Inhibition of succinate:ubiquinone reductase and decrease of ubiquinol in nephrotoxic cysteine S-conjugate-induced oxidative cell injury.
Inhibition of succinate:ubiquinone reductase and decrease of ubiquinol in nephrotoxic cysteine S-conjugate-induced oxidative cell injury.
复制标题
抑制琥珀酸：泛醌还原酶并减少泛醇在肾毒性半胱氨酸 S-缀合物诱导的氧化细胞损伤中的作用。
作者：
B. van de Water;J. Zoeteweij;H. D. de Bont;J. Nagelkerke
在新鲜分离的大鼠肾近端小管细胞（PTC）中，使用肾毒素S-（1，2-二氯乙烯基）-L-半胱氨酸（DCVC）研究复合物II在细胞抗氧化应激保护中的作用。DCVC引起PTC的氧化应激，通过流式细胞术与二氢罗丹明-123测定;这种荧光探针很容易被初级氢过氧化物氧化，如脂质过氧化过程中形成的。氧化应激可以通过抑制β-裂解酶介导的反应性DCVC代谢物的形成和与细胞大分子的共价结合来防止，用氨基氧乙酸（AOA），或通过抗氧化剂N，N '-二苯基-对苯二胺。AOA和DPPD都能防止细胞死亡。DCVC诱导的氧化应激与复合物II的琥珀酸：泛醌还原酶（SQR）活性的降低有关，而复合物I的NADH：泛醌还原酶活性不受影响。AOA阻止了对SQR活性的影响，而N，N '-二苯基-对苯二胺则没有。用噻吩甲酰三氟丙酮（TTFA）抑制SQR活性增强DCVC诱导的氧化性细胞损伤，表明SQR活性参与抗氧化途径。为了更详细地研究这一点，在含有甘氨酸的缓冲液中用细胞色素c氧化酶抑制剂KCN处理PTC，所述甘氨酸防止KCN引起的细胞死亡。甘氨酸不影响DCVC引起的细胞死亡。KCN可防止DCVC诱导的氧化应激和细胞死亡。KCN的细胞保护作用可以通过抑制SQR的活性草酰乙酸或TTFA，而无论是复合物I或III与鱼藤酮和抗霉素，分别抑制，并没有阻止它。DCVC对复合物II的影响与减少细胞的量减少泛醌（QH 2）; KCN介导的细胞保护作用与细胞QH 2增加60%。鱼藤酮几乎完全抑制泛醌还原，即使在KCN的存在下，而草酰乙酸与KCN的组合导致QH 2水平与对照相当。这表明复合物II的SQR活性而不是还原泛醌（QH 2）的细胞含量作为细胞抗氧化机制的一部分在抗氧化应激的细胞保护中是重要的。
The role of complex II in the cellular protection against oxidative stress was investigated in freshly isolated rat renal proximal tubular cells (PTC) with the use of the nephrotoxin S-(1,2-dichlorovinyl)-L-cysteine (DCVC). DCVC caused oxidative stress in PTC as determined by flow cytometry with dihydrorhodamine-123; this fluorescent probe is readily oxidized by primary hydroperoxides such as those formed during lipid peroxidation. The oxidative stress could be prevented by inhibition of the beta-lyase-mediated formation and covalent binding to cellular macromolecules of reactive DCVC metabolites, with amino oxyacetic acid (AOA), or by the antioxidant N,N'-diphenyl-p-phenylenediamine. Both AOA and DPPD also prevented cell death. The DCVC-induced oxidative stress was associated with a decrease in the succinate:ubiquinone reductase (SQR) activity of complex II, whereas NADH:ubiquinone reductase activity of complex I remained unaffected. AOA prevented the effect on SQR activity, whereas N,N'-diphenyl-p-phenylenediamine did not. Inhibition of SQR activity with thenoyl trifluoracetone (TTFA) potentiated the DCVC-induced oxidative cell injury, suggesting the involvement of SQR activity in an antioxidant pathway. To investigate this in greater detail, PTC were treated with an inhibitor of cytochrome-c-oxidase, KCN, in a buffer containing glycine, which prevents cell death by KCN. Glycine did not affect cell death by DCVC. KCN prevented the DCVC-induced oxidative stress and cell death. KCN cytoprotection could be prevented by inhibition of SQR activity with oxaloacetate or TTFA, whereas inhibition of either complex I or III with rotenone and antimycin, respectively, did not prevent it. The effect of DCVC on complex II was associated with a decrease in the cellular amount of reduced ubiquinone (QH2); the KCN-mediated cytoprotection was related to a 60% increase of cellular QH2. Rotenone almost completely inhibited ubiquinone reduction even in the presence of KCN, whereas oxaloacetate in combination with KCN resulted in QH2 levels comparable to control. This suggests that the SQR activity by complex II rather than the cellular content of reduced ubiquinone (QH2) is important as a part of the cellular antioxidant machinery in the cyto-protection against oxidative stress.