The Inhibition Effects Of Curcumin On Acrylamide-induced Apoptosis In SH-SY5Y Cells

Objective: To investigate the effect of CUR on the apoptosis induced by acrylamide in SH-SY5 Y cells,to provide new ideas and clues for the prevention and treatment of ACR poisoning.Methods:The dose of ACR and CUR were determined by the result of MTT assay.Cells were divided into 5 groups: control group,ACR group and 3 CUR pretreatment groups.Cells in ACR group were treated with ACR at final concentration of 2 mmol/L for 24 h.Cells in CUR pretreatment groups were treated with CUR at final concentration of 1?5?10?mol/L respectively for 2h,and then treated with 2 mmol/L ACR for 24 h.Cell viability was determined by MTT assay.Cell morphological changes were observed under an optical microscopy.ROS generation was detected with DCFH-DA fluorescence probe.MDA levels ? GSH contents and GSH-Px activity weredetermined using a spectrophotometer.The protein expression level of nuclear and cytosolic NF-?B,Bax,Bcl-2 and P53 were quantified by Western blot analysis.The rate of cell apoptosis was determined by flow cytometry.Results:(1)a:Cell viability: Compared with control group,cells treated with 2?4 and 8 mmol/L ACR exhibited neurite shrink,adherence reduction,and ball float,Cell viability in 2?4and 8 mmol/L ACR treatment group significantly decreased in a dose-dependent manner(P<0.001).Therefore,the final dose of ACR in subsequent experiments was 2 mmol/L.b:In CUR pretreatment group,80?mol/LCUR showed a significantly decrease in cell viability(P<0.001),compared with control group.Therefore,the final doses of CUR in subsequent experiments were 1 ?mol/L,5 ?mol/L and 10 ?mol/L.c:Compared with ACR group,5 ?mol/L and 10 ?mol/L CUR pretreatment groups significantly increased cell viability(P<0.001),indicating that CUR could attenuate the reduction of cell viability caused by ACR.(2)Oxidative stress: a: Compared with control group,ACR group significantly increased ROS and MDA levels(P<0.001),and significantly decreased GSH contents,indicating that ACR could induce oxidative stress.b: Compared with ACR group,5?mol/L and 10 ?mol/L CUR pretreatment groups significantly decreased MDA and ROS levels(P<0.001),and increased the GSH contents(P<0.001).It indicated that CUR could attenuate the ACR-induced oxidative stress levels.(3)Effects on apoptosis: a: As shown in flow cytometry and western blot,compared with control group,apoptosis rate in ACR group was significantly increased(P<0.001),the expression of Bax as well as P53 protein levels were significantly increased(P<0.001),and Bcl-2 protein expression levels were significantly decreased(P<0.001).b: Compared with ACR group,apoptosis rate in CUR groups(1?mol/L,5?mol/L)were significantly decreased,expression of Bax were significantly decreased in CUR pretreatmentgroups(1,5,10 ?mol/L)(P<0.001),and expression of P53 were significantly decreased in CUR pretreatment group(5?mol/L)(P<0.01),pretreatmentwith 5 and 10 ?mol/L CURsignificantly increased expression of Bcl-2.(P<0.01)(4)Effects on NF-?B : As shown in western blot,compared with control group,expression of both nuclear and cytosolic NF-?B markedly increased;compared with ACR group,CUR pretreatment groups(1,5,10 ?mol/L)significantly decreased the expression of NF-?B(P<0.001).(5)The role of NF-?B pathway in apoptosis-induced by ACR: a: Compared with control group,the expression of Bax were significantly increased(P<0.001),and the expression of Bcl-2 were significantly decreased in ACR group.b: Compared with ACR group,BAY11-7082 pretreatment groups(2,5 ?mol/L)significantly increased the expression of Bax protein,and markedly decreased the expression of Bcl-2 protein in a dose-dependent manner.c: Flow cytometry showed that inhibition of NF-?B pathway led to a markedly increased in cells apoptosis rate,compared with ACR group.It indicated that inhibition of NF-?B pathway could aggravate the apoptosis induced by ACR in SH-SY5 Ycells.Conclusion: Treatment with ACR at 2 mmol/L for 24 h could decrease cell viability,increase oxidative stress and induce neuron apoptosisin SH-SY5 Y cells,CUR pretreatment at 1-10?mol/Lcould reverse the cytotoxicity of ACR and reduce the neruron apoptosis.In addition,thecompensatory elevation of NF-?B caused by ACR could protect cells against the ACR-induced apoptosis.