黄芩素和LY294002对人肝癌细胞系SMMC-7721细胞增殖和凋亡的影响研究

发布时间：2018-03-22 01:12

本文选题：肝肿瘤　切入点：细胞增殖　出处：《中国全科医学》2017年03期 　论文类型：期刊论文

【摘要】：目的探讨黄芩素和LY294002对人肝癌细胞系SMMC-7721细胞增殖和凋亡的影响。方法 2015年3月—2016年1月,取人肝癌细胞系SMMC-7721,制备细胞悬液。加入黄芩素(1、2、5、10、20、50、100、200、300μmol/L,分别命名为A1、A2、A3、A4、A5、A6、A7、A8、A9组)或LY294002(1、2、5、10、20、30μmol/L,分别命名为B1、B2、B3、B4、B5、B6组),设定空白对照组和二甲基亚砜(DMSO)对照组,采用CCK8试剂盒检测各组细胞增殖水平;采用20μmol/L黄芩素(C1组)单独处理,20μmol/L黄芩素联合10μmol/L LY294002(C2组)处理人肝癌细胞系SMMC-7721,设定空白对照组和DMSO组,采用流式细胞术检测各组细胞周期;采用2、5、10、20μmol/L黄芩素(分别命名为D1、D2、D3、D4组)处理人肝癌细胞系SMMC-7721,设定空白对照组和DMSO组,采用显微摄影检测各组细胞数量;采用20μmol/L黄芩素(E1组)单独处理,20μmol/L黄芩素联合10μmol/L LY294002(E2组)处理人肝癌细胞系SMMC-7721,设定空白对照组和DMSO组,采用流式细胞术检测各组早期凋亡和晚期凋亡情况;采用20μmol/L黄芩素(F1组)、10μmol/L LY294002(F2组)或20μmol/L黄芩素联合10μmol/L LY294002(F3组)处理人肝癌细胞系SMMC-7721,设定空白对照组和DMSO组,采用实时荧光定量PCR(Realtime PCR)法检测细胞外调节蛋白激酶(ERK)1/2、周期素D1(Cyclin D1)、糖原合成酶激酶-3β(GSK-3β)、丝氨酸/苏氨酸蛋白激酶(AKT)mRNA表达水平;采用20μmol/L黄芩素(G1组)、10μmol/L LY294002(G2组)或20μmol/L黄芩素联合10μmol/L LY294002(G3组)处理人肝癌细胞系SMMC-7721,设定空白对照组和DMSO组,采用Western blotting法检测磷酸化ERK1/2(P-ERK1/2)、Cyclin D1、磷酸化GSK-3β(P-GSK-3β)、磷酸化AKT(P-AKT)表达水平。结果 A8组、A9组、B6组细胞增殖水平低于空白对照组、DMSO对照组(P0.05)。C2组G0/G1期、G2/M期细胞比例高于空白对照组、DMSO组,S期细胞比例低于空白对照组、DMSO组(P0.05)。D4组细胞数量少于空白对照组、DMSO组(P0.05)。空白对照组、DMSO组、E1组、E2组早期凋亡和晚期凋亡比较,差异无统计学意义(P0.05)。F3组ERK1/2、Cyclin D1、GSK-3β、AKT mRNA表达水平均低于空白对照组、DMSO组(P0.05)。G3组P-ERK1/2、Cyclin D1、P-GSK-3β、P-AKT表达水平低于空白对照组、DMSO组(P0.05)。结论黄芩素和LY294002可抑制人肝癌细胞系SMMC-7721细胞增殖,但不影响其凋亡。
[Abstract]:Objective to investigate the effects of baicalin and LY294002 on proliferation and apoptosis of human hepatoma cell line SMMC-7721. Human hepatoma cell line SMMC-7721 was used to prepare cell suspension, and the cell suspension was added to baicalein (SMMC-7721). The cell suspension was added to baicalein 5101020050200300 渭 mol / L, named as A1A1A2OA3A3A3A4A5A5OA7A7A7 (A9) or LY294002121010102030 渭 mol / L, named B1OB2B3B4B4B5B6 group, the blank control group and dimethyl sulfoxide (DMSOO) control group were set up. The cell proliferation levels in each group were measured by CCK8 kit. Human hepatoma cell line SMMC-7721 was treated with 20 渭 mol/L baicalein and 10 渭 mol/L LY294002(C2 alone (control group and DMSO group). The cell cycle of each group was detected by flow cytometry. Human hepatoma cell line SMMC-7721 was treated with SMMC-7721 with 20 渭 mol/L baicalein (D1D2D2OD3D4 group). The control group and DMSO group were set up, and the number of cells in each group was detected by microphotography. Human hepatoma cell line SMMC-7721 was treated with 20 渭 mol/L baicalin and 10 渭 mol/L LY294002(E2 alone. The control group and DMSO group were set up. The early apoptosis and late apoptosis were detected by flow cytometry. Human hepatoma cell line SMMC-7721 was treated with 20 渭 mol/L baicalein F _ 1 group (10 渭 mol/L LY294002(F2) or 20 渭 mol/L baicalin combined with 10 渭 mol/L LY294002(F3 group. The expression levels of extracellular regulated protein kinase (ERK) 1 / 2, cyclin D1(Cyclin D1, glycogen synthase kinase 3 尾 -GSK-3 尾 and serine / threonine protein kinase (TK) were detected by real-time fluorescence quantitative PCR(Realtime assay. Human hepatoma cell line SMMC-7721 was treated with 20 渭 mol/L baicalein G 1 group (10 渭 mol/L LY294002(G2) or 20 渭 mol/L baicalin combined with 10 渭 mol/L LY294002(G3 group. Western blotting method was used to detect the expression of phosphorylated ERK1 / 2pERK1 / 2pERK1 / 2 cyclin D1, phosphorylated GSK-3 尾 -P-GSK-3 尾, phosphorylated AKT1-P-AKT1.Results the proliferative level of B6 cells in A8 group was lower than that in DMSO control group P0.05.C2 group was higher than that in control group DMSO group. The percentage of cells in S phase was lower than that in DMSO group. The number of cells in DMSO group was lower than that in DMSO group (P 0.05). The early apoptosis and late apoptosis in E 2 group in E 2 group were compared with those in DMSO group. There was no significant difference in the expression of ERK1 / 2Cyclin D1GSK-3 尾 -AKT mRNA in P0.05U. F3 group compared with the control group. The expression level of P-ERK1 / 2Cyclin D1P-GSK-3 尾 P-AKT was lower than that of DMSO group. Conclusion baicalin and LY294002 can inhibit the proliferation of human hepatoma SMMC-7721 cell line, but do not affect the apoptosis of PERK1 / 2Cyclin D1- P-GSK-3 尾 -AKT. Conclusion baicalin and LY294002 can inhibit the proliferation of human hepatoma cell line SMMC-7721 cell line, but do not affect the apoptosis of PERK1 / 2Cyclin D1PGSK-3 尾 -AKT group.
【作者单位】：西南医科大学附属医院教务部;西南医科大学附属中医医院妇产科;西南医科大学附属医院医学实验中心;西南医科大学科技处;西南医科大学附属医院消化内科;
【基金】：四川省科技厅-泸州市人民政府-泸州医学院2014年联合科研项目(14JC01383-LH53)
【分类号】：R735.7

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