Exendin-4对甲基乙二醛诱导PC12细胞氧化应激的影响

Effects of exendin-4 on methylglyoxal-induced oxidative stress in PC12 cells

目的:探讨肠促胰岛素类似物(Ex-4)对甲基乙二醛(MG)诱导PC12细胞氧化应激的影响及其机制。方法:传代培养PC12细胞,不同浓度MG(0、0.25、0.50、0.75、1.0、2.0 mmol/l)处理PC12细胞12~48 h,或用不同浓度Ex-4(25、50、100、200 nmol/L)预处理24 h后加用MG(0.75 mmol/L)干预24 h后,MTT比色法检测细胞存活率;荧光探针法检测活性氧(ROS)含量,黄嘌呤氧化酶法检测超氧化物歧化酶(SOD)活力。Ex-4(100 nmol/L)预处理PC12细胞24 h加用MG(0.75 mmol/L)干预1 h后,Western blot检测蛋白P-IκB-α、IκB-α表达情况。结果:随着MG浓度的增加和作用时间的延长,PC12细胞存活率逐渐降低;加用不同浓度Ex-4预处理后,PC12细胞存活率较单独MG处理组逐渐升高。100 nmol/L的Ex-4预处理PC12细胞后,ROS表达量较MG单独处理组下降65.30%(P<0.01);NAC预处理组(阳性对照)ROS表达量下降107.40%(P<0.01);Ex-4预处理组SOD活力增加5.30 U/mg prot(P<0.01);NAC预处理组SOD活力增加8.53 U/mg prot(P<0.01)。Ex-4预处理组P-IκB-α/IκB-α表达比例下降25.50%(P<0.01);NAC预处理组P-IκB-α/IκB-α表达比例下降35.14%(P<0.01)。结论:Ex-4浓度依赖性地增加MG诱导的PC12细胞的存活率。Ex-4能够减轻MG诱导的PC12细胞的氧化应激,其机制可能涉及抑制蛋白IκB-α的活化。

Objective: To study whether Exendin-4( Ex-4) could influence oxidative stress in PC12 cells induced by methylglyoxal and its underlying mechanism. Methods: PC12 cells were cultured with methylglyoxal( 0,0. 25,0. 50,0. 75,1. 0,2. 0 mmol / L) for12 ~ 48 h,or PC12 cells were pretreated with Ex-4( 25,50,100,200 nmol / L) for 24 h then incubated with methylglyoxal( 0. 75 mmol / L) for 24 h. MTT assay was used to measure cell viability. Fluorescent probe method was used to detect reactive oxygen species( ROS) expression. Xanthine oxidase method was used to detect superoxide dismutase( SOD) activity. With pretreatment of Exendin-4( 100 nmol / L) for 24 h,the expressions of P-IκBα,Inhibitor of NF-κB-α IκBα were detected by Western blot after PC12 cells were exposed to methylglyoxal( 0. 75 mmol / L) for 1 h. Results: Following methylglyoxal administration,cell viability was gradually decreased in a dose- and time-dependent manner. Pretreatment with Ex-4 for 24 hours,cell viability were gradually increased compared with methylglyoxal-alone group. Pretreatment with Ex-4( 100 nmol / L) for 24 hours,ROS expression was reduced by 65. 30%( P < 0.01) compared with methylglyoxal-alone group,ROS expression in NAC-pretreatment group was reduced by 107. 40%( P < 0. 01);SOD activity in the Ex-4 pretreatment group was increased by 5. 30 U / mg prot( P < 0. 01),SOD activity in the NAC pretreatment group was increased by 8. 53 U / mg prot( P < 0. 01); the ratio of P-IκB-α / IκB-α in the Ex-4 pretreatment group was reduced by 25.50%( P < 0. 01),the ratio of P-IκB-α / IκB-α in the NAC pretreatment group was reduced by 35. 14%( P < 0. 01). Conclusion:This study demonstrates that Ex-4 can increase the viabilities of PC12 cells and protect PC12 cells from oxidative stress induced by methylglyoxal,the mechanism may involve in suppressing the activation of protein IκB-α.