PI3K/Akt/NF-κB通路调控ABCB1/P-gp介导的人结肠癌细胞多药耐药的研究  

PI3K/Akt/NF-κB regulate ABCB1/P-glycoprotein–mediated multidrug resistance in colon carcinoma cells

作　　者：隋华[1] 付晓伶[1] 潘树芳[2] 石晓兰[2] 靳宝辉[1] 朱惠蓉[1] 任建琳[3] 李琦[1] 

机构地区：[1]上海中医药大学附属曙光医院肿瘤科,上海201203 [2]上海中医药大学附属普陀医院肿瘤科,上海200062 [3]上海市中医医院肿瘤科,上海200071

出　　处：《中国癌症杂志》2014年第2期106-111,共6页China Oncology

基　　金：国家自然科学基金(No:81373862;81202812);上海市科委资助项目(No:13140902500);上海市教委资助项目(No:2011JW57;12ZZ118);上海市卫生局科研项目(No:20114Y013;2010019);上海市中医药事业发展三年行动计划(No:ZYSNXD-CC-YJXYY-JS20)

摘　　要：背景与目的:肿瘤的多药耐药(multidrug resistance,MDR)基因的表达是目前化疗失败的主要原因,磷脂酰肌醇-3-激酶(phosphoinositide 3-kinases,PI3K)信号通路参与肿瘤多药耐药的发生,但PI3K信号通路与多药耐药的机制尚不明确。本研究旨在探讨PI3K/Akt信号通路及其下游靶点对ATP结合蛋白亚家族1抗体(ATP-binding cassette sub-family B member 1,ABCB1)基因编码的P-糖蛋白(P-glycoprotein,P-gp)介导的人结肠癌耐奥沙利铂细胞株HCT-116/L-OHP细胞多药耐药性的调控作用。方法:将PI3K特异性抑制剂LY294002(20μmol/L)处理人结肠癌HCT-116/L-OHP细胞2 h后,用细胞计数试剂盒-8(cell counting kit-8,CCK-8)检测细胞对奥沙利铂的敏感性;蛋白质印迹法(Western blot)检测相关耐药蛋白P-gp、肺耐药蛋白(lung resistance-related protein,LRP)、多药耐药相关蛋白-2(multidrug resistance-related-2,MRP-2)以及PI3K/Akt信号通路下游蛋白Akt、p-Akt、IκB、p-IκB的表达变化;CHIP—PCR法检测核转录因子κB(NF-κB)对ABCB1基因启动子的影响。结果:阻断PI3K/Akt信号通路激活后,奥沙利铂对HCT-116/L-OHP细胞的半数抑制浓度(IC50)由(157.48±16.73)μg/mL降至(53.68±3.18)μg/mL,逆转指数为2.93(P<0.01)。HCT-116/L-OHP细胞的p-Akt、p-IκB和P-gp的表达水平明显下降(P<0.01),Akt、IκB、LRP和MRP-2表达变化不明显。NF-κB能够与ABCB1基因启动子区域结合。结论:阻断PI3K/AKT信号通路可增强人结肠癌HCT-116/L-OHP耐药细胞的药物敏感性,抑制p-Akt和p-IκB的磷酸化表达水平,逆转P-gp介导的肠癌多药耐药。Background and purpose： Multidrug resistance （MDR） is the dominating obstacle to the chemotherapy. There is strong evidence that the phosphoinositide 3-kinases （PI3Ks） signaling pathway is involved in MDR phenotype, however, the mechanism of MDR occurrence is still unknown. This study tended to investigate the regulating effect of PI3K/Akt signaling pathway and its downstream target genes in P-glycoprotein （P-gp） （ABCB1 gene encoding）-mediated MDR in human colon carcinoma HCT-116/L-OHP cells. Methods： Pretreatment with PI3Kselective inhibitor LY294002 （20 μmaol/L） for 2 h, the sensitivity of L-OHP was evaluated by the CCK-8 （cell counting kit-8） assay in HCT-116/L-OHP cells, and the expressions of P-gp, LRP, MRP-2, Akt, p-Akt, IκB and p-InB were evaluated by Western blot. The activity of ABCB 1 promoter was evaluated by chromatin immunoprecipitation analysis （CHIP）. Results： After inhibiting the activity of PI3K/Akt signaling pathway, the IC50 value of L-OHP decreased from（157.48±16.73） μg/mL to （53.68±3.18） μg/mL in HCT- 116/L-OHP cells, and the reversal index was 2.93 （P〈0.01）. The expressions of P-gp, p-Akt and p-IvJ3 were down-regulation compared with the concrol group （P〈0.01）, but the expressions of LRP, MRP-2, Akt and IKB didn＇t change significantly. CHIP result has confirmed that NF-nB protein could bind to the region ofABCB 1 gene promoter in HCT 116/L-OHP cells. Conclusion： Blocking of PI3K/Akt/NF-kB signal pathway could increase the drug sensitivity to MDR cells, inhibit the phosphorylation of p-Akt and p-IκB, and reversing ABCB 1/P-glycoprotein-mediated multidrug resistance in colon carcinoma cells.

关 键 词：结肠癌 多药耐药 P-糖蛋白 磷脂酰肌醇-3-激酶 核转录因子ΚB 

分 类 号：R735-35[医药卫生—肿瘤]