Method for kinetic analysis of drug-induced cell cycle perturbations.
A method is described for quantitative study of the flux of cells through the cell cycle phases in in vitro systems perturbed by chemicals, such as chemotherapeutic agents. The method utilizes cell count and the flow cytometric technique of bromodeoxyuridine (BrdUrd) labeling, according to an optimized strategy. Cells are exposed to BrdUrd during the last minutes of drug treatment and fixed for analysis at 0, 1/3Ts, 2/3Ts, Ts, and Tc + TG1 recovery times, where Ts, TG1, Tc are the mean durations of phases S and G1 and of the whole cycle of control cells. As an example of application of the proposed procedure, a kinetic study of the effect of 1-(2-chloroethyl)-1-nitrosourea (CNU) on the L1210 cell cycle is described. Simple data analysis, requiring only a pocket calculator, showed that cells in phases G1 and G2M at the end of a 1 h treatment with 1 microgram/ml CNU were fully able to leave these phases but were destined to remain blocked in the following G2M phase (G1 for a minority of them). We also found that cells initially in S phase were slightly delayed in completing their S phase and that 50% of them remained temporarily blocked in the subsequent G2M phase, irrespective of their position in the S phase.
Ubezio
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Istituto di Ricerche Farmacologiche Mario Negri, Milano, Italy.
Filippeschi
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Spinelli
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D000818:Animals; D001973:Bromodeoxyuridine; D002452:Cell Count; D002453:Cell Cycle / Q000187:drug effects*; D004247:DNA / Q000032:analysis; D005038:Ethylnitrosourea / Q000031:analogs & derivatives / Q000494:pharmacology; D005434:Flow Cytometry; D007700:Kinetics; D007939:Leukemia L1210 / Q000473:pathology*; D051379:Mice; D013194:Staining and Labeling; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000473:pathology
C017139:1-(2-chloroethyl)-1-nitrosourea; D004247:DNA; D001973:Bromodeoxyuridine; D005038:Ethylnitrosourea