环状RNA_0054537联合连翘苷调控肝癌细胞生物学特性的分子机制  

Molecular mechanism of circ_0054537 combined with phillyrin to regulate the proliferation, migration, invasion and apoptosis of liver cancer cells

作　　者：高志强[1] 陈君[2] 李丁洋 刘兆臣 葛鹏磊 张弓[1] 党晓卫[1] Gao Zhiqiang;Chen Jun;Li Dingyang;Liu Zhaochen;Ge Penglei;Zhang Gong;Dang Xiaowei(Department of Hepatic and Biliary Pancreatic Surgery,the First Affiliated Hospital of Zhengzhou University,Henan Key Laboratory of Digestive Organ Transplantation,Zhengzhou 450052,China;Department of Cardiology,the First Affiliated Hospital of Zhengzhou University,Zhengzhou 450052,China)

机构地区：[1]郑州大学第一附属医院肝胆胰外科河南省消化器官移植重点实验室,450052 [2]郑州大学第一附属医院心内科,450052

出　　处：《中华实验外科杂志》2020年第8期1464-1468,共5页Chinese Journal of Experimental Surgery

基　　金：河南省高等学校重点科研项目(21A320061)。

摘　　要：目的探讨连翘苷对肝癌细胞增殖、迁移、侵袭、凋亡的影响及其对环状RNA_0054537(circ_0054537)/微小RNA(miRNA,miR)-409-3p的调控作用。方法体外培养肝癌细胞Hep3B,使用不同剂量(5、10、20μmol/L)的连翘苷处理Hep3B细胞;采用细胞计数试剂盒(CCK-8)法检测细胞增殖;流式细胞术检测细胞凋亡率;Transwell小室实验检测细胞迁移及侵袭能力;采用实时定量反转录聚合酶链反应(RT-qPCR)法检测circ_0054537、miR-409-3p的表达量;双荧光素酶报告实验检测circ_0054537、miR-409-3p的靶向关系;Hep3B细胞中分别转染si-circ_0054537、pcDNA-circ_0054537,采用上述方法检测细胞增殖、凋亡、迁移及侵袭。组间比较采用独立样本t检验,多组间比较采用单因素方差分析。结果连翘苷可明显降低细胞活力[(1.276±0.110)个比(1.031±0.090)、(0.637±0.050)、(0.507±0.040)个,F=244.841,P<0.05],增高凋亡率[(7.34±0.62)%比(12.44±1.03)%、(22.86±2.12)%、(28.16±2.71)%,F=244.841,P<0.05],减少迁移细胞数[(154.15±12.76)个比(124.06±10.04)、(78.05±7.25)、(61.13±5.83)个]及侵袭细胞数[(112.04±10.08)个比(89.43±8.13)、(57.11±5.12)、(43.08±4.05)个,F=186.018、166.514,P<0.05],抑制circ_0054537的表达[(1.00±0.10)比(0.40±0.04),t=16.713,P<0.05],促进miR-409-3p的表达[(1.02±0.11)比(3.42±0.31),t=21.889,P<0.05];转染si-circ_0054537可明显抑制细胞活力[(1.264±0.10)比(0.686±0.06)]、迁移[(151.96±11.17)比(79.03±7.21)]及侵袭能力[(115.28±11.05)比(68.14±6.43),t=14.869、16.457、11.062,P<0.05],增高凋亡率[(7.26±0.71)%比(23.79±2.06)%,t=22.759,P<0.05];双荧光素酶报告实验证实circ_0054537可靶向结合miR-409-3p;转染pcDNA-circ_0054537可明显逆转连翘苷对Hep3B细胞增殖、凋亡、迁移、侵袭的调控作用。结论连翘苷可通过下调circ_0054537的表达而上调miR-409-3p的表达从而抑制肝癌细胞增殖、迁移、侵袭及促进细胞凋亡。Objective To explore the effect of phillyrin on the proliferation,migration,invasion and apoptosis of hepatoma cells and its regulatory effect on circ_0054537/miR-409-3p.Methods Hep3B liver cancer cells were cultured in vitro,and Hep3B cells were treated with phillyrin at different doses(5,10,20μmol/L).Cell counting kit-8(CCK-8)method was used to detect cell proliferation.Flow cytometry was used to detect the apoptosis rate.Transwell cell test was used to detect cell migration and invasion ability.The expression levels of circ_0054537 and miR-409-3p were detected by real-time quantitative reverse transcriptase-polymerase chain reaction(RT-qPCR)method.The dual luciferase report experiment detected the targeting relationship of circ_0054537 and miR-409-3p.Hep3B cells were transfected with si-circ_0054537,pcDNA-circ_0054537,respectively,and cell proliferation,apoptosis,migration and invasion were detected by the above methods.SPSS 21.0 statistical software was used to analyze the data.The measurement data were expressed as(Mean±SD)and all accorded with normal distribution.The comparison between the two groups was performed by independent sample t test.The comparison between groups was performed by one-way analysis of variance,with P<0.05 The difference was statistically significant.Results Phillyrin could significantly reduce cell viability[(1.276±0.110)vs.(1.031±0.090),(0.637±0.050),(0.507±0.040),F=244.841,P<0.05],increased apoptosis rate[(7.34±0.62)%vs.(12.44±1.03)%,(22.86±2.12)%,(28.16±2.71)%,F=244.841,P<0.05],reduce the number of migrating cells[(154.15±12.76)vs.(124.06±10.04),(78.05±7.25),(61.13±5.83)]and the number of invasive cells[(112.04±10.08)vs.(89.43±8.13),(57.11±5.12),(43.08±4.05),F=186.018,166.514,P<0.05],inhibit the expression of circ_0054537[(1.00±0.10)vs.(0.40±0.04),t=16.713,P<0.05],promote the expression of miR-409-3p[(1.02±0.11)vs.(3.42±0.31),t=21.889,P<0.05].Transfection of si-circ_0054537 could significantly inhibit cell viability[(1.264±0.10)vs.(0.686±0.06)],migration[(1

关 键 词：肝癌 连翘苷 环状RNA_0054537 微小RNA-409-3p 磷脂酰肌醇3激酶/蛋白激酶B信号通路 

分 类 号：R735.7[医药卫生—肿瘤]