Cinobufacini induces the apoptosis of U937 cells and its mechanism / 肿瘤

ABSTRACT

Objective: To investigate the effect of cinobufacini on apoptosis of U937 cells and its possible mechanism. Methods: Cell viability was measured by MTT assay. The morphological changes were observed by Wrightś staining and immunofluorescence staining. DNA fragmentation was visualized by agarose gel electrophoresis. Apoptotic rate was evaluated by teminal deoxynucleotidyl transferase (TdT) labeling in situ. Expression of bel-2 protein was analyzed by flow cytometry. The levels of Fas and Fas-1 mRNA were measured by semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR). Results: Compared with the control group, treatment with cinobufacini at 0.225 to 1.8 μg/ mL for 1-3 d significantly inhibited growth of U937 cells in a time and dose dependent manner. The IC50 value was 1.36 μg/ mL at 24 h. The typical morphological changes of apoptosis and typical apoptotic DNA ladder were observed after incubation with cinobufacini at 0.9 μg/mL for 1 d. The apoptotic rate evaluated by TdT labeling in situ were 4.8%, 13.57%, and 24.33% after exposure to cinobufacini at 0.225, 0.45, and 0.9 μg/ mL for 1 d, respectively. The expression levels of bcl-2 protein and Fas-1 mRNA significantly decreased and the expression of Fas mRNA significantly increased in apoptotic cells. Conclusions: Cinobufacini inhibits growth and inducs apoptosis of U937 cells via inhibition of bcl-2 and Fas-1 genes and activation of Fas gene.