Effects of tea polyphenols combined with epirubicin on T24 bladder cancer cell line and its mechanism

Abstract:

Objective：To investigate the effect of tea polyphenols（TP） on epirubicin（EPI）-induced cell proliferation and cell apopto－sis of human bladder cancer cells T24 in vitro and its mechanism. Methods：MTT assay was applied to investigate the effects of differ－ent levels of tea polyphenols or epirubicin on the T24 cells. The experiment was divided into 4 groups：control group（PBS），TP group（88.8 μmol/L），EPI group（4.6 μmol/L），and TP（88.8 μmol/L） plus EPI（4.6 μmol/L） group. The proliferation inhibition rate was as－sayed by MTT；Annexin V-FITC/PI double staining flow cytometry was used to analyze apoptosis. The ultra-structural cellular changes were observed by transmission electron microscope（TEM）；The formation of MAP1LC3-Ⅱ puncta was examined after transfection of a GFP-LC3 plasmid. The levels of LC-3-Ⅱ，P62，CASP3，PARP proteins were detected by Western blot，respectively. Results：Treatment with TP（88.8 μmol/L） and EPI（4.6 μmol/L） leaded to a significant increase of inhibiting rate（P=0.000） and a dramatic improvement in apoptotic changes compared to single EPI or TP group（P=0.000）. Autophagy was observed obviously in bladder cancer cells T24 treated with EPI after 8 h. TEM examination revealed the appear－ance of autophagosome with double-membrane structure. Im－munofluorescence showed that EPI strongly augmented the number of cells with increased GFP-LC3 puncta and GFP-LC3 puncta formation within the cell. Western blot showed the ex－pression of LC-3-Ⅱ was increased in T24 cells after treated with EPI，while the expression of P62 was decreased. TP com－bined with EPI and single groups both induced the expression of active apoptosis proteins，and the effect was more obvious in TP plus EPI group. Conclusion：TP combined with EPI can synergisti－cally inhibit the proliferation and promote cell apoptosis，which may be associated with the reduction of EPI-induced autophagy by TP.