Phenethyl isothiocyanate-induced cytoskeletal changes and cell death in lung cancer cells |
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Authors: | Andrzej Pawlik Mariusz Andrzej Szczepanski Anna Klimaszewska Lidia Gackowska Agnieszka Zuryn Alina Grzanka |
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Institution: | 1. Department of Histology and Embryology, Collegium Medicum in Bydgoszcz, Nicolaus Copernicus University, Bydgoszcz, Poland;2. Department of Immunology, Collegium Medicum in Bydgoszcz, Nicolaus Copernicus University, Bydgoszcz, Poland |
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Abstract: | Isothiocyanates are known for their anticarcinogenic and antitumor potential, however, the exact mechanism of their action has not been fully elucidated. The present study was designed to investigate and compare the effects of phenethyl isothiocyanate on cell morphology, the cytoskeleton and induction of cell death in human non-small cell lung cancer cell lines A549 and H1299 differing in p53 status. Cell viability tests (MTT assay, xCELLigence system) showed that PEITC exhibits lower cytotoxicity to A549 cells containing wild-type p53. The observed growth-inhibitory effect of PEITC was dose-dependent, but time-dependence was observed only at higher concentrations. The results of flow-cytometric and fluorescence-microscopic analyses indicate that PEITC induced disassembly of actin stress fibers and degradation of tubulin which, most likely, contributed to the induction of cell death. Although, 24-h incubation caused G2/M cell cycle arrest, the fraction of G2/M cells decreased in a dose- and time-dependent manner in favor of cells with sub-G1 DNA content. Further experiments (Annexin V staining, electron microscopic observations) confirmed that the apoptosis-inducing potency of PEITC is probably the main factor responsible for cell growth inhibition. However, PEITC treatment also resulted in the appearance of an increased proportion of H1299 cells exhibiting morphological features of mitotic catastrophe. |
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Keywords: | BSA bovine serum albumin CI cell index DAPI 4&prime 6-diamidino-2-phenyindole DMEM Dulbecco&rsquo s Modified Eagle Medium DTSP 3 3&prime -dithiodipropionic acid di(N-hydroxysuccinimide ester) FITC fluorescein isothiocyanate GSH glutathione GST glutathione-S-transferase ITC isothiocyanate MTOC microtubule-organizing centers MTSB microtubule stabilizing buffer MTT thiazolyl blue tetrazolium bromide NNK nicotine-derived nitrosamine ketone NSCLC non-small cell lung cancer PEITC phenethyl isothiocyanate PI propidium iodide ROS reactive oxygen species RTCA real-time cell analyzer TBS tris-buffered saline |
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