A hydrogen peroxide-generating agent, 6-formylpterin, enhances heat-induced apoptosis
A hydrogen peroxide-generating agent, 6-formylpterin, enhances heat-induced apoptosis
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DOI：
10.1080/02656730400025404
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发表时间：
2005-05
影响因子：
3.1
通讯作者：
S. Wada;Zheng-Guo Cui;Takashi Kondo;Qing‐Li Zhao;R. Ogawa;M. Shoji;Toshiyuki Arai;Keisuke Makino;Isao Furuta
中科院分区：
文献类型：
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作者：
S. Wada;Zheng-Guo Cui;Takashi Kondo;Qing‐Li Zhao;R. Ogawa;M. Shoji;Toshiyuki Arai;Keisuke Makino;Isao Furuta
在人骨髓单核细胞淋巴瘤U937细胞中，检测了6-甲酰蝶呤（一种细胞内过氧化氢（H2 O2）发生器）对热诱导凋亡的增强作用。将细胞单独用无毒浓度为300 µM（37°C）的6-甲酰蝶呤处理，单独用热休克（44°C/20 min）处理或两者联合处理，然后在37°C下孵育6 h。通过流式细胞术评估细胞凋亡、线粒体膜电位和caspase-3活化。此外，caspase-8激活和细胞内Ca 2+浓度（[Ca 2 +]i）的变化进行了检查。Western blotting检测Bax、Bcl-2、Bcl-XL、Bid、细胞色素c和PKCδ的表达。通过形态学观察和DNA片段化评估，6-甲酰蝶呤的加入促进了热诱导凋亡的诱导。线粒体膜电位降低，caspase-3和-8的活化增强，在细胞与联合处理。尽管联合治疗后未观察到Bax、Bcl-2和Bcl-XL表达的显著变化，但注意到Bid表达降低。此外，加入6-甲酰蝶呤可增强热休克诱导的细胞色素c从线粒体向胞浆的释放和PKCδ从胞浆向线粒体的转位。加入6-甲酰蝶呤后，细胞内[Ca 2 +]i升高。这些结果表明，[Ca ~（2+）]i升高、PKCδ向线粒体转位以及caspase-1依赖性通路的激活在6-FP增强热诱导的细胞凋亡中起主要作用。
The enhancement of heat-induced apoptosis by 6-formylpterin, an intra-cellular generator of hydrogen peroxide (H2O2), was examined in human myelomonocytic lymphoma U937 cells. The cells were treated with either 6-formylpterin alone at a nontoxic concentration of 300 µM (37°C), heat shock (44°C per 20 min) alone or a combination of the two, then incubated at 37°C for 6 h. Assessments of apoptosis, mitochondrial membrane potential and caspase-3 activation were performed by flow cytometry. Moreover, caspase-8 activation and changes in the intra-cellular Ca2+ concentration ([Ca2+]i) were examined. Bax, Bcl-2, Bcl-XL, Bid, cytochrome c and PKCδ were detected by Western blotting. The induction of heat-induced apoptosis evaluated by morphological observation and DNA fragmentation were promoted by the addition of 6-formylpterin. Mitochondrial membrane potential was decreased and the activation of caspase-3 and -8 was enhanced in the cells treated with the combination. A decreased-expression of Bid was noted, although no significant changes in Bax, Bcl-2 and Bcl-XL expression were observed after the combined treatment. Furthermore, both the release of cytochrome c from mitochondria to cytosol and the translocation of PKCδ from cytosol to mitochondria, which were induced by heat shock, were enhanced by the addition of 6-formylpterin. The number of cells with a higher [Ca2+]i was also increased by the addition of 6-formylpterin. These findings suggest that the increase in [Ca2+]i, the activation of the mitochondria–caspase dependent pathway and the translocation of PKCδ to mitochondria play principal roles in the enhancement of heat-induced apoptosis by 6-FP.