去甲基脱氧鬼臼毒素衍生物DPPC诱导A549细胞凋亡的研究

目的 探讨鬼臼毒素衍生物L-N-甲酰-4′-去甲基脱氧鬼臼-甲硫-4-N-哌嗪酰胺(DPPC)抑制人非小细胞肺癌A549细胞的增殖作用,阐述DPPC诱导A549细胞凋亡的相关分子机制.方法 体外培养A549细胞,Ho-chest33258检测DPPC处理A549细胞后细胞凋亡现象;Annexin V-FITC/PI双染定量检测DPPC处理后细胞凋亡数量;蛋白免疫印迹实验检测凋亡基因p53、caspase-3、Bax的表达.并与依托泊苷(阳性对照组)和未给予任何药物处理的A549细胞做对照.结果 DPPC组A549细胞中γ-H2AX的表达量较阴性对照组和阳性对照组都有明显的增加.荧光显微镜下见DPPC处理后的A549细胞核呈现萎缩,并出现细胞凋亡小体.与阴性对照组比较,不同浓度DPPC处理A549细胞后,抑制肿瘤发生发展的p53基因表达增加,并呈剂量依赖性,其下游的凋亡相关蛋白caspase-3以及Bax表达增加(P<0.05,P<0.01).相同浓度的DPPC和依托泊苷抑制凋亡相关蛋白有明显的差异(P<0.05).结论 DPPC能够引起A549细胞DNA损伤,并诱导细胞凋亡,DPPC能够使细胞内凋亡相关蛋白表达异常,抑制A549细胞增殖;此外,DPPC体外表现出优越的抗肿瘤细胞增殖作用.

A Study Demethyl -deoxypodophyllotoxin Derivative DPPC Induced A549 Cells Apoptosis

Objective To investigate effect of podophyllotoxin derivative L-N-formycl-4'-demethyldeoxypodophyl-lotoxin-Methyl-4-N-Piperazine acidamide ( DPPC) in inhibition of proliferation of human non-small cell lung cancer A549 cells, and the molecular mechanism of DPPC induced apoptosis in A549 cells. Methods The A549 cells were cultured in vitro, and apoptoses were detected using Hochest33258 after A549 cells were treated with DPPC. Apoptosis number af-ter DPPC treatment was detected by double-dye quantitative detection of Annexin V-FITC/PI. Expressions of apoptosis genes P53, caspase-3 and Bax were detected by protein immunoblot test. The above values in DPPC group were compared with those in Etoposide ( positive control) group and A549 cells without any medication treatment group ( negative control group). Results γ-H2AX expression of A549 cells in DPPC group were significantly increased compared with those in negative and positive control groups. A549 cells after DPPC treatment were nucleus atrophy and cell apoptotic body under fluorescence microscope. Compared with those in negative control group, inhibited tumorigenesis and development p53 gene expressions of A549 cells after different concentrations of DPPC treatment were increased, and the expression showed dose dependent, and expressions of apoptosis related protein caspase-3 and Bax were increased in the downstream (P<0. 05, P<0. 01). The same concentration of DPPC and VP16 inhibited the apoptosis related proteins showed signif-icant difference (P<0. 05). Conclusion DPPC can cause DNA damage of A549 cells, induce apoptosis, make abnor-mal expressions of apoptosis related proteins in cells and inhibit proliferation of A549 cells. In addition, DPPC shows su-perior effect of anti-tumor cell proliferation in vitro.