刺槐素影响人骨肉瘤MG-63细胞增殖和凋亡分子机制  

Effect of acacetin on proliferation and apoptosis in human osteosarcoma MG-63 cells and its mechanism

作　　者：陈伟达[1] 袁磊[3] 王桁扬 谭嗣伟 刘文庆 苗成利[1] CHEN Wei-da;YUAN Lei;WANG Heng-yang;TAN Si-wei;LIU Wen-qing;MIAO Cheng-li(Peking University International Hospital,Beijing102206,P.R.China;Key Laboratory of Medical Bioengineering,Luohe Medical College,Luohe 462002,P.R.China;First Department of General Surgery,Ninth Hospital of Xi'an,Xi'an 710054,P.R.China)

机构地区：[1]北京大学国际医院腹膜后肿瘤外科,北京102206 [2]北京大学国际医院肛肠外科,北京102206 [3]漯河医学高等专科学校医学生物工程重点实验室,河南漯河462002 [4]西安市第九医院普外一科,陕西西安710054

出　　处：《中华肿瘤防治杂志》2020年第20期1616-1621,共6页Chinese Journal of Cancer Prevention and Treatment

基　　金：首都科委特色临床应用研究项目(Z161100000516025)。

摘　　要：目的刺槐素(acacetin,Aca)是从豆科植物刺槐的花中提取的一种黄酮类化合物,具有抗感染、抗菌和抗肿瘤等药理作用,但是否具有抗骨肉瘤活性尚不明确。本研究旨在探讨Aca对人骨肉瘤MG-63细胞增殖和凋亡的影响及其可能的分子机制。方法体外常规培养MG-63细胞,用0、15、30、60、120和240μmol/L Aca处理MG-63细胞24或48h后,CCK-8法检测细胞活力,集落形成实验检测细胞增殖,AnnexinⅤ-PI染色检测细胞凋亡,PI染色检测细胞周期,蛋白质印迹法分析Survivin、Cdc25C、c-Met、p-c-Met、Akt和p-Akt蛋白的表达。结果 Aca可抑制MG-63细胞活力,24和48h的IC50分别为77.94和57.36μmol/L,且具有剂量和时间依赖性。MG-63细胞经0、30、60和120μmol/L Aca作用24h后,平板克隆形成实验结果显示,克隆形成率分别为(62.73±4.59)%、(50.42±3.83)%、(37.65±3.14)%和(25.18±2.67)%,差异有统计学意义,F=59.64,P<0.001;AnnexinⅤ-PI染色结果显示,细胞凋亡率分别为(2.41±0.68)%、(9.72±1.43)%、(20.36±1.92)%和(31.85±2.27)%,差异有统计学意义,F=174.89,P<0.001;PI染色结果显示,G2/M期细胞构成比分别为(10.73±1.81)%、(20.36±2.05)%、(31.58±1.72)%和(42.11±2.16)%,差异有统计学意义,F=138.42,P<0.001;蛋白质印迹法结果显示,Survivin和Cdc25C蛋白表达水平以及c-Met和Akt蛋白磷酸化水平随药物剂量增加而降低,两两比较差异均有统计学意义,F值分别为110.49、70.85、93.72和96.14,均P<0.001。结论 Aca可诱导MG-63细胞发生G2/M期阻滞和凋亡,这可能与其下调Survivin和Cdc25C蛋白表达,以及抑制c-Met/Akt信号通路有关。OBJECTIVE Acacetin(Aca),a flavone isolated from Robinia pseudoacacia,has been reported to have anti-inflammatory,antibacterial and antineoplastic properties.The objective of this study was to investigate the effect of Aca on proliferation and apoptosis in human osteosarcoma MG-63 cells and its possible mechanism.METHODS MG-63 cells were cultured in vitro and treated with CTS(0,15,30,60,120 and 240μmol/L)for 24 hor 48 hand then the cell viability were detected by CCK-8 assay.The proliferation was observed by plate clone formation assay.Quantification of apoptosis was measured by Annexin Ⅴ-PI assay.The cell cycle was determined by PI assay.The protein expression of Survivin,Cdc25 C,c-Met,p-c-Met,Akt and p-Akt were measured by western blot.RESULTS Aca remarkably reduced the cell viability of MG-63 cells with IC50 during 24 hand 48 hof 77.94μmol/L and 57.36μmol/L respectively,in a dose-dependent and time-dependent manner.Plate clone formation assay showed that Aca inhibited proliferation of MG-63 cells in a dose-dependent manner[(50.42±3.83)%,(37.65±3.14)%,(25.18±2.67)% vs(62.73±4.59)%,F=59.64,P<0.001].AnnexinⅤ-PI assay showed that Aca promoted apoptosis of MG-63 cells in a dose-dependent manner[(9.72±1.43)%,(20.36±1.92)%,(31.85±2.27)%vs(2.41±0.68)%,F=174.89,P<0.001].PI assay showed that Aca induced G2/M phase arrest of MG-63 cells in a dose-dependent manner[(20.36±2.05)%,(31.58±1.72)%,(42.11±2.16)%vs(10.73±1.81)%,F=138.42,P<0.001].Western blot results showed that Aca down-regulated the protein levels of Survivin and Cdc25 C(Fvalues were 110.49 and 70.85,both P<0.001),and suppressed the protein phosphorylation of c-Met and Akt in a dose-dependent manner(Fvalues were 93.72 and 96.14,both P<0.001).CONCLUSIONS Aca may induce G2/M phase arrest and apoptosis of MG-63 cells through down-regulating the protein levels Survivin and Cdc25 C,and suppressing the c-Met and Akt signal pathway.

关 键 词：刺槐素 骨肉瘤 MG-63细胞 G2/M期阻滞 凋亡 

分 类 号：R738.6[医药卫生—肿瘤]