摘要
目的:研究洋川芎内酯A(SEA)联合表柔比星(EPI)干预膀胱癌5637细胞的作用机制,探讨SEA对EPI敏感性的影响。方法:采用CCK-8试剂盒检测人输尿管上皮永生化细胞SV-HUC-1和人膀胱癌细胞5637的细胞活力,选用人膀胱癌细胞5637进行后续实验,计算半数抑制浓度(IC50值)。将细胞随机分为对照组、SEA低剂量组(SEA10组)、SEA中剂量组(SEA20组)、SEA高剂量组(SEA40组)、EPI单独处理组(EPI组)和联合处理组(SEA40+EPI组)。流式细胞仪检测细胞凋亡;CCK-8试剂盒检测细胞光密度(OD)值;免疫蛋白质印迹法(Western blot)检测多药耐药关联蛋白1(MRP1)、肺耐药蛋白(LRP)、拓扑异构酶-Ⅱ(Topo-Ⅱ)蛋白水平。建立裸鼠移植瘤模型,随机分为对照组、SEA药物组、EPI药物组和SEA+EPI组,并采用相应药物灌胃进行干预。检测肿瘤体积;Western blot检测裸鼠移植瘤模型MRP1、增殖细胞核抗原(PCNA)、裂解的半胱氨酸天冬氨酸蛋白酶-3(cleaved caspase-3)蛋白水平。结果:体外实验结果表明,与对照组比较,SEA20组、SEA40组和EPI组细胞凋亡率升高,OD值降低,MRP1、LRP蛋白水平降低,Topo-Ⅱ蛋白水平升高,差异均有统计学意义(P<0.05);与EPI组比较,SEA40+EPI组细胞凋亡率升高,OD值降低,MRP1、LRP蛋白水平降低,Topo-Ⅱ蛋白水平升高,差异均有统计学意义(P<0.05)。体内实验结果表明,与对照组比较,SEA药物组和EPI药物组裸鼠肿瘤体积缩小,PCNA蛋白水平降低、cleaved caspase-3蛋白水平升高,SEA药物组MRP1水平降低,EPI药物组MRP1水平升高,差异均有统计学意义(P<0.05);与EPI药物组比较,SEA+EPI组裸鼠肿瘤体积缩小,MRP1、PCNA蛋白水平降低,cleaved caspase-3蛋白水平升高,差异均有统计学意义(P<0.05)。结论:SEA可通过抑制膀胱癌5637细胞增殖,诱导细胞凋亡,下调MRP1、LRP蛋白表达,上调Topo-Ⅱ蛋白表达,抑制裸鼠移植瘤肿瘤生长,从而增强对EPI的敏感性。
Objective:To investigate the effect mechanism of Senkyunolide A(SEA)combined with epirubicin(EPI)on bladder cancer 5637 cells,and to explore the sensitivity of Senkyunolide A to epirubicin.Methods:The activity of human ureteral epithelial immortalized cell SV-HUC-1 and human bladder cancer cell 5637 were detected by CCK-8 kit.The latter was selected for follow-up experiment,and the IC50 value was calculated.The cells were randomly divided into control group,Senkyunolide A low-dose group(SEA10 group),Senkyunolide A medium-dose group(SEA20 group)and Senkyunolide A High-dose group(SEA40 group),epirubicin alone(EPI group)and combined treatment group(SEA40+EPI group).Cell apoptosis was detected by flow cytometry.CCk-8 kit was used to detect the optical density(OD)of cells.Multidrug resistance associated protein 1(MRP1),lung resistance protein(LRP)and topoisomerase-Ⅱ(Topo-Ⅱ)protein levels were detected by Western blot.The transplanted tumor model was established in nude mice and randomly divided into control group,SEA drug group,EPI drug group and SEA+EPI group,and intervened with the correspending drug gavage.To detect tumor volume.Protein western blot was used to detect MRP1,PCNA,and Cleaved caspase-3 protein levels in nude mouse transplanted tumor models.Results:Compared with control group,the apoptosis rate of SEA20 group,SEA40 group and EPI group was significantly increased(P<0.05),OD value was significantly decreased(P<0.05),MRP1 and LRP protein levels were significantly decreased(P<0.05).The level of Topo-Ⅱ protein was significantly increased(P<0.05).Compared with EPI group,apoptosis rate of SEA40+EPI group was significantly increased(P<0.05),OD value was significantly decreased(P<0.05),protein levels of MRP1 and LRP were significantly decreased(P<0.05),and protein level of Topo-Ⅱ was significantly increased(P<0.05).In vivo results showed that compared with control group,tumor volume of SEA group and EPI group was significantly decreased(P<0.05),protein levels of PCNA were significantly decreased(P<0.05)
作者
林磊
杨树立
孙超群
李宝山
LIN Lei;YANG Shuli;SUN Chaoqun;LI Baoshan(Department of Urology Surgery,First People’s Hospital of Pingdingshan,Pingdingshan 467000,China;Department of Urology Surgery,First Affiliated Hospital of Henan University,Kaifeng 475000,China)
出处
《山东中医药大学学报》
2022年第5期618-625,共8页
Journal of Shandong University of Traditional Chinese Medicine
基金
河南省科技厅科技攻关项目(编号:182102311205)。
关键词
洋川芎内酯A
膀胱癌
表柔比星
耐药性
耐药蛋白
裸鼠移植瘤
Senkyunolide A
bladder cancer
Epirubicin
drug resistance
drug resistant protein
transplanted tumor in nude mice
