PI3K/AKT通路在低氧环境下对结肠癌细胞HIF-1α及糖酵解的作用

摘要

目的 探讨缺氧环境培养下,阻断磷脂酰肌醇-3激酶(phosphatidylinositol 3-kinase,PI3K)/丝氨酸苏氨酸激酶(serine-threoninekinase,AKT)信号途径后结肠癌细胞中缺氧诱导因子(hypoxia inducible factor-1α,HIF-1α)及糖酵解相关蛋白葡萄糖转运蛋白(glucose transporter 1,GLUT-l)和乳酸脱氢酶A(lactate dehydrogenase A,LDH-A)的表达,分析PI3K/AKT信号通路与结肠癌细胞糖酵解之间的关系.方法 选择人结肠癌SW480细胞予以缺氧环境培养,分为4组:缺氧对照组、PI3K抑制剂组、AKT抑制剂组、PI3K+ AKT 抑制剂组,培养时间为24 h.采用 RT-PCR和Western blot法检测各组结肠癌细胞 HIF-1α及糖酵解相关基因(GLUT-1,LDH-A)的表达.比色法测定各组结肠癌细胞培养上清液中的乳酸浓度.结果 与缺氧对照组相比较,PI3K抑制剂组和 AKT 抑制剂组结肠癌细胞中 HIF-1α、GLUT-1、LDH-A表达下降(均 P<0.05),结肠癌细胞上清液中乳酸浓度降低(P<0.05).PI3K+ AKT 抑制剂组与缺氧对照组及单项抑制剂组相比,各项指标下降明显(P<0.01).结论 阻断PI3K/AKT 信号途径,结肠癌细胞中 HIF-1α表达下调,并可导致糖酵解相关基因GLUT-1、LDH-A的表达下降,提示PI3K/AKT信号途径参与了结肠癌细胞的糖酵解过程.%Objective To examine the protein expression of hypoxia inducible factor-1(HIF-1α)and glycolysis-related pro-teins including glucose transporter 1(GLUT-l)and lactate dehydrogenase A(LDH-A)in colon cancer cells after blockage of phosphatidylinositol 3-kinase(PI3K)/serine-threonine kinase(AKT)signal pathway under hypoxia,and to explore the relation-ship between PI3K/AKT pathway and glycolysis.Methods Human colon cancer cells SW480 were cultured under hypoxia and divided into 4 groups:hypoxia group,PI3K inhibitor group,AKT inhibitor group,and PI3K+ AKT inhibitor group.Cells were treated for 24 h.The expression levels of HIF-1α and glycolysis-related proteins(GLUT-1,LDH-A)were detected by RT-PCR and Western blotting,respectively.The concentrations of lactic acid in the supernatant of cells were determined by colorimetric assay.Results The expression levels of HIF-1α,GLUT-1 and LDH-A,and the concentration of lactic acid in the supernatant of colon cancer cells were significantly decreased in PI3K inhibitor group and AKT inhibitor group as compared with hypoxia group(P<0.05 for all).These indexes were the lowest in the PI3K+ AKT inhibitor group(P<0.01).Conclusion Blocking PI3K/AKT signal pathway inhibits the expression of HIF-1α and glycolysis-related proteins(GLUT-1,LDH-A),which sug-gests that PI3K/AKT signal pathway is involved in the glycolysis of colon cancer cells.