羟基丁酸与羟基辛酸共聚物一体化骨软骨组织工程支架的制备及性能研究

发布时间：2018-07-30 08:37

【摘要】：关节骨软骨承载了体重、滑动、减震等生理功能,在动物体中起着非常重要的作用,同时由于软骨组织缺乏血管、淋巴细胞,导致了关节软骨损伤很难自愈。经多年临床研究,建立了包括抛削术、清创术、异体移植术和自体移植术在内的传统治疗方法。然而,这些方法的治疗效果尚不能令人满意。抛削术、清创术会造成二次伤害,且新生软骨易发生退化；异体软骨移植会引起免疫反应,导致损伤修复失败；自体软骨移植受材料来源限制,而不能用于大面积软骨损伤的修复。因此,关节骨软骨损伤的治疗成为世界性的医疗难题,组织工程研究为关节骨软骨修复带来了希望。 目前,关节组织工程研究集中在种子细胞、支架制备、细胞生长因子三个方面,其中制备支架是开展此项研究最基本的条件。在以往骨组织工程支架的研究中研制出了多种骨或软骨单层支架,单层软骨支架忽视了软骨下骨层作为软骨支撑点并为软骨提供营养的重要功能。为此,必需设计一种仿生骨软骨一体化支架以满足骨软骨损伤组织工程修复的需要。 本文进行了羟基丁酸与羟基辛酸共聚物(PHBHOx)一体化骨软骨支架的研制,从乳兔提取了骨髓间充质干细胞,并初步进行了支架与细胞的复合培养研究。 采用溶剂浇注/颗粒沥滤法,以三氯甲烷为溶剂、PHBHOx为主材料和连续相,通过分别添加不同粒径与比例致孔剂,一次性制备由骨层-中间过渡层-软骨层组成的一体支架。仿照骨软骨解剖学特征与物理学特性,一体支架的各层应该结构不同,性能有别,且三层之间彼此连通。经过制备条件优化,成功制备出骨软骨一体化支架,其三层孔径分别约为280μ m,200μ m,≤60μ m,孔隙率分别约为75%,85%,60%,其力学强度分别为4.23MPa,2.75MPa,6.30MPa,基本上达到了组织工程修复骨软骨损伤的要求。 本文采用全骨髓法提取兔骨髓间充质干细胞(BMSCs),用贴壁法将BMSCs提纯。经过72小时的培养,贴壁率达到了60%,六天后BMSCs布满瓶底,细胞生长状态旺盛,细胞形态稳定。 本文还进行了一体支架与BMSCs细胞的复合培养试验,在24h后,细胞粘附率达到了76%。细胞形态良好,表明了一体化支架具有良好的生物相容性。 本文应用专利材料PHBHOx首次成功制备了仿生骨层-中间过渡层-软骨层相连续的一体化支架,建立了分步操作一次成型支架制作新工艺,获得了三层结构不同、功能有别的骨软骨组织工程支架新产品。并进行了一体支架与BMSCs细胞的复合培养试验,证明了该支架有着良好的生物相容性。本文的研究进展为关节损伤修复的深入研究提供了一定的理论和实验基础。
[Abstract]:Articular bone cartilage bears physiological functions such as weight sliding shock absorption and plays a very important role in animals. At the same time because of the lack of blood vessels and lymphocytes in cartilage tissue it is difficult to heal the injury of articular cartilage. After many years of clinical research, traditional treatments including excision, debridement, allograft and autologous transplantation have been established. However, the therapeutic effect of these methods is not satisfactory. Dissection, debridement will cause secondary injury, and new cartilage will degenerate easily; allogenic cartilage transplantation will cause immune response, resulting in damage repair failure; autologous cartilage transplantation is limited by the source of materials. It can not be used to repair large area cartilage injury. Therefore, the treatment of articular cartilage injury has become a worldwide medical problem, tissue engineering research has brought hope for articular cartilage repair. At present, the research of joint tissue engineering focuses on seed cells, scaffold preparation and cell growth factor, among which the preparation of scaffolds is the most basic condition for this study. In previous studies of bone tissue engineering scaffolds, a variety of bone or cartilage monolayer scaffolds have been developed. Monolayer cartilage scaffolds have neglected the important function of subchondral bone layer as the supporting point of cartilage and providing nutrition for cartilage. Therefore, a biomimetic osteochondral scaffold must be designed to meet the need of tissue engineering repair of bone cartilage injury. The bone marrow mesenchymal stem cells (BMSCs) were extracted from neonatal rabbits using hydroxybutyric acid and hydroxyoctanoic acid copolymers (PHBHOx) as bone cartilage scaffolds. The co-culture of bone marrow mesenchymal stem cells (BMSCs) was studied in this paper. A monolithic scaffold composed of bone layer, intermediate transition layer and cartilage layer was prepared by solvent pouring / particle leaching method, using trichloromethane as solvent PHBHOx as main material and continuous phase, respectively, by adding different particle size and proportion of pore-forming agent. According to the anatomical and physical characteristics of osteochondral, each layer of the monolithic scaffold should have different structure and performance, and the three layers should be connected with each other. After optimization of the preparation conditions, the osteochondral scaffolds were successfully prepared. The three-layer pore sizes of the scaffolds were about 280 渭 m ~ 200 渭 m, 鈮，

本文编号：2154436