BH3 Profiling: A Functional Assay to Measure Apoptotic Priming and Dependencies
BH3 Profiling: A Functional Assay to Measure Apoptotic Priming and Dependencies
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DOI：
10.1007/978-1-4939-8861-7_4
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发表时间：
2019-01-01
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影响因子：
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通讯作者：
Sarosiek, Kristopher
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文献类型：
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作者：
Fraser, Cameron;Ryan, Jeremy;Sarosiek, Kristopher
细胞凋亡(程序性细胞死亡)是由多种细胞应激或侮辱所激活的，对哺乳动物的正常发育以及维持机体的动态平衡至关重要。细胞凋亡途径也被许多常见的抗癌治疗和电离辐射所涉及，这有助于肿瘤的消退或治疗的毒副作用。由于维持健康的细胞存活或有效清除癌细胞的重要性，BH3图谱分析被开发用于从功能上测量任何给定细胞中的凋亡途径的状态。这种检测包括将细胞线粒体暴露于模拟内源性促凋亡蛋白活性的促凋亡BH3肽，其中bcl2蛋白家族驻留并控制细胞对凋亡的承诺。通过使用激活剂或敏感肽，可以确定线粒体凋亡启动的水平(接近细胞死亡的阈值)或对存活的bcl2家族蛋白的依赖程度。这里描述了两种BH3图谱的方法，它们可以使用户进行这些功能测量，这对于预测细胞对抑制原存活蛋白活性的促凋亡应激源或疗法(BH3模拟)的反应是有用的。
Apoptosis (programmed cell death) is activated by a wide variety of cellular stresses or insults and is vital for proper mammalian development as well as the maintenance of organismal homeostasis. The apoptosis pathway is also engaged by many common types of anticancer therapies and ionizing radiation, which contributes to the regressions of tumors or the toxic side effects of treatment. Due to the importance of maintaining healthy cell survival or the efficient clearance of cancer cells, the BH3 profiling assay was developed to functionally measure the state of the apoptosis pathway in any given cells. This assay involves the exposure of cellular mitochondria, where the BCL-2 family of proteins resides and controls the commitment to apoptosis, to proapoptotic BH3 peptides that mimic the activity of endogenous proapoptotic proteins. By using either activator or sensitizer peptides, the level of mitochondrial apoptotic priming (proximity to the threshold at which a cell commits to cell death) or dependence on prosurvival BCL-2 family proteins can be determined. Described here are two methods of BH3 profiling that can enable a user to make these functional measurements, which can be useful for predicting cellular responses to proapoptotic stressors or therapeutics (BH3 mimetics) that inhibit the activity of prosurvival proteins.