盐酸罗哌卡因对骨肉瘤细胞增殖、侵袭、凋亡的影响及其机制  

Effects of ropivacaine hydrochloride on the proliferation,invasion and apoptosis of osteosarcoma cells and its mechanism

作　　者：于鹏 尹天翔 罗彩云[1] 曾萍[1] YU Peng;YIN Tian-xiang;LUO Cai-yun;ZENG Ping(Anesthesiology Department of Yichang Central People's Hospital,Yichang 443008,China)

机构地区：[1]宜昌市中心人民医院麻醉科,湖北宜昌443008

出　　处：《中国应用生理学杂志》2021年第6期654-659,664,共7页Chinese Journal of Applied Physiology

摘　　要：目的:探讨盐酸罗哌卡因对骨肉瘤细胞增殖、侵袭、凋亡的影响及分子机制。方法:采用逐步增加药物剂量诱导法建立骨肉瘤多柔比星耐药细胞株(U2OS/DOX),用浓度分别为0、20、50、100μg/ml的盐酸罗哌卡因处理U2OS/DOX细胞,作为不同浓度盐酸罗哌卡因处理组;将pcDNA3.1、pcDNA3.1-Livin转染至U2OS/DOX细胞中再用浓度为100μg/ml的盐酸罗哌卡因处理,记为盐酸罗哌卡因100μg/ml+pcDNA3.1组、盐酸罗哌卡因100μg/ml+pcDNA3.1-Livin组。MTT检测细胞增殖抑制率及细胞半数抑制浓度(IC_(50));蛋白质印迹(Western blot)法检测细胞周期蛋白依赖性激酶抑制剂1A(P21)、活化的半胱氨酸天冬氨酸蛋白酶-3(Cleaved Caspase-3)、上皮钙黏蛋白(E-cadherin)、基质金属蛋白酶2(MMP-2)、Livin蛋白表达;克隆形成实验检测细胞克隆形成数;流式细胞术检测细胞凋亡;Transwell检测细胞迁移和侵袭;实时荧光定量PCR(RT-qPCR)检测Livin mRNA表达水平。结果:多柔比星浓度大于1μg/ml时,骨肉瘤细胞U2OS增殖抑制率显著升高,且具有剂量依赖性(P<0.05);多柔比星浓度大于10μg/ml时,骨肉瘤细胞骨肉瘤耐药细胞U2OS/DOX增殖抑制率显著升高,且具有剂量依赖性(P<0.05)。盐酸罗哌卡因处理的U2OS/DOX细胞中P21、Caspase-3、E-cadherin表达水平显著升高,MMP-2表达水平显著降低,细胞增殖抑制率显著升高,克隆形成数显著降低,细胞凋亡率显著升高,细胞迁移、侵袭数显著降低,Livin表达水平显著降低,且呈浓度依赖性(P<0.05)。过表达Livin部分逆转了盐酸罗哌卡因对细胞U2OS/DOX增殖、迁移、侵袭的抑制作用及凋亡的促进作用。结论:盐酸罗哌卡因能明显抑制对多柔比星具有耐药性的骨肉瘤细胞的增殖,迁移和侵袭,明显促进骨瘤细胞凋亡,其机制可能与Livin有关。Objective:To investigate the effects and molecular mechanism of ropivacaine hydrochloride on osteosarcoma cell proliferation,invasion,and apoptosis.Methods:The osteosarcoma doxorubicin-resistant cell line(U2OS/DOX)was established by gradually increasing the drug doses.U2OS/DOX cells were treated with ropivacaine hydrochloride at the concentrations of 0,20,50 and 100μg/ml,respectively;as different concentrations treatment groups of ropivacaine hydrochloride.pcDNA3.1 and pcDNA3.1-Livin were transfected into U2OS/DOX cells and then treated with 100μg/ml ropivacaine hydrochloride,which were defined as ropivacaine hydrochloride 100μg/ml+pcDNA3.1 group,ropivacaine hydrochloride 100μg/ml+pcDNA3.1-Livin group.MTT was used to detect the cell proliferation inhibition rate and inhibitory concentration(IC 50).Western blot was used to detect the expressions of cyclin-dependent kinase inhibitor 1A(P21)and activated cysteine aspartic protease-3(Cleaved Caspase-3),E-cadherin,matrix metalloproteinase 2(MMP-2)and Livin;clone formation experiments were used to detect the number of cell clones formed;flow cytometry was used to detect apoptosis;Transwell was used to detect cell migration and invasion;real-time fluorescent quantitative PCR(RT-qPCR)was used to detect Livin mRNA expression.Results:When the concentration of doxorubicin was more than 1μg/ml,the proliferation inhibition rate of osteosarcoma cells U2OS was significantly increased in a concentration-dependent manner(P<0.05);when the concentration of doxorubicin was more than 10μg/ml,the proliferation inhibition rate of osteosarcoma resistant cell U2OS/DOX was significantly increased,and it was dose-dependent(P<0.05).In U2OS/DOX cells treated with ropivacaine hydrochloride,the expressions of P21,Cleaved Caspase-3,and E-cadherin were increased significantly,the expression of MMP-2 was decreased significantly,the cell proliferation inhibition rate was increased significantly,the number of colony formation was decreased significantly,and the cells apoptosis rate was increa

关 键 词：盐酸罗哌卡因 LIVIN 骨肉瘤 增殖 侵袭 凋亡 化疗敏感性 

分 类 号：R738.1[医药卫生—肿瘤]