Effects Of Ursolic Acid On Human Erythroleukemia K562 Cells

Ursolic acid (UA) is a kind of triterpenoid compound which exists widely in natural plants. It has multiple biological functions. Here we studied effects of UA on K562 cells and primary mechanisms. The concentration of UA used in cell culture system was from 0 umol/L to 50 umol/L and 24, 48, 60 hour were selected as the termination point of cell culture. Inhibition of cell proliferation was determined by MTT assay and FCM. Cytotoxic effect was analysed by single cell electrophoresis assay. Nuclear morphological changes were observed by Hoechst33258 fluorescent staining. Expression of Bcl-2 and Bax were assessed by immunochemical staining. Activation of Caspase-3 and expression of p210 phosphotyrosine were detected by Western blotting.MTT assay showed UA had a moderate anti-proliferation effect on K562 cells at the concentration above 30 umol/L in time-and-dose-dependant manner. After treated with 40 umol/L of UA, K562 cells showed S phase decline and G2/M phase arrest: Cells at G2/M phase were 11.0%, 40.5% and 48.1% in control group, 24-hour-treated group and 48-hour-treated group, respectively. single cell electrophoresis assay indicated UA-treated cells presented clear leaked DNA tails. After treatment with UA, nuclei shrunk and the chromosomes were condensed and decomposed into fragments, which showed intensive fluorescence. 40 umol/L of UA caused up-regulation of Bax in K562 cells. Compared with the control group, inactive form of Caspase-3, which was a 32KDa protein, decreased while 17KDa active cleaved fragments increased in UA-treated groups, p210 Phosphotyrosine as well as other phosphotyrosines also decreased, as detected in treated groups by Western blotting.UA has moderate anti-proliferation and cytotoxicity effects on K562 cells in time-and-dose-dependent manner. It is very possible that UA mainly acts on K562 cells at G2/M phase by causing DNA injury to suppress cell proliferation. It can up-regulate Baxexpression so as to promote sensitivity for apoptosis in K562 cells. In addition, UA lead to activation of Caspase-3 in a time-dependent manner, suggesting its ability of inducing apoptosis is one mechanism of its effects. Down-regulation of the expression of the 210KDa phosphotyrosine by UA may be a cue that PTK activity of BCR-ABL infusion protein, which is regarded as a characteristic of K562 cell line, is inhibited. Then K562 proliferation and survival were interrupted.