Quantitation of mitochondrial alterations associated with apoptosis
Quantitation of mitochondrial alterations associated with apoptosis
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DOI：
10.1016/s0022-1759(02)00069-8
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发表时间：
2002-07-01
影响因子：
2.2
通讯作者：
Kroemer, G
中科院分区：
文献类型：
--
作者：
Castedo, M;Ferri, K;Kroemer, G
线粒体在早期凋亡过程中经历两个主要变化。一方面，线粒体外膜变得可渗透蛋白质，导致可溶性膜间蛋白（SIMPs）从线粒体中释放。另一方面，线粒体内膜跨膜电位（DeltaPsi（m））降低。这些变化发生在大多数（如果不是全部的话）细胞死亡模型中，并且可以利用这些变化在早期阶段检测细胞凋亡。在这里，我们描述了用于检测DeltaPsi（m）中的改变的方法，该方法基于细胞与阳离子亲脂性荧光染料的孵育，其摄取由DeltaPsi（m）驱动。某些DeltaPsi（m）敏感染料可以与其他荧光染料组合，以同时检测细胞活力、磷脂酰丝氨酸残基的质膜暴露或活性氧物质（ROS）的线粒体产生。此外，我们描述了一种免疫荧光方法检测两个功能重要的蛋白质易位从线粒体，即半胱天冬酶共激活细胞色素c和半胱天冬酶独立的死亡效应细胞凋亡诱导因子（AIF）。(C)出版社：Elsevier Science B.V.
Mitochondria undergo two major changes during early apoptosis. On the one hand, the outer mitochondrial membrane becomes permeable to proteins, resulting in the release of soluble intermembrane proteins (SIMPs) from the mitochondrion. On the other hand, the inner mitochondrial membrane transmembrane potential (DeltaPsi(m)) is reduced. These changes occur in most, if not all, models of cell death and can be taken advantage of to detect apoptosis at an early stage. Here, we delineate methods for the detection of alterations in the DeltaPsi(m), based on the incubation of cells with cationic lipophilic fluorochromes, the uptake of which is driven by the DeltaPsi(m). Certain DeltaPsi(m)-sensitive dyes can be combined with other fluorochromes to detect simultaneously cellular viability, plasma membrane exposure of phosphatidylserine residues, or the mitochondrial production of reactive oxygen species (ROS). In addition, we describe an immunofluorescence method for the detection of two functionally important proteins translocating from mitochondria, namely, the caspase co-activator cytochrome c and the caspase-independent death effector apoptosis inducing factor (AIF). (C) 2002 Published by Elsevier Science B.V.