摘要
目的探讨紫草素(SHI)调节Notch信号通路对肝癌细胞恶性生物学活性的影响。方法采用Western blot分别检测肝癌组织、癌旁组织、肝癌细胞(HepG2、Hep3B、HCCLM3、Huh-7、SMMC-7721细胞)及正常肝细胞(HL-7702细胞)中Notch、发状分裂相关增强子-1(HES1)、hairy相关转录因子1(HEY1)蛋白表达;将Huh-7细胞分为对照组、L-SHI组(1μmol/L SHI)、M-SHI组(2μmol/L SHI)、H-SHI组(4μmol/L SHI)、DAPT组(50μmol/L Notch信号抑制剂DAPT)、H-SHI+丙戊酸(VPA)组(4μmol/L SHI和8 mmol/L Notch通路激活剂VPA)。CCK-8法和平板克隆实验检测Huh-7细胞增殖;流式细胞术检测Huh-7细胞凋亡以及细胞周期;细胞划痕实验以及Transwell侵袭实验分别检测Huh-7细胞迁移和侵袭情况;Western blot检测上皮间质转化(EMT)及凋亡相关蛋白表达。结果Notch、HES1、HEY1在肝癌组织和细胞中表达水平明显升高,且Huh-7细胞差异最明显,以Huh-7细胞为研究对象。与对照组相比,L-SHI组、M-SHI组、H-SHI组、DAPT组Notch、HES1、HEY1、Bcl-2蛋白表达水平下调,S期和G2/M期细胞比例、OD450值及克隆数、迁移率、侵袭细胞数目、N-cadherin、Vimentin水平降低(P<0.05),G1/G0期细胞比例、凋亡率、Bax、cleaved-Caspase-3、E-cadherin水平升高(P<0.05),且SHI的作用效果呈现剂量依赖性;与H-SHI组相比,H-SHI+VPA组以上指标趋势相反,VPA减弱了SHI降低肝癌细胞恶性生物学活性的作用。结论SHI可能通过抑制Notch信号通路抑制Huh-7细胞增殖、迁移和侵袭,促进Huh-7细胞凋亡。
Objective To investigate the impact of shikonin(SHI)on the malignant biological activity of liver cancer cells by regulating Notch signaling pathway.Methods Western blot assay was used to detect the expression of Notch,Hairy mitosis-related enhancer-1(Hes1),hairy-related transcription factor-1(HEY1)protein in liver cancer tissue,paracancerous tissue,hepatoma cells(HepG2 cells,Hep3B cells,HCCLM3 cells,Huh-7 cells and SMMC-7721 cells)and normal liver cells(HL-7702 cells).Huh-7 cells were divided into the control group,the L-SHI group(1μmol/L SHI),the M-SHI group(2μmol/L SHI),the H-SHI group(4μmol/L SHI),the DAPT group(50μmol/L Notch signal inhibitor DAPT)and the H-SHI+VPA group[4μmol/L SHI and 8 mmol/L Notch pathway activator Valproic acid(VPA)].The proliferation of Huh-7 cells was detected by CCK-8 method and plate cloning test.The apoptosis and cell cycle of Huh-7 cells were detected by flow cytometry.Cell scratch test and Transwell invasion test were used to detect migration and invasion of Huh-7 cells.Western blot assay was used to detect the expression of epithelial-mesenchymal transformation(EMT)and apoptosis related proteins.Results The expression levels of Notch,HES1 and HEY1 were obviously increased in liver cancer tissue and cells,and Huh-7 cells showed the most obvious difference,therefore,Huh-7 cells were taken as the research object.Compared with the control group,the protein levels of Notch,HES1,HEY1 and Bcl-2 decreased,and the proportions of S phase and G2 phase cells,OD450 value,number of clones,migration rate,number of invasive cells and levels of N-cadherin and Vimentin decreased significantly in the L-SHI group,the M-SHI group,the H-SHI group and the DAPT group(P<0.05).The proportion of G1/G0 phase cells,apoptosis rate and levels of Bax,cleaved Casase-3,and E-cadherin increased obviously(P<0.05).The effect of SHI was dose-dependent.Compared with the H-SHI group,the above indexes showed the opposite trend in the H-SHI+VPA group.VPA attenuated the effect of SHI on reducing the malignant biolog
作者
吴德建
杨秋
谢桂丹
彭鑫
WU Dejian;YANG Qiu;XIE Guidan;PENG Xin(Department of Gastroenterology,Danzhou People's Hospital,Danzhou 571799,China;Department of Laboratory,Danzhou People's Hospital,Danzhou 571799,China)
出处
《天津医药》
CAS
北大核心
2023年第12期1293-1299,共7页
Tianjin Medical Journal
关键词
紫草素
癌
肝细胞
受体
NOTCH
细胞增殖
细胞运动
细胞凋亡
Shikonin
carcinoma,hepatocellular
receptors,Notch
cell proliferation
cell movement
apoptosis
