间充质源性软骨种子细胞与脱细胞软骨复合培养构建软骨组织  

Construction of cartilage with cocultures of mesenchymal-derived cartilage seed cells and acellular allogenic cartilage in vitro

作　　者：刘萌萌[1] 栾保华[1] 孙良[1] 李中华[2] 王小霞 

机构地区：[1]山东大学附属省立医院烧伤整形科,济南250021 [2]济南市第四人民医院烧伤科,济南250021 [3]济南护理职业学院,济南250000

出　　处：《山东大学学报（医学版）》2011年第7期24-28,34,共6页Journal of Shandong University:Health Sciences

基　　金：山东省自然科学基金资助项目(Y2002C19)

摘　　要：目的兔外周血间充质源性软骨种子细胞与同种异体脱细胞软骨体外复合培养,构建新型软骨组织,为临床修复器官缺损及再造提供实验基础。方法提取兔外周血间充质干细胞,行体外诱导培养14 d获得软骨种子细胞,显微镜下观察细胞生长特性,制作细胞爬片行甲苯胺蓝染色、Ⅱ型胶原免疫细胞化学染色。联合去垢剂—酶法获得兔脱细胞肋软骨支架,软骨种子细胞与同种异体脱细胞软骨支架体外复合培养7 d获得复合软骨,脱细胞软骨及复合软骨固定后行扫描电镜观察及组织学HE、甲苯胺蓝染色。结果兔外周血间充质干细胞体外诱导培养14 d,胞浆内较多软骨细胞特异性Ⅱ型胶原分泌;兔脱细胞软骨呈乳白色,结构完整、孔隙均匀,仍保存大量酸性黏多糖及胶原成分,其孔隙率(61.31±8.45)%,孔径长度(32.80±5.15)μm。支架与软骨细胞黏附良好,并伴有多量基质分泌。结论间充质源性软骨种子细胞与同种异体脱细胞软骨体外复合培养可构建新型软骨组织,本研究为临床修复器官缺损及再造提供了实验依据。Objective To build new cartilage tissues by means of co-cultures of rabbit mesenchymal-derived cartilage seed cells and allograft acellular cartilage cells in vitro,and to provide scientific basis for repair and reconstruction of organ defects.Methods Rabbit peripheral blood stem cells were extracted and induced in vitro for 14 days.Then,cartilage seed cells were harvested.Characteristics of cell growth were observed under an inverted phase contrast microscope.Toluidine blue and Type Ⅱ collagen immunocytochemistry staining were applied on cellular creep plates.Acellular allogenic rib cartilage was obtained by the detergent-enzyme method.Composite cartilage was collected after 7 days of co-culturing of cartilage seed cells and acellular allogenic cartilage in vitro.Acellular cartilage and composite cartilage were fixed with HE and toluidine blue staining for observation,as well as scanning electron micrographs（SME）.Results After 14 days of induction,lots of chondrocyte-specific type Ⅱ collagens were expressed in the cytoplasm of rabbit peripheral blood mesenchymal stem cells.Rabbit acellular allogenic cartilage was milky white and integrally structured,in which the pores were homogeneous and large amounts of acid mucopolysaccharide and collagen remained.The porosity was（61.31±8.45）% and aperture length was（32.80±5.15） μm.Chondrocytes well adhered to the stent,and large amounts of matrix were secreted by the composite cartilage.Conclusion Cartilage tissue can be constructed by means of co-culturing of mesenchyma-derived cartilage seed cells and acellular allogenic cartilage in vitro,which may provide an experimental basis for organ defect repair and reconstruction.

关 键 词：间充质 诱导 脱细胞 软骨组织 

分 类 号：R318[医药卫生—生物医学工程]