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摘要:ClC-3 is a member of the voltage-gated chloride channel superfamily.It has been suggested by us and others to be closely related to cell proliferation,migration,apoptosis and cell proliferation,cell cycle progression,and migration in cancer cells.However,less studies on the relationship between ClC-3 and tumor drug resistance was reported.The aim of this study was to explore the different expression and function of ClC-3 chloride channels between parental tumor cells and drug resistant tumor cells and furtherly replenish the effect of ClC-3 chloride channel in drug resistance.Methods: In this study,the parental cell line(MCF-7,sensitive to Adriamycin) and the Adriamycin-resistant cell line(MCF-7/Adr) were used.The real-time PCR,immunofluorescence,and immunoblotting(Western-blotting) techniques were used to detect the expression of ClC-3 channels in m RNA and protein levels.The whole-cell patch-clamp technique was used to record chloride currents.Up-regulation of the expression of chloride channel proteins was achieved by transfection of the plasmid containing the ClC-3 gene.The si RNA interference technique was used to down-regulate the expression of chloride channel proteins.Results: 1.Expression of ClC-3 channel proteins were more superior in parental MCF-7 cells than in drug resistant MCF-7/Adr cells.The result showed that the fluorescence intensity of ClC-3 channels(973.1 ± 28.3) in MCF-7 cells was significantly higher than that(411.5 ± 14.3) in MCF-7/Adr cells(n = 20,P <0.01).Western-blotting results showed that the expression ratio of ClC-3/β-actin(2.1±0.2) in MCF-7 cells was significantly higher than that(0.7±0.2) in MCF-7/Adr cells(P<0.05),consistent with the changes of m RNA level between the two cells.(P<0.01).2.Chloride currents are inferior in MCF-7/Adr cells.The result of whole cell patch-clamp recordings showed that under isotonic solution,only a small amount of chloride channels in the two kinds of cells were open,but the density of the basal chloride current at +80 m V in MCF-7 cells(5.5 ± 1.1 pA/pF) was significantly greater than that(3.0 ± 0.4 pA/pF) in MCF-7/Adr cells.3.The Adriamcyin-activated chloride current was stronger in MCF-7 cells than in MCF-7/Adr cells.The amplitude of chloride currents was increased by Adriamycin treatment in both cells,but such an increment was more obvious in MCF-7 cells than in MCF-7/Adr cells.The current density was increased from 5.5 ± 1.1 pA/pF to 61.8 ± 10.4 pA/pF at + 80 m V,and from-4.3 ± 1.1 pA/pF to-41.4 ± 8.6 pA/pF at-80 m V(P<0.01,n=6) in MCF-7 cells.In MCF-7/Adr cells,the current density was increased from 3.0 ± 0.4 pA/pF to 29.1 ± 5.2 pA/pF at+80 mV,and from-2.9 ± 0.4 pA/pF to-25.2 ± 4.6 pA/pF at-80 mV(n = 6,P<0.05).The chloride currents induced by Adriamycin in both cells were blocked by the chloride channel blocker NPPB,suggesting Adriamycin-sensitive chloride channels are expressed in both cells,especially in MCF-7 cells.4.Down regulation of ClC-3 expression decreased the Adriamycin
会议名称:

2016年国际生理学学术大会

会议时间:

2016-09-25

会议地点:

中国北京

  • 专辑:

    医药卫生科技

  • 专题:

    肿瘤学

  • 分类号:

    R73-3

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