Yi-Ping Huang1, Yi-Wen Jiang2, Hsin-Yu Chen2, Yung-Ting Hsiao2, Shu-Fen Peng2, Yu-Cheng Chou3,4, Jiun-Long Yang2, Te-Chun Hsia5,6, Jing-Gung Chung7,8. 1. Department of Physiology, College of Medicine, China Medical University, Taichung, Taiwan, R.O.C. 2. Department of Biological Science and Technology, China Medical University, Taichung, Taiwan, R.O.C. 3. Division of Neurosurgical Oncology, Neurological Institute, Taichung Veterans General Hospital, Taichung, Taiwan, R.O.C. 4. School of Medicine, National Defense Medical Center, Taipei, Taiwan, R.O.C. 5. Department of Respiratory Therapy, China Medical University, Taichung, Taiwan, R.O.C. jgchung@mail.cmu.edu.tw hsiatc@mail.cmu.edu.tw. 6. Department of Internal Medicine, China Medical University Hospital, Taichung, Taiwan, R.O.C. 7. Department of Biological Science and Technology, China Medical University, Taichung, Taiwan, R.O.C. jgchung@mail.cmu.edu.tw hsiatc@mail.cmu.edu.tw. 8. Department of Biotechnology, Asia University, Taichung, Taiwan, R.O.C.
Abstract
BACKGROUND/AIM: Gefitinib is used to treat patients with lung cancer, but in some patients, the disease becomes gefitinib-resistant. Benzyl isothiocyanate (BITC), found in cruciferous vegetables, has shown anticancer activity in many human cancer cell lines. However, the effects of BITC on gefitinib-resistant NCI-H460 lung cancer cells in vitro have not been investigated. MATERIALS AND METHODS: The effects of BITC on gefitinib-resistant NCI-H460 lung cancer cells were investigated in vitro. Flow cytometric assay was used for determining the total viable cell number, apoptotic cell death, the production of reactive oxygen species (ROS) and Ca2+, mitochondriaI membrane potential (Ψm) and caspase-3, -8 and -9 activities. Furthermore, 4', 6-diamidino-2-phenylindole staining was used to examine chromatin condensation in NCI-H460 and NCI-H460/G cells. RESULTS: BITC reduced total viable cell number via the induction of apoptotic cell death, that was also confirmed by annexin V/propidium iodide double staining assay. BITC increased ROS and Ca2+ production, reduced Ψm and increased caspase-3, -8 and -9 activities in both NCI-H460 and NCI-H460/G cells. Western blotting assay also showed that BITC increased expression of cleaved caspase-3 and -9, cytochrome c, BCL2-associated X protein, endonuclease G, poly (ADP-ribose) polymerase, growth arrest and DNA-damage protein 153, caspase-7 and activating transcription factor 6 alpha, but reduced apoptosis-inducing factor and caspase-9, BH3-interacting domain death agonist, calpain 1, glucose-regulated protein 78 and inositol requiring enzyme 1 alpha in NCI-H460/G cells. CONCLUSION: BITC-induced apoptotic cell death appears to occur via caspase- and mitochondria-dependent pathways in both cell lines. Copyright
BACKGROUND/AIM: Gefitinib is used to treat patients with lung cancer, but in some patients, the disease becomes gefitinib-resistant. Benzyl isothiocyanate (BITC), found in cruciferous vegetables, has shown anticancer activity in many humancancer cell lines. However, the effects of BITC on gefitinib-resistant NCI-H460 lung cancer cells in vitro have not been investigated. MATERIALS AND METHODS: The effects of BITC on gefitinib-resistant NCI-H460 lung cancer cells were investigated in vitro. Flow cytometric assay was used for determining the total viable cell number, apoptotic cell death, the production of reactive oxygen species (ROS) and Ca2+, mitochondriaI membrane potential (Ψm) and caspase-3, -8 and -9 activities. Furthermore, 4', 6-diamidino-2-phenylindole staining was used to examine chromatin condensation in NCI-H460 and NCI-H460/G cells. RESULTS:BITC reduced total viable cell number via the induction of apoptotic cell death, that was also confirmed by annexin V/propidium iodide double staining assay. BITC increased ROS and Ca2+ production, reduced Ψm and increased caspase-3, -8 and -9 activities in both NCI-H460 and NCI-H460/G cells. Western blotting assay also showed that BITC increased expression of cleaved caspase-3 and -9, cytochrome c, BCL2-associated X protein, endonuclease G, poly (ADP-ribose) polymerase, growth arrest and DNA-damage protein 153, caspase-7 and activating transcription factor 6 alpha, but reduced apoptosis-inducing factor and caspase-9, BH3-interacting domain death agonist, calpain 1, glucose-regulated protein 78 and inositol requiring enzyme 1 alpha in NCI-H460/G cells. CONCLUSION:BITC-induced apoptotic cell death appears to occur via caspase- and mitochondria-dependent pathways in both cell lines. Copyright