目的 :研究阻断磷脂酰肌醇3-激酶(phosphatidylinositol 3-kinase,PI3K)/丝氨酸-苏氨酸激酶(seride-threonine protein kinase,又称Akt)信号通路对逆转耐羟基喜树碱(hydroxycamptothecin,HCPT)人结肠癌SW1116/HCPT细胞多药耐药性的影响。方法:采用蛋白质印迹法检测亲本SW1116细胞和耐HCPT细胞株SW1116/HCPT中Akt及磷酸化Akt(phospho-Akt,p-Akt)蛋白的表达水平。应用针对PI3K/Akt信号通路的小分子抑制剂LY294002或siR NA干扰的方法分别抑制或沉默Akt蛋白的表达;MTT法检测抑制或沉默Akt蛋白表达后,HCPT对SW1116/HCPT细胞生存率的影响;实时荧光定量PCR及蛋白质印迹法检测对耐药转运体三磷酸腺苷结合盒转运体G(2ATP-binding cassette transporter G2,ABCG2)mR NA及蛋白表达的影响;罗丹明123(rhodamine123,Rh123)法检测对SW1116/HCPT细胞药物外排功能的影响。结果 :蛋白质印迹法检测结果发现,耐药SW1116/HCPT细胞中p-Akt的表达水平明显高于亲本SW1116细胞(P<0.01)。小分子抑制剂LY294002及靶向Akt基因的Akt-siR NA均能明显降低p-Akt的表达水平,增加细胞对HCPT的药物敏感性(P值均<0.01),有明显的多药耐药逆转作用。LY294002作用48 h可使耐药转运体ABCG2 mR NA的表达水平下调(74.82±4.71)%,蛋白表达水平下调(58.24±4.78)%(P值均<0.01),并且耐药蛋白体的转运功能也明显受到抑制,细胞内Rh123的浓度增加了(1.45±0.12)倍(P<0.01)。沉默Akt基因后ABCG2 mR NA的表达水平下降了(59.63±5.14)%,蛋白表达水平下降了(44.41±2.56)%(P值均<0.01),细胞内Rh123的浓度增加了(1.22±0.10)倍(P<0.01)。结论:PI3K/Akt信号通路可正向调节耐药基因ABCG2的表达,在调节HCPT诱导的多药耐药过程中发挥重要作用。 OBJECTIVE: To investigate the effects of blocking the phosphorylation of phosphatidylinositol 3-kinase (PI3K) / serine-threonine protein kinase (Akt) signaling pathway on the reversal of hydroxycamptothecin HCPT) multidrug resistance in human colon cancer SW1116 / HCPT cells. Methods: Western blotting was used to detect the expression of Akt and phospho-Akt (p-Akt) protein in SW1116 cells and SW1116 / HCPT cells. Akt protein expression was inhibited or silenced by the small molecule inhibitor PI3K / Akt signaling pathway, LY294002 or siR NA, respectively. The effect of HCPT on the survival rate of SW1116 / HCPT cells was detected by MTT assay. Real-time fluorescent quantitative PCR and Western blotting were used to detect the mRNA and protein expressions of mR NA and protein of drug-resistant transporter ATP-binding cassette transporter G2 (ABCG2); rhodamine123 (Rh123) / HCPT cell drug efflux function. Results: The results of Western blotting showed that the expression of p-Akt in SW1116 / HCPT cells was significantly higher than that in SW1116 cells (P <0.01). The small molecule inhibitors LY294002 and Akt-siR NA targeting the Akt gene could significantly decrease the expression of p-Akt and increase the drug sensitivity to HCPT (all P <0.01), and obvious reversal of multidrug resistance effect. The expression of ABCG2 mR NA was down-regulated by LY294002 (74.82 ± 4.71)% and decreased by 58.24 ± 4.78% (P <0.01) at 48 h after transfection, and the translocation of drug-resistant protein Was also significantly inhibited, intracellular Rh123 concentration (1.45 ± 0.12) times (P <0.01). The expression of ABCG2 mR NA was decreased by 59.63 ± 5.14% and the protein expression was decreased by 44.41 ± 2.56% (P <0.01), while the intracellular Rh123 concentration was increased by (1.22 ± 0.10) Times (P <0.01). Conclusion: The PI3K / Akt signaling pathway can positively regulate the expression of drug resistance gene ABCG2 and play an important role in the regulation of HCPT-induced multidrug resistance.