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研究新合成的N~1-(正-丁基)-7-氮杂异靛蓝(N~1-(n-butyl)-7-azaisoindigo,7-AI-b)抑制人非小型肺癌细胞A549的增殖,初步探索其抗肿瘤的机制。以不同浓度的7-AI-b作用于A549,MTT检测与相差显微镜观察相结合,分析7-AI-b对细胞增殖的影响;MDC染色和Western印迹检测自噬标志蛋白(LC3)的表达,研究药物处理后细胞是否发生了自噬。PI单染,流式检测细胞周期的变化;Fluo-3-AM荧光探针检测细胞中Ca~(2+)的含量。结果发现,7-AI-b以时间和剂量依赖性方式抑制细胞的增殖;MDC染色后,自噬小泡增多,LC3的总表达量增加,且由LC3-Ⅰ向LC3-Ⅱ的转变量升高。流式细胞检测发现细胞被阻滞在G_0/G_1期;同时,药物处理后,细胞内Ca~(2+)出现了超载。由此,新合成的7-AI-b有很好的抑制肿瘤细胞增殖的作用,其机制与引起细胞周期阻滞、诱导细胞自噬有关,胞内Ca~(2+)超载也参与了该作用。
The newly synthesized N ~ 1- (n-butyl) -7-azaisoindigo, 7-AI-b was used to inhibit the growth of human non-small lung cancer cell line A549 Proliferation, preliminary explore its anti-tumor mechanism. The effect of 7-AI-b on the cell proliferation was analyzed by MTT assay and phase contrast microscopy. The expression of autophagy marker protein (LC3) was detected by MDC staining and Western blot. To investigate whether autophagy occurs in cells after drug treatment. PI single staining was used to detect the changes of cell cycle. Fluo-3-AM fluorescence probe was used to detect the content of Ca 2+ in the cells. The results showed that 7-AI-b inhibited cell proliferation in a time-and dose-dependent manner. After MDC staining, the autophagic vesicles increased and the total expression of LC3 increased, and the change from LC3-Ⅰ to LC3-Ⅱ high. Flow cytometry showed that the cells were arrested in G_0 / G_1 phase. At the same time, Ca 2+ in cells were overloaded after drug treatment. Thus, the newly synthesized 7-AI-b has a good inhibitory effect on tumor cell proliferation, the mechanism of which is related to cell cycle arrest and induction of autophagy, and intracellular Ca 2+ overload is also involved effect.