大黄酚对氨诱导小鼠星形胶质细胞c-fos和c-jun mRNA表达下调的影响及其相关机制

Effect of chrysophanol on down-regulated expression of c-fos and c-jun mRNA in mouse astrocytes exposed to ammonia and related mechanisms

目的探讨大黄酚对氨诱导小鼠星形胶质细胞的影响及相关机制。方法原代小鼠星形胶质细胞共分5组,分别同时加入氯化铵5 mmol·L^(-1)+大黄酚0.1,1.0和10.0 mg·L^(-1)(共3组),氯化铵5 mmol·L^(-1)+细胞外信号调节激酶1/2(ERK1/2)抑制剂UO126(10μmol·L^(-1))组和氯化铵5 mmol·L^(-1)+p38丝裂原活化蛋白激酶(p38 MAPK)抑制剂SB239063(10μmol·L^(-1))组,处理48 h。紫外分光法测定氧化应激指标丙二醛(MDA)、一氧化氮(NO)的含量以及谷胱甘肽过氧化物酶(GSH-Px)、超氧化物歧化酶(SOD)和一氧化氮合酶(NOS)的活性;Western蛋白印迹法检测ERK1/2和p38 MAPK磷酸化水平,RT-PCR检测c-fos和c-jun m RNA的表达。结果与氯化铵5 mmol·L^(-1)相比,大黄酚1.0和10.0 mg·L^(-1)显著改善氨诱导的细胞氧化应激,降低了MDA和NO的含量及NOS的活性(P<0.05),明显升高了GSH-Px和SOD的活性(P<0.05);大黄酚1.0和10.0 mg·L^(-1)也明显降低氨诱导ERK1/2和p38 MAPK的磷酸化增加(P<0.05);大黄酚0.1,1.0和10.0 mg·L^(-1),UO126和SB239063均可逆转氨诱导c-fos和c-jun m RNA表达下调的作用(P<0.05)。结论大黄酚通过抗氧化机制影响ERK1/2和p38 MAPK的磷酸化,进而逆转氨诱导c-fos和c-jun m RNA表达下调。

OBJECTIVE To investigate the effect and mechanisms of chrysophanol on the expression of c-fos and c-jun mRNA induced by ammonia chloride（NH4CI） in mouse astrocytes. METHODS Primary mouse astrocytes were cultured with NH,CI 5 mmol-L-1＋ chrysophanol 0.1, 1.0 or 10.0 mgo L-1 , or NH,CI 5 mmol. L-1 ＋extracellular regulated kinasel/2 （ERK1/2） inhibitor UO126 10 mmol-L-1 and NH,CI 5 mmol. L-1＋p38 mitogen-activated protein kinase （p38 MAPK） inhibitor SB239063 10 mmolo L-1 for 48 h. The levels of glutathione peroxidase （GSH-Px）, superoxide dismutase （SOD）, malondaldehyde （MDA）, nitric oxide （NO） and nitric oxide synthase （NOS） were detected by UV spectrophotometry. The phos- phorylation levels of ERK1/2 and p38 MAPK were detected by Western blotting. The expression of c-fos and c-jun mRNA was detected by RT-PCR. RESULTS Compared with NH,CI 5 mmolo L-1 group, ammonia-induced astrocytes oxidative stress was improved by chrysophanol （1.0 and 10.0 mg-L-1）. The content of MDA and NO and the activity of NOS were reduced （P〈0.05）. The activity GSH-Px and SOD was increased （P〈0.05）. The phosphorylation level of ERK1/2and p38 MAPK induced by ammonia was reduced in chrysophanol groups （P〈0.05）. Ammonia-induced c-fos and c-jun mRNA expression downregulation was reversed by chrysophanol （0.1, 1.0 and 10.0 mg. L-1） and UO126 and SB239063 （P〈0.05）. CONCLUSION Chrysophanol may improve the downregulated expresion of c-fos and c-jun mRNA in mouse astrocytes exposed to NH,CI by anti-oxidative stress by inhibiting the ERK1/2 and p38 MAPK phosphorylation.