Saito Yoshiro, Nishio Keiko, Yoshida Yasukazu, Niki Etsuo
Human Stress Signal Research Center (HSSRC), National Institute of Advanced Industrial Science and Technology (AIST), 1-8-31 Midorigaoka, Ikeda, Osaka 563-8577, Japan.
Toxicology. 2005 Jun 1;210(2-3):235-45. doi: 10.1016/j.tox.2005.02.006.
It is well known that formaldehyde (HCHO) and reactive oxygen species (ROS), such as free radicals, are cytotoxic as well as potentially carcinogenic. Although the individual effects of these reactants on cells have been investigated, the cytotoxicity exerted by the coexistence of HCHO and reactive radicals is poorly understood. The present study using Jurkat cells demonstrated that the coexistence of HCHO with water-soluble radical initiator, 2,2'-azobis-[2-(2-imidazolin-2-yl)propane] dihydrochloride (AIPH) dramatically decreased cell viability, and that under such conditions scant cell death was observable induced by either of the reactants alone. Based on the results of phosphatidylserine exposure and caspase activation, this observed cell death, in fact, was apparently necrotic rather than apoptotic. To understand the mechanisms of the cell toxicity of HCHO and AIPH, we assessed two kinds of oxidative stress markers such as cellular glutathione (GSH) content and cellular ROS, and the DNA-protein cross-links, which formed as the result of HCHO treatment. A marked decrease in total cellular GSH was observed not only in the case of the coexistence conditions but also with AIPH alone. Dichlorodihydrofluorescein (DCF) assay revealed that cellular ROS were synergistically increased before cell death. The formation of DNA-protein cross-links was observed in the presence of HCHO and AIPH, and the extent was similar to HCHO alone. Co-incubation with semicarbazide, which inactivates HCHO, prevented this cell death induced by a combination of HCHO and AIPH. Semicarbazide also exhibited an inhibitory effect on the synergistic increment of cellular ROS and the formation of DNA-protein cross-links. These results suggest that the free radicals from AIPH induced GSH reduction, while HCHO resulted in the formation of DNA-protein cross-links, eventuating in a synergistic, incremental increase of cellular ROS and cell death brought about by this combination.
众所周知，甲醛（HCHO）和活性氧（ROS），如自由基，具有细胞毒性且有潜在致癌性。尽管已经研究了这些反应物对细胞的单独作用，但对HCHO与活性自由基共存时所产生的细胞毒性了解甚少。本研究使用Jurkat细胞表明，HCHO与水溶性自由基引发剂2,2'-偶氮双-[2-(2-咪唑啉-2-基)丙烷]二盐酸盐（AIPH）共存会显著降低细胞活力，并且在这种条件下，单独任何一种反应物都几乎观察不到细胞死亡。基于磷脂酰丝氨酸暴露和半胱天冬酶激活的结果，这种观察到的细胞死亡实际上显然是坏死性的而非凋亡性的。为了了解HCHO和AIPH的细胞毒性机制，我们评估了两种氧化应激标志物，如细胞内谷胱甘肽（GSH）含量和细胞内ROS，以及HCHO处理后形成的DNA-蛋白质交联。不仅在共存条件下，而且单独使用AIPH时，都观察到细胞内总GSH显著降低。二氯二氢荧光素（DCF）测定表明，细胞内ROS在细胞死亡前协同增加。在HCHO和AIPH存在的情况下观察到了DNA-蛋白质交联的形成，其程度与单独使用HCHO时相似。与能使HCHO失活的氨基脲共同孵育，可防止由HCHO和AIPH组合诱导的这种细胞死亡。氨基脲对细胞内ROS的协同增加和DNA-蛋白质交联的形成也表现出抑制作用。这些结果表明，来自AIPH的自由基诱导了GSH减少，而HCHO导致了DNA-蛋白质交联的形成，最终导致细胞内ROS的协同、渐进增加以及这种组合所带来的细胞死亡。
