Triclosan Regulates Nrf2/HO-1 Pathway Through The PI3K/Akt/JNK Signalling Cascade To Induce Oxidative Damage In Neurons

Objective: Triclosan(TCS),a broad-spectrum antimicrobial agent,was widely applied.TCS has caused a wide range of environmental,water and soil pollution.It has been detected in food successively and can enter the human body through biological enrichment.Since its high lipophilicity and stability,TCS can accumulate in organisms for a long time and pose a potential threat to human health.Therefore,the human health problems caused by TCS contamination have become the focus of increasing attention.Studies have confirmed that mice exposed to TCS produce anxious behaviours,significant changes in autonomous activities and coordination ability,resulting in behaviour disorders and central nervous system toxicity.But the exact mechanism remains unclear.Therefore,this study took the mouse hippocampal neuron cell line(HT-22)as the research object,aiming to explore the specific mechanism of TCS-induced oxidative damage of HT-22.That is,the effect of TCS in HT-22 cells on PI3K/Akt and MAPK regulation of the Nrf2/HO-1 signalling pathway.Methods: HT-22 cells were treated with control and TCS of different concentrations(0.1 μM,1 μM,10 μM).Then ROS level was detected by flow cytometry,and SOD activity,MDA and GSH-Px level were detected by test kit.The expression levels of Nrf2 and HO-1 were detected by Western blot.It was also suitable for PI3 K,Akt,ERK,JNK,P38 proteins and apoptosis-related proteins Caspase-3,Bcl-2,and BAX.RT-q PCR was used to detect the m RNA changes of apoptosis-related proteins in HT-22 cells.PI3K/Akt inhibitor LY294002 and JNK inhibitor SP600125 were used to observe the changes of Nrf2 and HO-1 in cells.Results: TCS reduced the cell viability of HT-22 in a dose and time-dependent manner and increased intracellular ROS levels compared to control.The protein and m RNA levels of Caspase-3 were significantly increased when treated with TCS.And the ratio of BAX and Bcl-2 was also showed an upward trend.Moreover,the levels of MDA and SOD were obviously increased,while the activity of GSH-Px was obviously decreased,which may further aggravate cell injury.Western blot results showed that after treated with 10 μM TCS,the levels of Nrf2 and HO-1 in HT-22 cells were upregulated.In addition,TCS also stimulated the expression of phosphorylated PI3 K,Akt and JNK proteins,and this statistical effect was downregulated after the use of corresponding pharmacological inhibitors.The expression of Nrf2/HO-1 was also downregulated,suggesting that TCS regulates Nrf2/HO-1 through the PI3K/Akt and JNK pathways.Meanwhile,compared to the individual TCS treatment group,intracellular p-Akt level did not change significantly after co-treatment with TCS and JNK inhibitor;while the level of p-JNK was significantly downregulated after co-treatment with TCS and PI3K/Akt inhibitor.This suggests that JNK was regulated by PI3K/Akt.In conclusion,the neurotoxic mechanism of TCS was associated with the activation of PI3K/Akt,JNK and Nrf2/HO-1 signalling pathways,in which TCS mediated PI3K/Akt/JNK regulation of Nrf2/HO-1 signalling pathway played a pivotal role.Conclusion: TCS regulated oxidative damage of HT-22 neurons mediated by Nrf2/HO-1 through PI3K/Akt/JNK signalling pathway.