吴茱萸碱对人结肠癌细胞自吞噬的影响

目的 观察吴茱萸碱对结肠癌Lovo细胞自噬的影响,探讨吴茱萸碱对Lovo细胞的生长抑制作用及其机制.方法 联合应用MTT法和形态学变化观察吴茱萸碱对Lovo细胞的生长抑制作用,借助激光共聚焦显微镜观察自噬现象,并用丹(磺)酰戊二胺染色测定对自噬进行定量分析,Western blot方法检测微管相关蛋白轻链3的表达,确认自噬的发生.同时检测吴茱萸碱和3-甲基腺嘌呤对Lovo细胞活力及凋亡的影响.结果 吴茱萸碱呈浓度依赖性抑制结肠癌Lovo细胞的生长活力(P＜0.05),尤其在浓度为60 μmol/L时最为明显(60%),且在光镜下形态表现为细胞裂解和细胞间隙增宽.吴茱萸碱可诱导结肠癌Lovo细胞发生自噬(P＜0.05),阻断自噬后,吴茱萸碱抑制结肠癌Lovo细胞生长活力的能力进一步增强(P＜0.01).结论 吴茱萸碱可诱导结肠癌Lovo细胞发生自噬,但自噬却拮抗吴茱萸碱的抗肿瘤活性.联合应用吴茱萸碱和自噬凋节剂可增强其抗肿瘤作用,逆转其对化疗药物的耐药性.

Abstract：

Objective To investigate the effects of evodiamine on autophagy of human colon a cleno carcinoma lovo cells, and to explore the role and mechanism of autophagy which was induced by evodiamine (EVO). Methods MTT assay combined with the morphologic changes were used to observe the cell viability. Monodansylcadaverine was used to detect autophagy by fluorospectrophotometer and the confocal laser fluorescence microscopy respectively. Immunoblotting assay was used to observe the microtubule-associated protein 1 light chain 3. Finally, evodiamine combined with 3-methyladenine to detect the cell viability with MTT assay and the apoptosis with the flow cytometry, respectively.Results Evodiamine inhibited the viability of Lovo cells in dose-dependent manner ( P ＜ 0. 05 ), especially in 60 μmol/L that was obviously(60% ). Further more, the cell lysis and cell gap widened was observed by the light microscope. Evo triggered the autophagy, and after inhibition the autophagy by 3-MA, the killing capacities of the Evo was enhanced ( P ＜ 0. 01 ). However, autophagy prohibited the apoptosis pathways.Conclusions Evodiamine can trigger the autophagy, which might play a self-defense role in evodiamineinduced cell death. The cytototoxicity of evodiamine can be augmented by the autophagy inhibitors. The joint application of autophagy regulators with the chemotherapeutic agents might enhance the cell killing effects of chemotherapeutic drugs and show a potent role in cancer drug resistance.

The effects of evodiamine on autophagy in human colon adenocarcinoma Lovo cells

Objective To investigate the effects of evodiamine on autophagy of human colon a cleno carcinoma lovo cells, and to explore the role and mechanism of autophagy which was induced by evodiamine (EVO). Methods MTT assay combined with the morphologic changes were used to observe the cell viability. Monodansylcadaverine was used to detect autophagy by fluorospectrophotometer and the confocal laser fluorescence microscopy respectively. Immunoblotting assay was used to observe the microtubule-associated protein 1 light chain 3. Finally, evodiamine combined with 3-methyladenine to detect the cell viability with MTT assay and the apoptosis with the flow cytometry, respectively.Results Evodiamine inhibited the viability of Lovo cells in dose-dependent manner ( P ＜ 0. 05 ), especially in 60 μmol/L that was obviously(60% ). Further more, the cell lysis and cell gap widened was observed by the light microscope. Evo triggered the autophagy, and after inhibition the autophagy by 3-MA, the killing capacities of the Evo was enhanced ( P ＜ 0. 01 ). However, autophagy prohibited the apoptosis pathways.Conclusions Evodiamine can trigger the autophagy, which might play a self-defense role in evodiamineinduced cell death. The cytototoxicity of evodiamine can be augmented by the autophagy inhibitors. The joint application of autophagy regulators with the chemotherapeutic agents might enhance the cell killing effects of chemotherapeutic drugs and show a potent role in cancer drug resistance.