Hauser Paul, Wang Sha, Didenko Vladimir V
Baylor College of Medicine, and Michael E. DeBakey Veterans Affairs Medical Center, Houston, TX, 77030, USA.
Methods Mol Biol. 2017;1554:193-200. doi: 10.1007/978-1-4939-6759-9_12.
Normal and dying cells release various types of membrane-bound vesicles including microvesicles, exosomes, and apoptotic bodies. These vesicles play important roles in intercellular communication and signal transduction. However, their diverse forms and subtypes fluctuate in size and other properties. In result current purification approaches do not fully discriminate between different categories of extracellular vesicles. Here, we present a fluorescence technique that specifically identifies apoptotic bodies in preparations of microvesicles, exosomes, and other extracellular vesicles.The approach exclusively labels the vesicles that contain DNA with 5'PO blunt-ended DNA breaks, such as those produced by the apoptotic CAD nuclease during apoptotic DNA degradation. The technique can be useful in studies of apoptosis involving microvesicles and exosomes.
正常细胞和濒死细胞会释放各种类型的膜结合囊泡,包括微囊泡、外泌体和凋亡小体。这些囊泡在细胞间通讯和信号转导中发挥着重要作用。然而,它们多样的形式和亚型在大小及其他特性方面存在波动。因此,目前的纯化方法无法完全区分不同类别的细胞外囊泡。在此,我们提出一种荧光技术,可特异性识别微囊泡、外泌体及其他细胞外囊泡制剂中的凋亡小体。该方法专门标记含有5'PO平端DNA断裂的囊泡,比如凋亡DNA降解过程中凋亡CAD核酸酶产生的那些断裂。这项技术在涉及微囊泡和外泌体的凋亡研究中可能会很有用。