Acetaminophen hepatotoxicity and mechanisms of its protection by N-acetylcysteine: a study of Hep3B cells
Acetaminophen hepatotoxicity and mechanisms of its protection by N-acetylcysteine: a study of Hep3B cells
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DOI：
10.1078/0940-2993-00215
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发表时间：
2002-02-01
影响因子：
--
通讯作者：
Iancu, TC
中科院分区：
文献类型：
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作者：
Manov, I;Hirsh, M;Iancu, TC
对乙酰氨基酚 (AAP) 的肝毒性，导致小叶中心坏死，经常在自杀企图后出现，尤其是青少年，但在婴儿过度使用后也会出现这种情况。导致肝细胞死亡的亚细胞和分子序列尚不清楚，因此我们通过使用暴露于 AAP 和用作保护剂的 N-乙酰半胱氨酸 (NAC) 的培养的肝癌来源细胞 (Hep3B) 来研究 AAP 的肝毒性。具体来说，我们研究了细胞凋亡和氧化损伤作为 AAP 相关细胞毒性的推定机制的作用。 Hep3B 细胞暴露于 AAP (5-25 mM) 和 NAC (5 mM) 不同的时间段。通过 Alamar Blue 还原试验和 LDH 评估细胞活力。通过测量活性氧（ROS）和谷胱甘肽来评估氧化损伤。通过流式细胞术和透射电子显微镜研究AAP诱导的细胞凋亡。我们发现： 1. 在 Hep3B 细胞中，AAP 会产生时间和浓度依赖性的细胞毒作用，导致氧化应激、线粒体功能障碍、膜通透性改变和细胞凋亡； 2. 在AAP细胞毒性过程中，ROS的产生作为早期事件出现，先于活力下降、LDH渗漏、谷胱甘肽耗竭和细胞凋亡； 3. NAC保护Hep3B细胞免受AAP诱导的氧化损伤，但不能阻止细胞凋亡。
Acetaminophen (AAP) hepatotoxicity, resulting in centrilobular necrosis, is frequently encountered following suicidal attempts, especially by adolescents, but also after its excessive use in infants. The subcellular and molecular sequences leading to hepatocellular cell death are not yet clear, We therefore investigated AAP hepatotoxicity by using cultured hepatoma-derived cells (Hep3B) exposed to AAP and N-acetylcysteine (NAC), used as a protective agent. Specifically, we studied the role of apoptosis and oxidative damage as putative mechanisms of AAP-associated cytotoxicity. Hep3B cells were exposed to AAP (5-25 mM) and NAC (5 mM) for different time periods. Cell viability was assessed by the Alamar Blue Reduction Test and LDH. Oxidative damage was evaluated by measuring reactive oxygen species (ROS) and glutathione. AAP-induced apoptosis was investigated by flow cytometry and transmission electron microscopy. We found that: 1. In Hep3B cells, AAP causes a time- and concentration-dependent cytotoxic effect, leading to oxidative stress, mitochondrial dysfunction, alterations of membrane permeability and apoptosis; 2. In the course of AAP cytotoxicity, the generation of ROS appears as an early event which precedes decrease of viability, LDH leakage, glutathione depletion and apoptosis; 3. NAC protects Hep3B cells from AAP-induced oxidative injury, but does not prevent apoptosis.