溴氰菊酯及6-羟基多巴胺对PC12细胞Nrf2/ARE通路影响

Effect of deltamethrin and 6-hydroxydopamine on Nrf2/ARE pathway in PC12 cells

目的探讨溴氰菊酯(DM)与6-羟基多巴胺(6-OHDA)对转录因子NF-E2相关因子2(Nrf2)/抗氧化反应元件(ARE)通路作用的影响及差异。方法用10μmol/L溴氰菊酯处理PC12细胞1h或用100μmol/L6-OHDA处理PC12细胞17h后,用RT-PCR方法检测细胞Nrf2基因和其驱动的靶基因-γ-谷氨酰半胱氨酸合成酶(GCS,由重亚单位GCSh和轻亚单位GCS1组成)和血红素氧合酶(HO-1)基因 mRNA表达水平,蛋白印迹(Westernblot)方法检测细胞Nrf2蛋白水平,免疫荧光细胞化学技术检测细胞HO-1蛋白水平。结果 DM染毒诱导转录因子Nrf2 mRNA表达、激活Nrf2,Nrf2可能参与DM对Nrf2下游基因GCSh、GCS1以及HO-1表达的调控;6-OHDA能诱导PC12细胞Nrf2 mRNA和蛋白的表达以及激活Nrf2;6-OHDA也诱导细胞HO-1或GCSh mRNA和蛋白的表达。结论 DM与6-OHDA对Nrf2/ARE通路的作用相似,DM是可干扰Nrf2/ARE通路的神经毒物。

Objective To explore the effect of deltamethrin（DM） and 6-hydroxydopamine（6-OHDA） on NF-E2 related factor 2（Nrf2） /（ARE） pathway in PC12 cells in vitro.Methods PC12 cells were treated with 10 μmol/L of DM for 1 hr or 100 μmol/L of 6-OHDA for 17 hr,respectively.The relative amount of mRNA expresion of Nrf2 gene and it＇s target gene was measured with reverse transcription polymerase chain reaction（RT-PCR）.The Nrf2 protein level was detected with western blot.The hemeoxygenase1（HO-1） protein level was detected with immunocytochemistry combined with laser scan confocal microscopy（LSCM）.Results The results showed that DM enhanced cellular expression of Nrf2 at transcriptional and protein level.In addition,DM treatment caused nuclear accumulation of Nrf2 in association with downstream activation of Nrf2 mediated oxidative response genes such as glutamylcysteine synthetase h（GCSh）,and HO-1.6-OHDA could induce Nrf2 and expressions of both HO-1 mRNA and protein and activate Nrf2.6-OHDA could also induce mRNA expression of GCSh gene.Conclusion The study suggests that DM and 6-OHDA exhibit similar activation of Nrf2 /ARE pathway and DM is a neurotoxic agent disturbing Nrf2 /ARE pathway.