大鼠胫骨生长板软骨细胞的分离与培养鉴定  

Primary culture and identification of rat tibial growth plate chondrocytes

作　　者：余振华[1] 朱顺叶[2] 张珑娟[3] 

机构地区：[1]中山大学附属第一医院神经外科,广州510080 [2]中山大学附属第三医院儿科 [3]中山大学附属第一医院外科实验室,广州510080

出　　处：《中华实验外科杂志》2008年第10期1309-1310,共2页Chinese Journal of Experimental Surgery

摘　　要：目的探讨高效生长板软骨细胞分离方法和体外培养条件。方法采用二步酶消化法对6只SD大鼠胫骨生长板的软骨细胞进行分离，采用含炭吸附过的胎牛血清培养基培养，按5×10^5个／瓶的密度接种细胞并对软骨细胞进行形态学观察和鉴定，描绘原代软骨细胞在无激素培养基中的生长曲线。结果软骨细胞贴壁较慢，12h后开始附壁，第8天时90％融合，互相连接成“铺路石”样结构。原代软骨细胞胞质Ⅱ型胶原免疫着色强阳性，传代后染色减弱。结论本研究所采用的方法能高效快速获得原代软骨细胞，原代软骨细胞最接近体内生理状态，最适合进行实验研究。Objective To establish the methods of isolating and cultivating chondrocytes from rat growth plate. Methods Through two-step enzymatic digestion （ digested by 0.25 % trypsin for 30 min, and then treated with 0.15% collagenase Ⅱ for 3 h） , tibial growth plate chondrocytes of Sprague-Dawley rats （n = 6） were isolated and the primary chondrocytes was adjusted to a density of 5 ×10^5/bottle. The chondrocytes were incubated for 8-10 days in culture medium （DMEM with L-Glutamine, Hepes ,without phenol red） supplemented with 10% fetal bovine serum. Collagen type Ⅱ expression was detected by histochemistry. Viability ,multiplication ,cell growth kinecties and morphologic changes were observed. Results Chondrocytes adhered after 12 h and formed confluent growth on day 8. Collagen type Ⅱ expression in the primary passage was positive by histochemistry, and subsequently with the extension of time,the synthesis effect of collagen Ⅱ gradually weakened. Conclusion The primary passage chondrocytes have fine biological activity and are suitable for potential experimental model for studying in vitro.

关 键 词：软骨细胞 细胞培养 生长板 

分 类 号：R329[医药卫生—人体解剖和组织胚胎学]