拉帕替尼诱导人鼻咽癌CNE-2Z细胞周期阻滞和细胞凋亡观察

Lapatinib induced cell cycle arrest and apoptosis in CNE-2Z cells

目的探讨拉帕替尼对人鼻咽癌CNE-2Z细胞周期和凋亡的影响及其作用机制。方法采用CCK-8法检测拉帕替尼对CNE-2Z细胞增殖的影响,PI染色后以流式细胞术进行细胞周期分析,Annexin V/PI双标记法检测细胞凋亡,实时定量PCR检测细胞周期相关分子Cyclin D1、P21和P53的m RNA表达水平,Western blotting检测Cyclin D1、P21、P53及凋亡相关蛋白Mcl-1和Bax的表达,利用Apo-ONE Homogeneous caspase-3/7检测试剂盒检测Caspase-3/7酶活性。结果拉帕替尼呈剂量依赖性地抑制CNE-2Z细胞增殖,并可诱导CNE-2Z细胞发生明显的细胞周期G0/G1期阻滞,促进细胞凋亡,下调Cyclin D1并上调P21、P53 m RNA和蛋白表达,下调Mcl-1和上调Bax蛋白表达,并诱导CNE-2Z细胞内caspase-3/7激活。结论拉帕替尼可有效抑制人鼻咽癌CNE-2Z细胞增殖,诱导细胞周期阻滞并促进其凋亡,可作为治疗鼻咽癌的潜在药物。

Objective To explore the effect oflapatinib on the cell cycle and apoptosis of human CNE-2Z nasopharyngeal carcinoma （NPC） cells, and to study the related mechanisms. Methods CCK-8 assay was used to assess the proliferation of CNE- 2Z cells treated by lapatinib. After PI staining, the cell cycle distribution was determined by flow cytometry. Apoptosis was analyzed using Annexin V/PI double binding assay. The mRNA expression of cell cycle related molecular Cyclin D 1, P21 and P53 was assessed with real time PCR. The expression of protein Cyclin D1, P21, P53, and apoptosis related protein Mcl-1 and Bax was determined by Western blotting. The enzymatic activity of caspase-3/7 was measured by using Apo-ONE Homogeneous Caspase-3/7 Assay kit. Results Lapatinib inhibited the proliferation of CNE-2Z cells in a dose-dependent manner. Various concentrations of lapatinib induced significant G0/G1 phase arrest and promoted apoptosis of CNE-2Z cells. Lapatinib significantly down-regulated Cyclin D1 expression, but up-regulated P21 and P53 expression at mRNA and protein levels. Lapatinib also induced down-regulation of Mcl- 1 with up-regulation of Bax protein expression, and activated Caspase-3/7 in CNE-2Z cells. Conclusion Lapatinib may effectively inhibit proliferation, induce cell cycle arrest and promote apoptosis of CNE-2Z ceils, so it may serve as a potent therapeutic agent against nasopharyngeal carcinoma.