代谢型谷氨酸受体5亚型介导U251细胞增殖及机制的研究

Effect of metabotropic glutamate receptor 5 subtype on proliferation of U251 cells and its mechanism

目的通过观察代谢型谷氨酸受体5亚型（mGluR5）的特异性拮抗剂2-甲基-6-苯基乙炔嘧啶（MPEP）对人胶质瘤细胞U251的影响间接探讨mGluR5对U251增殖的影响并对其机制进行分析。方法体外常规培养人胶质瘤细胞株U251，噻唑蓝（MTT）检测100μmol／LMPEP、20μmol/LSP600125（JNK特异性拮抗剂）、400感染复数（MOI）Ad-△169（c-Jun负显性病毒1作用48h后U251细胞增殖的变化；Westernblotting检测5、10、20、50、100μmol／LMPEP作用24h及100μmol／LMPEP作用4、8、12、24h后U251细胞表达磷酸化C-Jun及磷酸化JNK的变化。结果100μmol／LMPEP、20μmol／LSP600125、400MOIAd—A169处理48h后U251细胞的吸光度值均明显低于对照组，差异有统计学意义（P〈0．051；5、10、20、50、100μmol／LMPEP处理人胶质瘤细胞U25124h后的磷酸化C-Jun、磷酸化JNK表达量均明显低于对照组，而且随着用药浓度的增加，u251细胞磷酸化C-Jun、磷酸化JNK的表达越低，差异有统计学意义伊〈0．05）；100μmol／LMPEP处理U251细胞4、8、12、24h后磷酸化C-Jun、磷酸化JNK表达量均降低，而且随着用药时间的延长，U251细胞磷酸化c-Jun、磷酸化JNK的表达越低，差异有统计学意义（P〈0．05）。结论mGluR5特异性拮抗剂MPEP能抑制人胶质瘤细胞U251增殖，其机制可能与JNK通路中磷酸化c—Jun及磷酸化JNK的表达水平改变有关，并且MPEP引起的这种改变表现出浓度及时间依赖性。

Objective To discuss the effects of metabotropic glutamate receptor 5 subtype （mGluRS） on proliferation of U251 cells and the underlying mechanism by observing the effect of antagonist 2-methyl-6- （phenylethynyl）-pyridine （MPEP） on proliferation ofU251 cells. Methods Human glioma cell line U251 was conventionally cultured in vitro; MTT assay was employed to detect the effects of 100 μmol/L MPEP, 20 μ mol/L SP600125 （antagonist to JNK） and 400 multiplicity of infection （MOI） adenoviruses-A169 （negatively dominant virus of c-Jun） for 48 h on U251 cells. Western blotting was used to observe the expressions of phosphorylated c-Jun and phosphorylated JNK in U251 cells after being treated with different concentrations of MPEP （5, 10, 20, 50 and 100 μmol/L） for 24 h and with 100μ mol/L MPEP for 4, 8, 12 and 24 h. Results MTT assay showed that U251 cells being treated with 100 μmol/L MPEP, 20 μmol/L SP600125 and 400MOI Ad-A169 for 48 h had significantly lower absorbance value than their control groups （P〈0.05）. Western blotting indicated that the expressions of phosphorylated c-Jun and phosphorylated JNK in U251 cells being treated with 5, 10, 20, 50 and 100 p, mol/L MPEP for 24 h were significantly lower than those in normal controls （P〈0.05）; the higher the MPEP concentration, the lower the expression of phosphorylated c-Jun. The expressions ofphosphorylated c-Jun and phosphorylated .INK in U251 cells being treated with 100 μmol/L MPEP for 4, 8, 12 and 24 h were significantly lower than those in normal controls （P〈0.05）; the longer the MPEP given time, the lower the expression ofphosphorylated c-Jun. Conclusions MPEP, the antagonist mGluR5, could significantly promote cell proliferation of U251, which might be relevant to the expressions of phosphorylated c-Jun and phosphorylated JNK in JNK pathway, and the trend is concentration-dependent and time-dependent.