In vivo multiphoton microscopy of NADH and FAD redox states, fluorescence lifetimes, and cellular morphology in precancerous epithelia
In vivo multiphoton microscopy of NADH and FAD redox states, fluorescence lifetimes, and cellular morphology in precancerous epithelia
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DOI：
10.1073/pnas.0708425104
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发表时间：
2007-12-04
影响因子：
11.1
通讯作者：
Ramanujam, Nirmala
中科院分区：
文献类型：
--
作者：
Skala, Melissa C.;Riching, Kristin M.;Ramanujam, Nirmala
还原型NADH和FAD的相对量以及这些代谢电子载体的微环境的代谢成像可用于非侵入性地监测代谢的变化，这是致癌的标志之一。这项研究结合了细胞氧化还原率，NADH和FAD寿命，以及三维亚细胞形态成像，以确定癌症发展最早阶段体内代谢和结构对比的内在来源。在正常组织中，随着上皮深度的增加，核质比（NCR）显著增加（P < 0.05），而在癌前组织中，NCR随上皮深度的增加无显著变化。与正常复层鳞状上皮浅层中的更成熟细胞相比，分化较低的基底上皮细胞中的氧化还原比显著降低，表明NCR增加的细胞中的代谢活性增加。而癌前组织中浅表细胞和基底细胞的氧化还原率无明显差异。与正常上皮组织相比，在低级别和高级别上皮癌前病变中观察到蛋白结合的NADH的贡献和寿命（整个上皮的平均值）显著降低。此外，仅在高级别癌前病变中观察到蛋白结合FAD寿命的显著增加和蛋白结合FAD贡献的减少。与正常细胞相比，在癌前细胞中观察到氧化还原比、NADH和FAD荧光寿命的细胞内变异性增加。
Metabolic imaging of the relative amounts of reduced NADH and FAD and the microenvironment of these metabolic electron carriers can be used to noninvasively monitor changes in metabolism, which is one of the hallmarks of carcinogenesis. This study combines cellular redox ratio, NADH and FAD lifetime, and subcellular morphology imaging in three dimensions to identify intrinsic sources of metabolic and structural contrast in vivo at the earliest stages of cancer development. There was a significant (P < 0.05) increase in the nuclear to cytoplasmic ratio (NCR) with depth within the epithelium in normal tissues; however, there was no significant change in NCR with depth in precancerous tissues. The redox ratio significantly decreased in the less differentiated basal epithelial cells compared with the more mature cells in the superficial layer of the normal stratified squamous epithelium, indicating an increase in metabolic activity in cells with increased NCR. However, the redox ratio was not significantly different between the superficial and basal cells in precancerous tissues. A significant decrease was observed in the contribution and lifetime of protein-bound NADH (averaged over the entire epithelium) in both low- and high-grade epithelial precancers compared with normal epithelial tissues. In addition, a significant increase in the protein-bound FAD lifetime and a decrease in the contribution of protein-bound FAD are observed in high-grade precancers only. Increased intracellular variability in the redox ratio, NADH, and FAD fluorescence lifetimes were observed in precancerous cells compared with normal cells.