胰岛素样生长因子Ⅰ对三维培养兔关节软骨细胞增殖和表型的影响  

Effects of IGF-Ⅰ on proliferation and phenotype of three-dimensionally cultured rabbit articular chondrocytes

作　　者：吴剑宏[1] 王秋根[1] 张秋林[1] 陆晴友[1] 王万宗[1] 张素贞[2] 

机构地区：[1]第二军医大学长海医院骨科,上海200433 [2]长海医院血管外科

出　　处：《第二军医大学学报》2005年第4期418-421,共4页Academic Journal of Second Military Medical University

基　　金：上海市科技发展基金攻关计划项目(034119830).

摘　　要：目的:研究胰岛素样生长因子Ⅰ(IGF -Ⅰ)对藻酸盐凝胶三维培养条件下兔关节软骨细胞的影响。方法:利用机械与酶消化的方法获得均一性的兔关节软骨细胞,在单层培养条件下增殖至P2代;将P2代细胞高密度条件下转入藻酸盐凝胶培养介质,进行三维培养,另一组在进行三维培养的同时,在培养液中加入IGF -Ⅰ50 ng/ml,绘制各组细胞生长曲线,并与单层培养对照组细胞进行比较。将培养细胞行Ⅱ型胶原免疫组织化学检测,H E染色观察细胞形态。进行不同浓度IGF- Ⅰ作用下的三维培养软骨细胞MTT检测,观察0、25、50、75、100 ng/ml IGF0 Ⅰ对三维培养软骨细胞各时间点的促增殖作用。结果:IGF 0Ⅰ可明显刺激三维培养条件下关节软骨细胞增殖及集落形成;不同浓度的IGF 0Ⅰ均可促进关节软骨细胞增殖,以50 ng/ml浓度时D490值最高。培养6周后从凝胶体系中收获细胞的软骨特异性Ⅱ型胶原表达水平较转入三维体系前未见降低。结论:IGF Ⅰ可明显刺激藻酸盐三维培养条件下的关节软骨细胞增殖及集落形成,且细胞表型稳定。Objective:To explore the effects of IGF-Ⅰ on the proliferation and phenotype of three-dimensionally cultured rabbit articular chondrocytes. Methods: The cultured articular chondrocytes of 4-week old rabbits were obtained released by mechanical dissection and enzyme digestion. The chondrocytes were first replicated under monolayer culture condition until the second generation, and then suspended in the aqueous sodium alginate beads with high density. The chondrocytes were cultured in Dulbecco's modified Eagle medium and some were treated with IGF-Ⅰ at 50 ng/ml for certain period. Chondrocytes cultured in alginate with IGF-Ⅰ at the concentrations of 0,25,50,75,100 ng/ml were detected by MTT. Results: IGF-Ⅰ stimulated the proliferation and colony formation of chondrocytes. The chondrocytes cultured with different concentrations of IGF-Ⅰ were significantly more than that of control group,with the best concentration being 50 ng/ml. Compared with the pre-seeding state of chondrocytes 6 weeks ago, the harvested cells from alginate medium showed an equivalent level of type Ⅱ collagen expression. Conclusion: Under the three dimensional culture circumstance, IGF-Ⅰ can significantly stimulate the proliferation and colony formation of chondrocytes. The chondrocytes cultured in alginate system maintain the excellent differentiation phenotype.

关 键 词：胰岛素样生长因子Ⅰ 关节 软骨细胞 三维培养 藻酸盐 细胞增殖 表型 

分 类 号：R684[医药卫生—骨科学]