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目的:研究氯离子通道在华蟾酥毒基(CBG)诱导低分化鼻咽癌CNE-2Z细胞凋亡和凋亡性容积减小中的作用。方法:实验设立空白对照组、CBG高、中、低剂量组及CBG与氯通道阻断剂NPPB联合作用组。Hoechst 33342染色检测细胞的凋亡率,活细胞图像分析法检测细胞容积变化,全细胞膜片钳法记录氯电流。结果:(1)CBG浓度依赖性诱导鼻咽癌细胞凋亡,高剂量CBG(8μmol/L)诱导的细胞凋亡率为43.2%;(2)CBG可诱导细胞产生凋亡性容积减小,在CBG作用早期(50 min),细胞容积减小约9.9%。(3)CBG可激活氯通道,产生一个无明显外向优势的氯电流。(4)氯通道阻断剂NPPB可抑制CBG诱导的细胞凋亡、凋亡性细胞容积减小和氯电流。NPPB对CBG诱导的细胞凋亡和凋亡性容积减小的抑制率分别达80.1%和74.8%。结论:以上结果提示CBG可能通过激活氯通道而诱导细胞凋亡,氯通道的激活可能在CBG抗肿瘤机制中起重要作用。
OBJECTIVE: To study the role of chloride channel in the apoptosis and apoptotic volume reduction of CNE-2Z induced by cinobufagin (CBG) in poorly differentiated nasopharyngeal carcinoma. Methods: The blank control group, CBG high, medium and low dose groups and the combination of CBG and chloride channel blocker NPPB were established. Hoechst 33342 staining was used to detect the apoptotic rate of cells. Changes of cell volume were measured by image analysis of live cells. Whole-cell patch clamp was used to record the chloride current. Results: (1) The apoptosis of nasopharyngeal carcinoma cells was induced by CBG in a concentration-dependent manner. The apoptotic rate induced by high dose of CBG (8μmol / L) was 43.2%. (2) CBG induced apoptotic volume decrease, In the early phase of CBG (50 min), the cell volume decreased by about 9.9%. (3) CBG activates the chloride channel to produce a chlorine current with no apparent outward advantage. (4) NPPB, a chloride channel blocker, inhibited CBG-induced apoptosis, decreased apoptotic cell volume and chloride current. The inhibitory rates of NPPB on CBG-induced apoptosis and apoptotic volume reduction were 80.1% and 74.8%, respectively. Conclusion: The above results suggest that CBG may induce apoptosis through activation of chloride channel. The activation of chloride channel may play an important role in anti-tumor mechanism of CBG.