乌苏酸对人胰腺癌细胞PANC-1增殖和凋亡的影响  

Effects of Ursolic Acid on Proliferation and Apoptosis of Human Pancreatic Cancer Cell PANC-1

作　　者：金俊华[1] 赵承伟 付佳 郑桂茹[1] JIN Junhua;ZHAO Chengwei;FU Jia;ZHENG Guiru(Department of Pharmacy,Jinhua People's Hospital,Jinhua,Zhejiang,China 321000;Institute of Biomedicine,School of Pharmaceuti-cal Science,Wenzhou Medical University,Zhuji,Zhejiang,China 311800)

机构地区：[1]浙江省金华市人民医院药剂科,浙江金华321000 [2]温州医科大学药学院<诸暨>生物医药研究院,浙江诸暨311800

出　　处：《中国药业》2024年第2期46-51,共6页China Pharmaceuticals

基　　金：浙江省药学会医院药学专项科研资助项目[2019ZYY52];浙江省金华市科学技术研究计划项目[2020-4-026]。

摘　　要：目的探讨乌苏酸对人胰腺癌细胞PANC-1增殖、凋亡的影响。方法以1.25,2.5,5,10,25,50μmol/L乌苏酸培养PANC-1细胞24,48,72 h,采用四氮唑盐(MTT)法测定细胞活性。实验分为对照1组(等体积二甲基亚砜)及乌苏酸低、中、高剂量组(5,10,20μmol/L乌苏酸),显微镜下观察细胞形态,采用Western blot法检测磷脂酰肌醇3激酶(PI3K),磷酸化的蛋白激酶B(p-Akt),磷酸化哺乳动物雷帕霉素靶蛋白(p-m TOR),活化半胱氨酸蛋白酶3(Cleaved Caspase-3),B淋巴细胞瘤-2(Bcl-2),Bcl-2关联X蛋白(Bax)的蛋白表达水平,采用细胞集落形成实验观察细胞增殖情况。实验分为对照2组(等体积二甲基亚砜)和乌苏酸组(10μmol/L乌苏酸),采用细胞划痕实验观察细胞培养48,72 h的迁移情况。利用分子对接实验模拟乌苏酸与PI3K和Akt2的相互作用。结果随着乌苏酸浓度的升高,PANC-1细胞活性逐渐减弱,24,48,72 h时的半数抑制浓度(IC50)分别为7.89,6.26,5.06μmol/L。与对照1组比较,乌苏酸各剂量组细胞逐渐失去原有形态,且随着浓度的增加,变形细胞数目随之增加,且细胞边界模糊不清;细胞数量显著减少(P<0.05);乌苏酸各剂量组细胞Cleaved Caspase-3、Bax蛋白的表达水平均显著升高,乌苏酸中、高剂量组细胞Bcl-2蛋白表达水平显著降低,乌苏酸各剂量组细胞p-m TOR,中、高剂量组细胞p-Akt,高剂量组细胞PI3K蛋白表达水平均显著降低(P<0.05)。与对照2组比较,乌苏酸组细胞48 h,72 h的迁移距离缩短。乌苏酸的乌苏烷型三萜类结构可进入PI3K与Akt2中的三磷酸腺苷(ATP)结合位点竞争性结合疏水口袋,从而影响PI3K和Akt2与ATP的结合,抑制其激活。结论乌苏酸可通过抑制PI3K/Akt/m TOR信号通路的激活而抑制PANC-1细胞的增殖,促进其凋亡。Objective To investigate the effects of ursolic acid on the proliferation and apoptosis of human pancreatic cancer cell PANC-1.Methods PANC-1 cells were cultured with 1.25,2.5,5,10,25 and 50μmol/L ursolic acid for 24,48 and 72 h,and the cell viability was measured by the methyl thiazolyl tetrazolium(MTT)method.PANC-1 cells were divided into the control group one(equal volume of dimethyl sulfoxide)and the ursolic acid low-,medium-and high-dose groups(5,10 and 20μmol/L ursolic acid),the cell morphology was observed by microscope,the expression levels of phosphatidylinositol 3-kinase(PI3K),phosphorylated-protein kinase B(p-Akt),phosphorylated-mammalian target of rapamycin(p-mTOR),Cleaved Caspase-3,B-cell lymphoma-2(Bcl-2),Bcl-2-associated X(Bax)proteins were detected by the Western blot,and the cell proliferation was observed by the colony formation assay.PANC-1 cells were divided into the control group two(equal volume of dimethyl sulfoxide)and the ursolic acid group(10μmol/L ursolic acid),the cell migration was observed by the wound healing assay after 48 and 72 h of culture.The interaction of ursolic acid with PI3K and Akt2 proteins was simulated by the molecular docking.Results As the concentration of ursolic acid increased,the viability of PANC-1 cells gradually weakened,with the half-inhibitory concentrations(IC50)of 7.89,6.26 and 5.06μmol/L at 24,48 and 72 h,respectively.Compared with those in the control group one,the cell morphology in the ursolic acid low-,medium-and high-dose groups was abnormal,with the increase of concentration,the number of deformed cells increased,and the cell boundaries became blurred;the number of cells also significantly decreased(P<0.05).Compared with those in the control group one,the expression levels of Cleaved Caspase-3 and Bax proteins in each dose group of ursolic acid significantly increased;the expression level of Bcl-2 protein in the ursolic acid medium-and high-dose groups significantly decreased;the expression levels of p-mTOR protein in the ursolic acid low-,medium

关 键 词：乌苏酸 人胰腺癌细胞PANC-1 PI3K/Akt/mTOR信号通路 细胞凋亡 

分 类 号：R965[医药卫生—药理学]