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Application of Laser Micro-irradiation for Examination of Single and Double Strand Break Repair in Mammalian Cells
Published on: September 5, 2017
DNA double-strand breaks and apoptosis after in vitro exposure to toluene diisocyanate
B Marczynski1, A B Czuppon, G H Schreiber
1Research Institute of Occupational Medicine at the Ruhr-University of Bochum, Gilsingstrasse 14, 44789 Bochum, Germany.
Abstract:
The immediate effect of toluene diisocyanate (TDI) on genomic DNA of sheep and rabbit white blood cells has been investigated. Following the treatment of sheep blood by TDI, the isolated DNA was analysed by anion exchange chromatography (FPLC) and melting behaviour before and after denaturation. The results of FPLC indicated that TDI induces DNA double-strand breaks in sheep white blood cells. Following denaturation of TDI-treated DNA, the results demonstrated that at least one DNA fragment, representing about 10-20% of total DNA, is cross-linked by TDI or TDI metabolite. About 39% more of double-stranded structures are reconstituted in DNA extracted from sheep white blood cells 3 hr after incubation with TDI. In addition, chromosomal DNA from rabbit blood was investigated with the 'agarose-plugs' method by pulsed-field gel electrophoresis 2 hr after incubation with TDI, and the agarose gels were quantitatively evaluated with a laser densitometer. Further studies with prednisone were conducted to correlate these results with the phenomenon of apoptosis. The in vitro results show that TDI can induce degradation of mitochondrial DNA and large DNA fragments, which result from apoptosis, to small DNA fragments.
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