兔关节软骨块和骨髓间充质干细胞共培养  

Co-culture of bone marrow mesenchymal stem cells with articular cartilage block in rabbits

作　　者：崔小明 吴佳奇 熊小天 华强[4] 左宇 石厚银 CUI Xiao-ming;WU Jia-qi;XIONG Xiao-tian;HUA Qiang;ZUO-Yu;SHI Hou-Yin(Department of Orthopaedics,Yongchuan Traditional Chinese Medicine Hospital,Chongqing 402160,China;Department of Orthopaedics,The Affiliated TCM Hospital,Southwest Medical University,Luzhou 646000,China;Chongqing Orthopaedic Hospital of Traditional Chinese Medicine,Chongqing 643000,China;Department of Orthopaedics,Sport Hospital,Chengdu Sport University,Chengdu 610041,China)

机构地区：[1]重庆市永川区中医院骨科,重庆市402160 [2]西南医科大学附属中医医院骨科,四川省泸州市646000 [3]重庆市中医骨科医院,重庆市643000 [4]成都体育学院附属体育医院骨科,四川省成都市610041

出　　处：《中国矫形外科杂志》2020年第4期349-355,共7页Orthopedic Journal of China

基　　金：四川省卫计委资助项目(编号:130280)。

摘　　要：[目的]探讨与兔关节软骨块共培养对兔骨髓间充质干细胞(RBMSCs)分化的影响。[方法]全骨髓贴壁培养法分离培养RBMSCs,对P3代细胞进行氯甲基苯甲酰氨荧光染料(CM)标记。用4周龄雄性新西兰大白兔股骨头关节面制备软骨块。分为3组,分别为单纯细胞组、细胞+软骨组和单纯软骨组,于第7、14、21 d细胞爬片,行甲苯胺蓝染色法检测和II型胶原免疫组化染色。[结果]经过3周培养,单纯细胞组细胞仍保持骨髓间充质干细胞的特点;细胞+软骨组RBMSCs与兔关节软骨块混合共培养后,RBMSCs细胞形态由长梭形逐渐变为短小的三角形或不规则形,表现出典型的"铺路石"样改变;甲苯胺蓝染色阳性和II型胶原免疫组化阳性,符合软骨细胞特点;单纯软骨组软骨块在培养液中成活,无细胞生长。[结论]兔关节软骨块所营造的微环境能够促进RBMSCs向软骨细胞方向分化,此种RBMSCs能够在兔关节软骨块表面贴附生长。[Objective] To explore the effect of co-culture of rabbit bone marrow mesenchymal stem cells(RBMSCs) with articular cartilage blocks on the differentiation of the cells in rabbits. [Methods] The RBMSCs was obtained from whole bone marrow by adherent culture isolation. The 3 rdgeneration of cells were labeled with chloromethyl-benzamidodial-kyl carbocyanine(CM), and further cultured in 3 groups, including the RBMSCs only, the co-culture of RBMSCs with 2 cartilage blocks which harvested from a 4-week-old male New Zealand white rabbit and the articular cartilage blocks only. The toluidine blue staining and immunohistochemical staining for collagen II were conducted on the cell-slips at 7, 14 and 21 days. [Results] At21 days, the cells in the RBMSCs only group remained the characteristics of bone marrow mesenchymal stem cells with negative results of toluidine blue staining and immunohistochemistry staining for type II collagen. By contrast, the cells in the co-culture group became short triangular or irregular shaped from long spindle shaped with positive results of toluidine blue staining and immunohistochemistry staining for type II collagen, adhered and grew on the surface of cartilage block. However, the tissue in the cartilage block only group remained survival without cell outgrowth. [Conclusion] The microenvironment in co-culture with articular cartilage block does improve chondrogenic differentiation of RBMSCs, which can adhere and grow on the surface of cartilage block in this experimental study.

关 键 词：软骨 骨髓间充质干细胞 软骨块 共培养 

分 类 号：R318[医药卫生—生物医学工程]