintered ceramic products
    • C04B2235/65—Aspects relating to heat treatments of ceramic bodies such as green ceramics or pre-sintered ceramics, e.g. burning, sintering or melting processes
    • C04B2235/656—Aspects relating to heat treatments of ceramic bodies such as green ceramics or pre-sintered ceramics, e.g. burning, sintering or melting processes characterised by specific heating conditions during heat treatment
    • C04B2235/6567—Treatment time

Definitions

  • the invention comprises a bone tissue engineering biological scaffold material and a preparation method and application thereof, in particular to a bioceramic scaffold and a preparation method and application thereof.
  • Tissue engineering is an interdisciplinary study of the biologically active alternatives for repairing or improving the structure and function of a diseased tissue or organ in human body, based on the principles of ffl cell biology, biomaterials, and engineering. His research mainly includes one aspect: seed cells, biomaterial scaffolds and growth signals. Bioscaffold materials are the key factors in tissue engineering, so the development and construction of ideal extracellular scaffold materials is one of the research focus and hotspots of tissue engineering at this stage.
  • Bone tissue engineering is a branch of tissue engineering. Bone tissue engineering refers to the isolation of osteoblasts, bone marrow stromal stem cells or chondrocytes, which are cultured in vitro and cultured on a scaffold material with good biocompatibility: After the cells grow and proliferate on a three-dimensional scaffold in a prefabricated form, The cell-material composite is implanted into the bone defect site, and the bone cells are continuously proliferated while the biological material is gradually degraded, and the bone tissue is gradually formed and plays a corresponding functional role, thereby achieving the purpose of repairing the bone tissue defect.
  • tissue engineering scaffold material should have the following characteristics: (1) It is non-toxic and has good bio-tissue compatibility. The scaffold material body and its degradation products will not cause immune rejection of the body; (2) High porosity Structure to support and induce cell attachment, proliferation, and production of extracellular matrix; (3) biodegradable, and controllable degradation rate: (4) appropriate plasticity and mechanical strength; to maintain scaffold material microenvironment Stable.
  • bone tissue engineering scaffold materials there are three main types of bone tissue engineering scaffold materials; one is natural bio-derived tree material, which has the advantages of abundant source, simple production, good histocompatibility and certain biodegradability.
  • the second is synthetic materials such as calcium strontium ceramics and bioactive glass.
  • It is a composite material such as an inorganic polymer composite material or a metal polymer composite material.
  • Bioceramics have good biocompatibility and osteoinductivity, and at the same time have good mechanical properties and plasticity, which can meet the requirements of bone tissue engineering for scaffold material formation, mechanical properties and osteoinductivity. Has received a lot of attention and application.
  • porous bioceramic scaffolds generally lack sufficient strength, hydroxyl
  • the compressive strength of the sootstone porous bioceramic scaffold generally does not exceed 10 MPa
  • the compressive strength of the tricalcium phosphate porous scaffold generally does not exceed 15 MPa, and it can barely reach the strength requirement of cancellous bone.
  • Dicalcium silicate and the active glass belong to the CaO-SiO 2 system and are considered to be biologically active. Studies have shown that osteoblasts can grow normally on the surface of the dicalcium silicate coating and proliferate and differentiate, indicating that the dicalcium silicate coating has good biocompatibility. Therefore, the ffl-dicalcium silicate bioceramic stent is prepared to solve the above problems and is used for bone tissue engineering.
  • the technical problem to be solved by the present invention is to overcome the deficiencies of the existing bone tissue engineering scaffold materials, and to provide a porous calcium sulphate porous material with high mechanical strength, high open porosity, no toxicity to organisms, and good biocompatibility.
  • the bioceramic stent and correspondingly, provides a preparation method and application of the ⁇ -silicate dicalcium porous bioceramic stent.
  • the technical solution proposed by the present invention is a ⁇ -silicate dicalcium porous bioceramic stent, and the ⁇ -silicate dicalcium porous bioceramic stent is prepared as follows. Specifically, the following steps are included -
  • Precursor template treatment The porous scaffold template is obtained by processing with organic foam according to the desired shape and size of the product.
  • Step (2) The prepared powder is stirred evenly by adding binder and distilled water to form a slurry.
  • the sponge precursor prepared in the step (1) is placed in a slurry and repeatedly subjected to extrusion impregnation, and the excess slurry in the sponge precursor is extruded.
  • the precursor template is preferably a polyurethane sponge.
  • the preferred molar ratio of the Ca source to the Si source is from 1:1 to 3:1, particularly preferably in a molar ratio of 2:1.
  • the stabilizer used is preferably a calcium hydride salt; it is preferably used in an amount of 0.5% by mass of the calcium carbonate and silica mixed powder.
  • the Ca source is preferably CaCO 3 , CaO or Ca(HC 0 3 ) 2
  • the Si source is preferably SiO 2 , white carbon or air-soluble silicon.
  • the binder is preferably sodium carboxymethylcellulose (CMC), preferably used in an amount of from 2% to 4% by mass of the powder obtained by ball milling.
  • the water content in the slurry to be formulated is preferably from 30% to 70%.
  • the sintering furnace is preferably a silicon molybdenum furnace.
  • the blast drying temperature is preferably controlled to be 80 to 120 Torr, and the drying time is preferably 9 to 15 hours.
  • the firing system is preferably carried out in stages, and the heating rates in different temperature ranges are different, and the firing temperature is preferably controlled to be 12001 > 1400, and is preferably kept at the firing temperature for 2 to 4 hours.
  • the sample is immediately taken out of the furnace ⁇ and quenched.
  • the present invention also provides a dicalcium silicate bioceramic stent prepared according to the foregoing method.
  • the present invention provides the above-mentioned ⁇ -silicate dicalcium porous bioceramic stent in bone tissue engineering, which can be used to grow osteoblasts of ⁇ -silicate dicalcium porous bioceramics.
  • the stent is implanted into the body to repair or replace the defect or necrotic bone tissue in the body.
  • the compressive strength of the dicalcium silicate porous bioceramic stent prepared by the invention is 18 ⁇ 30 MPa, which meets or exceeds the mechanical strength of the cancellous bone in the human body. , can be used to repair cancellous bone damage.
  • the invention adopts a template method to prepare a process, and has the characteristics of strong plasticity, controllable product shape and size.
  • the ⁇ -silicate dicalcium porous bioceramic stent described in the present invention is a tissue engineering support material which is highly promising.
  • Fig. 1 is a photomicrograph of a microscopic morphology of a dicalcium silicate porous bioceramic stent prepared in Example 1 of the present invention.
  • 2 is a photomicrograph of the ⁇ -silicate dicalcium porous bioceramic stent implanted in the first embodiment of the present invention; wherein, the map and the B map are micrographs of the cells cultured for 7 days and U days, respectively.
  • Fig. 3 is a photomicrograph of a microscopic morphology of a ⁇ -silicate dicalcium porous bioceramic stent prepared in Example 2 of the present invention.
  • Fig. 4 is a photomicrograph of the seed cell of the ⁇ -silicate dicalcium porous bioceramic stent prepared in Example 2 of the present invention 7 days after. detailed description
  • the dicalcium silicate porous bioceramic scaffold is sintered by impregnating a porous organic polyurethane foam as a precursor.
  • the preparation method of the dicalcium silicate porous bioceramic stent comprises the following steps:
  • Precursor template treatment According to the shape and size required of the product, the polyurethane sponge is processed to obtain a cylindrical porous scaffold template with a diameter of about 19 mm and a thickness of about 10 mm; the template is immersed in a 10% NaOH solution. After soaking for 6 hours, the template was immersed in distilled water for 12 hours, and taken out for drying.
  • step (3) ⁇ was placed in an oven and dried at 90 ° C for 12 hours.
  • the ⁇ -silicate dicalcium porous bioceramic stent prepared in this embodiment has a high pupil ratio, the stent is a connected open pore structure, the pore size is 300-550 ⁇ , and the distribution is uniform, and the compressive strength is 18-28 ⁇ . »
  • the ⁇ -silicate dicalcium porous bioceramic scaffold prepared in this example was used for cell culture experiments, and osteoblastoma cells (MG 63 cells) were implanted to observe cell growth. The SEM images after 7 days of cell culture showed that MG-63 cells had proliferated in a large amount, and the cells were normally attached in the scaffold (Fig.
  • This embodiment adopts another preparation method of the ⁇ -silicate dicalcium porous bioceramic stent of the present invention, which comprises the following steps.
  • Precursor template treatment According to the shape and size required of the product, the polyurethane sponge is processed to obtain a cylindrical porous scaffold template with a diameter of about 19 mm and a thickness of about 10 mm; the template is immersed in a 10% NaOH solution. After soaking for 6 hours, the template was immersed in distilled water for 12 hours, and taken out for drying.
  • Firing The dried sample is sintered in a silicon molybdenum furnace to obtain a ⁇ -silicate dicalcium porous bioceramic stent.
  • the firing system is carried out in stages, and the heating rates in different temperature sections are different and strictly controlled.
  • the firing temperature was 350 ° C and the temperature was kept at the firing temperature for 4 hours.
  • the specific heating system is shown in the following table:
  • the ⁇ -silicate dicalcium porous bioceramic stent prepared in this embodiment has a high opening ratio as shown in FIG. 3, and the stent is a connected open pore structure with a pore size of 300-550 ⁇ m, and the distribution is uniform and resistant.
  • the compressive strength is 24 to 30 MPa.
  • the ⁇ -silicate dicalcium porous bioceramic scaffold prepared in the present example was used for cell culture experiments, and the growth of the cells was observed after implanting osteosarcoma cells (MG-63 cells).
  • MG-63 cells could grow normally on the scaffold, the morphology of the cells in the scaffold was normal, and the cells proliferated 7 days after culture, and the cells were observed to be continuous in the scaffold. Covering the stent in a wide range of extracellular matrix, the cells exhibit good biological activity, which indicates that the stent is not only mechanically strong but also has good biocompatibility.

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Abstract

一种β-硅酸二钙多孔生物陶瓷支架及其制备方法和应用。该β-硅酸二钙多孔生物陶瓷支架制备方法为采用多孔有机泡沫作为前驱体，泡沫前驱体经处理后加入陶瓷浆料中浸渍挂浆，去除多余陶瓷浆料后烘干、烧成制得β-硅酸二钙多孔生物陶瓷支架。该生物陶瓷支架可应用于骨组织工程中。

Description

β -硅酸二钙多孔生物陶瓷支架及其制备方法和应用 技术领域
本发明包括一种骨组织工程生物支架材料及其制备方法和应用， 尤其涉及一种生物陶瓷 支架及其制备方法和应用。
背景技术
组织工程是应 ffl细胞生物学、 生物材料和工程学的原理， 研究开发用于修复或改善人体 病损组织或器官的结构、 功能的生物活性替代物的交叉学科。 其研究主要包括≡个方面： 种 子细胞、 生物材料支架和生长信号。 生物支架材料是组织工程的关键因素， 因而开发构建理 想的细胞外支架材料是现阶段组织工程的研究重点和热点之一。
骨组织工程是组织工程的一个分支。骨组织工程是指将分离的成骨细胞、 骨髓基质干 细胞或软骨细胞， 经体外扩增培养后种植于具有良好生物相容性的支架材料上： 细胞在 预制形态的三维支架上生长增殖后， 将这种细胞-材料复合体植入骨缺损部位， 在生物 材料逐步降解的同时，种植的骨细胞不断增殖，逐步形成骨组织并发挥相应的功能作用， 从而达到修复骨组织缺损的目的。
支架材料在组织工程研究 Φ的作用为：为组织或器官体外构建提供细胞生长的 维支架， 使细胞形成适宜的空间分布和细胞阆的信号传导与相互作用； 提供特殊的生长和分化信号， 维持细胞的定^分化， 是组织工程研究及其临床应用的关键。 理想的组织工程支架材料应具 有以下特点： ( 1 ) 无毒性且具有良好的生物组织相容性， 支架材料本体及其降解产物均不会 引起机体的免疫排斥反应； （2 ) 具有高孔隙率的结构来支持和诱导细胞的贴附、 增殖以及细 胞外基质的产生； （3 )具有生物可降解性， 且降解速率可控： （4 )适宜的可塑性和机械强度； 以维持支架材料微环境的稳定。
目前骨组织工程支架材料主要有三类； 其一是天然生物衍生树料， 具有来源丰富， 制作 简便， 组织相容性良好以及具有一定的生物降解性的优点。 其二是人工合成材料， 如钙璘陶 瓷、 生物活性玻璃等。 其 为复合材料， 如无机高分子复合材料、 金属高分子复合材料等。 生物陶瓷以其良好的生物相容性以及骨诱导性， 同时兼具良好的机械性能可塑性， 能够满足 骨组织工程对支架材料成形、 力学性能和骨诱导性等方面的要求， 在骨组织工程中受到了较 大的关注和应用。
尽管目前己对多种生物支架材料进 了不少硏究， 但生物支架材料存在的种种技术难题 如细胞贴附性、 机械强度等依然没有得到很好的解决。 尤其在骨组织工程中， 支架材料的机 械强度往往成为制约其应用的关键因素。 目前多孔生物陶瓷支架普遍缺乏足够的强度， 羟基 磯灰石多孔生物陶瓷支架的抗压强度一般不超过 lOMPa, 磷酸三钙多孔支架的抗压强度一般 不超过 15MPa, 也只能勉强达到松质骨的强度要求。
硅酸二钙与活性玻璃同属 CaO- Si02体系， 被认为具有生物活性。 己有研究表明成 骨细胞能够在硅酸二钙涂层表面正常贴 生长， 并增殖分化， 表明硅酸二钙涂层具有良 好的生物相容性。因而拟 ffl -硅酸二钙生物陶瓷支架来解决上述难题并将其用于骨组织工 程。
发明内容
本发明拟解决的技术问题是克服现有骨组织工程支架材料的不足，提供一种机械强度高、 开孔率高、 对生物体无毒性、 生物相容性良好的 β -硅酸二钙多孔生物陶瓷支架， 并相应地提 供了一种该 β -硅酸二钙多孔生物陶瓷支架的制备方法和应用。
为实现上述目标， 本发明提出的技术方案为一种 β -硅酸二钙多孔生物陶瓷支架， 所述 β -硅酸二钙多孔生物陶瓷支架按如下的方法制备而成。 具体包括以下歩骤-
(】）前驱体模板处理： 根据产品所需的形状和尺寸， 用有机泡沫加工处理得到多孔支架 模板。
(2)陶瓷原料制备： 将 Ca源和 Si源按适当的比例配比， 同时添加适量水和稳定剂进行 球磨， 球磨后的粉料烘干过筛备用。
(3 ) 浸渍挂浆： 称取步骤 （2) 所制备粉料， 按比^加入粘结剂和蒸馏水搅拌均匀， 配 成浆料。 将歩骤（1 )制备的海绵前驱体置于浆料中反复挤压浸渍， 并挤压出海绵前驱体中多 余浆料。
(4) 千燥： 将步骤 (3 ) Φ得到的样品进行鼓风千燥。
(5 ) 烧成： 干燥后的样品放入烧结炉中高温烧结， 并于烧成温度下保温。
(6) 冷却： 保温结束立郎将样品从炉中取出冷却。
为了更好的解决上述技术问题， 所述前驱体模板优选为聚氨脂海绵。
所述 Ca源和 Si源的优选的摩尔比为 1 :1至 3: 1 , 特别优选为摩尔比 2:1的比例。
所用稳定剂优选为 II酸钙盐； 其优选用量为碳酸钙和二氧化硅混合粉料质量的 0.5%〜
3%<>
所述 Ca源优选为 CaC03、 CaO或 Ca(HC03)2， 所述 Si源优选为 Si02、 白炭黑或者气溶 硅。
所述粘结剂优选为羧甲基纤维素钠 (CMC),其用量优选为球磨所得粉料质量的 2%〜4%。 所配浆料中的含水率优选为 30%〜70%。
所述烧结炉优选为硅钼炉。 所述鼓风千燥的温度优选控制在 80〜120Ό , 干燥时间优选为 9〜15小时。 所述烧成制度优选为分段进行， 不同温度段的升温速度不同， 烧成温度优选控制在 12001>〜1400 ， 并优选于烧成温度下保温 2〜4小时。
所述保温结束后， 优选将样品立即由炉 Φ取出并急冷。
本发明还提供一种根据前述方法制备的 -硅酸二钙生物陶瓷支架。
作为一个完整的技术构思， 本发明提供一种上述的 β -硅酸二钙多孔生物陶瓷支架在骨组 织工程中的应用， 即可将种有成骨细胞的 β -硅酸二钙多孔生物陶瓷支架植入体内以修复或替 代体内缺损或坏死的骨组织。
与现有技术相比； 本发明的优点在于： （】）本发明所制备的 硅酸二钙多孔生物陶瓷支 架的抗压强度为 18〜30MPa, 达到甚至超过了人体内松质骨的机械强度， 可用于修复松质骨 损伤。 （2) 本发明采用模板法制备工艺， 具有可塑性强、 产品形状和尺寸可控的特点。 （3 ) 通过工艺控制， 本发明制备的 β -硅酸二钙多孔生物陶瓷支架内部和表面均为连通孔结构， 大 大提高了支架中的物质交换速率和效率， 为细胞的增殖和组织的修复再生提供了更为有效且 真实的微环境， 细胞可在支架里外良好的贴 ffl-生长并增殖。
由此， 本发明所阐述的 β -硅酸二钙多孔生物陶瓷支架是一种极具应 ffl前景的组织工程支 架材料。
附图说明
图 1为本发明实施例 1制得的 -硅酸二钙多孔生物陶瓷支架的微观形貌显微照片。 图 2为本发明实施例 1制得的 β -硅酸二钙多孔生物陶瓷支架种植细胞后的显微照片； 其 中， Α图、 B图分别为细胞培养 7天、 U天后的显微照片。
图 3为本发明实施例 2制得的 β -硅酸二钙多孔生物陶瓷支架的微观形貌显微照片。 图 4为本发明实施例 2制备的 β -硅酸二钙多孔生物陶瓷支架种楦细胞后 7天的显微照片。 具体实施方式
实施例一
-种如图 1 所示的本发明的 13 -硅酸二钙多孔生物陶瓷支架； 该 硅酸二钙多孔生物陶 瓷支架是以多孔有机聚氨酯泡沫作为前驱体经浸渍挂浆后烧结而成。
如图 2所示的本发明的！ 3 硅酸二钙多孔生物陶瓷支架的制备方法， 包括以下步骤：
( 1 ) 前驱体模板处理： 根据产品所需的形状和尺寸， 将聚氨脂海绵加工得到直径约为 19mm, 厚约为 10mm的圆柱体多孔支架模板； 再将该模板浸入 10%的 NaOH溶液中浸泡 6 小时， 此后将模板浸入蒸馏水中浸泡清洗 12小时， 并取出烘干备用。
(2)粉体的制备： 将（ (〕()3和 S )2摩尔比 2 : 1配比， 同时添加上述粉料质量 0.8%的磷 酸三钙 （TCP) 作为稳定剂。 球磨采用 星球磨机球磨， 球磨机转速为 600r/min， 球磨时间 为 4.5小时。 球磨后的粉料立即取出烘千过 100目筛备用。
(3 )浸渍挂浆： 称取步骤 （2 ) 中的混合粉料， 加入其质量分数 2.5%的羧甲基纤维素钠 (CMC)作为粘结剂。 搅拌混合均匀后， 加入双重蒸馏水配成含水率为 40%的浆料， 搅拌均 匀。 将步骤（1 )制备的海绵前驱体浸入浆料中反复挤压浸渍， 浸渍完成后取出并挤去多余浆 料。
(4) 千燥： 将步骤 （3 ) Φ得到的样品放入烘箱中于 90°C下千燥 12小时。
(5 ) 烧成： 千燥后的样品放入硅钼炉中烧结即可得到 β -硅酸二钙多孔生物陶瓷支架。 烧成制度分段进行， 不同温度段的升温速度不同且进行严格控制。 烧成温度为 1250°C , 在烧 成温度下保温 4小时。 具体升温制度如下表所示- 表 1 : 升温制度
序号 温度 /°c 时间 /min
00 0 10
01 100 10
02 200 10
03 300 80
04 410 130
05 800 100
06 950 100
07 1250 180
08 1250 0
： 保温结束后: ί 即将样品^炉 Φ取出， 置于空气环境中冷却。
本实施例制得的 β-硅酸二钙多孔生物陶瓷支架具有较高的幵孔率， 支架为连通性开孔结 构, 孔径大小为 300-550μηι, 且分布均匀， 抗压强度为 18~28ΜΡα» 将本实施例制得的 β-硅 酸二钙多孔生物陶瓷支架用于细胞培养实验， 种植骨肉瘤细胞 （MG 63 细胞） 后观察细胞 的生长情况。 细胞培养 7天后的 SEM图片显示 MG- 63细胞己有大量增殖， 细胞在支架中正 常贴附（如图 2A); 培养 11天后支架中连续范围的大面积区域均可观察到细胞， 细胞在支架 中充分覆盖， 细胞与细胞间接触良好,细胞分泌大量细胞外基质 （如图 2Β ), 呈现好良好的生 实施例二
本实施例采用另外一种本发明的 β -硅酸二钙多孔生物陶瓷支架的制备方法， 包括以下步 ( 1 ) 前驱体模板处理： 根据产品所需的形状和尺寸， 将聚氨脂海绵加工得到直径约为 19mm, 厚约为 10mm的圆柱体多孔支架模板； 再将该模板浸入 10%的 NaOH溶液中浸泡 6 小时， 此后将模板浸入蒸馏水中浸泡清洗 12小时， 并取出烘干备用。
(2 )粉体的制备： 将 CaC03和 Si02摩尔比 2 : 1配比， 同时添加上述粉料质量 2.2%的磷 酸三钙 （TCP) 作为稳定剂。 球磨后立即取出烘千过 100目筛备用。
( 3 )浸渍挂浆： 称取步骤 （2) 中的混合粉料， 加入其质量分数 3.5%的羧甲基纤维素钠 ( CMC)作为粘结剂。 搅泮混合均匀后， 加入双重蒸馏水配成含水率为 60%的浆料， 搅稃均 匀。 将歩骤（1 )制备的海绵前驱体浸入浆料中反复挤压浸渍， 浸渍完成后取出并挤去多余浆 料。
(4 ) 干燥： 将步骤 （3 ) 中得到的样品放入烘箱中于 110°C下干燥 14小时。
( 5 ) 烧成： 干燥后的样品放入硅钼炉中烧结即可得到 β -硅酸二钙多孔生物陶瓷支架。 烧成制度分段进行， 不同温度段的升温速度不同且进行严格控制。 烧成温度为】 350°C， 在烧 成温度下保温 4小时。 具体升温制度如下表所示：
表 2: 升温制度
序号 温度八〕 时间 rrdn
00 0 10
01 100 10
02 200 10
03 300 80
04 410 130
05 800 100
06 950 115
07 1350 180
08 1350 0
( 6) 冷却： 保温结束后立即将样品从炉中取出， 置于空气环境 Φ冷却。
本实施例制得的 β -硅酸二钙多孔生物陶瓷支架如图 3 所示， 具有较高的开孔率， 支架 为连通性开孔结构， 孔径大小为 300- 550μηι, 且分布均匀， 抗压强度为 24〜30MPa。 将本实 施例制得的 β -硅酸二钙多孔生物陶瓷支架用于细胞培养实验， 种植骨肉瘤细胞 （MG- 63细 胞） 后观察细胞的生长情况。 将细胞固定后通过 SEM 电镜观察发现， MG-63细胞在支架上 能够正常贴^生长， 细胞在支架丄的形态正常， 且培养 7天后细胞出现明显的增殖， 在支架 中可观察到细胞呈连续范围地覆盖在支架中， 细胞分泌大量的细胞外基质， 呈现出良好的生 物活性， 由此也说明本支架不仅机械强度高， 而且具有良好的生物相容性。

Claims

WO 2014/090035 权 利 要 求 书 PCT/CN2013/085130
1. 一种 β -硅酸二钙生物陶瓷支架的制备方法， 其特征在于， 包括以下步骤：
( 1 )前驱体模板处理： 根据产品所需的形状和尺寸， 用有机泡沫加工处理得到多孔支架 模板：
(2)陶瓷原料制备： 将 Ca源和 Si源按适当的比例配比， 同时添加适量水和稳定剂进行 球磨， 球磨后的粉料烘干过筛备用；
(3 ) 浸渍挂浆： 称取步骤 （2) 所制备粉料； 按比例加入粘结剂和蒸馏水搅拌均匀， 配 成浆料， 将步骤（1 )制备的海绵前驱体置于浆料中反复挤压浸渍， 并挤压出海绵前驱体中多 余浆料；
(4) 干燥： 将步骤 （3 ) 中得到的样品进行鼓风干燥；
(5 ) 烧成： 干燥后的样品放入烧结炉中高温烧结， 并于烧成温度下保温；
(6) 冷却： 保温结束立即将样品从炉中取出冷却。
2. 根据权利要求 1所述的 硅酸二钙多孔生物陶瓷支架的制备方法， 其特征在于：所述 前驱体模板为聚氨脂海绵。
3. 根据权利要求 1 所述的 β -硅酸二钙多孔生物陶瓷支架的制备方法， 其特征在于： Ca 源和 Si源的摩尔比为 1 :1至 3:1 ,所 ffl稳定剂为磷酸钙盐， 其用量为碳酸钙和二氧化硅混合粉 料质量的 0.5%〜3%。
4. 根据权利要求 1所述的 硅酸二钙多孔生物陶瓷支架的制备方法， 其特征在于：所述 粘结剂为羧甲基纤维素钠 （CMC), 其用量为球磨所得粉料质量的 2%〜4%。
5. 根据权利要求 1所述的 β -硅酸二钙多孔生物陶瓷支架的制备方法， 其特征在于：所配 浆料中的含水率为 30%〜70%。
6. 根据权利要求 i所述的 β -硅酸二钙多孔生物陶瓷支架的制备方法， 其特征在于：样品 的鼓风干燥的温度控制在 80〜120°C , 干燥时间为 9〜15小时。
7. 根据权利要求 1所述的 硅酸二钙多孔生物陶瓷支架的制备方法， 其特征在于：烧成 制度分段进行， 不同温度段的升温速度不同， 烧成温度控制在 1200°C〜i400°C， 并于烧成温 度下保温 2〜4小时。
8. 根据权利要求 1所述的 β -硅酸二钙多孔生物陶瓷支架的制备方法， 其特征在于： 保温 结束后立即由炉中取出急冷。
9. 一种根据权利要求 1至 8任何一项所述方法制备的 β -硅酸二钙生物陶瓷支架。
10. 一种根据权利要求 9中所述的 β -硅酸二钙多孔生物陶瓷支架在组织工程中的应用， 可应用于骨组织工程中硬骨组织的修复和替代。
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