Department of Biology and Ecology, Faculty of Sciences, University of Novi Sad, Trg Dositeja Obradovića 2, 21000 Novi Sad, Serbia.
Department of Cytology, Institute for Biological Research "Siniša Stanković"-National Institute of Republic of Serbia, University of Belgrade, Bulevar despota Stefana 142, 11060 Belgrade, Serbia.
Int J Mol Sci. 2022 May 28;23(11):6062. doi: 10.3390/ijms23116062.
Acrylamide (AA) toxicity is associated with oxidative stress. During detoxification, AA is either coupled to gluthatione or biotransformed to glycidamide by the enzyme cytochrome P450 2E1 (CYP2E1). The aim of our study was to examine the hepatotoxicity of AA in vivo and in vitro. Thirty male Wistar rats were treated with 25 or 50 mg/kg b.w. of AA for 3 weeks. Qualitative and quantitative immunohistochemical evaluation of inducible nitric oxide synthase (iNOS), CYP2E1, catalase (CAT), superoxide dismutase 1 (SOD1), and SOD2 expression in liver was carried out. Bearing in mind that the liver is consisted mainly of hepatocytes, in a parallel study, we used the rat hepatoma cell line H4IIE to investigate the effects of AA at IC20 and IC50 concentrations on the redox status and the activity of CAT, SOD, and glutathione-S-transferase (GST), their gene expression, and CYP2E1 and iNOS expression. Immunohistochemically stained liver sections showed that treatment with AA25mg induced a significant decrease of CYP2E1 protein expression (p < 0.05), while treatment with AA50mg led to a significant increase of iNOS protein expression (p < 0.05). AA treatment dose-dependently elevated SOD2 protein expression (p < 0.05), while SOD1 protein expression was significantly increased only at AA50mg (p < 0.05). CAT protein expression was not significantly affected by AA treatments (p > 0.05). In AA-treated H4IIE cells, a concentration-dependent significant increase in lipid peroxidation and nitrite levels was observed (p < 0.05), while GSH content and SOD activity significantly decreased in a concentration-dependent manner (p < 0.05). AA IC50 significantly enhanced GST activity (p < 0.05). The level of mRNA significantly increased in a concentration-dependent manner for iNOS, SOD2, and CAT in AA-treated H4IIE cells (p < 0.05). AA IC50 significantly increased the transcription of SOD1, GSTA2, and GSTP1 genes (p < 0.05), while AA IC20 significantly decreased mRNA for CYP2E1 in H4IIE cells (p < 0.05). Obtained results indicate that AA treatments, both in vivo and in vitro, change hepatocytes; drug-metabolizing potential and disturb its redox status.
丙烯酰胺 (AA) 毒性与氧化应激有关。在解毒过程中，AA 要么与谷胱甘肽偶联，要么被细胞色素 P450 2E1 (CYP2E1) 酶转化为环氧丙酰胺。我们的研究目的是体内和体外研究 AA 的肝毒性。30 只雄性 Wistar 大鼠用 25 或 50mg/kg b.w. 的 AA 处理 3 周。对肝组织中诱导型一氧化氮合酶 (iNOS)、CYP2E1、过氧化氢酶 (CAT)、超氧化物歧化酶 1 (SOD1) 和 SOD2 表达进行定性和定量免疫组织化学评估。鉴于肝脏主要由肝细胞组成，在一项平行研究中，我们使用大鼠肝癌细胞系 H4IIE 研究 AA 在 IC20 和 IC50 浓度下对氧化还原状态和 CAT、SOD 和谷胱甘肽-S-转移酶 (GST) 的活性、基因表达以及 CYP2E1 和 iNOS 表达的影响。免疫组织化学染色的肝组织切片显示，用 25mg AA 处理会显著降低 CYP2E1 蛋白表达（p<0.05），而用 50mg AA 处理会显著增加 iNOS 蛋白表达（p<0.05）。AA 处理剂量依赖性地增加 SOD2 蛋白表达（p<0.05），而 SOD1 蛋白表达仅在 50mg AA 时显著增加（p<0.05）。CAT 蛋白表达不受 AA 处理的显著影响（p>0.05）。在 AA 处理的 H4IIE 细胞中，观察到脂质过氧化和亚硝酸盐水平的浓度依赖性显著增加（p<0.05），而 GSH 含量和 SOD 活性则呈浓度依赖性显著降低（p<0.05）。AA IC50 显著增强 GST 活性（p<0.05）。AA 处理的 H4IIE 细胞中，iNOS、SOD2 和 CAT 的 mRNA 水平呈浓度依赖性显著增加（p<0.05）。AA IC50 显著增加 SOD1、GSTA2 和 GSTP1 基因的转录（p<0.05），而 AA IC20 显著降低 H4IIE 细胞中 CYP2E1 的 mRNA（p<0.05）。结果表明，AA 处理无论是在体内还是体外，都会改变肝细胞；改变其药物代谢能力并扰乱其氧化还原状态。
