Intrinsic fluorescence and redox changes associated with apoptosis of primary human epithelial cells
Intrinsic fluorescence and redox changes associated with apoptosis of primary human epithelial cells
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DOI：
10.1117/1.2401149
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发表时间：
2006-11-01
影响因子：
3.5
通讯作者：
Georgakoudi, Irene
中科院分区：
文献类型：
--
作者：
Levitt, Jonathan M.;Baldwin, Amy;Georgakoudi, Irene
细胞凋亡在人体组织的发育和维持中起着关键作用。这一过程传统上是在用外源荧光团染色的细胞中研究的。这些方法会影响细胞的活性，因此不适合在体内应用。我们提出了一种成像方法，它可以基于对细胞自发荧光强度和位置的明显变化的检测和量化来识别活细胞群体中的凋亡细胞。具体地说，我们获得了顺铂处理1、9、14和18h后原代角质形成细胞的NAD(P)H、FAD和氧化还原比自体荧光图像。顺铂是一种已知的诱导细胞凋亡的化疗药物。我们发现，在顺铂治疗下，强烈的自发荧光与低氧化还原荧光率结合在一起，逐渐限制在一个逐渐变小的核周细胞质区域。对外源性核荧光团的研究表明，这些自体荧光变化发生在细胞凋亡的早期阶段。另外的共染色实验表明，这个强烈的自体荧光、高度代谢活性的核周环代表了线粒体的一个亚群，这些线粒体是为了响应凋亡刺激而被动员的，并可能提供执行最终凋亡步骤所需的能量。因此，自体荧光定位变化可以作为体内早期细胞凋亡的敏感的、非侵入性的指标。(C)2006年光学仪器工程师学会。
Apoptosis plays a key role in the development and maintenance of human tissues. This process has been studied traditionally in cells that are stained with exogenous fluorophores. These approaches affect cell viability, and thus are ill-suited for in vivo applications. We present an imaging approach that can identify apoptotic cells in living cell populations based on detection and quantification of distinct changes in the intensity and localization of cellular autofluorescence. Specifically, we acquire NAD(P)H, FAD, and redox ratio autofluorescence images of primary keratinocytes following 1, 9, 14, and 18 h of treatment with cisplatin, a known apoptosis-inducing chemotherapy agent. We find that intense autofluorescence combined with a low redox fluorescence ratio is progressively confined to a gradually smaller perinuclear cytoplasmic region with cisplatin treatment. Studies with exogenous nuclear fluorophores demonstrate that these autofluorescence changes occur at early stages of apoptosis. Additional costaining experiments suggest that this strongly autofluorescent, highly metabolically active perinuclear ring represents a subpopulation of mitochondria that are mobilized in response to the apoptotic stimulus and may provide the energy required to execute the final apoptotic steps. Thus, autofluorescence localization changes could serve as a sensitive, noninvasive indicator of early apoptosis in vivo. (c) 2006 Society of Photo-Optical Instrumentation Engineers.