Study of apoptosis in vitro using the Xenopus egg extract reconstitution system.
Study of apoptosis in vitro using the Xenopus egg extract reconstitution system.
复制标题
使用非洲爪蟾卵提取物重构系统进行体外细胞凋亡研究。
DOI：
--
复制
发表时间：
2006
影响因子：
--
通讯作者：
S. Kornbluth
中科院分区：
文献类型：
--
作者：
Paula B Deming;S. Kornbluth
20世纪90年代，Newmeyer和他的同事首次证明，利用非洲爪哇卵提取液可以在体外重建细胞凋亡的分子事件。当鸡蛋提取物在室温下孵育较长时间时，细胞凋亡的生化事件就会自发地被激活。非洲爪哇重组系统中的细胞凋亡特征与哺乳动物细胞相似：细胞色素c从线粒体中释放，半胱氨酸天冬氨酸酶被激活，细胞底物被切割，添加的细胞核碎裂。此外，这些凋亡事件可以通过添加抗凋亡蛋白如bcl2和常规的caspase抑制剂(如zVAD-fmk)来抑制。线粒体在提取物中被未知的信号触发释放细胞色素c，对于诱导这些提取物中的自发凋亡程序是必不可少的。然而，人们可以通过制备不含膜成分(包括线粒体)的提取物来研究发生在线粒体细胞色素c释放下游的凋亡事件。当将纯化的细胞色素c加入这种胞浆提取物中时，可以监测到凋亡的生化标记物(例如，半胱氨酸氨基转移酶的激活)。因此，鸡蛋提取物为单独或共同研究线粒体上游和下游发生的凋亡事件提供了一个易于处理的系统。
It was first shown by Newmeyer and colleagues in the 1990s that the molecular events of apoptosis could be reconstituted in vitro using Xenopus egg extracts. When the egg extract is allowed to incubate at room temperature for an extended time, the biochemical events of apoptosis are activated spontaneously. The features of apoptosis in the Xenopus reconstitution system mimic those that occur in mammalian cells: Cytochrome c is released from the mitochondria, caspases are activated, cellular substrates are cleaved, and added nuclei are fragmented. Moreover, these apoptotic events can be inhibited by addition of antiapoptotic proteins such as Bcl-2 and conventional caspase inhibitors (e.g., ZVAD-fmk). The mitochondria, which are triggered to release cytochrome c by as-yet-unknown signals in the extract, are essential for induction of the spontaneous apoptotic program in these extracts. However, one can study the apoptotic events that occur downstream of mitochondrial cytochrome c release by preparing extracts devoid of membrane components (including mitochondria). When purified cytochrome c is added to such cytosolic extracts, biochemical markers of apoptosis (e.g., activation of caspases) can be monitored. The egg extract therefore offers a tractable system for studying either separately or together the events of apoptosis occurring upstream and downstream of the mitochondria.
影响因子：
3.3
作者：
Tashker, JS;Olson, M;Kornbluth, S
通讯作者：
Kornbluth, S