【摘要】 目的探讨大鼠剑突软骨细胞的培养方法,以及多聚2-羟乙基甲基丙烯酸脂(poly-2-hydroxyethylmethacrylate,poly-HEMA)在培养过程中对软骨细胞生物学特性的影响。方法无菌条件下取Wistar大鼠的剑突,用剪刀将其剪成1 mm3大小,加入0.25%的胰酶消化30 min,然后用0.15%的胶原Ⅱ消化3~5 h。将分离得到的细胞分别接种在预先用poly-HEMA包被的平皿和普通的平皿内,并将普通平皿的细胞进行差速贴壁。通过形态学及甲苯胺蓝染色对细胞进行鉴定。结果①从剑突软骨可获得大量软骨细胞,每克软骨可获得软骨细胞(3.1±0.5)×107个细胞,活性率为(95.2±0.3)%。②在普通平皿内经过两次差速贴壁得到的软骨细胞较纯,甲苯胺蓝染色阳性率达90%。③软骨细胞在poly-HEMA包被的平皿内始终悬浮生长,且能长期保持软骨细胞的特性。结论从剑突获得大量软骨细胞的方法是可行的,poly-HEMA能使体外培养的软骨细胞未经传代的情况下长期保持其生物学特性,为体外实验及组织工程研究提供优质种子细胞。更多还原

【Abstract】 ObjectiveTo establish the methods for isolation and culture of xiphoid chondrocytes of rat in vitro,and to explore the possible effect of poly-2-hydroxyethylmethacrylate(poly-HEMA) on the biological features of chondrocytes.MethodsXiphoid of Wistar rat as segregated under sterile condition.Xiphoid cartilage as cut into 1 mm~3 fragments and digested by(0.25)% trypsin for 30 minutes,and then treated with(0.15)% collagenase Ⅱ for 3～5 hours.Chondrocytes were implanted in culture dishes with poly-HEMA and without poly-HEMA respectively.Chondrocytes were indentified with morphological observation and toluidine blue staining.Results① The method of digestion with 0.25% trypsin and 0.15% collagenase II can acquire pure xiphoid chondrocyte isolation.The number of chondrocytes gained from wet xiphoid cartilage was((3.1)±(0.5))×10~7/g with a viability of((95.2)±(0.3))%.② Chondrocytes could be purified after differential attachment twice in general dishes.The cells stained by toluidine blue were 90%.③ Chondrocytes in culture dishes coated with poly-HEMA can grow in suspension.Poly-HEMA can keep chondrocytes primary morphology and biological features.ConclusionThe method of isolation and culture of chondrocytes from rat’s xiphoid is feasible.Poly-HEMA can keep chondrocytes feature for a long time,which allow to provid good seed cells for experiment in vitro and tissue engineering.更多还原