Zhu Jian-Hong, Lei Xin Gen
Department of Animal Science, Cornell University, 252 Morrison Hall, Ithaca, NY 14853, USA.
Exp Biol Med (Maywood). 2006 May;231(5):545-52. doi: 10.1177/153537020623100508.
This study was conducted to determine the impact of knockout of selenium (Se)-dependent glutathione peroxidase-1 (GPX1-/-) or double knockout of GPX1 and copper, zinc (Cu,Zn)-super-oxide dismutase (SOD1) on cell death induced by acetaminophen (APAP) and its major toxic metabolite N-acetyl-P-benzoquinoneimine (NAPQI). Primary hepatocytes were isolated from GPX1-/-, double knockout of GPX1 and SOD1 (DKO), and their wild-type (WT) mice and were treated with 5 mM APAP or 100 microM NAPQI for 0, 6, and 12 hrs. Compared with the WT cells, the GPX1-/- and DKO hepatocytes were more resistant (P < 0.05) to the APAP-induced cell death but less resistant to the NAPQI-induced cell death. The APAP-mediated glutathione (GSH) depletion was greater (P < 0.05) at 6 hrs in the WT cells than in the GPX1-/- and DKO cells, whereas there was no genotype effect on the NAPQI-mediated GSH depletion. The DKO cells had lower (P < 0.05) microsomal cytochrome P450 2E1 activities, but higher (P < 0.05) glutathione reductase and thioredoxin reductase activities than the WT cells at 0 hrs, and they responded differently to the APAP and NAPQI treatments. Glutathione-S-transferase activity was not affected by genotypes or treatments. Neither APAP nor NAPQI induced nitric oxide production or protein nitration in cells of any genotype. However, the GPX1-/- and DKO cells were more resistant to peroxynitrite-mediated protein nitration than were the WT cells. In conclusion, double null of GPX1 and SOD1 enhanced the resistance of mouse primary hepatocytes to APAP toxicity by affecting events prior to or at NAPQI formation. While the double knockout attenuated the peroxynitrite-mediated protein nitration in hepatocytes, no protein nitration was detected in these cells treated with APAP or NAPQI.
本研究旨在确定硒（Se）依赖性谷胱甘肽过氧化物酶-1基因敲除（GPX1-/-）或GPX1与铜锌（Cu,Zn）超氧化物歧化酶（SOD1）双基因敲除对乙酰氨基酚（APAP）及其主要毒性代谢产物N-乙酰-p-苯醌亚胺（NAPQI）诱导的细胞死亡的影响。从GPX1-/-、GPX1和SOD1双基因敲除（DKO）小鼠及其野生型（WT）小鼠中分离出原代肝细胞，并用5 mM APAP或100 microM NAPQI处理0、6和12小时。与WT细胞相比，GPX1-/-和DKO肝细胞对APAP诱导的细胞死亡更具抗性（P < 0.05），但对NAPQI诱导的细胞死亡抗性较低。WT细胞在6小时时APAP介导的谷胱甘肽（GSH）消耗比GPX1-/-和DKO细胞更大（P < 0.05），而NAPQI介导的GSH消耗没有基因型效应。DKO细胞在0小时时微粒体细胞色素P450 2E1活性较低（P < 0.05），但谷胱甘肽还原酶和硫氧还蛋白还原酶活性高于WT细胞（P < 0.05），并且它们对APAP和NAPQI处理的反应不同。谷胱甘肽-S-转移酶活性不受基因型或处理的影响。APAP和NAPQI均未诱导任何基因型细胞产生一氧化氮或蛋白质硝化。然而，GPX1-/-和DKO细胞比WT细胞对过氧亚硝酸盐介导的蛋白质硝化更具抗性。总之，GPX1和SOD1双基因缺失通过影响NAPQI形成之前或之时的事件增强了小鼠原代肝细胞对APAP毒性的抗性。虽然双基因敲除减弱了肝细胞中过氧亚硝酸盐介导的蛋白质硝化，但在用APAP或NAPQI处理的这些细胞中未检测到蛋白质硝化。
