蟾蜍灵对宫颈癌细胞的增殖抑制作用及其机制研究

Research on the cytostatic effect of bufalin on cervical carcinoma cells and the related mechanism

背景与目的:宫颈癌仍然是妇科肿瘤致死病因的第二位,部分原因为化疗耐药。蟾蜍灵是传统中药蟾酥的成分之一,目前在国内广泛应用于肿瘤的治疗中。该研究旨在探讨蟾蜍灵对宫颈癌细胞ME180和C33A的增殖抑制作用及相关机制。方法:采用CCK8(cell counting kit-8)法检测蟾蜍灵对宫颈癌细胞增殖的抑制作用,采用葡萄糖检测分析试剂盒(glucose assay kit)进行细胞内糖代谢检测分析,采用实时荧光定量聚合酶链反应(quantitative real-time polymerase chain reaction,q RT-PCR)检测细胞内葡萄糖转运蛋白1(glucose transporter1,GLUT1)和己糖激酶2(hexokinase 2,HK2)的m RNA的表达水平,采用蛋白[质]印迹法(Western blot)检测原癌基因C-MYC和缺氧诱导因子1α(Hif-1α)表达的变化。结果:CCK8法结果显示,蟾蜍灵能明显抑制宫颈癌细胞ME180和C33A细胞增殖(P=0.027,P=0.018)。糖代谢检测结果显示,蟾蜍灵处理组葡萄糖代谢水平显著降低(P=0.034,P=0.036)。q RT-PCR检测结果显示,蟾蜍灵处理组糖代谢相关指标GLUT1(P=0.019,P=0.016)和HK2(P=0.039,P=0.041)的表达水平明显下调。Western blot检测结果显示,蟾蜍灵处理后宫颈癌细胞内C-MYC和Hif-1α蛋白表达水平显著下调。结论:蟾蜍灵可以通过降低宫颈癌细胞ME180和C33A糖代谢水平,从而抑制细胞增殖。

Background and purpose：Cervical cancer remains the second leading cause of death in gynecologic malignancies partially because of resistance to chemotherapy. Bufalin, a component of the traditional Chinese medicine Chansu, has been widely used in cancer treatment in China. This study aimed to investigate the effects of bufalin on inhibiting the proliferation of ME180 and C33A and explore its possible mechanism.Methods：The cytostatic effects of bufalin on ME180 and C33A cells were evaluated by CCK8 assay （cell counting kit-8）. Glucose levels in ME180 and C33A cells were measured using glucose assay kit. Then the alterations of GLUT1 （glucose transporter 1） and HK2 （hexokinase 2） gene expression were detected by quantitative real-time polymerase chain reaction （qRT-PCR）. The expressions of proto-oncogene C-MYC and HIF1α （hypoxia-inducible factor 1α） were determined by Western blot.Results：According to the results of CCK-8, bufalin can significantly inhibit the proliferation of carcinoma cells ME180 and C33A （P=0.027,P=0.018）. Test on glycometabolism indicated that glucose uptake in cells treated with bufalin decreased （P=0.034,P=0.036）. Results from real-time PCR showed that the expression of glycometabolism related indicators GLUT1 （P=0.019） and HK2 （P=0.016） levels were signiifcantly down-regulated in bufalin treated group. Western blot showed that the expression of C-MYC and HIF1αin cells with bufalin treatment was down-regulated markedly.Conclusion：Bufalin can inhibit the proliferation of the cervical carcinoma cells ME180 and C33A through inhibition of their glucose metabolism.