艾司氯胺酮对肝癌细胞恶性生物学行为的影响及作用机制实验研究  

Effect of esketamine on malignant biological behavior of hepatocellular carcinoma cells and its mechanism

作　　者：吴洁 田鑫越 秦耐宇 朱爱玲 孙佳琦 朱颖静 WU Jie;TIAN Xinyue;QIN Naiyu;ZHU Ailing;SUN Jiaqi;ZHU Yingjing(Department of Gastroenterology,the People’s Hospital of Rugao,Rugao 226500,China)

机构地区：[1]如皋市人民医院消化内科,江苏如皋226500

出　　处：《陕西医学杂志》2024年第8期1041-1046,1050,共7页Shaanxi Medical Journal

基　　金：江苏省卫生健康委员会医学科研项目(ZD20210079)。

摘　　要：目的:探讨艾司氯胺酮对肝癌细胞恶性生物学行为的影响及可能的作用机制。方法:收集26例肝癌患者的癌组织和癌旁组织。实时荧光定量聚合酶链反应(RT-qPCR)检测肝癌组织和细胞微小RNA(miR)-449b-5p和转化生长因子-β诱导因子同源盒2(TGIF2)mRNA表达。选取上述基因变化最显著的细胞进行后续实验。将肝癌SK-Hep-1细胞分为对照组(正常培养)、艾司氯胺酮-L组(1 mmol/L艾司氯胺酮处理)、艾司氯胺酮-H组(2 mmol/L艾司氯胺酮处理)、艾司氯胺酮-H+inhibitor NC组(转染inhibitor NC+2 mmol/L艾司氯胺酮处理)、艾司氯胺酮-H+miR-449b-5p inhibitor组(转染miR-449b-5p inhibitor+2 mmol/L艾司氯胺酮处理)。细胞计数试剂盒(CCK-8)法检测细胞增殖。划痕实验检测细胞迁移能力。Transwell实验检测细胞侵袭能力。流式细胞术检测细胞凋亡情况。Western blot检测细胞中肿瘤增殖细胞核抗原(Ki-67)、半胱氨酸蛋白酶-3(Cleaved caspase-3)、基质金属蛋白酶-9(MMP-9)、血管内皮生长因子A(VEGF-A)、TGIF2蛋白表达量。双荧光素酶报告基因实验验证miR-449b-5p和TGIF2的靶向关系。结果:肝癌组织和细胞TGIF2 mRNA表达水平升高,miR-449b-5p表达水平降低(均P<0.05)。与对照组比较,艾司氯胺酮-L组、艾司氯胺酮-H组以及艾司氯胺酮-H+inhibitor NC组SK-Hep-1细胞中miR-449b-5p表达水平、细胞凋亡率及Cleaved caspase-3蛋白表达水平升高,TGIF2 mRNA表达水平、OD_(450)值(24、48 h)、细胞划痕愈合率、细胞侵袭数及Ki-67、MMP-9、VEGF-A、TGIF2蛋白表达水平降低(均P<0.05)。与艾司氯胺酮-H+inhibitor NC组比较,艾司氯胺酮-H+miR-449b-5p inhibitor组SK-Hep-1细胞miR-449b-5p表达水平、细胞凋亡率及Cleaved caspase-3蛋白表达水平降低,TGIF2 mRNA表达水平、OD_(450)值(24、48 h)、细胞划痕愈合率、细胞侵袭数及Ki-67、MMP-9、VEGF-A、TGIF2蛋白表达水平升高(均P<0.05)。miR-449b-5p mimic+TGIF2-WT组荧光素酶活性低�Objective:To investigate the effect of esketamine on the malignant biological behavior of hepatocellular carcinoma cells and its possible mechanism.Methods:Cancer tissues and adjacent tissues of 26 patients with liver cancer were collected.Real-time quantitative polymerase chain reaction(RT-qPCR)was used to detect the expression of miR-449b-5p and transforming growth factor-β-induced factor homeobox 2(TGIF2)mRNA in liver cancer tissues and cells.The cells with the most significant changes in the above genes were selected for subsequent experiments.SK-Hep-1 cells were divided into control group(normal culture),esketamine-L group(1 mmol/L esketamine treatment),esketamine-H group(2 mmol/L esketamine treatment),esketamine-H+inhibitor NC group(transfected with inhibitor NC+2 mmol/L esketamine treatment),and esketamine-H+miR-449b-5p inhibitor group(transfected with miR-449b-5p inhibitor+2 mmol/L esketamine treatment).Cell proliferation was detected by the cell counting kit(CCK-8).The scratch assay was used to detect cell migration ability.The Transwell assay was used to detect cell invasion ability.Flow cytometry was used to detect apoptosis ability.Western blot was used to detect the protein expressions of tumor proliferating cell nuclear antigen(Ki-67),Cleaved caspase-3,matrix metalloproteinase-9(MMP-9),vascular endothelial growth factor A(VEGF-A)and TGIF2 in cells.Dual luciferase reporter gene assay verified the targeting relationship between miR-449b-5p and TGIF2.Results:The mRNA expression level of TGIF2 in hepatocellular carcinoma tissues and cells was increased,and the expression level of miR-449b-5p decreased(all P<0.05).Compared with the control group,the expression level of miR-449b-5p,the apoptosis rate and the expression level of Cleaved caspase-3 protein in SK-Hep-1 cells in the esketamine-L group,esketamine-H group and esketamine-H+inhibitor NC groups were increased,and the expression level of TGIF2 mRNA,OD_(450) value(24 and 48 hours),cell scratch healing rate,cell invasion number,and the protein expres

关 键 词：肝癌 艾司氯胺酮 微小RNA-449b-5p 转化生长因子-β诱导因子同源盒2 细胞增殖 细胞迁移 细胞侵袭 细胞凋亡 

分 类 号：R36[医药卫生—病理学]