Demonstration of an S phase population of cells without DNA synthesis generated by cisplatin and pentoxyifylline

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Abstract
Dual parameter flow cytometry measuring DNA and BrdUrd uptake in "stripped" nuclei was used as a sample preparation procedure. The utility of this method is demonstrated in a human ovarian cell line (BG-1) by two parameter flow cytometric analysis of DNA and bromodeoxyuridine (BrdUrd) incorporated into the DNA. BG-1 cells, treated with cisplatin followed by constant exposure to pentoxifylline, produce G0/G1, S phase, and G2/M perturbations. The G0/G1 population is greatly diminished. Large numbers of cells are distributed in S phase in which a large portion is incapable of DNA synthesis. The G2/M perturbations show a continually increasing block over a time course of 168 h. The identification of two types of S phase cells (DNA synthesizing and DNA nonsynthesizing) is clearly shown with the use of BrdUrd incorporation as a second parameter in addition to flow cytometric analysis of DNA. This increase in the non-DNA-synthesizing S phase population corresponds to an enhancement of cytotoxicity when cells are treated with the combination of cisplatin and pentoxifylline. It is felt that this technique provides a useful method to investigate the dynamics of the cell cycle perturbations affected by modulators of chemotherapeutic agents.

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