Analysis of apoptosis by DNA end labeling method (TDT) in leukemia cell lines HL-60 and U937

Abstract

Antitumor-drug-induced apoptosis in leukemia cell lines HL-60 and U937 was quantitatively analyzed with TDT (terminal deoxynuleotidyl transferase-mediated nick-end labeling) approach, which allows to label the ends of DNA broken strands in apoptotic cells by biotinlated dUTP which to avidin-FITC. In this way the apoptotic cells show fluorescent when the labeling cells were emitted by UV light microscope or laser-activated flow cell sorting at r=480. In our study, HL-60 and U937 cell lines were cocultured respectively with cis-diaminodichloroplatinum (CDDP), hydroxycamptothecinum (HCT) and vindesini sulfa (VCR) for 18 hours. By calculating percentages of apoptotic cells with TDT method, we were able to show that the two cell lines gave different sensitivity to the drugs. HL-60 showed high sensitivity to CDDP but U937 cells were more sensitive to other two drugs, HCT and VCR. Meanwhile we compared the results of obtained by DNA gel electrophoresis with that by TDT. We found that gel electrophoresis is not sensitive enough to reveal apoptosis since there was no ladder structure, a typical electrophoresis pattern for apoptosis, appeared until the apoptotic cells reached or over 13%. And we report in this paper as first time that three forms of apoptotic cells could be detected under fluorescent microscope, which we called as spot form and crescent form and assembling form in terms of distribution of light spots within cell nuclei. It seemed that the spot form was at an early stage of apoptosis and the crescent form represented a later stage of apoptosis.