蛇床子素对TRAIL抗乳腺癌活性的影响及其作用机制

Effect of Osthole on the Anti-breast Cancer Activity of TRAIL and Its Mechanism

目的探讨中药活性成分蛇床子素对抗肿瘤药物TRAIL抗乳腺癌活性的影响并研究其机制。方法用蛇床子素联合TRAIL体外治疗乳腺癌细胞系BT-20,MTT法检测肿瘤细胞的细胞活力,Annexin V/PI染色检测肿瘤细胞的凋亡,免疫共沉淀法检测RIP1-FADD-caspase-8复合物的形成。Western blot法检测BT-20细胞caspase-8的活化及蛇床子素对BT-20细胞c IAP2蛋白表达的影响。结果联用蛇床子素显著提高TRAIL对BT-20细胞活力的抑制率和凋亡诱导活性。免疫共沉淀及Western blot结果发现联合蛇床子素后,TRAIL治疗的BT-20细胞内的RIP1-FADD-caspase-8复合物水平及caspase-8的活化程度显著提高。Western blot结果发现蛇床子素对BT-20细胞内的c IAP2蛋白表达有抑制作用。在BT-20细胞中转染c IAP2表达质粒后,蛇床子素对TRAIL抗肿瘤活性的促进作用受到抑制。结论蛇床子素通过促进死亡受体复合物的形成增强TRAIL对乳腺癌细胞的凋亡诱导效应。

OBJECTIVE To investigate the effect of osthole on the anti-breast cancer activity of TRAIL and study its mechanism. METHODS After the BT-20 cells were treated with TRAIL combined with osthole, the cell viability, cell apoptosis, and the formation of RIP1-FADD-caspase-8 complex were detected by MTT assay, Annexin Ⅴ/PI staining, and co-immunoprecipitation, respectively. The activation of caspase-8 and the expression of c IAP2 in BT-20 cells were evaluated by Western blot assay. RESULTS Addition of osthole significantly enhanced the cell viability inhibition rate and apoptosis inducing activity in BT-20 cells treated with TRAIL. The results of co-immunoprecipitation and Western blot indicated that the formation of RIP1-FADD-caspase-8 complex and the activation of caspase-8 in TRAIL-treated BT-20 cells were significantly increased due to the combination of osthole. The results of Western blot assay demonstrated that the expression of c IAP2 could be significantly inhibited due to the osthole treatment. Moreover, the transfection of c IAP2 vector abolished the promotion of osthole on TRAIL-induced cell death in BT-20 cells. CONCLUSION Osthole promotes TRAIL-induced apoptosis by the formation of RIP1-FADD-caspase-8 complex in breast cancer.