Mitochondrial function, apoptosis and cell cycle delay in the WEHI-3B leukaemia cell line and its variant Ciprofloxacin-resistant WEHI-3B/CPX.
OBJECTIVE
The susceptibility of two cell lines, WEHI-3B myelomonocytic leukaemia and its variant Ciprofloxacin-resistant WEHI-3B/CPX to undergo apoptosis induced by Ciprofloxacin was studied and compared.


METHODS
Apoptosis was checked by measuring the DNA fragmentation and determining the ratio of apoptotic/necrotic cells. The relationship between the induction of apoptosis and G(1), S or G(2) block in the cell cycle has also been investigated and cytogenetical evaluation of chromosomal aberrations in both cell lines has been carried out. The regulation of expression of Bax and Bcl-2 was also checked by western blotting after Ciprofloxacin treatment.


RESULTS
We observed that the resistance of the subline was caused by a small percentage of cells that underwent apoptosis during continuous exposure to Ciprofloxacin in comparison with the parental cell line, whereas the percentage of necrotic cells remained unchanged. The WEHI-3B cells showed a G(2) block and a higher degree of cytogenetic damage after drug exposure. The two cell lines expressed the same level of Bax and Bcl-2 following stimulation by Ciprofloxacin. Only in the resistant subclone, the ratio Bcl-2/Bax reversed in the anti-apoptotic gene expression.


CONCLUSIONS
The resistance to ciprofloxacin observed is not related to mitochondrial function and although Bcl-2/Bax ratio behaviour does not fully explain the resistance of the WEHI3B/CPX subclone it is consistent with phenotypic character of resistance to CPX. The toxic effect on sensitive cells could be mediated by the cell cycle arrest whereas in the resistant clone, the prolonged G(2) phase could play a key role to favour cell cycle progression and proliferation.
Pessina
A
A
Department of Public Health, Microbiology, Virology, University of Milan, Milan, Italy. augusto.pessina@unimi.it
Bonomi
A
A
Casati
S
S
Collotta
A
A
Croera
C
C
Marafante
E
E
Palitti
F
F
Gribaldo
L
L
D000818:Animals; D000970:Antineoplastic Agents / Q000633:toxicity*; D017209:Apoptosis; D002453:Cell Cycle / Q000187:drug effects*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002939:Ciprofloxacin / Q000633:toxicity*; D019008:Drug Resistance, Neoplasm; D007951:Leukemia, Myeloid / Q000378:metabolism / Q000473:pathology; D051379:Mice; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D051028:bcl-2-Associated X Protein / Q000378:metabolism
D000970:Antineoplastic Agents; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D002939:Ciprofloxacin