兔骨髓间充质干细胞/壳聚糖-胶原支架复合体构建软骨组织工程支架  

Construction of cartilage tissue engineered scaffold by rabbit bone marrow mesenchymal stem cells/chitosan-collagen composite

作　　者：初殿伟 王玮[2] 陈立军[2] 王文良[3] 张华亮[3] 杨亮[3] 李建鑫[3] 

机构地区：[1]武装警察部队天津总队医院骨科,天津市300162 [2]武警医学院生物化学教研室,天津市300162 [3]武警医学院附属医院骨科,天津市300162

出　　处：《中国组织工程研究与临床康复》2009年第42期8248-8252,共5页Journal of Clinical Rehabilitative Tissue Engineering Research

基　　金：国家自然科学基金资助项目(30740089)~~

摘　　要：背景:软骨组织工程的核心是利用少量的细胞经体外培养、扩增后,在一定环境下附着在三维多孔支架上,再将细胞/支架复合体移植到体内形成新的组织。目的:拟构建兔骨髓间充质干细胞/壳聚糖-胶原支架复合体,探讨该支架作为软骨组织工程支架的可行性。设计、时间及地点:细胞-支架学体外观察,于2008-03/2009-02在武警医学院生物化学教研室完成。材料:日本大耳白兔6只用于分离培养骨髓间充质干细胞。医用壳聚糖粉末为山东奥康生物科技有限公司产品。Ⅰ型胶原为Sigma公司产品。方法:取3%的壳聚糖和2%的胶原混合的醋酸溶液,倒入72孔板内,-20℃预冷冻10h,冻干机冷冻抽干24h制作成壳聚糖-胶原支架。取第3代兔骨髓间充质干细胞,以1×109L-1密度接种于支架内,构建细胞/支架复合体。主要观察指标:傅里叶红外光谱、扫描电镜、液体置换法测定支架的理化性质,观察三维培养后细胞在支架上的生长情况。结果:壳聚糖-胶原支架孔径为160～380μm,平均为270μm,孔相通性好,支架孔隙率为(86.00±5.12)%。傅里叶红外光谱仪测定数据表明复合支架组成物有典型的壳聚糖、胶原峰,未发现有聚乙二醇峰。细胞/支架共培养24,48,72h后,骨髓间充质干细胞可渗入支架多孔结构内,并黏附在支架上成簇生长,部分细胞已与支架融合。结论:壳聚糖-胶原支架基本符合软骨组织工程支架要求,能够作为种子细胞的承载体。BACKGROUND： The core of cartilage tissue engineering is that a few of cells following in vitro culture and amplification are attached to three-dimensional porous scaffold, and cell/scaffold composite is then implanted to form new tissues. OBJECTIVE： To study the feasibility of rabbit bone marrow mesenchymal stem cells （BMSCs）/chitosan-collagen scaffolds to considered as cartilage tissue engineered scaffold. DESIGN, TIME AND SETTING： An in vitro cell-scaffold observation was performed at Research Room of Biochemistry, Medical College of Chinese Peoples Armed Police Force from March 2008 to February 2009. MATERIALS： Six Japanese rabbits were used for separation and culture BMSCs. Chitosan was provided by Aokang Biotechnology Co., Ltd., Shangdong; type I collagen was provided by Sigma, USA. METHODS： 3% chitosan and 2%collagen were mixed in acetic acid solution, and the mixture was then put in 72-well plate, pre-frozen at -20 ℃ for 10 hours, and frozen and dried for 24 hours to obtain chitosan-collagen scaffold. The third-passage rabbit BMSCs were inoculated in the scaffold at the density of 1×109/L to construct cell/scaffold composite. MAIN OUTCOME MEASURES： Physico-chemical properties of scaffold were detected using Fourier transform infrared spectroscopy, scanning electron microscopy, and liquid substitution method, while cell growth was observed after three-dimensional culture. RESULTS： Pore of chitosan-collagen scaffold was 160-380 μm, with the mean value of 270 μm. the permeability was well, and the porosity was （86.00±5.12）%. Fourier transform infrared spectroscopy indicated that the products in the cell/scaffold composite group contained chitosan and collagen peaks, but polyethylene glycol peak was not found. At 24, 48, and 72 hours after cell/scaffold co-culture, BMSCs permeated into porous structure of the scaffold and adhered to the scaffold. Partial BMSCs were in fusion with the scaffold. CONCLUSION： The novel chitosan-collagen scaffold is satisfactory for cartilage tissue en

关 键 词：壳聚糖-胶原支架 骨髓间充质干细胞 冻干法 软骨组织工程 

分 类 号：R318[医药卫生—生物医学工程]