神经生长因子及其受体调节胶质瘤细胞Warburg效应促进其转移的作用机制研究  

Mechanism of nerve growth factor and its receptor regulating glioma cell Warburg effect on promoting metastasis

作　　者：杜鹏程[1] 张海军[1] 武焕颖[1] 郭伟伦[1] Du Pengcheng;Zhang Haijun;Wu Huanying;Guo Weilun(Department of Neurosurgery,Chengde Central Hospital)

机构地区：[1]河北省承德市中心医院神经外科

出　　处：《重庆医科大学学报》2019年第12期1571-1576,共6页Journal of Chongqing Medical University

基　　金：2016年河北省级重大医学科研课题资助项目(编号:zd2016008)

摘　　要：目的:探究神经生长因子(nerve growth factor,NGF)及其受体调节胶质瘤细胞Warburg效应促进其转移的作用机制。方法:Western blot和实时荧光定量PCR检测3株胶质瘤细胞中NGF以及TrkA的蛋白及m RNA水平。以U251为实验细胞,将U251分为Control组和NGF组,50μg/L的NGF干预。CCK-8检测细胞活力,PI染色后流式细胞术检测细胞周期的变化,Transwell小室实验检测细胞的侵袭能力,Western blot检测神经生长因子受体(growth factor receptor,TrkA)、AKT、PI3K以及代谢酶磷酸果糖激酶(Phosphofructokinase,PFK)、丙酮酸激酶M2(pyruvate kinase M2,PKM2)的表达水平,检测葡萄糖摄取水平、乳酸水平、细胞氧耗水平。将U251细胞分为为Control组、AKT inhibitor组和NGF组,Control组为常规培养细胞,AKT inhibitor组为AKT抑制剂预处理后再使用50μg/L的NGF干预,而NGF组为50μg/L的NGF干预。同样进行上述检测。结果:胶质瘤细胞中NGF和TrkA水平高于胶质细胞,其中U251水平最高。NGF干预后,U251细胞活力增高,NGF组细胞中G2期明显高于Control组,NGF组细胞的侵袭数目明显高于Control组,NGF组细胞中TrkA、AKT、PI3K的蛋白水平明显高于Control组。而代谢酶PFK、PKM2的表达水平明显低于Control组。NGF组细胞葡萄糖摄取水平、乳酸水平、细胞氧耗水平相比Control组具有明显差异。AKT信号抑制后,NGF组细胞活力高于Control组、AKT inhibitor组,NGF组细胞中G2期比例明显高于Control组和AKT inhibitor组。NGF组细胞的侵袭数目明显高于Control组和AKT inhibitor组,NGF组细胞中TrkA、AKT、PI3K的蛋白水平明显高于Control组和AKT inhibitor组。而代谢酶PFK、PKM2的表达水平明显低于Control组和AKT inhibitor组。NGF组细胞葡萄糖摄取水平、乳酸水平、细胞氧耗水平相比Control组和AKT inhibitor组具有明显差异。结论:NGF可以通过TrkA-PI3K/AKT信号促进胶质瘤细胞Warburg效应,促进其转移。Objective:To investigate the mechanism of action of nerve growth factor(NGF)and its receptor in regulating Warburg effect and promoting metastasis of glioma cells.Methods:Western blot and RT-qPCR were used to measure the protein and m RNA expression of NGF and TrkA in three glioma cell lines.U251 cells were divided into control group and NGF group(treated with 50μg/L NGF).CCK-8 assay was used to measure cell viability;flow cytometry after PI staining was used to observe the change in cell cycle;Transwell chamber was used to measure cell invasion ability;Western blot was used to measure the expression of TrkA,AKT,PI3 K,phosphofructokinase(PFK),and pyruvate kinase M2(PKM2).The levels of glucose uptake,lactic acid,and cell oxygen consumption were measured.U251 cells were divided into control group,AKT inhibitor group,and NGF group.The cells in the control group were given routine treatment,those in the AKT inhibitor group were pretreated with AKT inhibitor and then treated with 50μg/L NGF,and those in the NGF group were treated with 50μg/L NGF.The above tests were also performed.Results:Glioma cells had significantly higher levels of NGF and TrkA than glial cells,and U251 glioma cells had the highest levels.U251 cells showed a significant increase in cell viability after NGF intervention.Compared with the control group,the NGF group had a significantly higher number of cells in G2 phase,a significantly higher number of invasive cells,and significantly higher protein expression of TrkA,AKT,and PI3 K,as well as significantly lower expression of the metabolic enzymes PFK and PKM2.There were significant differences between the NGF group and the control group in the levels of glucose uptake,lactic acid,and oxygen consumption.After AKT signal suppression,the NGF group had significantly higher cell viability and number of cells in G2 phase than the control group and the AKT inhibitor group.Compared with the control group and the AKT inhibitor group,the NGF group had significantly higher number of invasive cells and prot

关 键 词：神经生长因子 胶质瘤细胞 Warburg效应 转移 

分 类 号：R735.7[医药卫生—肿瘤]