Lee V M, Keefer L K, Archer M C
Department of Medical Biophysics, University of Toronto, Canada.
Chem Res Toxicol. 1996 Dec;9(8):1319-24. doi: 10.1021/tx960077u.
N-Nitrosodimethylamine (NDMA) is a potent hepatotoxic agent in rats. NDMA causes cell death that does not correlate with known mechanisms of toxicity such as the production of oxidative stress or covalent binding to proteins. The following studies were designed to determine whether NDMA cytotoxicity is the result of metabolic denitrosation or alpha-hydroxylation of the nitrosamine. We determined the toxicity of various metabolites of NDMA in monolayer cultures of primary rat hepatocytes. NDMA was toxic at 0.1 mM in our cultures, but the metabolites formaldehyde, methanol, and methylamine were not toxic at this concentration. We used diazeniumdiolates that spontaneously release nitric oxide (NO) in aqueous media to deliver NO to hepatocytes in culture. The results show that, while NO released from diazeniumdiolates causes death in hepatocytes, the levels of NO produced during NDMA metabolism are insufficient to account for the toxicity of the nitrosamine. NDMA-d6, the fully deuteriated form of NDMA that undergoes approximately twice as much denitrosation in vivo as NDMA, was significantly less cytotoxic than NDMA. In contrast, N-nitroso(acetoxymethyl)methylamine (AcO-NDMA), a stable precursor of the methanediazonium ion, was found to cause toxicity equivalent of NDMA on a molar basis. Altogether, our results with methylamine, formaldehyde, methanol, the diazeniumdiolates, and NDMA-d6 indicate that NDMA toxicity is not the result of metabolic denitrosation, while the toxicity of AcO-NDMA provides strong strong evidence that the formation of the methanediazonium ion via alpha-hydroxylation leads to cell death.
N-亚硝基二甲胺（NDMA）是一种对大鼠有强肝毒性的物质。NDMA导致的细胞死亡与已知的毒性机制（如氧化应激的产生或与蛋白质的共价结合）无关。以下研究旨在确定NDMA的细胞毒性是亚硝胺代谢脱亚硝化作用还是α-羟基化作用的结果。我们测定了NDMA各种代谢产物在原代大鼠肝细胞单层培养物中的毒性。在我们的培养物中，NDMA在0.1 mM时具有毒性，但代谢产物甲醛、甲醇和甲胺在此浓度下无毒性。我们使用在水性介质中自发释放一氧化氮（NO）的重氮二醇盐将NO递送至培养的肝细胞。结果表明，虽然从重氮二醇盐释放的NO会导致肝细胞死亡，但NDMA代谢过程中产生的NO水平不足以解释亚硝胺的毒性。NDMA-d6是NDMA的完全氘代形式，其在体内的脱亚硝化作用大约是NDMA的两倍，但其细胞毒性明显低于NDMA。相比之下，发现N-亚硝基（乙酰氧基甲基）甲胺（AcO-NDMA），一种甲重氮离子的稳定前体，在摩尔基础上引起的毒性与NDMA相当。总之，我们使用甲胺、甲醛、甲醇、重氮二醇盐和NDMA-d6的研究结果表明，NDMA的毒性不是代谢脱亚硝化作用的结果，而AcO-NDMA的毒性提供了有力证据，表明通过α-羟基化形成甲重氮离子会导致细胞死亡。
