IL-1β对小鼠软骨细胞中c-myc蛋白表达的影响  

Effects of IL-1β on protein expression of c-myc in chondrocytes of mouse

作　　者：王宁[1] 龙迪[1] 孟晓娜[2] 

机构地区：[1]沈阳医学院附属中心医院骨外一科,110024 [2]中国医科大学生物化学与分子生物学教研室

出　　处：《中华临床医师杂志（电子版）》2013年第24期222-224,共3页Chinese Journal of Clinicians(Electronic Edition)

摘　　要：目的通过观察不同浓度IL-1β对软骨细胞c-myc蛋白表达的影响,探讨IL-1β在骨关节炎软骨损伤中的作用及机制并寻找有效治疗骨关节炎的方法。方法选用20只C57BL/6小鼠,体外进行关节软骨细胞的分离及原代培养;将原代培养的软骨细胞分为4组:A组为空白对照组,采用含10%FBS的RPMI-1640常规培养基培养;B、C、D组为IL-1β处理组,分别用含有1、10和100 ng/ml IL-1β的常规培养基培养,12 h后进行实验分析。采用Western blot和流式细胞仪检测法,观察各组c-myc蛋白表达情况及细胞凋亡情况。结果原代培养细胞24 h后开始贴壁,呈圆形或多角形,传至第4代、第5代细胞体积变大,逐渐成为梭形,甲苯胺蓝染色为软骨细胞内见蓝紫色异染颗粒。与对照组相比较,不同浓度IL-1β处理组软骨细胞c-myc蛋白表达水平逐渐增高,呈剂量依赖性(P<0.05);与对照组相比较,不同浓度IL-1β处理组软骨细胞凋亡率逐渐增高,呈剂量依赖性(P<0.05)。结论实验体外分离小鼠关节软骨,可成功获得高纯度,且传至3代以内活性率超过90%的软骨细胞;IL-1β可以按照剂量依赖方式上调软骨细胞c-myc蛋白的表达,同时增加软骨细胞凋亡率,说明IL-1β可能通过c-myc蛋白诱导软骨细胞凋亡。Objective Through the investigation on the effect of IL-1βon c-myc protein expression in chondrocytes of mouse, to explore the role and mechanism of IL-1βin osteoarthritic cartilage injury and to find an effective method for the treatment of osteoarthritis. Methods Chondrocytes were isolated from twenty C57BL/6 mice and cultured in vitro;the primary cultured chondrocytes were divided into 4 groups：A group, which was the control group, was cultured in conventional medium （RPMI-1640 containing 10% FBS）; B, C, D groups, which were the IL-1β-treated groups, were cultured in conventional culture medium containing 1, 10 and 100 ng/ml IL-1β, respectively. After 12 h culture, experimental analysis were performed. Western Blot and flow cytometry method were used to observe the expression changes of c-myc protein and the apoptosis rate of each group. Results After 24 h, the primary cultured cells started to be adherent with rounded or polygonal shapes;Volume of cells began to be exaggerated at 4th or 5th cell passage, spindle-shaped. The blue-violet iso dye particles could be seen in the cells by toluidine blue staining; Compared with control group, the c-myc protein levels were gradually increased in a dose-dependent manner（P〈0.05） when the cells were treated with IL-1β of different concentrations; compared with control group, the apoptosis rate of chondrocytes challenged to IL-1β of different concentrations were increased significantly, in a dose-dependent manner（P〈0.05）. Conclusion Chondrocytes can be successfully obtained with high purity and the viability of them is 90% above at 3rd cell passage; c-myc protein can be up-regulated by IL-1β in a dose dependent manner and simultaneously the apoptosis rate are also increased correspondingly, indicating that chondrocyte apoptosis could be induced by IL-1βthrough c-myc protein changes.

关 键 词：软骨细胞 白细胞介素1Β 印迹法 蛋白质 流式细胞术 分离培养 

分 类 号：R684.3[医药卫生—骨科学]