Involvement of hydrogen peroxide in mistletoe lectin-II-induced apoptosis of myeloleukemic U937 cells Myung-Sunny Kim a,e , Jienny Lee b , Kang-Min Lee c , Sei-Hoon Yang a , Sujinna Choi d , Sang-Young Chung d , Tae-Young Kim a , Won-Hun Jeong f , Raekil Park a,b, * a Institute of Medical Science, School of Medicine, Korea b Professional graduate school of Oriental Medicine, Wonkwang University, Iksan 570-749, Korea c Department of Molecular Biology, Chonbuk National University, Jeonju Chonbuk 561-756, Korea d Department of Surgery, Chonnam National University Medical School, Kwangju 501-758, Korea e Korea Food Research Institute, Kyongi-do 463-746, Korea f Wonkwang Health Science College, Iksan 570-750, Korea Received 13 August 2002; accepted 9 October 2002 Abstract Mistletoe lectin-II, a major component of Korean mistletoe (Viscum album var. coloratum) induces apoptotic death in cancer cells. In this study, we demonstrated that lectin-II induced the generation of pro- oxidants and thus resulted in the apoptotic death of human myeloleukemic U937 cells. We observed that lectin-II-induced apoptotic death was inhibited by antioxidants including reduced glutathione (GSH), N- acetylcysteine (NAC), ebselen, mnTBP, catalase and pyrrolidine dithiocarbamate (PDTC). GSH and NAC also abolished the apoptotic DNA ladder pattern fragmentation of U937 cells after lectin-II stimulation. Obviously, lectin-II treatment of cells resulted in a remarkable generation of intracellular hydrogen peroxide (H 2 O 2 ) as an early event, which was monitored fluorimetrically using scopoletin-horse radish peroxidase (HRP) assay and peroxide-sensitive fluorescent probe, DCF-DA. In addition, antioxidants inhibited the activation of c-Jun N- terminal kinase (JNK)/stress-activated protein kinase (SAPK) as well as cytosolic release of cytochrome c by mistletoe lectin-II. Moreover, lectin-II-induced activation of caspase-9 and 3-like protease and cleavage of poly(ADP-ribose) polymerase (PARP) were inhibited by pretreatment of cells with thiol antioxidants, GSH and NAC. 0024-3205/03/$ - see front matter D 2003 Elsevier Inc. All rights reserved. doi:10.1016/S0024-3205(03)00418-1 * Corresponding author. Department of Microbiology, Wonkwang University School of Medicine, 344-2 Shinyong-dong, Iksan, Chonbuk 570-749, South Korea. Tel.: +82-63-850-6777; fax: +82-63-852-0220. E-mail address: rkpark@wonkwang.ac.kr (R. Jeong). www.elsevier.com/locate/lifescie Life Sciences 73 (2003) 1231 – 1243