Kim B S, Yang K H, Haggerty H G, Holsapple M P
Department of Pharmacology and Toxicology, Medical College of Virginia/Virginia Commonwealth University, Richmond 23298.
Mutat Res. 1989 Aug;213(2):185-93. doi: 10.1016/0027-5107(89)90150-4.
In an attempt to elucidate the mechanism whereby primary hepatocytes, but not liver S9 homogenates, generate immunosupprssive metabolites of dimethylnitrosamine (DMN), the production of DNA single-strand breaks (SSB) in unstimulated splenocytes was investigated with alkaline-elution analysis. Both hepatocytes and S9 homogenates induced SSB in cultured splenocytes by DMN - minimum detectable doses with the two metabolic activation systems (MAS) were 1 microM and 5 mM, respectively. DNA elution profiles were linear in splenocytes co-cultured with DMN and hepatocytes and convex in splenocytes incubated with DMN and S9 homogenates. Aminoacetonitrile (AAN; 10 mM), a DMN demethylase inhibitor, reversed SSB in splenocytes when incubated with either MAS. Addition of exogenous calf-thymus DNA to the hepatocyte co-culture medium did not affect the production of SSB. Rocking the hepatocyte-splenocyte cultures changed the elution profile from linear to convex. All of these treatments have been previously shown to block the immunosuppression by DMN in the hepatocyte co-culture system. These results indicate that the immunosuppression by DMN is not related to DNA damage, as measured by the production of SSB, and suggest that the metabolism of DMN to intermediates capable of producing genotoxicity and immunotoxicity may be qualitatively and/or quantitatively different.
为了阐明原代肝细胞而非肝脏S9匀浆产生二甲基亚硝胺（DMN）免疫抑制性代谢产物的机制，采用碱性洗脱分析法研究了未刺激脾细胞中DNA单链断裂（SSB）的产生情况。肝细胞和S9匀浆均通过DMN在培养的脾细胞中诱导SSB——两种代谢活化系统（MAS）的最低可检测剂量分别为1 microM和5 mM。在与DMN和肝细胞共培养的脾细胞中，DNA洗脱曲线呈线性，而在与DMN和S9匀浆孵育的脾细胞中呈凸形。氨基乙腈（AAN；10 mM），一种DMN脱甲基酶抑制剂，在与任何一种MAS孵育时均可逆转脾细胞中的SSB。向肝细胞共培养基中添加外源性小牛胸腺DNA并不影响SSB的产生。摇晃肝细胞 - 脾细胞培养物可使洗脱曲线从线性变为凸形。所有这些处理先前已被证明可在肝细胞共培养系统中阻断DMN的免疫抑制作用。这些结果表明，DMN的免疫抑制作用与通过SSB产生所衡量的DNA损伤无关，并提示DMN代谢为具有遗传毒性和免疫毒性的中间体的过程在质量和/或数量上可能存在差异。
