辛伐他汀对胆管癌细胞周期相关蛋白的作用  

Effect of simvastatin on cell cycle-related proteins in cholangiocarcinoma cells

作　　者：蔡建鹏[1] 张昆松[1] 陈伟[1] 王曦域 陈流华[1] Cai Jianpeng;Zhang Kunsong;Chen Wei;Wang Xiyu;Chen Liuhua(Department of Pancreaticobiliary Surgery,the First Affiliated Hospital of Sun Yat-sen University,Guangzhou 510080,China)

机构地区：[1]中山大学附属第一医院胆胰外科,广州510080

出　　处：《中华普通外科学文献（电子版）》2021年第1期5-10,共6页Chinese Archives of General Surgery(Electronic Edition)

基　　金：广东省医学科研基金项目(A2017318)。

摘　　要：目的了解辛伐他汀对胆管癌细胞周期相关蛋白Cyclin D1、Cyclin E、CDK4及pRB的作用。方法体外实验使用辛伐他汀处理胆管癌细胞EGI-124 h和48 h后,利用CCK-8试剂盒测定肿瘤细胞增殖情况,其后实时荧光定量PCR(qPCR)检测细胞周期相关基因Cyclin D1、Cyclin E、CDK4及pRB的mRNA,根据结果再用Western blotting验证相应蛋白的表达水平。体内实验使用EGI-1接种于12只NOD-SCID小鼠皮下,成瘤后随机分为实验组和对照组,每组6例。实验组每天按照20 mg/kg的辛伐他汀灌胃,对照组每天喂予同剂量的药物溶剂。24 d后取出移植瘤测量体积并进行免疫组织化学染色,明确上述蛋白的体内表达水平。结果辛伐他汀处理对胆管癌细胞EGI-1有增殖抑制作用,呈现药物的浓度依赖性,其24 h及48 h的半抑制浓度数值分别为(8.27±0.77)μmol/L和(5.90±1.81)μmol/L。辛伐他汀处理48 h的EGI-1细胞内Cyclin D1、CDK4和pRB mRNA表达显著下调(P<0.05),而Cyclin E mRNA表达差异无统计学意义。相应地,EGI-1细胞内Cyclin D1、CDK4、pRB蛋白也被抑制,与mRNA的结果保持一致;同时随药物时间延长,抑制作用增强。体内实验显示,辛伐他汀可抑制小鼠移植瘤生长,免疫组织化学分析显示实验组小鼠移植瘤细胞Cyclin D1、CDK4、pRB蛋白下调(P<0.05)。结论辛伐他汀可通过下调细胞周期相关蛋白Cyclin D1、CDK4、pRB抑制胆管癌细胞增殖。Objective To investigate the effect of simvastatin on cell cycle related proteins Cyclin D1,Cyclin E,CDK4 and pRB in cholangiocarcinoma.Methods In vitro,after 24 and 48 h of treatment with simvastatin,the proliferation of cholangiocarcinoma cells EGI-1 was detected by cell counting kit-8(CCK-8).Then qPCR were employed to determine the expressions of cell cycle-related genes Cyclin D1,Cyclin E,CDK 4 and pRB.And related proteins were further analyzed by Western blotting according to the result of qPCR.In vivo,twelve NOD-SCID mice were inoculated with EGI-1 cell,then they were randomly divided into experimental group and control group after tumor formation,with 6 rats in each group.The experimental group took 20 mg/kg simvastatin every day,while the control group took simvastatin solvent every day.Simvastatin were given at a dose of 20 mg/kg once a day for the experimental group,while the solvent were given for the control group.24 days later,the volume of tumors were measured and immunohistochemical staining was performed to determine the expression level of the above proteins in vivo.Results The proliferation of EGI-1 cell was suppressed by simvastatin in a dosedependent manner.The half maximal inhibitory concentration of 24 and 48 h were(8.27±0.77)μmol/L and(5.90±1.81)μmol/L,respectively.Simvastatin significantly suppressed mRNA expressions of Cyclin D1,CDK4 and pRB after 48 h(P<0.05).No significant difference was found in mRNA expression of Cyclin E.Correspondingly,proteins of Cyclin D1,CDK4 and pRB were downregulated,which were consistent with the results of mRNA.And increasing simvastatin treated time could elevated this suppression.Simvastatin treatment yielded a significant growth inhibition in vivo.Immunohistochemical staining illustrated that Cyclin D1,CDK4 and pRB expressions were markedly downregulated by simvastatin in transplanted tumor(P<0.05).Conclusion Simvastatin inhibits cholangiocarcinoma cell by down-regulating cyclins such as Cyclin D1,CDK4,pRB.

关 键 词：胆管肿瘤 辛伐他汀 细胞周期蛋白类 

分 类 号：R735.8[医药卫生—肿瘤]