Everett S A, Smith K A, Patel K B, Dennis M F, Stratford M R, Wardman P
Gray Laboratory Cancer Research Trust, Mount Vernon Hospital, Northwood, Middlesex, UK.
Br J Cancer Suppl. 1996 Jul;27:S172-6.
The free-radical intermediates and the stable products formed on one-electron oxidation of hydroxyguanidine (HOG) were investigated in order to suggest a mechanistic basis for HOG-induced cytotoxicity and cytostasis in leukaemia HL60 cells. The azide radical (generated radiolytically) reacted with HOG to produce a carbon-centred radical which in the absence of oxygen decays by a first-order process (k = 3.2 x 10(3) s-1) to yield nitric oxide (NO) and urea. Although the HOG radical reacts rapidly with oxygen (rate constant for O2 addition, k = 4.2 x 10(8) dm3 mol-1 s-1) this neither prevented the elimination of NO. nor generated alternative nitrogen oxides (e.g. peroxynitrite) capable of contributing to cellular oxidative stress. The detection of NO. in HL60 cells corroborated mechanistic studies that oxidative denitrification of HOG does not require catalysis by nitric oxide synthase. Quantitation of NO. by electron paramagnetic resonance (EPR) spectroscopy (utilising a NO. -selective probe) shows higher amounts of NO. under anoxic conditions, reflecting competition for NO. with molecular oxygen in oxic cells. Inhibition of cytochrome P450 and myeloperoxidase activity decreased NO. production thereby identifying these enzyme systems as capable of oxidizing HOG in vitro. A correlation exists between the intracellular levels of NO. with both the cytotoxic and cytostatic effects of HOG within HL60 cells. A higher toxicity was observed with hypoxic than with oxic cells. The lower levels of NO. associated with aerobic conditions caused a G1 --> S block in the cell cycle which under anoxia potentiated NO. -induced apoptotic cell death.
为了揭示羟基胍（HOG）诱导白血病HL60细胞产生细胞毒性和细胞生长停滞的机制基础，对HOG单电子氧化形成的自由基中间体和稳定产物进行了研究。叠氮自由基（通过辐射分解产生）与HOG反应生成一个以碳为中心的自由基，该自由基在无氧条件下按一级反应过程衰变（k = 3.2×10³ s⁻¹），生成一氧化氮（NO）和尿素。尽管HOG自由基与氧气反应迅速（氧气加成的速率常数，k = 4.2×10⁸ dm³ mol⁻¹ s⁻¹），但这既没有阻止NO的消除，也没有产生能够导致细胞氧化应激的其他氮氧化物（如过氧亚硝酸盐）。在HL60细胞中检测到NO，证实了HOG的氧化脱氮不需要一氧化氮合酶催化的机制研究。通过电子顺磁共振（EPR）光谱（使用NO选择性探针）对NO进行定量分析表明，在缺氧条件下NO的含量更高，这反映了在有氧细胞中NO与分子氧之间的竞争。细胞色素P450和髓过氧化物酶活性的抑制降低了NO的产生，从而确定这些酶系统能够在体外氧化HOG。HL60细胞内NO的水平与HOG的细胞毒性和细胞生长停滞作用之间存在相关性。观察到缺氧细胞比有氧细胞具有更高的毒性。与有氧条件相关的较低水平的NO导致细胞周期中G1期向S期的阻滞，在缺氧条件下这种阻滞增强了NO诱导的凋亡性细胞死亡。
