以交联透明质酸钠为支架体外构建组织工程软骨  

Construction of tissue-engineered cartilage with cross-linked sodium hyaluronate as scaffold materials in vitro

作　　者：刘少英[1] 陈建英[1] 陈倩倩[1] 刘飞[1] 刘霞[1] 朱希强[1] 凌沛学[1,2] 

机构地区：[1]山东省生物药物研究院山东省生物药物重点实验室,山东省济南市250101 [2]山东福瑞达医药集团公司山东省黏膜与皮肤给药技术重点实验室,山东省济南市250101

出　　处：《中国组织工程研究》2014年第8期1191-1197,共7页Chinese Journal of Tissue Engineering Research

基　　金：山东省优秀中青年科学家科研奖励基金(BS2011YY050);济南市高校院所自主创新计划(201202038)~~

摘　　要：背景:采用组织工程技术再生和重建软骨是目前修复软骨组织缺损效果最好、最有应用前景的方法。目的:以体外培养的软骨细胞和交联透明质酸钠为支架材料,开发一套体外构建组织工程软骨的完整方案。方法:分离新西兰兔膝关节软骨细胞,制成细胞悬液滴加于交联透明质酸钠支架上,体外复合培养21 d,提取RNA进行RT-PCR检测,制备冰冻切片进行显微观察和免疫组织化学观察。结果与结论:软骨细胞接种于交联透明质酸钠支架材料后,可贴附于支架上生长,并且大量细胞聚集成团,在支架材料的纤维间隙中生长或呈单层细胞附着于支架材料纤维。细胞-支架复合物表达软骨组织特异性蛋白聚糖基因和Ⅱ型胶原α1基因,以及软骨组织特异性蛋白Ⅱ型胶原蛋白,可维持软骨细胞表型。表明培养的细胞-支架复合物在体外培养可形成软骨细胞外基质,有望获得组织工程软骨组织。BACKGROUND：Tissue engineering provides new ideas and approaches for repair of cartilage defects. OBJECTIVE：To develop a complete set of solutions for construction of tissue engineered cartilagein vitro, with chondrocytes as seed cels and cross-linked sodium hyaluronate as scaffold materials. METHODS： New Zealand rabbit articular chondrocytes were isolated, counted, and then cultured and passaged to prepare cellsuspension. Toluidine blue staining, RT-PCR and immunocytochemistry were exerted to evaluate the cultured cels. Chondrocytes were seeded and co-cultured with cross-linked sodium hyaluronate scaffold for 21 days. Then, RNA was isolated for RT-PCR, and frozen sections were prepared for morphological observation and immunohistochemistry study. RESULTS AND CONCLUSION：The chondrocytes could adhere to the cross-linked sodium hyaluronate scaffold and aggregate, growing between fibers or adhering to the scaffold in a monolayer manner. The transcripts of cartilage specific aggrecan gene and colagen type II alpha 1 gene and cartilage specific protein colagen type II were expressed in cel-scaffold complexes to maintain the phenotype of chondrocytes. Cel-scaffold complexes co-cultured in vitro can form cartilage extracelular matrix, by which tissue engineered cartilage is expected to be obtained.

关 键 词：生物材料 软骨生物材料 组织工程 关节软骨 支架材料 软骨细胞 分离培养 免疫细胞化学 RT-PCR 形态学观察 交联透明质酸钠 体外 

分 类 号：R318[医药卫生—生物医学工程]