新型旋转壁式生物反应器内三维组织工程骨的构建  

Three-dimensional Fabrication of Engineered Bone in Rotating Wall Vessel Bioreactor

作　　者：宋克东[1] 刘天庆[1] 李香琴[1] 崔占峰 葛丹[1] 孙相玉[1] 马学虎[1] 

机构地区：[1]大连理工大学精细化工国家重点实验室,干细胞与组织工程研究室,大连116024 [2]Department of Engineering Science,Oxford University,Parks Road,Oxford OX1 3PJ,UK

出　　处：《生物化学与生物物理进展》2004年第11期996-1005,共10页Progress In Biochemistry and Biophysics

基　　金：国家自然科学基金两个基地国际合作项目 ( 2 0 0 2 0 0 8);辽宁省科学技术基金资助项目 ( 2 0 0 2 2 14 0 )~~

摘　　要：利用微载体悬浮培养法将成骨细胞在旋转壁式生物反应器内进行大规模扩增 ,并检测细胞的组织形态和生物功能 .然后以此作为种子细胞 ,分别以 2× 10 6个 /ml和 1× 10 6个 /ml两种密度接种到支架材料上 ,于旋转壁式生物反应器 (RWV)内进行三维组织工程骨的构建 .并将所构建的骨组织分别进行倒置显微镜 (invertedmicroscope)、扫描电镜 (SEM )、碱性磷酸酶 (ALP)、矿化结构和AO/EB双重荧光染色等生物学性能检测 ,以及对培养过程的营养物质代谢情况进行监控和分析 .结果表明 ,在RWV中培养的骨组织生长良好 ,分泌大量胶原纤维 ,并有矿化基质和新骨样组织形成 .由上述结果可断定 ,通过RWV内部流体对流所产生的应力刺激 ,可提高成骨细胞碱性磷酸酶的活性表达 ,并加速矿化结节的形成 ,从而完成成骨细胞的快速增殖与分化以及工程化组织的三维构建 .The osteoblasts were expanded in large-scale by microcarrier suspension culture in rotating wall vessel bioreactor (RWV), then the biological functions of the cells were detected by histomorphometry and the cells were seeded at 2 X 10(6) cells/ml and 1 X 10(6) cells/ml onto scaffolds respectively to culture for 2 weeks. The biological properties of the fabricated bone tissue were detected by inverted microscope, scanning electron microscope (SEM), alkaline phosphatase (ALP), von-Kossa staining on mineralized nodules and AO/EB fluorescence staining. Meanwhile the cells' metabolism of nutrients was monitored and analyzed during the whole culture process. The bone tissue fabricated in RWV with two different seeded cell densities grew well. Osteoblasts in the scaffolds secreted much collagen fibers and formed mineralized nodules and new osteoid tissue. It can be conclueled that: With the stress stimulation inside the fluid in the RWV, the active expression of ALP can be increased, the formation of mineralized nodules can be accelerated. The rapid proliferation and differentiation of osteoblasts are possible and the three-dimensional fabrication of engineered bone could be realized.

关 键 词：组织工程骨 成骨细胞 生物反应器 支架材料 细胞培养 

分 类 号：R782[医药卫生—口腔医学]