兔骨髓源成骨细胞作为骨组织工程实验细胞的研究  

Rabbit Marrow Stromal Osteoblast Used as Seed Cell of Bone Tissue Engineering Experiment

作　　者：龚泰芳[1] 何少斌[1] 陈文[1] 卢云[1] 王栋锋[1] 王平年[1] 

机构地区：[1]郧阳医学院附属太和医院骨关节科,湖北省十堰市442000

出　　处：《中国骨与关节损伤杂志》2008年第12期999-1002,共4页Chinese Journal of Bone and Joint Injury

基　　金：湖北省教育厅项目(B200524005);十堰市科技攻关项目(2006050D)

摘　　要：目的探讨兔骨髓来源成骨细胞体外分离培养的条件及生长特征,为骨组织工程学实验研究奠定基础。方法穿刺抽取4个月龄新西兰兔的骨髓,采用离心和贴壁相结合方法,用矿化诱导培养液培养贴壁的骨髓细胞并传代。观察其生物学特征,绘制生长曲线、分裂指数曲线和贴壁率曲线,测定细胞贴壁及延展时间,并将培养细胞进行冻存、复苏,比较细胞特性有无改变。同时,电镜观察细胞在藻酸钙凝胶载体内的生长情况。结果培养细胞在第12代以前生长性状稳定,具有典型的成骨细胞特性;第4天为细胞生长倍增时间,第5～6天细胞达生长高峰;第4天细胞分裂最为旺盛,达18%;传代后10h贴壁率最高,为90%;冻存复苏后的细胞生物学特性无改变。电镜观察细胞在藻酸钙凝胶载体内生长良好,有骨基质分泌。结论本实验方法体外培养的成骨细胞,生长性状稳定,增殖速度快,可保证相关实验的可靠性和准确性;第12代以前,生长3～6d的细胞可作为理想的实验细胞。Objective To observe culture condition and growth characteristics of osteoblasts in vitro, which were developed from bone marrow stromal of rabbits, contributing to the bone tissue engineering experiment. Methods Osteoblasts, which were developed from bone marrow stromal of New Zealand rabbits with a mean age of 4 months, were cultured in a conditional medium including dexametha- sone, vitamine C and 0634 - glycerophosphate, arid subcultured. Their biological features were investigated. The ALP activity was evaluat- ed. Their strapping and expanding time were accessed and the graphs of growth, split index and adhesion rate assess were drawn. Frozen and resuscitated cells, after culturing, were compared to understand the differences of their features. Results Features of cul- tured cells, which could be used as experiment cells, with typical features of osteoblasts, were steady before the 12th passage, doubling time of cells was the 4th day. The highest proliferation was on the 5th to 6th day. The mitosis index of the cells was the highest on the 4th day （18 % ）, adhesion rate of the cells after passage of 10 hours was the highest （90%）. The biological features of cells had no changes after freezing and resuscitation. Conclusion Osteoblasts cultured in vitro by this method, characterized of steady growth, rapid proliferation and the same biological features with osteoblasts in vivo, are proved to assure experiment of authenticity and accuracy and to be applied to the study of bone tissue engineering as seed cells.

关 键 词：骨髓 成骨细胞 体外培养 生物学特性 藻酸盐 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学] R318.08[医药卫生—基础医学]