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Objective: To monitor the cisplatin-induced autophagy and investigate the function of autophagy in bladder cancer cells., Methods: The transmission electron microscope was used to detect autophagic vacuoles and the fluorescence microscope to detect GFP-LC3. The expressions of proteins, such as LC3, PARP, mTOR, P70S6K were analyzed by immunoblotting. Cell viability was analyzed by MTS assay, in which rapamycin was used to inhibit mTOR phosphorylation and enhance autophagy. LC3 expression was knocked down by RNA interference., Results: In bladder cancer cell T24, autophagic vacuoles were observed by the transmission electron microscope and GFP-LC3 aggregation was viewed by the fluorescence microscope after cisplatin treatment. The LC3-II accumulation was enhanced by cispaltin treatment. Particularly at the concentrations of 50, and 100 μmol/L for 48 h , the gray value of LC3-II/Actin(%) increased 30 and 44, respectively. Cisplatin treatment inhibited the phosphorylation of mTOR/P70S6K, which was most significant at the concentration of 100 μmol/L for 48 h. Cisplatin also induced cell viability loss, which was 12% and 45% at the concentrations of 50, and 100 μmol/L for 24 h. This effect could be enhanced by rapamycin (F=74.890,P<0.01). Furthermore, knocking down LC3 by RNA interference reduced PARP cleavage., Conclusion: Cisplatin could induce autophagy in bladder cancer cell T24, which promoted cisplatin-induced apoptosis.

Panax Notoginseng Saponins (PNS) have been well known to have anti-tumor activity and enhance cytotoxicity of some cancer chemotherapy agents, but the mechanisms underlying these effects are still unknown. This study investigates the effect of PNS on cytotoxicity of cisplatin and the relationship between this effect and the modulation of gap junctions (GJ) function by PNS in a transfected cell line. The cytotoxicity of cisplatin (0.25-1 µg/mL) was increased in the presence of GJ. Inhibition of gap junction by either GJ blocker or interception of Connexin (Cx) expression decreased the cytotoxicity of cisplatin. Increasing GJ function enhanced cytotoxicity of cisplatin, only in the cells with functional GJ. PNS (50-200 µg/mL) significantly enhanced cisplatin cytotoxicity, but this effect required functional gap junctions between the cells. Exposure of the cells to PNS (50-200 µg/mL) for 4 h leads to a significant enhance in dye coupling of GJ in a dose-dependent manner. These results suggest that PNS increases the cytotoxicity of cisplatin through enhancement of GJ activity.