Quercetin Protects HUVECs From Iron Overload Damages Via ROS/ADMA/DDAH?/eNOS/NO Pathway

Objective:Iron is one of the trace elements necessary for the human body,while iron overload can be harmful to our health.The aberrant accumulation of excessive iron in blood vessels often causes vascular endothelial cells(VECs)damage,which is initially thought to be related to the increase in reactive oxygen species(ROS).Asymmetric dimethylarginine(ADMA),an endogenous nitric oxide synthase(NOS)inhibitor,is an important substance regulating NOS-NO system to reduce NO synthesis and cause endothelial dysfunction.The main metabolic pathway of ADMA is degradated by dimethylarginine dimethylaminohydrolase(DDAH),and DDAH is extremely sensitive to ROS.Iron overload can lead to an increase in oxidative stress,while Quercetin,as a flavonoid,has a certain ability to scavenge free radicals.Therefore,this study aims to explore the underlying pathway,focusing on ROS/ADMA/DDAH ?/eNOS/NO pathway,to investigate the protection mechanism of Que on iron overload induced HUVECs injury.Method:In this paper,HUVECs were treated with 50?M iron dextran for 48 h to construct an iron overload injury model.Then,the cell viability and LDH activity,apoptosis and the expression of cleaved-caspase 3,the activities of SOD and GSH-PX,the content of MDA,the level of ADMA and NO,the activity of DDAH?,and the expression of p-eNOS/ eNOS and DDAH?,the level of ROS,mitochondrial membrane potential(MMP),mPTP opening was detected.At the same time,Que was added to co-incubation with iron to detect changes in the above indicators.In addition,L-Arg,the ADMA competition substrate,CsA,the mPTP blocking agent,and Eda,the free radical scavenger,were established as positive control groups.Results:1.Treatment with 50?M iron dextran significantly decreased cell viability and evaluated LDH activity when compared to the control group.Additionally,the apoptosis rate was increased,and the expression of cleaved caspase-3 was increased as well.2.20?M Que could increased the cell viability and decreased the LDH activity.Moreover,the SOD and GSH-PX activities improved,and the content of MDA reduced.Subsequently,the DDAH? activity and expression increased,the ADMA concentration decreased,the content of NO and the ratio of p-eNOS/eNOS increased.Furthermore,intracellular ROS generation decreased,MMP increased,and mPTP opening was inhibited,eventually leaded to apoptosis rate decreased.Its effects were similar to that of L-Arg,CsA and Eda treatment.3.After infected HUVECs with pAD/DDA?-shRNA,the effects of Que mentioned above were reversed.Conclusion:Quercetin could alleviate oxidative stress and mitochondrial dysfunction induced by iron overload in HUVECs,and its mechanism may be related to ROS/ADMA/DDAH?/eNOS/NO pathway.