硒化壳聚糖逆转K562/ADM细胞耐药与PI-3K/Akt信号通路的关系
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摘要
目的研究硒化壳聚糖对体外培养多药耐药白血病细胞株K562/ADM的耐药逆转作用,探讨其与PI-3K/Akt信号通路的关系。方法 K562/ADM细胞培养于含0.6μg/ml阿霉素的培养液中维持其耐药性,应用MTT法检测硒化壳聚糖对K562/ADM细胞增殖的作用,计算逆转倍数;应用流式细胞法检测细胞凋亡;应用免疫印迹法检测磷酸化Akt(p-Akt)和P糖蛋白(P-gp)表达的改变。结果 K562/ADM细胞对阿霉素(ADM)耐药,硒化壳聚糖可有效抑制K562/ADM细胞增殖,呈一定的剂量和时间效应关系(P<0.05,P<0.01);硒化壳聚糖能够对K562/ADM细胞耐ADM产生一定的逆转作用,明显增强ADM对K562/ADM细胞的诱导凋亡作用(P<0.05,P<0.01),下调p-Akt和P-gp水平(P<0.01)。结论硒化壳聚糖对耐药白血病细胞株K562/ADM可产生增殖抑制和多药耐药逆转作用,其部分逆转机制可能是通过诱导细胞凋亡,抑制细胞P-gp表达,阻断细胞PI-3K/Akt信号通路而实现。
Objective The study aimed to investigate the reversing effect of selenium chitosan on drug resistance of cultured multidrug resistant leukemia cell lines K562/ADM cells and its correlation with the PI-3K/Akt signaling pathway. Methods K562/ADM cells were cultured in medium supplemented with 0.6 μg/ml of adriamycin to maintain their drug resistance. MTT method was adopted to measure the effect of selenium chitosan on the proliferation of K562/ADM cells and to calculate the multiplicity of the reversal. Apoptosis of the cells was assessed by flow cytometry, and changes in the expression of phosphorylated Akt(p-Akt) and phosphorylated glycoprotein(P-gp) were determined by immunoblotting. Results K562/ADM cells were resistant against adriamycin(ADM). Selenium chitosan could effectively inhibit the proliferation of K562/ADM cells in a dose- and time-dependent manner(P< 0.05, P<0.01). Selenium chitosan could reverse the resistance of K562/ADM against ADM, significantly enhance the apoptosis of K562/ADM cells induced by ADM(P<0.05, P<0.01), and downregulate the expression of p-Akt and P-gp(P<0.01). Conclusion Selenium chitosan can inhibit proliferation and reverse multidrug resistance of drug-resistant leukemia cell lines K562/ADM. The reversal of drug resistance may partially involve the induction of cell apoptosis, inhibition of cellular expression of P-gp, and blockage of the PI-3K/Akt signaling pathway.
Objective The study aimed to investigate the reversing effect of selenium chitosan on drug resistance of cultured multidrug resistant leukemia cell lines K562/ADM cells and its correlation with the PI-3K/Akt signaling pathway. Methods K562/ADM cells were cultured in medium supplemented with 0.6 μg/ml of adriamycin to maintain their drug resistance. MTT method was adopted to measure the effect of selenium chitosan on the proliferation of K562/ADM cells and to calculate the multiplicity of the reversal. Apoptosis of the cells was assessed by flow cytometry, and changes in the expression of phosphorylated Akt(p-Akt) and phosphorylated glycoprotein(P-gp) were determined by immunoblotting. Results K562/ADM cells were resistant against adriamycin(ADM). Selenium chitosan could effectively inhibit the proliferation of K562/ADM cells in a dose- and time-dependent manner(P< 0.05, P<0.01). Selenium chitosan could reverse the resistance of K562/ADM against ADM, significantly enhance the apoptosis of K562/ADM cells induced by ADM(P<0.05, P<0.01), and downregulate the expression of p-Akt and P-gp(P<0.01). Conclusion Selenium chitosan can inhibit proliferation and reverse multidrug resistance of drug-resistant leukemia cell lines K562/ADM. The reversal of drug resistance may partially involve the induction of cell apoptosis, inhibition of cellular expression of P-gp, and blockage of the PI-3K/Akt signaling pathway.
引文
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[15]石小燕,蔡晓军,类健翔,等.PI-3K/Akt抑制剂LY294002对卵巢癌细胞A2780/Taxol多药耐药性的逆转作用[J].癌症,2008,27(4):343-347.
[16]Kim D,Dan HC,Park S,et al.AKT/PKB signaling mechanisms in Cancer and chemoresistance[J].Front Biosci,2005,10(10):975-987.
[2]穆玉兰,温泽清,汤春生.微量元素与肿瘤耐药性关系的研究进展[J].中国癌症杂志,2002,12(2):184-186.
[3]崔晶,丁璟,吴轶苹,等.亚硒酸钠对K562/ADR多药耐药的逆转作用及其机制[J].中国实验血液学杂志,2007,15(4):756-761.
[4]魏虎来,张哲文,高丽萍,等.硒酸酯多糖通过下调mdr1/P—gp表达逆转K562/ADM细胞凋亡抑制[J].现代肿瘤医学,2006,14(12):1489-1492.
[5]孙兰萍,张胜义,许晖.硒化壳聚糖的制备及理化性质的研究[J].食品工业科技,2006,27(2):145-146,151.
[6]邓守恒,李芳,李林均,等.硒化壳聚糖对慢性粒细胞白血病K562细胞bcr/abl融合基因表达的影响[J].中国实验方剂学杂志,2011,17(1):106-109.
[7]邓守恒,蔡晓军,佐志刚,等.硒化壳聚糖对NB4细胞增殖的影响及其与PML-RARα融合蛋白激活的Ras信号途径的关系[J].现代预防医学,2011,38(6):1074-1076.
[8]邓守恒,张春香,曾小华,等.硒化壳聚糖对HL60细胞增殖分化的影响[J].江苏医药,2010,36(10):1172-1175,封2.
[9]Comstock CE,Knudsen KE.The complex role of AR signaling after cytotoxic insult:implications for cell-cycle-based chemotherapeutics[J].Cell Cycle,2007,6(11):1307-1313.
[10]Lage H.An overview of Cancer multidrug resistance:a still unsolved problem[J].Cell Mol Life Sci,2008,65(20):3145-3167.
[11]Flens MJ,Zaman GJ,van der Valk P,et al.Tissue distribution of the multidrug resistance protein[J].Am J Pathol.1996,148(4):1237-1247.
[12]Mahadevan D,List AF.Targeting the multidrug resistance-1 transporter in AML:molecular regulation and therapeutic strategies[J].Blood,2004,104(7):1940-1951.
[13]张哲文,魏虎来,苏海翔,等.硒酸酯多糖诱导白血病多药耐药细胞凋亡的作用机制[J].中国临床药理学与治疗学,2005,10(5):505-508.
[14]Kim D,Cheng GZ,Lindsley CW,et al.Targeting the phosphatidylinositol-3 kinase/Akt pathway for the treatment of cancer[J].Curr Opin Investig Drugs,2005,6(12):1250-1258.
[15]石小燕,蔡晓军,类健翔,等.PI-3K/Akt抑制剂LY294002对卵巢癌细胞A2780/Taxol多药耐药性的逆转作用[J].癌症,2008,27(4):343-347.
[16]Kim D,Dan HC,Park S,et al.AKT/PKB signaling mechanisms in Cancer and chemoresistance[J].Front Biosci,2005,10(10):975-987.