Sequential events of apoptosis involving docetaxel, a microtubule-interfering agent: a cytometric study.
BACKGROUND
Despite the great advances in the understanding of programmed cell death, little attention has been paid to the sequence of the events that characterise it. In particular, the course of apoptotic events induced by microtubule-interfering agents such as taxanes is poorly understood. In order to increase such knowledge, we studied a number of independent biochemical and cytological modifications using cytometric methods in a bladder cancer cell line treated with the second generation taxane, docetaxel.


RESULTS
Within a few hours, drug treatment had induced mitochondrial membrane transition, cell shrinkage and a decrease in granularity. Cell cycle was almost completely blocked in G2/M phase within 24 hours. The hypodiploid peak started to become prominent 48 hours after the treatment. At the same time, the appearance of a DNA ladder demonstrated caspase-dependent chromatin fragmentation. Concurrently, specific cell surface modifications took place, involving at first glycoprotein syalilation and later phospholipid asymmetry. DNA fragmentation was subsequently detected by TUNEL assay. Over time, cell membranes became permeable to propidium iodide. A very similar time-course of apoptotic events was found after treatment of a myelomonocytic cell line with the same drug.


CONCLUSIONS
After discussing some characteristics of the methods employed and their limitations, a succession of apoptotic events over time is suggested, in which the collapse of mitochondrial transmembrane potential (Deltapsim) is the earliest sign of apoptosis.
Fabbri
Francesco
F
Department of Medical Oncology, Morgagni-Pierantoni Hospital, Via Forlanini 34, 47100 Forlì, Italy. francesco.fabbri@ausl.fo.it
Carloni
Silvia
S
Brigliadori
Giovanni
G
Zoli
Wainer
W
Lapalombella
Rosa
R
Marini
Marina
M
D000970:Antineoplastic Agents / Q000633:toxicity; D017209:Apoptosis; D002452:Cell Count; D002453:Cell Cycle / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D004249:DNA Damage; D000077143:Docetaxel; D005434:Flow Cytometry; D006801:Humans; D007700:Kinetics; D037102:Lectins / Q000032:analysis; D008564:Membrane Potentials / Q000187:drug effects; D008928:Mitochondria / Q000187:drug effects / Q000502:physiology; D009336:Necrosis; D010718:Phosphatidylserines / Q000032:analysis; D043823:Taxoids / Q000633:toxicity*; D050257:Tubulin Modulators / Q000633:toxicity*
D000970:Antineoplastic Agents; D037102:Lectins; D010718:Phosphatidylserines; D043823:Taxoids; D050257:Tubulin Modulators; D000077143:Docetaxel