CAJ | 학술논문

目的：观察去甲斑蝥素对人胆管癌RBE细胞周期的影响及对微管蛋白（α-Tubulin）的作用。方法体外培养胆管癌RBE细胞，分别用不同剂量去甲斑蝥素处理，流式细胞学检测细胞周期的影响，免疫组化染色法观察α-Tubulin结构变化，Real-time PCR检测α-Tubulin mRNA转录水平变化。结果流式细胞学检测显示胆管癌RBE细胞随着药物浓度的增加，G2/M期阻滞作用增强，10、20、40μg/ml组与对照组相比具有统计学意义（P＝0.000，P＜0.01）。免疫组化染色法观察到去甲斑蝥素作用下，胆管癌RBE细胞α-Tubulin骨架结构被破坏，Real-time PCR证实去甲斑蝥素使胆管癌RBE细胞α-Tubulin转录水平减少，差异具有统计学差异（P＝0.002，P＝0.000，P＜0.01）。结论去甲斑蝥素可以通过破坏胆管癌RBE细胞Tubulin骨架、抑制α-Tubulin mRNA的表达，阻止纺锤体的形成，将细胞周期阻滞在G2/M期，导致生长抑制，是其抗肿瘤机制之一。
목적：관찰거갑반모소대인담관암RBE세포주기적영향급대미관단백（α-Tubulin）적작용。방법체외배양담관암RBE세포，분별용불동제량거갑반모소처리，류식세포학검측세포주기적영향，면역조화염색법관찰α-Tubulin결구변화，Real-time PCR검측α-Tubulin mRNA전록수평변화。결과류식세포학검측현시담관암RBE세포수착약물농도적증가，G2/M기조체작용증강，10、20、40μg/ml조여대조조상비구유통계학의의（P＝0.000，P＜0.01）。면역조화염색법관찰도거갑반모소작용하，담관암RBE세포α-Tubulin골가결구피파배，Real-time PCR증실거갑반모소사담관암RBE세포α-Tubulin전록수평감소，차이구유통계학차이（P＝0.002，P＝0.000，P＜0.01）。결론거갑반모소가이통과파배담관암RBE세포Tubulin골가、억제α-Tubulin mRNA적표체，조지방추체적형성，장세포주기조체재G2/M기，도치생장억제，시기항종류궤제지일。
ObjectiveTo explore the mechanisms of Norcantharidin's anti-tumor effect by investigating cell cycle andα-Tubulin cytoskeleton changes in cholangiocarcinoma cell line RBE after treated by Norcantharidin. MethodsThe RBE cells were culturedin vitro, followed by treating with various concentration of norcantharidin for 24 h. Cell cycle changes were detected by flow cytometric analysis. We used immunohistochemical (IHC) to observe α-Tubulin cytoskeleton changes, and real-time PCR was used to detectα-Tubulin mRNA transcript levels. ResultsThe result of flow cytometric analysis shown the cell cycle arrested at G2/M. The damages ofα-Tubulin cytoskeleton under the treatment of Norcantharidin were observed by IHC, and real-time PCR determined thatα-Tubulin was decreased at mRNA level.Conclusions Norcantharidin can prevent the formation of spindle to arrest the cell cycle in G2/M phase, by disrupting Tubulin cytoskeleton, inhibitingα-Tubulin mRNA expression, which is one of the mechanisms of Norcantharidin's anti-tumor effect.