Measuring "free" iron levels in Caenorhabditis elegans using low-temperature Fe(III) electron paramagnetic resonance spectroscopy.
Measuring "free" iron levels in Caenorhabditis elegans using low-temperature Fe(III) electron paramagnetic resonance spectroscopy.
复制标题
使用低温 Fe(III) 电子顺磁共振波谱测量秀丽隐杆线虫中的“游离”铁水平。
DOI：
10.1016/j.ab.2006.08.025
复制
发表时间：
2006
影响因子：
2.9
通讯作者：
Srinivasan,Chandra
中科院分区：
文献类型：
--
作者：
Pate,KiraT;Rangel,NatalieA;Fraser,Brian;Clement,MatthewHS;Srinivasan,Chandra
由体内自由基引起的氧化应激与衰老过程和许多人类疾病有关。由于自由基，特别是超氧化物，很难测量，一种替代的间接方法来测量氧化应激水平已被成功地用于大肠杆菌和酵母。该方法基于超氧化物水平升高与暴露于溶剂的[4 Fe-4S]酶簇释放铁之间的拟议联系，最终导致羟基自由基产生的增加。在过去使用细菌和酵母的研究中，发现超氧化物产生或氧化应激之间的正相关性，由于超氧化物在生物体和电子顺磁共振（EPR）可检测的“游离”铁水平。在目前的研究中，我们已经开发出一种可靠和有效的方法，用于测量“游离”的铁水平在秀丽隐杆线虫使用低温Fe（III）EPR在g=4.3。该方法使用在平板上生长的同步蠕虫培养物，在填充EPR管之前，将平板均质化并用去铁胺（一种Fe（III）螯合剂）处理。均质化被认为不会改变“游离”铁水平，而去铁胺治疗显着提高这些水平，表明存在的Fe（II）和Fe（III）的“游离”铁池。自由基水平和观察到的“游离”铁水平之间的相关性进行了检查，通过使用热应激和百草枯处理。的Fe（III）EPR信号的强度，从而浓度的“自由”铁池，改变自由基水平而不改变总铁水平的治疗。这项研究提供了揭示氧化应激，“游离”铁水平和C寿命之间的相关性所需的基础。优雅的。
Oxidative stress, caused by free radicals within the body, has been associated with the process of aging and many human diseases. Because free radicals, in particular superoxide, are difficult to measure, an alternative indirect method for measuring oxidative stress levels has been used successfully in Escherichia coli and yeast. This method is based on a proposed connection between elevated superoxide levels and release of iron from solvent-exposed [4Fe–4S] enzyme clusters that eventually leads to an increase in hydroxyl radical production. In past studies using bacteria and yeast, a positive correlation was found between superoxide production or oxidative stress due to superoxide within the organism and electron paramagnetic resonance (EPR) detectable “free” iron levels. In the current study, we have developed a reliable and efficient method for measuring “free” iron levels in Caenorhabditis elegans using low-temperature Fe(III) EPR at g=4.3. This method uses synchronized worm cultures grown on plates that are homogenized and treated with desferrioxamine, an Fe(III) chelator, prior to packing the EPR tube. Homogenization was found not to alter “free” iron levels, whereas desferrioxamine treatment significantly raised these levels, indicating the presence of both Fe(II) and Fe(III) in the “free” iron pool. The correlation between free radical levels and the observed “free” iron levels was examined by using heat stress and paraquat treatment. The intensity of the Fe(III) EPR signal, and thus the concentration of the “free” iron pool, varied with the treatments that altered radical levels without changing the total iron levels. This study provides the groundwork needed to uncover the correlation among oxidative stress, “free” iron levels, and longevity in C. elegans.