Measuring the lifetime of singlet oxygen in a single cell: addressing the issue of cell viability
Measuring the lifetime of singlet oxygen in a single cell: addressing the issue of cell viability
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DOI：
10.1039/b707313e
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发表时间：
2007-01-01
影响因子：
3.1
通讯作者：
Ogilby, Peter R.
中科院分区：
文献类型：
--
作者：
Hatz, Sonja;Lambert, John D. C.;Ogilby, Peter R.
在时间分辨光学实验中，通过掺入细胞中的光敏剂照射后的1270 nm磷光（a（1）Delta（g）-> X-3 Sigma（-）（g）），从单个神经元和HeLa细胞中检测到单线态分子氧O-2（a（1）Delta（g））。将细胞维持在缓冲培养基中，并通过活/死测定评估其活力。为了便于检测单线态氧，细胞内的H2O被替换为D2 O的渗透脱水和再水化过程。同样评估了这种损伤对细胞的影响。这些数据表明，在从“活”细胞到“死”细胞的复杂过渡中，我们的大多数细胞具有活细胞的代谢活性和形态特征。淬灭实验表明，在我们的细胞中的单线态氧的寿命主要是由与细胞内水的相互作用，而不是由与其他细胞成分的相互作用。数据表明，在一个可行的，代谢功能，和含H2O的细胞，单线态氧的寿命是类似于3 μ s。这与我们以前的报告一致，并证实了单态氧在电池中的寿命比迄今为止认为的要长得多。这意味着，在细胞中，单线态氧最好被表征为选择性而不是反应性中间体。当考虑单线态氧作为信号传导剂和作为导致细胞死亡的事件中的组分所发挥的作用时，这是重要的。本文报道的数据还表明，空间分辨光学探针可用于监测光诱导的单线态氧介导的单细胞死亡中的选定事件。
Singlet molecular oxygen, O-2(a(1)Delta(g)), has been detected from single neurons and HeLa cells in time-resolved optical experiments by its 1270 nm phosphorescence (a(1)Delta(g) -> X-3 Sigma(-)(g)) upon irradiation of a photosensitizer incorporated into the cell. The cells were maintained in a buffered medium and their viability was assessed by live/ dead assays. To facilitate the detection of singlet oxygen, intracellular H2O was replaced with D2O by an osmotic de-and rehydration process. The effect of this insult on the cells was likewise assessed. The data indicate that, in the complicated transition from a "live" to "dead" cell, the majority of our cells have the metabolic activity and morphology characteristic of a live cell. Quenching experiments demonstrate that the singlet oxygen lifetime in our cells is principally determined by interactions with intracellular water and not by interactions with other cell constituents. The data indicate that in a viable, metabolically-functioning, and H2O-containing cell, the lifetime of singlet oxygen is similar to 3 mu s. This is consistent with our previous reports, and confirms that the singlet oxygen lifetime in a cell is much longer than hitherto believed. This implies that, in a cell, singlet oxygen is best characterized as a selective rather than reactive intermediate. This is important when considering roles played by singlet oxygen as a signaling agent and as a component in events that result in cell death. The data reported herein also demonstrate that spatially-resolved optical probes can be used to monitor selected events in the light-induced, singlet-oxygen-mediated death of a single cell.