Direct ESR measurement of mutagenic slow releasing long-lived radicals inind iated manmalian cells with adequate dose forradiaion biology

我们在没有任何自旋捕捉剂的情况下，用电子自旋共振波谱在4Gy射线辐照的叙利亚仓鼠胚胎(SHE)细胞中新发现了新的缓释长寿命自由基(SRLRs)。据我们所知，现在直接从整个哺乳动物细胞中报告ESR灵巧的自由基可能是很快的，而不需要任何提取或浓缩。在21℃群体倍增时，原代SHE细胞大约有1.8(±0.07)×10^6自旋/细胞。当细胞受到4Gy射线照射并保存1、5、12和20h后，自由基浓度分别增加到1.9(±0.09)×106、2.2(±0.15)×106、2.2(±0.12)×106和2.1(±0.05)×106Spins/cell，增量分别为1.0×105(+6%)、3.8×10^5(+21%)、3.6×10^5(+20%)和3.2×10^5(+17%)。由于电离辐射直接诱导的ROS或RN在照射后1至20小时内完全消失，这种增加不是由于细胞成分的直接或间接辐射分解，而是由于某些诱导的…辐射可能会增强更多的反应。抗坏血酸处理2 h可抑制SRLLRs水平的增加，幅度在70%以内。添加Myxthiiazol作为线粒体电子传递的抑制剂，可抑制受照射细胞中SRLLRs水平的增加。0.5μM中的霉唑主要阻断线粒体复合体III呼吸链b-c1段的电子传递。经辐射修饰的线粒体中的电子异常泄漏，在受辐射的细胞中与溶剂化的氧分子产生超氧化物，然后超氧化物歧变酶将超氧化物转化为过氧化氢。与过氧化氢的反应可能会产生缓慢释放的长寿命自由基。在培养基中直接加入过氧化氢可提高它们的水平。这些结果表明，SRLLRs很可能是由功能失调的线粒体中的过氧化氢产生的。由于SRLLR具有长达20h以上的寿命，并被L-抗坏血酸处理后减少，因此它们可能与辐射的延迟效应有关。较少

We newly found novel slow-releasing long-lived radicals (SRLLRs) in 4 Gy y-irradiated Syrian hamster embryo (SHE) cells by ESR spectroscopy without any spin-trapping agent. As far as we know, it might be fast time to report the radicals those were deft directly by ESR from whole mammalian cells without any extraction or concentration. Primary SHE cells at population doublings of 21^st have ca. 1.8 (±0.07) ×10^6 spins/cell. When the cell were y-irradiated in 4 Gy and stored for I, 5, 12, and 20 h, concentrations of radicals increased to 1.9(±0.09)×10^6, 2.2 (±0.15)×10^6, 2.2 (±0.12) ×10^6 and 2.1(±0.05)×10^6 spins/cell, respectively, and the increments were 1.0×10^5(+6 %), 3.8×10^5 (+21%), 3.6×10^5 (+20%), and 3.2×10^5(+17%), spins/cell, respectively. Because ROS or RNS directly induced by ionizing radiation are completely disappeared in the course of time as I to 20 h after irradiation, the increase is not due to direct or indirect radiolysis of cell components but due to some induced … More reactions those may be enhanced by the radiation. Pest-treatment of ascorbate 2 h suppressed the increase in the levels of SRLLRs within 70%. Addition of myxothiazol, as an inhibitor of electron transportation in mitochondria, suppressed the increase in the levels of SRLLRs in irradiated cells. The treatment of myxothiazol in 0.5 μM mainly blocks the electron transportation within the cytochrone b-c1 segment of the respiratory chain in complex III in mitochondria. Abnormal leaks of electron from radiation-modified studious in the irradiated mitochondria produce superoxide with solvated oxygen molecules in irradiated cells, then superoxide dysmutase convert superoxide to hydrogen peroxide. The reaction with the hydrogen peroxide is likely to produce the slow releasing long-lived radicals. Direct addition of hydrogen peroxide to the culture medium increased the levels of them. These results indicate that SRLLRs are likely to be produced by hydrogen peroxide from dysfunctional mitochondria. Because SRLLRs have a long lifetime over 20h and reduced by L-ascorbic acid treatment, they might be responsible of delayed effects of radiation. Less