J. Nygren et al., INDUCTION OF DMA STRAND BREAKS BY ETHYLENE-OXIDE IN HUMAN-DIPLOID FIBROBLASTS, Environmental and molecular mutagenesis, 24(3), 1994, pp. 161-167
In vitro exposure of normal human diploid fibroblasts blasts (strain V
H-10) to ethylene oxide (EtO) induced DNA strand breaks in the dose ra
nge of 2.5-30 mMh of EtO. Alkaline DNA unwinding (ADU), neutral filter
elution (NFE), pulsed field gel electrophoresis (PFGE), and the comet
assay were used to measure DNA single (SSBs) and double strand breaks
(DSBs). Different induction rates of SSBs and DSBs depending on appli
ed method and also on treatment conditions (cells in monolayer or in s
uspension were used), were found. A dose-dependent increase of DNA str
and breaks was found by the ADU method in the dose range of 2.5-20 mMh
of EtO when treatment wets performed in monolayer and in suspension.
DSBs were detected by NFE only when the cells were treated with EtO in
suspension (doses 10-30 mMh). The highest induction rate of DSBs (abo
ut 4 DSBs per 100 Mbp per 1 mMh of EtO) was detected in suspension wit
h PFGE applied. We have shown that heat-labile sites ore formed by EtO
. Presumably, the different DSB levels detected by PFGE and NFE result
from the conversion of these sites to DSBs during cell lysis at eleva
ted temperature in the PFGE method. The results of the comet assay con
firmed that apoptotic processes are not involved in the formation of D
SBs in our experimental conditions (less than 1% of apoptotic cells we
re observed at all doses studied). Possible mechanisms for the inducti
on of DNA strand breaks by EtO-treatment are discussed. The capacity t
o repair DSBs in EtO-exposed (5-7.5 mMh) cells was studied, and it was
found that a considerable part of the damage (about 50%) could be rep
aired during 18 hr of incubation. (C) 1994 Wiley-Liss, Inc.