石蒜碱对胃癌AGS细胞恶性生物学行为的影响及机制研究  

Effects of lycorine on malignant biological behaviors of AGS gastric cancer cells and its mechanism

作　　者：郑金蒂 赖铭裕[1] 程若溪 ZHENG Jindi;LAI Mingyu;CHENG Ruoxi(Department of Gastroenterology,the First Affiliated Hospital of Guangxi Medical University,Nanning 530021,Guangxi,China;Department of Gastroenterology,the Second Affiliated Hospital of Guangxi Medical University,Nanning 530007,Guangxi,China)

机构地区：[1]广西医科大学第一附属医院消化内科,南宁市530021 [2]广西医科大学第二附属医院消化内科,南宁市530007

出　　处：《内科》2024年第4期363-371,共9页Internal Medicine

基　　金：广西医疗卫生适宜技术开发与推广应用(S2019100)。

摘　　要：目的探讨石蒜碱对胃癌AGS细胞恶性生物学行为的影响并初步探讨其可能的作用机制。方法用不同浓度的盐酸石蒜碱(LH)(0μmol/L、0.015μmol/L、0.0625μmol/L、0.25μmol/L、1μmol/L、4μmol/L、16μmol/L)干预AGS细胞0 h、24 h和48 h,采用细胞计数试剂盒-8(CCK-8)试验计算干预48 h时药物半数抑制浓度(IC50),以确定后续试验的LH浓度。将AGS细胞分为对照组(0.1%二甲基亚砜处理)、LH-L组(0.125μmol/L LH处理)、LH-M组(0.5μmol/L LH处理),LH-H组(2μmol/L LH处理),以及微小RNA(miRNA)-675-nc组(转染慢病毒空白载体)、miRNA-675-mimic组(转染慢病毒miRNA-675过表达载体)和LH+miRNA-675-mimic组(转染慢病毒miRNA-675过表达载体+2μmol/L LH处理)。应用CCK-8试验、平板克隆试验、细胞划痕愈合试验和Transwell试验分别检测各组细胞的增殖、迁移和侵袭能力;应用实时荧光定量PCR检测miRNA-675和糖原合成酶激酶(GSK)-3βmRNA的相对表达水平;应用蛋白质印迹法检测GSK-3β和β-联蛋白(β-catenin)蛋白相对表达水平。结果在0.015~16μmol/L范围内,LH可时间-浓度依赖地抑制AGS细胞的增殖能力(P<0.05),干预48 h时,药物IC50为(0.81±0.13)μmol/L。对照组、LH-L组、LH-M组、LH-H组干预14 d后细胞克隆数,干预24 h、48 h后划痕愈合率,干预24 h后侵袭细胞数、迁移细胞数,干预48 h后miRNA-675相对表达水平均依次降低;干预48 h后LH-M组细胞的GSK-3βmRNA相对表达水平高于对照组,LH-H组细胞的GSK-3βmRNA相对表达水平高于对照组、LH-L组、LH-M组;干预48 h后LH-M组GSK-3β蛋白相对表达水平高于对照组,LH-H组GSK-3β蛋白相对表达水平高于对照组和LH-L组;干预48 h后LH-H组β-catenin蛋白相对表达水平低于对照组和LH-L组(均P<0.05)。miRNA-675-mimic组的光密度值(48 h)、细胞克隆数(14 d)、细胞侵袭数(24 h)、细胞迁移数(24 h)、β-catenin蛋白相对表达水平(48 h)均高于miRNA-675-nc组、LH+miRNA-675-mimic组;miRNA-675-mimic组GObjective To investigate the effects of lycorine on the malignant biological behaviors of AGS gastric cancer cells,and to preliminarily explore its possible action mechanism.Methods AGS cells were intervened with different concentrations of lycorine hydrochloride(LH)(0μmol/L,0.015μmol/L,0.062,5μmol/L,0.25μmol/L,1μmol/L,4μmol/L,16μmol/L)for 0 hour,24 hours,and 48 hours,and its inhibitory concentration 50(IC50)at the 48th hour of the intervention was calculated by the cell counting kit-8(CCK-8)test to determine the LH concentration in subsequent trials.AGS cells were divided into a control group(treated with 0.1%dimethyl sulfoxide),an LH-L group(treated with 0.125μmol/L LH),an LH-M group(treated with 0.5μmol/L LH),or an LH-H group(treated with 2μmol/L LH),as well as a microRNA(miRNA)-675-nc group(transfected with blank lentiviral vectors),a miRNA-675-mimic group(transfected with lentiviral vectors overexpressing miRNA-675),or an LH+miRNA-675-mimic group(transfected with lentiviral vectors overexpressing miRNA-675+treated with 2μmol/L LH).CCK-8 test,clonogenic assay in plates,cell scratch healing assay,and Transwell test were separately used to detect the proliferation,migration,and invasion abilities of cells in each group;real-time fluorogenic quantitative PCR was used to detect the relative expression levels of miRNA-675 and glycogen synthase kinase(GSK)-3βmRNA;western blotting was used to detect the relative protein expression levels of GSK-3βandβ-catenin.Results In the range of 0.015-16μmol/L,LH could inhibit the proliferation ability of AGS cells in a time-concentration dependent manner(P<0.05),and its IC50 was(0.81±0.13)μmol/L at the 48th hour of the intervention.The number of cell colonies after 14 days of intervention,the scratch healing rates 24 hours and 48 hours after the intervention,the numbers of invasive cells and migrated cells 48 hours after the intervention,and the relative expression level of miRNA-67548 hours after the intervention in the control group,LH-L group,LH-M group,and

关 键 词：胃癌 胃癌AGS细胞 石蒜碱 微小RNA-675 糖原合成酶激酶-3Β β-联蛋白 

分 类 号：R735.2[医药卫生—肿瘤]