CN114209890A - 一种亲骨性强的骨修复材料及其制备方法 - Google Patents
一种亲骨性强的骨修复材料及其制备方法 Download PDFInfo
- Publication number
- CN114209890A CN114209890A CN202111398397.0A CN202111398397A CN114209890A CN 114209890 A CN114209890 A CN 114209890A CN 202111398397 A CN202111398397 A CN 202111398397A CN 114209890 A CN114209890 A CN 114209890A
- Authority
- CN
- China
- Prior art keywords
- bone
- repair material
- bone repair
- rhbmp
- osteogenic
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 210000000988 bone and bone Anatomy 0.000 title claims abstract description 86
- 239000000463 material Substances 0.000 title claims abstract description 51
- 230000008439 repair process Effects 0.000 title claims abstract description 41
- 238000002360 preparation method Methods 0.000 title claims abstract description 12
- 239000004005 microsphere Substances 0.000 claims abstract description 41
- 229920001606 poly(lactic acid-co-glycolic acid) Polymers 0.000 claims abstract description 34
- 150000001875 compounds Chemical class 0.000 claims abstract description 22
- 239000006177 biological buffer Substances 0.000 claims abstract description 10
- 238000011068 loading method Methods 0.000 claims abstract description 4
- 230000000087 stabilizing effect Effects 0.000 claims abstract description 4
- 238000000034 method Methods 0.000 claims description 22
- 230000002188 osteogenic effect Effects 0.000 claims description 20
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 claims description 18
- 238000003756 stirring Methods 0.000 claims description 12
- 239000006172 buffering agent Substances 0.000 claims description 10
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 claims description 9
- 230000011164 ossification Effects 0.000 claims description 9
- DBLXOVFQHHSKRC-UHFFFAOYSA-N ethanesulfonic acid;2-piperazin-1-ylethanol Chemical group CCS(O)(=O)=O.OCCN1CCNCC1 DBLXOVFQHHSKRC-UHFFFAOYSA-N 0.000 claims description 8
- 239000000203 mixture Substances 0.000 claims description 7
- 238000005406 washing Methods 0.000 claims description 7
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 claims description 6
- KFZMGEQAYNKOFK-UHFFFAOYSA-N Isopropanol Chemical compound CC(C)O KFZMGEQAYNKOFK-UHFFFAOYSA-N 0.000 claims description 6
- 238000004108 freeze drying Methods 0.000 claims description 6
- 239000007853 buffer solution Substances 0.000 claims description 4
- 239000007864 aqueous solution Substances 0.000 claims description 3
- 230000001804 emulsifying effect Effects 0.000 claims description 3
- 239000000839 emulsion Substances 0.000 claims description 3
- 229940057995 liquid paraffin Drugs 0.000 claims description 3
- 239000003208 petroleum Substances 0.000 claims description 3
- 235000010482 polyoxyethylene sorbitan monooleate Nutrition 0.000 claims description 3
- 229920000053 polysorbate 80 Polymers 0.000 claims description 3
- 239000000843 powder Substances 0.000 claims description 3
- 230000010478 bone regeneration Effects 0.000 claims description 2
- 238000002156 mixing Methods 0.000 claims description 2
- 239000003814 drug Substances 0.000 abstract description 15
- 230000000694 effects Effects 0.000 abstract description 9
- 206010061218 Inflammation Diseases 0.000 abstract description 8
- 230000004054 inflammatory process Effects 0.000 abstract description 8
- 230000004071 biological effect Effects 0.000 abstract description 4
- 239000012567 medical material Substances 0.000 abstract description 2
- 230000001172 regenerating effect Effects 0.000 abstract description 2
- 239000000523 sample Substances 0.000 description 33
- 230000000052 comparative effect Effects 0.000 description 20
- 239000000047 product Substances 0.000 description 17
- 229940079593 drug Drugs 0.000 description 12
- 238000001179 sorption measurement Methods 0.000 description 11
- 241000699670 Mus sp. Species 0.000 description 7
- 238000001727 in vivo Methods 0.000 description 7
- 241000237536 Mytilus edulis Species 0.000 description 6
- 239000004098 Tetracycline Substances 0.000 description 6
- 235000003704 aspartic acid Nutrition 0.000 description 6
- OQFSQFPPLPISGP-UHFFFAOYSA-N beta-carboxyaspartic acid Natural products OC(=O)C(N)C(C(O)=O)C(O)=O OQFSQFPPLPISGP-UHFFFAOYSA-N 0.000 description 6
- 235000020638 mussel Nutrition 0.000 description 6
- 235000019364 tetracycline Nutrition 0.000 description 6
- 150000003522 tetracyclines Chemical class 0.000 description 6
- CKLJMWTZIZZHCS-REOHCLBHSA-N L-aspartic acid Chemical compound OC(=O)[C@@H](N)CC(O)=O CKLJMWTZIZZHCS-REOHCLBHSA-N 0.000 description 5
- 239000000872 buffer Substances 0.000 description 5
- 238000011156 evaluation Methods 0.000 description 5
- 230000035876 healing Effects 0.000 description 5
- 238000004519 manufacturing process Methods 0.000 description 5
- 230000008685 targeting Effects 0.000 description 5
- 210000001519 tissue Anatomy 0.000 description 5
- 208000010392 Bone Fractures Diseases 0.000 description 4
- PXIPVTKHYLBLMZ-UHFFFAOYSA-N Sodium azide Chemical compound [Na+].[N-]=[N+]=[N-] PXIPVTKHYLBLMZ-UHFFFAOYSA-N 0.000 description 4
- 230000002378 acidificating effect Effects 0.000 description 4
- 230000007547 defect Effects 0.000 description 4
- 230000008569 process Effects 0.000 description 4
- 229960002180 tetracycline Drugs 0.000 description 4
- 229930101283 tetracycline Natural products 0.000 description 4
- 206010067484 Adverse reaction Diseases 0.000 description 3
- 206010017076 Fracture Diseases 0.000 description 3
- 206010020649 Hyperkeratosis Diseases 0.000 description 3
- 206010067482 No adverse event Diseases 0.000 description 3
- 230000006838 adverse reaction Effects 0.000 description 3
- 230000008468 bone growth Effects 0.000 description 3
- 238000011161 development Methods 0.000 description 3
- 230000012010 growth Effects 0.000 description 3
- 230000002138 osteoinductive effect Effects 0.000 description 3
- 229940002612 prodrug Drugs 0.000 description 3
- 239000000651 prodrug Substances 0.000 description 3
- 230000001737 promoting effect Effects 0.000 description 3
- 239000000243 solution Substances 0.000 description 3
- 241000588724 Escherichia coli Species 0.000 description 2
- 102000015636 Oligopeptides Human genes 0.000 description 2
- 108010038807 Oligopeptides Proteins 0.000 description 2
- 239000002253 acid Substances 0.000 description 2
- 239000004480 active ingredient Substances 0.000 description 2
- 239000012227 artificial bone substitute Substances 0.000 description 2
- 230000009286 beneficial effect Effects 0.000 description 2
- 230000003115 biocidal effect Effects 0.000 description 2
- 230000017531 blood circulation Effects 0.000 description 2
- 239000012876 carrier material Substances 0.000 description 2
- 230000001186 cumulative effect Effects 0.000 description 2
- 230000018109 developmental process Effects 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 239000000706 filtrate Substances 0.000 description 2
- 229910052588 hydroxylapatite Inorganic materials 0.000 description 2
- 238000000338 in vitro Methods 0.000 description 2
- 230000001965 increasing effect Effects 0.000 description 2
- 230000007774 longterm Effects 0.000 description 2
- 210000003141 lower extremity Anatomy 0.000 description 2
- 210000003205 muscle Anatomy 0.000 description 2
- XYJRXVWERLGGKC-UHFFFAOYSA-D pentacalcium;hydroxide;triphosphate Chemical compound [OH-].[Ca+2].[Ca+2].[Ca+2].[Ca+2].[Ca+2].[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O XYJRXVWERLGGKC-UHFFFAOYSA-D 0.000 description 2
- 239000002861 polymer material Substances 0.000 description 2
- 238000000746 purification Methods 0.000 description 2
- 239000002994 raw material Substances 0.000 description 2
- 230000002459 sustained effect Effects 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 229940040944 tetracyclines Drugs 0.000 description 2
- QORWJWZARLRLPR-UHFFFAOYSA-H tricalcium bis(phosphate) Chemical compound [Ca+2].[Ca+2].[Ca+2].[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O QORWJWZARLRLPR-UHFFFAOYSA-H 0.000 description 2
- XPFJYKARVSSRHE-UHFFFAOYSA-K trisodium;2-hydroxypropane-1,2,3-tricarboxylate;2-hydroxypropane-1,2,3-tricarboxylic acid Chemical compound [Na+].[Na+].[Na+].OC(=O)CC(O)(C(O)=O)CC(O)=O.[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O XPFJYKARVSSRHE-UHFFFAOYSA-K 0.000 description 2
- 229940122361 Bisphosphonate Drugs 0.000 description 1
- 102000008143 Bone Morphogenetic Protein 2 Human genes 0.000 description 1
- 108010049931 Bone Morphogenetic Protein 2 Proteins 0.000 description 1
- 229910014455 Ca-Cb Inorganic materials 0.000 description 1
- 238000002965 ELISA Methods 0.000 description 1
- 102000010834 Extracellular Matrix Proteins Human genes 0.000 description 1
- 108010037362 Extracellular Matrix Proteins Proteins 0.000 description 1
- 239000006173 Good's buffer Substances 0.000 description 1
- 241000699666 Mus <mouse, genus> Species 0.000 description 1
- 229910019142 PO4 Inorganic materials 0.000 description 1
- 206010061363 Skeletal injury Diseases 0.000 description 1
- 102400001320 Transforming growth factor alpha Human genes 0.000 description 1
- 101800004564 Transforming growth factor alpha Proteins 0.000 description 1
- 208000027418 Wounds and injury Diseases 0.000 description 1
- 230000001133 acceleration Effects 0.000 description 1
- 238000009825 accumulation Methods 0.000 description 1
- 239000003463 adsorbent Substances 0.000 description 1
- 230000032683 aging Effects 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 230000003385 bacteriostatic effect Effects 0.000 description 1
- 150000004663 bisphosphonates Chemical class 0.000 description 1
- 230000014461 bone development Effects 0.000 description 1
- 239000000316 bone substitute Substances 0.000 description 1
- 230000003139 buffering effect Effects 0.000 description 1
- 239000011575 calcium Substances 0.000 description 1
- 239000001506 calcium phosphate Substances 0.000 description 1
- 229910000389 calcium phosphate Inorganic materials 0.000 description 1
- 235000011010 calcium phosphates Nutrition 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
- 230000015556 catabolic process Effects 0.000 description 1
- 210000004027 cell Anatomy 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 239000002131 composite material Substances 0.000 description 1
- 238000013270 controlled release Methods 0.000 description 1
- 238000006731 degradation reaction Methods 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 238000000502 dialysis Methods 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 239000003937 drug carrier Substances 0.000 description 1
- 230000035194 endochondral ossification Effects 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 238000011067 equilibration Methods 0.000 description 1
- 210000002744 extracellular matrix Anatomy 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 210000003414 extremity Anatomy 0.000 description 1
- 239000012065 filter cake Substances 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- 239000012530 fluid Substances 0.000 description 1
- 238000007490 hematoxylin and eosin (H&E) staining Methods 0.000 description 1
- 239000007943 implant Substances 0.000 description 1
- 238000002513 implantation Methods 0.000 description 1
- 230000001939 inductive effect Effects 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 229910052500 inorganic mineral Inorganic materials 0.000 description 1
- 238000004255 ion exchange chromatography Methods 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 239000007769 metal material Substances 0.000 description 1
- 239000011707 mineral Substances 0.000 description 1
- 239000002105 nanoparticle Substances 0.000 description 1
- 229920005615 natural polymer Polymers 0.000 description 1
- 210000000963 osteoblast Anatomy 0.000 description 1
- 230000004072 osteoblast differentiation Effects 0.000 description 1
- 210000002997 osteoclast Anatomy 0.000 description 1
- 229960001412 pentobarbital Drugs 0.000 description 1
- WEXRUCMBJFQVBZ-UHFFFAOYSA-N pentobarbital Chemical compound CCCC(C)C1(CC)C(=O)NC(=O)NC1=O WEXRUCMBJFQVBZ-UHFFFAOYSA-N 0.000 description 1
- 230000035699 permeability Effects 0.000 description 1
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 description 1
- 239000010452 phosphate Substances 0.000 description 1
- 230000002035 prolonged effect Effects 0.000 description 1
- 235000018102 proteins Nutrition 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 238000011084 recovery Methods 0.000 description 1
- 230000008929 regeneration Effects 0.000 description 1
- 238000011069 regeneration method Methods 0.000 description 1
- 239000012488 sample solution Substances 0.000 description 1
- 238000007493 shaping process Methods 0.000 description 1
- POECFFCNUXZPJT-UHFFFAOYSA-M sodium;carbonic acid;hydrogen carbonate Chemical compound [Na+].OC(O)=O.OC([O-])=O POECFFCNUXZPJT-UHFFFAOYSA-M 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 238000013268 sustained release Methods 0.000 description 1
- 239000012730 sustained-release form Substances 0.000 description 1
- 229920001059 synthetic polymer Polymers 0.000 description 1
- 230000009885 systemic effect Effects 0.000 description 1
- 229940126585 therapeutic drug Drugs 0.000 description 1
- 230000001225 therapeutic effect Effects 0.000 description 1
- 230000000699 topical effect Effects 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 230000007704 transition Effects 0.000 description 1
- 210000000689 upper leg Anatomy 0.000 description 1
Images
Classifications
-
- 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
- A61L27/54—Biologically active materials, e.g. therapeutic substances
-
- 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
- A61L27/14—Macromolecular materials
- A61L27/18—Macromolecular materials obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds
-
- 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
- A61L27/14—Macromolecular materials
- A61L27/22—Polypeptides or derivatives thereof, e.g. degradation products
-
- 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
-
- 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
- A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
- A61L2300/20—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices containing or releasing organic materials
- A61L2300/252—Polypeptides, proteins, e.g. glycoproteins, lipoproteins, cytokines
-
- 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
- A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
- A61L2300/40—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices characterised by a specific therapeutic activity or mode of action
- A61L2300/412—Tissue-regenerating or healing or proliferative agents
-
- 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
- A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
- A61L2300/60—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices characterised by a special physical form
- A61L2300/602—Type of release, e.g. controlled, sustained, slow
-
- 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
Landscapes
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Medicinal Chemistry (AREA)
- Animal Behavior & Ethology (AREA)
- Veterinary Medicine (AREA)
- Epidemiology (AREA)
- Oral & Maxillofacial Surgery (AREA)
- Dermatology (AREA)
- General Health & Medical Sciences (AREA)
- Public Health (AREA)
- Transplantation (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Engineering & Computer Science (AREA)
- Biomedical Technology (AREA)
- Molecular Biology (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Materials For Medical Uses (AREA)
Abstract
本发明属于生物再生医学材料领域,涉及一种亲骨性强的骨修复材料及其制备方法。一种亲骨性强的骨修复材料包括rhBMP‑2和PLGA微球,微球的表面通过亲骨性化合物修饰;还包括用于稳定载药体系微环境pH值的生物缓冲剂。该骨修复材料增强了rhBMP‑2对骨组织的定向作用以减少对非骨组织的副作用,并提高其在局部的利用度,同时稳定微球内部微环境,保护药物的生物活性并减少炎症反应。
Description
技术领域
本发明属于生物再生医学材料领域,涉及一种亲骨性强的骨修复材料及其制备方法,增强rhBMP-2对骨组织的定向作用以减少对非骨组织的副作用,并提高其在局部的利用度,同时稳定微球内部微环境,保护药物的生物活性并减少炎症反应。
背景技术
据统计我国每年骨缺损患者在300~700万人之间,随着交通设施的发展和人口老龄化进程的加速,这一数据还在逐年攀升。较大的骨缺损常难以自行修复,临床长需要行骨移植术来重建骨缺损部位的形态和功能,以缩短治疗周期,降低病残率,因此骨修复材料需求量巨大。随着材料科学及组织工程技术的迅猛发展,大量人工骨替代材料得以在临床广泛应用。目前应用于临床的人工骨替代材料主要包括细胞外基质材料、天然高分子材料、合成高分子材料、医用金属材料、磷酸钙类材料及其复合材料。但遗憾的是目前所有的人工骨替代材料都缺乏修复骨缺损所急需的骨诱导活性。目前主要通过在支架材料中引入骨诱导信号分子的方法来赋予骨植入材料以骨诱导活性。
重组骨形态发生蛋白-2(rhBMP-2)是促进骨生成的关键因子,属于TGF-α家族成员,rhBMP-2为细胞分化成矿物质沉积成骨细胞的主要信号分子,起到诱导成骨细胞分化的作用。在肢体生长、软骨内骨化、骨折时表达,在骨骼生长发育和再生修复起重要作用,在临床治疗骨伤中具有良好的应用前景。普通载rhBMP-2的骨修复材料在产品使用前期往往会集中释放rhBMP-2,不能控制rhBMP-2持续地、微量的释放,导致一段时间内局部组织的rhBMP-2含量富集,随血流或组织液流向待修复的组织区域以外的其他组织,可能导致不必要的异位成骨。此外,rhBMP-2在体内的半衰期短,局部使用很快就被代谢和吸收,很难发挥作用,即使大剂量持续给药也往往不能提供给靶部位持续有效的浓度,对于需要长时间生长和恢复的骨骼来说无法提供良好的治疗效果,并且容易引起患者成骨质量低、炎症反应以及其他潜在的风险。因此如何控制其持续、稳定、缓慢地释放,既达到有效促骨生长的作用同时又可以避免和防止不良反应的产生是用好rhBMP-2的一项关键技术。
聚乙丙交酯(PLGA)微球由于具有较好的生物相容性和生物可降解性,是应用和研究最广泛的药物载体材料之一,美国FDA已经批准其可以作为植入性医疗器械的原材料。如本专利申请人就公开了一种载rhBMP-2的骨修复材料微球,主要用于治疗骨折等疾病,其中本申请人还公开了一种定向吸附骨组织的方法是添加贻贝粘蛋白,但是由于贻贝粘蛋白的提取纯化较为困难,价格昂贵,目前难以将其应用于大量生产。因此需要采用研发一种便于大量生产应用且同时具有贻贝粘蛋白定向吸附骨组织功能的材料。
由于rhBMP-2存在缺乏亲骨性的问题,且因骨组织如血流量低、密度大、渗透性差等生物学特性,其在骨组织中的利用率受到严重影响。目前已报道了许多用骨靶向分子直接缀合活性成分的前药,其目的是减少全身剂量并增加骨组织的活性成分浓度。然而前药存在释放时间的问题,如果靶向基团和药物共价结合过于稳定而不利于药物的释放,但如果共价键不稳定又可能在药物达到骨组织之前提前释放,亦不能得到很好的靶向性。将骨靶向基团修饰的纳米颗粒来负载治疗药物,不仅可赋予药物的靶向性,而且可控制药物的释放速度及调节药物的溶解性。
另外,PLGA微球在体内释药过程中还会降解产生大量酸性产物,并在载药微球中富集,从而造成机体环境pH下降,导致产生炎症反应的同时又影响rhBMP-2的活性。
发明内容
本发明的目的一是为了解决rhBMP-2亲骨性差而造成的在骨组织的利用度低和易产生异位成骨不良反应的问题,通过提供一种由亲骨性化合物修饰后的PLGA微球负载rhBMP-2,大大增强rhBMP-2的亲骨性和利用率;二是为了解决所采用的PLGA载体材料在体内降解形成的酸性环境引发的炎症反应,通过加入生物缓冲剂,在释药过程中稳定载药体系微环境pH值,保护rhBMP-2的生物活性。
为了实现上述目的,本发明采用了以下的技术方案:
一种亲骨性强的骨修复材料,包括rhBMP-2和PLGA微球,微球的表面通过亲骨性化合物修饰;还包括用于稳定载药体系微环境pH值的生物缓冲剂。
优选,亲骨性化合物是天冬氨酸六肽。目前亲骨性化合物主要有四环素类、双膦酸盐类和酸性寡肽类等。其中,四环素类相对不易裂解会影响药物的释放,双膦酸盐类长期聚集在骨组织中会抑制破骨细胞活性,对骨转换过程可产生严重影响。本发明选择的天冬氨酸六肽属于酸性寡肽类,与其他亲骨性化合物相比,天冬氨酸六肽具备良好的亲骨性,能定向结合到骨组织,易于与载体连接,更利于药物的释放。
优选,生物缓冲剂是4-羟乙基哌嗪乙磺酸。与传统缓冲剂相比,如常用的磷酸盐缓冲剂、柠檬酸-柠檬酸钠缓冲剂和碳酸-碳酸氢钠缓冲剂,本发明选择的生物缓冲剂属于两性离子缓冲剂,具有更适宜的pK值,缓冲能力强,化学性质稳定,无生物毒性。其中4-羟乙基哌嗪乙磺酸的pK值与哺乳动物的缓冲体系最为接近。
优选,亲骨性化合物和PLGA微球的质量比为1:5~20。
优选,PLGA微球和生物缓冲剂的质量比为1:2~6。
进一步,本申请提供了一种所述的亲骨性强的骨修复材料的制备方法,该方法包括以下步骤:
1)制备负载rhBMP-2的PLGA微球;2)亲骨性化合物修饰PLGA微球;3)加入生物缓冲剂和其他辅料混合制备成骨修复材料。
优选,步骤1)具体操作方法为:采用W/O/W法制备,把rhBMP-2冻干粉溶解在醋酸缓冲溶液中,然后加入含PLGA的乙腈,3000rpm转速乳化30秒,加入液体石蜡和吐温-80的混合乳液中,以300rpm左右的转速搅拌2-4小时,完成后,静置30分钟,再用石油醚和异丙醇交替洗涤后冻干,得到干燥的rhBMP-2的PLGA微球。
优选,步骤2)具体操作方法为:将上述制备的微球悬浮于醋酸水溶液中,缓慢搅拌加入亲骨性化合物,缓慢搅拌3-4小时,静置12小时以上,所得修饰后的微球洗涤后冻干。
优选,步骤3)具体操作方法为:亲骨性化合物修饰后的PLGA微球加入生物缓冲剂中,缓慢搅拌2小时后加入其他辅料制成骨修复材料。
进一步,本申请提供了所述的骨修复材料及其制备方法在骨再生材料中的应用。
本发明以经亲骨性化合物表面修饰、负载有rhBMP-2的PLGA微球为主要原料,具有缓释、靶向控释rhBMP-2、定向结合骨组织以及修复因子促骨生长的作用,同时还具有稳定载药微球在体内释药过程中所引起的环境pH变化,是一种缓冲能力好、亲骨性好和无不良反应的骨修复材料。相对于本申请人现有产品中添加贻贝粘蛋白来实现定向吸附骨组织的方式,本专利的骨修复材料降低了生产成本。
附图说明
图1:小鼠体内成骨试验X光片;其中(a)为2号对比样品组,(b)为3号对比样品组,(c)为实施例1成品组,(d)为空白对照组,(e)为1号对比样品组。
具体实施方式
下面通过具体实施例对本发明的技术方案作进一步描述说明。
本申请中的所有样品以实施例1成品制备方法为例,仅组成成分不同。其中以申请人CN201910896495.3号中国专利申请中的rhBMP-2骨修复材料作1号对比样品。本实施例样品组成如下表:
表1 测试样品组成
组成 | 实施例1成品 | 1号对比样品 | 2号对比样品 | 3号对比样品 |
rhBMP-2 | + | + | + | + |
PLGA微球 | + | + | + | + |
四环素 | - | - | + | - |
天冬氨酸六肽 | + | - | - | + |
贻贝粘蛋白 | - | + | - | - |
柠檬酸-柠檬酸钠 | - | - | - | + |
4-羟乙基哌嗪乙磺酸 | + | - | + | - |
其他辅料 | + | + | + | + |
注:“+”表示含有某成分,“-”表示不含某成分。
实施例1 骨修复材料制备
主要实验材料:
rhBMP-2由企业自行生产,具体生产过程参照申请人CN201910030649.0号中国专利申请中的生产方法(大肠杆菌中表达,离子交换层析纯化,检测大肠杆菌宿主残留蛋白<0.005%),抑菌圈法检测抗生素残留<0.1ppm。聚乙丙交酯(PLGA)由美国DURECT公司生产。柠檬酸-柠檬酸钠、4-羟乙基哌嗪乙磺酸、天冬氨酸六肽和四环素均购自市售产品。
制备方法:
负载rhBMP-2的PLGA微球制备:采用W/O/W法制备,把rhBMP-2冻干粉溶解在醋酸缓冲溶液中,然后加入含PLGA的乙腈,3000rpm转速乳化30秒,加入液体石蜡和吐温-80的混合乳液中,以300rpm左右的转速搅拌2-4小时,完成后,静置30分钟,再用石油醚和异丙醇交替洗涤后冻干,得到干燥的rhBMP-2的PLGA微球;
亲骨性化合物修饰PLGA微球:将上述制备的微球悬浮于醋酸水溶液中,缓慢搅拌加入亲骨性化合物,缓慢搅拌3-4小时,静置12小时以上,所得修饰后的微球洗涤后冻干;
骨修复材料制备:亲骨性化合物修饰后的PLGA微球加入生物缓冲剂中,缓慢搅拌2小时后加入其他辅料制成骨修复材料。
实施例2 亲骨性评价
羟基磷灰石(nHAP)吸附法评价体外亲骨性:
具体方法是以纳米羟基磷灰石粉末作为吸附介质,使其与一定浓度(Ca)样品充分作用,除去吸附介质后用BCA法测定出平衡后样品浓度(Cb)。样品与羟基磷灰石亲和性的大小通过吸附率进行表征。计算公式为:
吸附率=(Ca-Cb)/Ca×100%。
nHAP吸附实验:
精密吸取25ug/mL的待测样品溶液5mL,分别加入精密称取的nHAP 30mg,设三个平行组,另设1×PBS溶液为空白对照组。超声分散5分钟,室温下分别剧烈振摇90分钟,取溶液过滤,洗涤滤饼并收集所有滤液,BCA法检测并计算滤液中未吸附的样品浓度值(ug/mL),根据上述公式计算吸附率。
表2 样品nHAP吸附率
样品 | 实施例1成品 | 1号对比样品 | 2号对比样品 |
吸附率(%) | 55.24±2.01 | 28.44±1.05 | 56.70±1.99 |
从表2的实验数据可以得到,经过亲骨性化合物修饰后的实施例1成品和2号对比样品的nHAP吸附率都较高,说明天冬氨酸六肽和四环素的亲骨能力相当。而未修饰的1号对比样品的吸附率较低,说明采用亲骨性化合物来修饰PLGA微球,能够明显增强载体材料对骨组织的定向结合作用。
实施例3 体外释放实验
将实施例1成品和1-3号对比样品置于透析袋中,透析袋置于20mL含0.2%叠氮化钠,pH7.0的PBS缓冲液中。静置于37℃下,在12h、24h、120h、168h、336h、504h、672h、840h分别取样1mL(随后补充含0.2%叠氮化钠,pH7.0的PBS缓冲液1mL),按照试剂盒的说明以ELISA法测定rhBMP-2浓度,计算累计释放百分数。结果如下表所示:
表3 rhBMP-2累计释放百分数
从表3实验数据可以得到,实施例1样品相较于1号对比样品的缓释时间略有延长,更适应骨修复的生长周期。3号对比样品与实施例1成品区别仅在于缓冲剂不同,其释放率较高,接近1号对比样品,说明添加4-羟乙基哌嗪乙磺酸作缓冲剂也可以提高其缓释效果。2号对比样品的rhBMP-2释放率较低,说明四环素修饰的微球不易降解和释放rhBMP-2,不利于骨修复时期的愈合。
实施例4 体内缓冲性能和骨愈合能力评价
分组设计:将10只正常ICR或KM雄性小鼠随机分为5组,每组2只;其中空白对照1组。
实验处理:小鼠用6%戊巴比妥钠麻醉后,后肢股部去毛消毒,切开皮肤,分离肌间隙,植入不同的样品,并加一定的抗生素防止感染,然后逐层缝合肌肉和皮肤,消毒创口,正常饲养。
大体观察:观察不同实验组有无不良反应。结果得到实施例1成品无任何不良反应,而1号和3号对比样品均出现不同程度的炎症,说明添加4-羟乙基哌嗪乙磺酸能够有效稳定PLGA微球载药体系微环境的pH值,从而避免酸性环境导致的炎症反应。
实验观察:埋植实验完成四周后,将小鼠进行放射学检查(X光),检查结果如图1所示,查看小鼠后肢是否出现骨组织。四周进行放射学检查后,将小鼠处死,去除埋植区的骨组织,称量里面所含有骨的湿重,并进行HE染色组织学评估。
X射线观察骨愈合评价标准:
1. 1962柴本甫骨折愈合衡量标准
2. Lane-Sandhu X射线及组织学评分标准:
实验结果如下表:
表4 1962柴本甫骨折愈合衡量表
样品 | 断端边缘 | 骨膜反应 | 骨痂量 | 骨痂密度 | 骨痂边缘 |
实施例1成品 | ++++ | ++++ | ++++ | ++++ | ++++ |
1号对比样品 | ++++ | ++++ | ++++ | ++++ | ++++ |
2号对比样品 | +++ | +++ | ++++ | ++++ | +++ |
3号对比样品 | ++ | +++ | ++ | ++ | ++ |
空白对照 | + | + | + | + | + |
表5 Lane-Sandhu X射线及组织学评分
样品 | 骨形成 | 骨连接 | 骨塑形 |
实施例1成品 | 4 | 4 | 4 |
1号对比样品 | 4 | 4 | 4 |
2号对比样品 | 4 | 3 | 3 |
3号对比样品 | 3 | 3 | 3 |
空白对照 | 1 | 2 | 1 |
从实验数据可以得到,实施例成品1的促骨生成性能较好,体内骨修复过程中无不良反应,说明采用天冬氨酸六肽修饰PLGA微球,并加入4-羟乙基哌嗪乙磺酸生物缓冲剂可以较好地提高rhBMP-2的骨组织利用度并大大降低炎症发生率,具有很好的应用前景。
Claims (10)
1.一种亲骨性强的骨修复材料,其特征在于,包括rhBMP-2和PLGA微球,微球的表面通过亲骨性化合物修饰;还包括用于稳定载药体系微环境pH值的生物缓冲剂。
2.根据权利要求1所述的亲骨性强的骨修复材料,其特征在于,亲骨性化合物是天冬氨酸六肽。
3.根据权利要求1所述的亲骨性强的骨修复材料,其特征在于,生物缓冲剂是4-羟乙基哌嗪乙磺酸。
4.根据权利要求1所述的亲骨性强的骨修复材料,其特征在于,亲骨性化合物和PLGA微球的质量比为1:5~20。
5.根据权利要求1所述的亲骨性强的骨修复材料,其特征在于,PLGA微球和生物缓冲剂的质量比为1:2~6。
6.一种权利要求1所述的亲骨性强的骨修复材料的制备方法,其特征在于,该方法包括以下步骤:
1)制备负载rhBMP-2的PLGA微球; 2)亲骨性化合物修饰PLGA微球;3)加入生物缓冲剂和其他辅料混合制备成骨修复材料。
7.根据权利要求6所述的亲骨性强的骨修复材料的制备方法,其特征在于,步骤1)具体操作方法为:采用W/O/W法制备,把rhBMP-2冻干粉溶解在醋酸缓冲溶液中,然后加入含PLGA的乙腈,3000rpm转速乳化30秒,加入液体石蜡和吐温-80的混合乳液中,以300rpm左右的转速搅拌2-4小时,完成后,静置30分钟,再用石油醚和异丙醇交替洗涤后冻干,得到干燥的rhBMP-2的PLGA微球。
8.根据权利要求6所述的亲骨性强的骨修复材料的制备方法,其特征在于,步骤2)具体操作方法为:将上述制备的微球悬浮于醋酸水溶液中,缓慢搅拌加入亲骨性化合物,缓慢搅拌3-4小时,静置12小时以上,所得修饰后的微球洗涤后冻干。
9.根据权利要求6所述的亲骨性强的骨修复材料的制备方法,其特征在于,步骤3)具体操作方法为:亲骨性化合物修饰后的PLGA微球加入生物缓冲剂中,缓慢搅拌2小时后加入其他辅料制成骨修复材料。
10.权利要求1-9任一所述的骨修复材料及其制备方法在骨再生材料中的应用。
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202111398397.0A CN114209890A (zh) | 2021-11-19 | 2021-11-19 | 一种亲骨性强的骨修复材料及其制备方法 |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202111398397.0A CN114209890A (zh) | 2021-11-19 | 2021-11-19 | 一种亲骨性强的骨修复材料及其制备方法 |
Publications (1)
Publication Number | Publication Date |
---|---|
CN114209890A true CN114209890A (zh) | 2022-03-22 |
Family
ID=80698133
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN202111398397.0A Pending CN114209890A (zh) | 2021-11-19 | 2021-11-19 | 一种亲骨性强的骨修复材料及其制备方法 |
Country Status (1)
Country | Link |
---|---|
CN (1) | CN114209890A (zh) |
Citations (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN102643330A (zh) * | 2011-07-09 | 2012-08-22 | 广东医学院 | 一种骨靶向药物天冬氨酸六肽丹参素的合成方法及其医疗用途 |
CN108586601A (zh) * | 2018-05-14 | 2018-09-28 | 四川大学 | 一种骨靶向性鲑鱼降钙素及其制备方法 |
CN110522946A (zh) * | 2019-09-20 | 2019-12-03 | 浙江瑞谷生物科技有限公司 | 一种载rhBMP-2的骨修复材料微球及其制备方法 |
CN110755638A (zh) * | 2019-10-30 | 2020-02-07 | 西南交通大学 | 一种具有骨靶向性的药物载体及其制备方法与应用 |
US20210212949A1 (en) * | 2018-04-23 | 2021-07-15 | Board Of Regents Of The University Of Nebraska | Nanofiber microspheres and methods os use thereof |
-
2021
- 2021-11-19 CN CN202111398397.0A patent/CN114209890A/zh active Pending
Patent Citations (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN102643330A (zh) * | 2011-07-09 | 2012-08-22 | 广东医学院 | 一种骨靶向药物天冬氨酸六肽丹参素的合成方法及其医疗用途 |
US20210212949A1 (en) * | 2018-04-23 | 2021-07-15 | Board Of Regents Of The University Of Nebraska | Nanofiber microspheres and methods os use thereof |
CN108586601A (zh) * | 2018-05-14 | 2018-09-28 | 四川大学 | 一种骨靶向性鲑鱼降钙素及其制备方法 |
CN110522946A (zh) * | 2019-09-20 | 2019-12-03 | 浙江瑞谷生物科技有限公司 | 一种载rhBMP-2的骨修复材料微球及其制备方法 |
CN110755638A (zh) * | 2019-10-30 | 2020-02-07 | 西南交通大学 | 一种具有骨靶向性的药物载体及其制备方法与应用 |
Non-Patent Citations (5)
Title |
---|
CTJAB, ET AL: "Poly aspartic acid peptide-linked PLGA based nanoscale particles: Potential for bone-targeting drug delivery applications", 《INTERNATIONAL JOURNAL OF PHARMACEUTICS》 * |
傅超美等: "《全国普通高等中医药院校药学类专业十三五规划教材 第二轮规划教材 中药药剂学 第2版》", 31 August 2018, 北京:中国医药科技出版社 * |
庄倩倩等: "《生物降解高分子材料及其应用现状研究》", 30 November 2019, 中国纺织出版社有限公司 * |
王志鑫等: "《骨折愈合学》", 31 August 1995, 武汉:湖北科学技术出版社 * |
边照阳等: "《 QuEChERS技术及应用》", 31 December 2017, 北京:中国轻工业出版社 * |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
Chang et al. | Liposomal dexamethasone–moxifloxacin nanoparticle combinations with collagen/gelatin/alginate hydrogel for corneal infection treatment and wound healing | |
Mi et al. | Injectable nanoparticles/hydrogels composite as sustained release system with stromal cell-derived factor-1α for calvarial bone regeneration | |
WO2010083778A1 (zh) | 注射用肺靶向脂质体药物组合物 | |
CN101756908A (zh) | 聚酯包衣的羟基磷灰石微球及其制备方法 | |
CN110522946B (zh) | 一种载rhBMP-2的骨修复材料微球及其制备方法 | |
CN103330680A (zh) | 纳米药物透皮制剂及其制备方法 | |
CN110339168B (zh) | 一种负载抗肺纤维化药物和免疫调节剂的纳米制剂及其制备方法 | |
CN109432498B (zh) | 一种用于骨结核治疗的骨修复支架及其制备方法 | |
CN111317709A (zh) | 一种可注射载双药物复合壳聚糖水凝胶及其制备方法 | |
Chen et al. | Preparation of recombinant human bone morphogenetic protein-2 loaded dextran-based microspheres and their characteristics | |
Chen et al. | Preparation and biological characteristics of recombinant human bone morphogenetic protein-2-loaded dextran-co-gelatin hydrogel microspheres, in vitro and in vivo studies | |
Chen et al. | Oxidative Stimuli‐Responsive “Pollen‐Like” Exosomes from Silver Nanoflowers Remodeling Diabetic Wound Microenvironment for Accelerating Wound Healing | |
CN104288093A (zh) | 纳米药物透皮制剂在肿瘤中的应用 | |
CN113651959A (zh) | 一种基于氨基酸-羟基酸共聚物的纳米载药体系及其制备方法和应用 | |
AU2008339100A1 (en) | Drug delivery system for administration of a water soluble, cationic and amphiphilic pharmaceutically active substance | |
JPH04507412A (ja) | 薬学上の化合物 | |
CN114209890A (zh) | 一种亲骨性强的骨修复材料及其制备方法 | |
CN111870739A (zh) | 一种多功能改性壳聚糖自愈合水凝胶的制备方法及应用 | |
Rasouli et al. | Multifunctional Hydroxyapatite-based Nanoparticles for Biomedicine: Recent Progress inDrug Delivery and Local Controlled Release | |
CN115105629B (zh) | 一种抗菌水凝胶及其制备方法和应用 | |
JP2011507840A (ja) | 水溶性、カチオン性および両親媒性の薬学的に活性な物質を投与するためのドラッグ・デリバリー・システム | |
CN114225046A (zh) | 一种抗菌/促愈合的双药递送系统、药物组合物及其制备方法和应用 | |
CN113995714A (zh) | 一种顺铂交联的蛋白水凝胶及制备方法 | |
Lee et al. | Preparation of collagen modified hyaluronan microparticles as antibiotics carrier | |
US20170112777A1 (en) | Protein-based particles for drug delivery |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
PB01 | Publication | ||
PB01 | Publication | ||
SE01 | Entry into force of request for substantive examination | ||
SE01 | Entry into force of request for substantive examination | ||
RJ01 | Rejection of invention patent application after publication |
Application publication date: 20220322 |
|
RJ01 | Rejection of invention patent application after publication |