A Study On The Uptake Of Adriamycin Promoted By Nature Borneol Via Chloride Channels In Nasopharyngeal Carcinoma Cells

Objectives:Borneol, a traditional Chinese medicine, is usually used as an ancillary drug to improve the absorption of medicine, but the underlying mechanism is unclear. It was found by us previously that the activation of chloride channels in the cell membrane promoted the cellular uptake of anticancer drugs. This action could be inhibited by down-regulation of the expression of chloride channel proteins and by chloride channel blockers. The aims of this study were to investigate the roles of chloride channels in transmembrane transport of adriamycin promoted by borneol and to explore the molecular identity of the chloride channels involved in the adriamycin transport in poorly-differentiated nasopharyngeal carcinoma cells(CNE-2Z).Methods:The natural borneol was used in this study. The fluorescence imaging technique was used to detect the transmembrane transport of adriamycin promoted by borneol. The patch clamp technique was used to record the whole-cell chloride currents activated by the borneol and to analyze the characteristics and volume sensitivity of the currents. Cell volume changes were detected using the time-lapse imaging technique and was analyzed by the imaging software. The intracellular drug concentration and its relationship with chloride channels were measured and analyzed using the microplate assay and flow cytometry. The expression of Cl C-3 chloride channel proteins was knocked using the si RNA technique, and was detected by the Western blotting. The fluorescence microscopy and flow cytometry were used to detect the effects of si RNA on the intracellular concentration of adriamycin. The MTT assay was used to evaluate the cytotoxicity of adriamycin in the presence and absence of borneol.Results:1) The uptake of adriamycin was significantly increased by about 29 % by borneol in nasopharyngeal carcinoma CNE-2Z cells. 2) Borneol could rapidly activate chloride channels and induce a chloride current in CNE-2Z cells. The current could be inhibited by the chloride channel blockers diisothiocyanato-stilbene-2,29-disulph-onicacid(DIDS) and tamoxifen. The current was also inhibited by the extracellular perfusion of 47 % hypertonic solution, indicating that the borneol-induced chloride current is volume sensitive. The ion selectivity was I-> Br- > Cl- > Gluconate-, demonstrated by the ion replacement experiment. 3) Extracellular perfusion of borneol solution could decrease the volume of CNE-2Z cells by about 9% in 30 min. This action could be inhibited by the chloride channel blocker DIDS. 4) The facilitative effect of borneol on the uptake of adriamycin was inhibited by the chloride channel blocker DIDS in CNE-2Z cells. 5)The expression of Cl C-3 chloride channels was decreased by about 57 % by the Cl C-3 si RNA. The down-regulation of Cl C-3 chloride channels induced by the Cl C-3 si RNA could inhibit the facilitative effect of borneol on the uptake of adriamycin in CNE-2Z cells. 6)There was no obvious cytotoxic effect of borneol on CNE-2Z cells, but borneol could decreased the 50 % inhibiting concentration(IC50) of adriamycin by about 50 % in its inhibitory action on CNE-2Z cell proliferation.Conclusions:1) Borneol can activate chloride channels and promote the uptake of adriamycin in poorly-differentiated nasopharyngeal carcinoma cells. 2) Chloride channels are one of the targets of borneol in promoting adriamycin transport across the cell membrane. 3) Cl C-3 chloride channels play important roles in the promotion of transmembrane transport of adriamycin by borneol. 4) The antineoplastic effect of adriamycin can be strengthened by borneol.