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目的研究吴茱萸碱(Evodiamine,Evo)对结肠癌Lo Vo细胞增殖的抑制作用及可能的分子机制。方法采用结晶紫染色分析Evo(0、0.5、1、2、4μmol/L)对Lo Vo细胞的增殖抑制作用;应用流式细胞术分析Evo(0、0.5、1、2μmol/L)对Lo Vo细胞凋亡的促进作用;Western blot法分析Evo(0、0.5、1μmol/L)对Lo Vo细胞中增殖细胞核抗原(proliferating cell nuclear antigen,PCNA)及Caspase-3蛋白表达水平的影响;利用缺氧诱导因子-1α(hypoxia inducible factor 1-alpha,HIF-1α)荧光素酶报告质粒检测Evo(0、0.5、1、2μmol/L)对HIF-1α转录活性的影响。采用Western blot法分析Evo(0、0.5、1、2μmol/L)对Lo Vo细胞中HIF-1α、Akt1/2和磷酸化Akt1/2蛋白水平的影响。结果与对照组相比,Evo明显抑制Lo Vo细胞增殖,下调PCNA蛋白水平,促进Lo Vo细胞凋亡;报告质粒统计结果显示,Evo呈浓度依赖性降低HIF-1α荧光素酶报告质粒活性(Evo为0.5μmol/L时,P<0.05;Evo为1μmol/L或2μmol/L时,P<0.01);Western blot检测结果显示,Evo能够明显降低HIF-1α蛋白水平,下调Akt1/2磷酸化水平。结论 Evo能够抑制Lo Vo细胞增殖并促进凋亡,其机制可能与Evo下调HIF-1α蛋白表达,抑制PI3K/Akt信号转导相关。
Objective To investigate the inhibitory effect of evodiamine (Evo) on the proliferation of colon cancer LoVo cells and its possible molecular mechanism. Methods The effect of Evo (0,0.5,1,2,4μmol / L) on the proliferation of LoVo cells was analyzed by crystal violet staining. The effects of Evo (0,0.5,1,2μmol / L) The effect of Evo (0, 0.5 and 1 μmol / L) on the expression of proliferating cell nuclear antigen (PCNA) and Caspase-3 in LoVo cells was analyzed by Western blot. The effect of Evo (0, 0.5, 1 and 2 μmol / L) on the transcriptional activity of HIF-1α was detected by using luciferase reporter plasmid of hypoxia inducible factor 1-alpha (HIF-1α) The effect of Evo (0,0.5,1,2 μmol / L) on the protein expression of HIF-1α, Akt1 / 2 and phosphorylated Akt1 / 2 in LoVo cells was analyzed by Western blot. Results Compared with the control group, Evo significantly inhibited the proliferation of LoVo cells, down-regulated the expression of PCNA protein and promoted the apoptosis of LoVo cells. Statistical analysis of the reporter plasmids showed that Evo reduced the activity of HIF-1alpha luciferase reporter plasmid in a concentration-dependent manner Evo was 1μmol / L or 2μmol / L, P <0.01). The result of Western blot showed that Evo could significantly decrease the level of HIF-1α protein and down-regulate the phosphorylation of Akt1 / 2 . Conclusion Evo can inhibit Lo Vo cell proliferation and promote apoptosis, which may be related to Evo down-regulating HIF-1α protein expression and PI3K / Akt signal transduction.