【摘要】 目的探讨双氢青蒿素对雄激素非依赖性前列腺癌C4-2B细胞的增殖和凋亡以及mTOR表达水平和分布定位的影响。方法体外培养C4-2B细胞给予不同浓度的双氢青蒿素处理,以MTT法检测细胞增殖率;以RT-PCR检测细胞mTOR mRNA表达的情况;流式细胞术检测细胞凋亡率;透射电镜观察细胞超微结构的变化;激光共聚焦显微镜免疫荧光分析法评价mTOR蛋白的表达水平和亚细胞定位分布。结果双氢青蒿素抑制C4-2B细胞的增殖呈时间和剂量依赖性(P<0.01);RT-PCR和免疫荧光细胞化学实验的结果表明,C4-2B细胞有mTOR mRNA和蛋白的表达,且随双氢青蒿素浓度的升高而增高(P<0.05);FCM显示C4-2B细胞的凋亡率随双氢青蒿素浓度的升高而增加(P<0.01);电镜观察到典型凋亡的形态学变化;激光共聚焦检测结果显示,mTOR不仅有胞膜和胞质的定位,胞核也有少量表达,随双氢青蒿素浓度的升高,胞核内的表达增加。结论双氢青蒿素抑制C4-2B细胞增殖并诱导其凋亡,呈时间和剂量依赖性。C4-2B细胞对双氢青蒿素的反应性可能与mTOR的表达和核易位激活有关。更多还原

【Abstract】 【Objective】To investigate the effects of dihydroartemisinin on the proliferation and apoptosis and the expression and distribution of mTOR in androgen independent prostate cancer cell line(C4-2B).【Methods】After in vitro cultured C4-2B cells were induced by different concentrations of dihydroartemisinin respectively.MTT assay was applied to determine the proliferation at different time points;mTOR mRNA expression was detected by RTPCR;Apoptosis rate was assayed by flow cytometry;the morphological changes were observed under the inverted electron microscopy;Laser scanning confocal microscope immunofluorescence analysis was performed to detect the expression and the sub-cellular localization of mTOR protein.【Results】The growth of C4-2B cells was suppressed obviously by dihydroartemisinin in a dose-and time-dependent manner(P <0.01).RT-PCR and fluorescent immunocytochemistry assay revealed that the expression of mTOR mRNA and protein was detected in C4-2B cells and increased with the increase of dihydroartemisinin concentration(P <0.05).The results of confocal microscopic immunofluorescence assay showed that the mTOR was not only localized in membrane and cytoplasm but also a little in nucleolus of C4-2B cells.The quantity of mTOR in the nuclei was gradually increased in a dose-dependent manner of dihydroartemisinin.【Conclusion】Dihydroartemisinin could suppress the growth of C4-2B cells and promote the apoptosis in a dose-and time-dependent manner.mTOR was located mainly in membrane and cytoplasm of C42B cells.The expression of mTOR and nuclear translocation was gradually increased with the increase of dihydroartemisinin.The reactivity of the C4-2B cells to dihydroartemisinin may be due to the intracellular location of mTOR receptors and specific signaling pathway.更多还原