Upregulation Of Peroxiredoxin 6 Expression By Curcumin In Cerebral Ischemic Injury

Background:Curcumin(Cur)is a phenolic compound that is extracted from Curcuma longa that displays neuroprotective effects as an antioxidant,antiapoptotic,and anti-inflammatory molecule in cerebral ischemic reperfusion.The mechanism of neuroprotective needs to be further discussed.Peroxiredoxin 6(Prdx6)is a functional protein with three kinds of protein activity,which has a protective effect on cerebral ischemia reperfusion,and is involved in the protective effect of curcumin on neurodegenerative diseases.Participation in the neuroprotective effect of curcumin on cerebral ischemia reperfusion injury has not been reported.Objective:To investigate whether curcumin has the neuroprotective effects on cerebral ischemia reperfusion injury with antioxidant and antiapoptotic effects by regulating Prdx6.Methods:(1)Transient cerebral ischemia and reperfusion was induced by middle cerebral artery occlusion(MCAO)using modified suture occlusion technique in rats.Brain tissue proteins were extracted from peri-infarct cortex at 6h,12 h,24h,48 h.Real-time PCR and Western blot were used to filter out the expression time peak of Prdx6.(2)Curcumin(300mg/kg)was injected intraperitoneal 1 hour after MCAO.Real-time PCR and Western blot were used to screen the expression of Prdx6.(3)Curcumin(300mg/kg)was injected intraperitoneal 1 hour after MCAO.Immunofluorescent Staining were used to screen the expression of Prdx6.(4)Small interfering RNA(siRNA)targeted to Prdx6(Prdx6-siRNA)and a scrambled RNA(scrRNA)controls were applied by intracerebroventricular injection 24 hours before MCAO injury.Real-time PCR and Western blot were used to test the interference efficiency of Prdx6-siRNA.(5)Prdx6-siRNA was applied by intracerebroventricular injection 24 hours before MCAO injury.Neurobehavioral evaluation and infarct volume were observed.(6)Prdx6-siRNA was applied by intracerebroventricular injection 24 hours before MCAO injury.Oxidative stress-related biochemical index including malondialdehyde(MDA),8-Hydroxy-2’-deoxyguanosine(8-OHdG)and nitrotyrosine were tested.(7)Prdx6-siRNA was applied by intracerebroventricular injection 24 hours before MCAO injury.TUNEL staining was observed and Bcl-2,Bax,Caspase-3 were tested.(8)Mithramycin A(MTM)(250g/kg),the blocker of specificity protein 1(SP1),was administered at the start of reperfusion.Real-time PCR and Western blot were used to test the expression of Prdx6.(9)MTM(250g/kg)was administered at the start of reperfusion.Neurobehavioral evaluation and infarct volume were observed.MDA,8-OHdG,nitrotyrosine,Bcl-2,Bax and Caspase-3 were tested.Results:(1)Prdx6 expression in the peri-infarct cortex peaked at 24 hours after reperfusion.(2)Curcumin treatment upregulated the expression of Prdx6 at 24 hours after reperfusion.(3)The co-expression of Prdx6 and NeuN was detected by immunofluorescence.Curcumin increased the numbers of the Prdx6/NeuN positive cells.(4)Compared with the I/R group,the efficiency of Prdx6-siRNA interference was up to 55.6%,and scrRNA has no interference effects.(5)Curcumin treatment reduced neurological outcome and cerebral infarct size,while Prdx6-si RNA reversed the effects.(6)Curcumin treatment significantly reduced the generation of MDA,8-OHdG and nitrotyrosine,while Prdx6-siRNA reversed the antioxidant effect of curcumin treatment.(7)The number of apoptotic cells decreased significantly after curcumin treatment,while the number of apoptotic cells after curcumin treatment was significantly increased by Prdx6-si RNA.After curcumin treatment,the expression of Bcl-2 was increased,the expression of Bax and Caspase-3 were decreased,while Prdx6-siRNA decreased the expression of Bcl-2 and increased the expression of Bax and Caspase-3.(8)Compared with the curcumin-treated model group,the expression of Prdx6 decreased after SP1 was inhibited.(9)Compared with the curcumin-treated model group,the neurological outcome,the cerebral infarct volume,the levels of MDA,8-OHdG and nitrotyrosine,the levels of Bax and Caspase-3 were increased and the levels of Bcl-2 were decreased after SP1 was inhibited.Conclusion:(1)Curcumin has neuroprotective effects against cerebral ischemic injury in vivo.(2)Curcumin has neuroprotective effects by antioxidant and antiapoptotic effects.These effects may be related to the upregulation of Prdx6 through SP1 induction.