山羊颈椎间盘纤维环成纤维细胞成骨潜能的体外观察  

IN VITRO STUDY ON OSTEOGENIC POTENTIAL OF ANNULUS FIBROSUS IN GOAT CERVICAL INTERVERTEBRAL DISC

作　　者：卢旭华[1] 赵定麟[1] 冯伟[1] 许卫兵[1] 陈德玉[1] 袁文[1] 

机构地区：[1]第二军医大学长征医院骨科,上海200003

出　　处：《中国修复重建外科杂志》2005年第12期977-981,共5页Chinese Journal of Reparative and Reconstructive Surgery

摘　　要：目的诱导培养椎间盘纤维环成纤维细胞,探讨其成骨潜能,比较重组人骨形成蛋白2(recom b inan thum an bone m orphogenetic prote in 2,rhBM P-2)和肿瘤坏死因子α(tum or necros is factor,αTNF-α)对其成骨的不同影响。方法利用体外细胞培养技术,建立实验山羊颈椎间盘纤维环成纤维细胞培养体系,根据培养液的不同,分为空白对照组、TNF-α组(加入50 U/m lTNF-α)、rhBM P-2组(加入0.1μg/m l rhBM P-2)和TNF-α+rhBM P-2组(加入50 U/m l TNF-α+0.1μg/m l rhBM P-2),培养3周观察纤维环成纤维细胞在条件培养液诱导下的成骨表型变化、碱性磷酸酶(a lka line phosphatase,ALP)和骨钙素(osteoca lc in,OC)等成骨指标变化。结果各实验组培养液对细胞增殖无明显影响。rhBM P-2组和TNF-α+rhBM P-2组细胞趋化性生长明显,矿化结节出现,茜素红染色呈深红色钙阳性反应。各实验组ALP活力与空白对照组比较差异均有统计学意义(P<0.05)。rhBM P-2组与TNF-+αrhBM P-2组成纤维细胞的OC分泌量与空白对照组比较差异均有统计学意义(P<0.05),TNF-α组与空白对照组比较差异无统计学意义(P>0.05)。结论体外培养的椎间盘纤维环成纤维细胞有成骨潜能,rhBM P-2对纤维环成纤维细胞有明确的诱导成骨作用。Objective To explore the osteogenic potential of cervical intervertebral disc fibroblasts in vitro, to investigate the regulatory factors of recombinant human bone morphogenetic protein 2（rhBMP-2） and tumor necrosis factor a（TNF-α） on osteogenic phenotype of fibroblasts and to discuss the condition that facilitates osteogenesis of fibroblasts. Methods The annulus fibroblasts cell lines of experiment goats were established in vitro and the biologic specificity was found. According to different medias, 4 groups were included in this experiment： control group, TNF-α group （ 50 U/ml TNF-α）, rhBMP 2 group （0. 1 μg/ml rhBMP-2） and TNF-α＋rhBMP-2 group （50 U/ml TNF-α＋ 0. 1μg/ml rhBMP-2）, The fibroblasts were incubated in the media for about 3 weeks,and then the markers for osteogenic features were investigated by biochemistry, histochemistry observations. Results rhBMP 2 and TNF-α had no effect on the proliferation of fibroblasts from the experiment goats, rhBMP-2or TNF a could stimulate fibroblaststo secrete alkaline phosphatase and collagen type 1 . The combined use of rhBMP 2 and TNF-α or the single use of rhBMP 2 could make fibroblasts to secrete osteocalin and the morphological changes of the fibroblasts were very obvious. Histochemical study of the nodules with specific new bone labeler（Alizarin red S） revealed positive reaction, denoting that the nodules produced by the fibroblasts were bone tissues. There was statistically significant difference （P〈0.05） in ALP activity between 3 experimental groups and control group and in secretion of osteocalcin between rhBMP-2 group, TNF-α＋rhBMP-2 group and control group. Conclusion The resuhs point out clearly that rhBMP-2 can induce the osteogenic potential of annulus fibroblasts in vitro.

关 键 词：椎间盘纤维环 成纤维细胞 成骨潜能 骨形成蛋白 山羊 

分 类 号：R681.5[医药卫生—骨科学] Q813[医药卫生—外科学]