摘要目的 观察姜黄素对人肝癌细胞HCC-LM3增殖和凋亡的影响并探究其相关机制.方法 采用CCK-8法检测经不同浓度姜黄素作用后的人肝癌细胞HCC-LM3的生长抑制率,初步观察姜黄素对人肝癌细胞HCC-LM3作用.参考相关文献,以0μmol/L浓度组为空白对照,以2.5、5.0、10.0、15.0、20.0、40.0、60.0μmol/L为浓度梯度加入姜黄素,培养48 h后进行检测.依据CCK-8法检测结果,选择其中生长抑制率差异较明显的10.0、20.0、40.0 μmol/L的浓度组为实验组,并以0 μmoL/L浓度组为对照组,细胞克隆形成实验检测细胞的克隆能力,流式细胞术检测细胞凋亡,Western blotting检测Mcl-1、Bax、Bcl-2和Bcl-xL蛋白的表达水平.计量资料用((x)±s)表示,组间比较用单因素方差分析.结果 CCK-8法检测显示,姜黄素浓度为2.5、5.0、10.0、15.0、20.0、40.0、60.0 μmol/L对应的人肝癌细胞HCC-LM3的生长抑制率分别为(6.71±3.45)％、(12.33±5.02)％、(20.07±5.60)％、(57.80±7.34)％、(78.37±6.53)％、(91.73±6.14)％和(96.18±3.45)％,提示姜黄素可抑制人肝癌细胞HCC-LM3的生长,并呈浓度依赖性.细胞克隆形成实验结果提示,姜黄素能够抑制人肝癌细胞HCC-LM3的克隆能力,当姜黄素浓度＞20.0 μmol/L时,人肝癌细胞HCC-LM3克隆形成受到显著的抑制.流式细胞术检测对照组和实验组的细胞凋亡率依次为(5.20±1.44)％、(9.90±3.31)％、(55.67 ±5.29)％、(79.63±4.71)％,各实验组凋亡率均高于对照组(P＜0.05),提示姜黄素可诱导人肝癌细胞HCC-LM3凋亡.Western blotting检测结果发现,经姜黄素作用后人肝癌细胞HCC-LM3中的Mcl-1蛋白表达降低(P＜0.05),Bax蛋白表达增加(P＜0.05),并呈浓度依赖性;而对Bcl-2抑凋亡蛋白家族另外2个主要蛋白Bcl-2和Bcl-xL则没有明显影响(P＞0.05).结论 姜黄素可抑制人肝癌细胞HCC-LM3的克隆、增殖,诱导人肝癌细胞HCC-LM3凋亡.

abstractsObjective To investigate the effects of curcumin on the growth and apoptosis of human hepatoma HCC-LM3 cells and explore the molecular mechanisms.Methods The human hepatoma HCC-LM3 cells were treated with different concentrations of curcumin and the cell growth inhibition rate was detected by CCK-8.The effect of curcumin on human hepatoma HCC-LM3 cells was observed.Refer to the relevant literature,the human hepatoma HCC-LM3 cells were treated with the concentration of 2.5,5.0,10.0,15.0,20.0,40.0,60.0 μmol/L of curcumin for 48 hours,taking the 0 μmol/L curcumin as control group,and the cell growth inhibition rate was detected by CCK-8.According to the results of CCK-8,selecting the concentration of 0 μmol/L as control group and the concentration of 10.0,20.0,40.0 μmol/L as experimental groups,which has significant difference on growth inhibition rates.Cell cloning assay was used to detect cell cloning ability,Flow cytometry was used to detect apoptosis,and Western blotting to detect the protein expression levels of Mcl-1,Bax,Bcl-2 and Bcl-xL.The measurement data were expressed in ((x) ± s),and the single factor analysis of variance was used for comparison between groups.Results CCK-8 assay showed that with treated by the concentration of 2.5,5.0,10.0,15.0,20.0,40.0,60.0 μmol/L,the growth inhibition rates were(6.71 ± 3.45)％,(12.33 ± 5.02)％,(20.07 ± 5.60)％,(57.80 ±7.34)％,(78.37 ±6.53)％,(91.73 ±6.14)％ and (96.18 ±3.45)％,suggesting that curcumin could inhibit the growth of human hepatoma HCC-LM3 cells in a dose-dependent manner.Cell clone formation experiment showed that curcumin could inhibit the clone of the human hepatoma HCC-LM3 cells,and the clone of the cells was inhibited significantly when the concentration of the curcumin was over 20.0μmol/L.The result of Annexin V-FITC/PI double staining analysis showed that the apoptotic rates of experimental groups and control groups were (5.20 ± 1.44) ％,(9.90 ± 3.31) ％,(55.67 ± 5.29) ％,(79.63 ±4.71)％,with all the apoptotic rates of experimental group over the control groups (P ＜0.05),suggesting curcumin could induce the apoptosis of human hepatoma HCC-LM3 cells.The Westen blotting showed that curcumin increased the expression of Bax protein while decreasing expression of Mcl-1 protein significantly in concentration-dependent manner (P ＜ 0.05),but have no effect on the expression of Bcl-2 and Bcl-xL proteins.Conclusion Curcumin could inhibit the proliferation and clone of human hepatoma HCC-LM3 cells,and induce apoptosis in a dose dependent manner.