Apoptotic effect of ethyl-4-isothiocyanatobutanoate is associated with DNA damage, proteasomal activity and induction of p53 and p21cip1/waf1
Autor: | Ivan Chalupa, Zdena Bartosova, Katarína Horáková, Lubomir Floch, Jana Jakubikova, Juraj Bodo, Jan Sedlak |
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Rok vydání: | 2006 |
Předmět: |
Cyclin-Dependent Kinase Inhibitor p21
Cancer Research Proteasome Endopeptidase Complex DNA damage Leupeptins Poly ADP ribose polymerase Clinical Biochemistry Pharmaceutical Science Apoptosis Cell Cycle Proteins Ataxia Telangiectasia Mutated Proteins Biology Protein Serine-Threonine Kinases Amino Acid Chloromethyl Ketones Cell Line chemistry.chemical_compound Isothiocyanates Caffeine MG132 medicine Humans Fragmentation (cell biology) Mitosis Pharmacology Chromosome Aberrations Membrane Potential Mitochondrial Biochemistry (medical) Cell Cycle Cell Biology Cell cycle Flow Cytometry Molecular biology Cell biology Butyrates chemistry Proteasome inhibitor Tumor Suppressor Protein p53 medicine.drug DNA Damage |
Zdroj: | Apoptosis : an international journal on programmed cell death. 11(8) |
ISSN: | 1360-8185 |
Popis: | The effect of synthetic isothiocyanate ethyl-4-isothiocyanatobutanoate (E-4IB) on survival of mismatch repair-proficient TK6 and -deficient MT1 cell lines as well as the influence of proteasomal inhibitor MG132, caspase inhibitor Z-VAD-fmk, and ATM inhibitor caffeine on E-4IB modulation of cell cycle and apoptosis was evaluated. Flow cytometric analyses of DNA double strand breaks (gamma-H2AX), mitotic fraction (phospho-histone H3), cell cycle modulation, apoptosis induction (sub-G(0) fraction and fluorescein diacetate staining), and dissipation of transmembrane mitochondrial potential (JC-1 staining) were performed. Western blotting was used for the evaluation of ERK activation, expression of p53, p21(cip1/waf1) and GADD45alpha proteins, as well as PARP fragmentation. Analysis of mitotic nuclei was performed for chromosomal aberrations assessment. MT1 cells were more resistant to E-4IB treatment then TK6 cells (IC(50) 8 muM vs. 4 muM). In both cell lines E-4IB treatment induced phosphorylation of H2AX, increase of p53 protein level, phospho-histone H3 staining, and G(2)/M arrest. The sub-G(0) fragmentation was accompanied by PARP degradation, decreased mitochondrial transmembrane potential, and diminished p21(cip1/waf1) protein expression in TK6 cells. Caspase inhibitor Z-VAD-fmk decreased E-4IB induced sub-G(0) fragmentation and extent of apoptosis in TK6 cells, while proteasome inhibitor MG132 increased number of apoptotic cells in both cell lines tested. A number of aberrant metaphases and clastogenic effect of high E-4IB concentration was observed. The synthetic isothiocyanate E-4IB induced DNA strand breaks, increased mitotic fraction and apoptosis potentiated by MG132 inhibitor in both mismatch repair-proficient and -deficient cell lines. |
Databáze: | OpenAIRE |
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