改良分步消化法培养原代软骨细胞

Improved stepwise digestion method for chondrocyte primary culture

背景:胰蛋白酶和细菌胶原酶结合使用消化关节软骨基质获得大量纯度高的软骨细胞的方法步骤繁琐、过程复杂,容易污染,但更好的简单易行、安全可靠的方法至今少有报道。目的:采用改良分步消化法进行软骨细胞培养,以获取大量纯净的软骨细胞。方法:将新西兰白兔6只随机分2组,酶消化法组运用酶消化法分两步获取原代软骨细胞,对照组用传统法进行原代软骨细胞培养。培养1周后观察两组培养的软骨细胞的生长状态,并进行细胞鉴定、计数,评估改良后的方法对细胞的影响。结果与结论:酶消化法组采用0.2%Ⅱ型胶原酶消化软骨细胞,与对照组相比,可将6h以上的消化时间缩短至3h。两组原代软骨细胞培养24h均多呈圆形,悬浮状态,48h后贴壁,培养1周后,两组软骨细胞可铺满培养瓶底。结果证实,采用改良分步消化法进行软骨细胞培养,在缩短了消化时间的同时其细胞生长及形态变化均无改变,可以顺利获取大量纯净的软骨细胞。

BACKGROUND：Combination of trypsin and bacterial collagenase to digest articulate cartilage matrix is widely used at home and abroad.Some studies have shown that the method steps is tedious,process is complicated,and cell is easy to pollution,but the better simple,safe and reliable method has rarely been reported.OBJECTIVE：To obtain highly purified and large amount of chondrocytes by improved stepwise digestion method.METHODS：Six 3-week-old New Zealand rabbits were divided into two groups：The experimental group was cultured by two-step enzyme digestion,and the control group was cultured by the traditional enzyme digestion method.The growth condition of chondrocytes was observed in the two group after 1 week,and cell counting and identification was performed.We estimated improved method to evaluate the influence of cells.RESULTS AND CONCLUSION：Compared with the traditional method,this method spent shorter time to digest chondrocytes.After 24 hours,the primary chondrocytes in the two groups were mostly round at a suspending state.After 48 hours,the cells began to adhere.The results demonstrated that the improved method shortens digestion time,and easy to obtain a great amount of purified chondrocytes.