人重组TNF相关凋亡诱导配体联合塞来昔布诱导胆囊癌细胞凋亡的作用研究

The proapoptotic effect of combined treatment with human recombinant TNF-related apoptosis-inducing ligand and Celecoxib on gallbladder carcinoma

目的 于体外实验观察人重组肿瘤坏死因子相关凋亡诱导配体（rhTRAIL）联合环氧化酶-2选择性抑制剂塞来昔布对胆囊癌的疗效，初步探讨产生疗效的机制。方法 用Western-blot法检测塞来昔布作用于胆囊癌细胞株后，抗凋亡蛋白c-FLIP和死亡受体DR4、DR5表达状况。rhTRAIL联合塞来昔布作用于胆囊癌细胞株SGC-996后，采用3种方法检测细胞凋亡状况：（1）细胞的相差显微镜观察；（2）caspase 3、7活性检测；（3）Annexin染色后凋亡细胞流式细胞仪检测。结果 塞来昔布可下调 SGC-996细胞c-FLIPs的表达，上调DR5的表达，且呈浓度、时间依赖性。rhTRAIL联合塞来昔布作用于胆囊癌细胞后，细胞凋亡水平明显高于单独用药组和对照组。结论 塞来昔布可通过下调c-FLIPs表达和上调DR5表达，显著增强rhTRAIL诱导胆囊癌SGC-996细胞凋亡的作用。

Objective To observe the effect of combined treatment with rhTRAIL（recombinant human TNF-related apoptosis inducing ligand） and selective Cox-2 inhibitor Celecoxib on gallbladder carcinoma in vitro and to explore the possible mechanism of the effect. Methods Western blot analysis was used to detect the expression of c-FLIP and death receptors after treatment by Celecoxib. Apoptosis of gallbladder cell line SGC-996 after the combined treatment with Celecoxib and rhTRAIL was detected in three ways： （1） phase microscopy of the cells,（2） detection of effector caspase-3 and caspase-7 activity, and （3） determination of the proportion of apoptotic cells labeled by Annexin V-PI flow cytometric analysis using CELLQUEST software. Results Celecoxib down-regulated the expression of c-FLIPs and up-regulated the expression of DR5 in a dose- and time-dependent mode on cell line SGC-996. Apoptotic levels in the combined treatment group in cell line SGC-996 were significantly higher than those in the single drug treatment group and control group. Conclusion Celecoxib markedly sensitized rhTRAIL-induced apoptosis through the down-regulation of c-FLIPs and up-regulation of DR5 in gallbladder carcinoma cell line SGC-996.