凋亡与增殖相关基因在损伤关节软骨细胞中的表达  

Expression of apoptotic and proliferation genes in articular chondrocyte injury model of rabbit

作　　者：马子君[1] 赵海洋[1] 任俊丽[1] 陈传好[1] 

机构地区：[1]蚌埠医学院人体解剖学教研室,蚌埠233030

出　　处：《解剖学杂志》2013年第5期884-887,共4页Chinese Journal of Anatomy

基　　金：安徽省自然科学基金（11040606M163）;博士研究生科研启动基金（2010BR030）

摘　　要：目的：建立兔关节软骨细胞损伤模型并分析细胞损伤后不同时间点的半胱氨酸的天冬氨酸蛋白水解酶3（caspase-3）与细胞增殖核抗原（PCNA）基因表达变化。方法：取4周龄健康新西兰兔膝关节软骨，以机械分离和酶消化分离相结合的方法获取膝关节软骨细胞。利用Ⅱ型胶原免疫荧光染色鉴定软骨细胞，建立机械性损伤模型。实时荧光定量PCR检测正常和划伤后caspase-3，PCNA和Ⅱ型胶原蛋白基因mRNA表达变化。结果：成功建立关节软骨细胞损伤模型。与正常组相比，损伤后1dcaspase-3mRNA表达开始上升。损伤后3d组与7d组caspase-3mRNA表达与正常组及损伤后1d组相比均明显下降。与正常组相比，损伤后1d组PCNAmRNA表达下降，损伤后3d组与1d组相比未见变化，7d组与1d组相比表达上升。与正常组相比，损伤后1d组Ⅱ型胶原蛋白mRNA表达明显下降，损伤后3d组上升，至第7天达到正常水平。结论：凋亡与增殖相关基因在关节软骨损伤后有不同的表达规律，为调节关节软骨内源性相关基因表达治疗关节软骨损伤提供实验依据。Objective： To establish injury model of rabbit articular chondrocyte in vitro and analyze the expression change of caspase-3 and PCNA in different time-point of injury model. Methods： Articular cartilage was aseptically dissected from the bilateral knee joints of 4-weekold healthy new Zealand white rabbits. Articular chondrocytes were harvested by the mechanical separation combined with enzyme digestion method, which were cultured and passaged in vitro. Type Ⅱ collagen immunofluorescence assay were employed to identify the articular chondrocytes. The expression of caspase-3, PCNA and collagen Ⅱ of injuried and normal articular chondrocytes at 1, 3, 7 days were studied by real-time quantitative polymerase chain reaction （qPCR）. Results： The injury model of rabbit articular chondrocyte was successfully established. Compared with the normal cell group, the levels of caspase-3 mRNA increased at day 1 after injury, and were obviously lower at day 3 and 7 than at day 1 and that of the normal group. Compared with the normal cell group, the levels of PCNA mRNA decreased at day 1 after injury, and changed not obviously at day 3 than at day 1, began to increase at day 7. Compared with the normal cell group, the levels of collagen Ⅱ mRNA decreased obviously at day 1 after injury, began to increase at day 3 and reached normal at day 7 after injury. Conclusion： The related genes of apoptosis and proliferation show different expression patterns, which can provide the experimental basis for gene therapy for articular cartilage injury.

关 键 词：关节软骨细胞 损伤模型 半胱氨酸的天冬氨酸蛋白水解酶-3 细胞增殖核抗原 Ⅱ型胶原蛋白 

分 类 号：R684[医药卫生—骨科学]