Aβ_(1-42)诱导U251细胞趋化因子RANTES的研究

Study on β-amyloid Peptide 1-42 Induced Chemokine RANTES in U251 Cell

目的探讨β淀粉样蛋白1-42(Aβ1-42)体外诱导人神经胶质瘤细胞(U251)细胞炎性反应的分子学机制。方法U251细胞经常规去血清培养,采用终浓度为0.2～4.0μmol/L Aβ1-42处理U251细胞24 h,以四唑盐比色(MTT)法测定细胞存活率;2μmol/L Aβ1-42处理U251细胞12、24、36、48 h后,采用硝酸还原酶法检测一氧化氮(NO)含量;采用双抗体夹心ELISA法测定细胞RANTES蛋白表达水平;免疫细胞化学染色法检测NF-κB和STAT1的表达。结果随着Aβ1-42剂量的增加,细胞的存活率降低(P<0.05);用2μmol/L Aβ1-42处理U251细胞12、24、36、48 h,结果显示NO的生成于24 h达高峰,此后逐渐下降;同浓度Aβ1-42处理U251细胞24 h后,RANTES的表达增加4倍(P<0.01);2μmol/L Aβ1-42刺激U251细胞24 h后细胞内NF-κB P65和STAT1从胞浆移至胞核且表达明显。结论Aβ1-42降低体外培养的U251细胞存活率,诱导NO和RANTES的释放,提示Aβ1-42诱导的U251细胞趋化因子RANTES产生可能与NF-κB和STAT1的活化有关。

Objective To study the molecular mechanisms of β-amyloid 1-42（Aβ 1-42） induced the inflammatory response in U251 cells.Methods U251 cells were treated with different concentration of Aβ 1-42（0.2-4.0 μmol/L） for 24 h,and the cell survival rate was evaluated by MTT.After treated with 2.0 μmol/L Aβ 1-42 for 12,24、36,48 hours,the nitric oxide content of U251 cells were detected with nitrate reductase method.The RANTES expression was tested by dual-antibody sandwich ELISA method.The expression of NF-κB and STAT1 were detected by the immunocytochemical staining.Results Aβ 1-42 decreased the survival rate of U251 cells in dose-dependent manner.The NO generated in 2 μmol/L Aβ 1-42 treated U251 cells reached its peak at 24 h and gradually decreased thereafter.The expression of RANTES increased 4-fold（P〈0.01）.The NFκB P65,the STAT1 moved from the cytoplasm to the nucleus and expressed significantly were also observed in the 2 μmol/L Aβ 1-42 treated U251 cells.Conclusion Aβ 1-42 decreases the survival rate of U251 cells and induce the releasing of NO and RANTES.This indicates that Aβ 1-42 induced chemokine RANTES in U251 cell may be close related to the activation of NF-κ B and STAT1.