田辉凯, 张红梅, 姚小皓, 李学军. 非洲爪蟾卵无细胞凋亡体系的建立非洲爪蟾卵无细胞凋亡体系的建立J. 药学学报, 2003, 38(1): 10-10.
引用本文: 田辉凯, 张红梅, 姚小皓, 李学军. 非洲爪蟾卵无细胞凋亡体系的建立非洲爪蟾卵无细胞凋亡体系的建立J. 药学学报, 2003, 38(1): 10-10.
TIAN Hui-kai, ZHANG Hong-mei, YAO Xiao-hao, LI Xue-jun. Foundation of cell-free apoptotic system from Xenopus eggsJ. Acta Pharmaceutica Sinica, 2003, 38(1): 10-10.
Citation: TIAN Hui-kai, ZHANG Hong-mei, YAO Xiao-hao, LI Xue-jun. Foundation of cell-free apoptotic system from Xenopus eggsJ. Acta Pharmaceutica Sinica, 2003, 38(1): 10-10.

非洲爪蟾卵无细胞凋亡体系的建立非洲爪蟾卵无细胞凋亡体系的建立

Foundation of cell-free apoptotic system from Xenopus eggs

  • 摘要: 目的建立一种简便、高效的无细胞凋亡体系,探讨外源细胞核在其中的凋亡过程及z-VAD-fmk对其的抑制作用。方法超速离心制备非洲爪蟾卵无细胞体系,用细胞色素C激活为凋亡体系。应用荧光显微镜、琼脂糖凝胶电泳、ApoAlert Caspase-3 Colorimetric Assay等方法检测外源细胞核在无细胞凋亡体系中的凋亡过程及z-VAD-fmk的抑制作用。结果制备的爪蟾卵无细胞体系可由细胞色素C激活为凋亡体系,外源加入的细胞核呈现典型的形态学变化,总DNA特异降解形成DNA ladder。体系内内源特异核酸酶被激活。z-VAD-fmk可抑制DNA ladder的形成。凋亡特异蛋白酶XCPP32活性明显升高。结论本实验建立的无细胞凋亡体系可以高效的诱导外源细胞核凋亡,重现体内过程。

     

    Abstract: AimTo find a convenient and highly effective cell-free apoptotic system and investigate the apoptotic process of mouse liver nuclei in the system and the inhibitory effects of z-VAD-fmk on apoptosis. MethodsThe cell-free system was prepared by ultracentrifugation of Xenopus eggs and activated by cytochrome C. Fluorescent microscope, argrose gel electrophoresis and apoalert Caspase-3 colorimetric assay were applied to detect the apoptotic process of isolated nuclei and the inhibitory effecs of z-VAD-fmk on the system. Results The prepared cell-free system was shown to be activated by cytochrome C. The exogenous mouse liver nuclei presented typical morphologic changes and DNA ladder formation was observed which could be inhibited by z-VAD-fmk. Specific endonuclease was activated and XCPP32 activity was significantly increased in the system. ConclusionThe prepared cell-free apoptotic system can effectively induce isolated nuclei apoptosis and mimick the process in vivo.

     

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