Research On The Role Of Prdx6 And Its Ralated IPLA2 Activity In Cerebral Ischemia-reperfusion Injury And Its Mechanism

Background:oxidative stress is the key factor in cerebral ischemia/reperfusion damage.Endogenous antioxidant systems containing glutathione reductase, superoxide disproportionation enzyme(SOD) system,antioxidants Peroxiredoxins(Prdxs) system,thioredoxin/thioredoxin reductase(Trx/Trx R) system,and so on.Prdx6 protein with NSGPx and i PLA2 dual activity.Jacques’ s disease,Parkinson’s disease,Peak disease,pulmonary malignant mesothelium cell tumor,squamous epithelial cancer,skin wound healing area,experimental premature aging cells etc,has been found related to the expression of Prdx6.They were mostly with peroxidase activity of Prdx6 by inhibition of lipid peroxidation.At present, the role of Prdx6-i PLA2 activity on cerebral ischemia disease has little research.Objective: To investigate the effect of Prdx6 and its related i PLA2 activity on cerebral ischemia-reperfusion injury and its related mechanism.Methods:(1) Rats were randomly divided into Sham group,MCAO group, Scramble group,Prdx6-si RNA group and Prdx6-si RNA+MJ33 group(n = 16);(2)A chemical synthesis Prdx6-si RNA,a out-of-order Prdx6-si RNA as the negative control had been constructed.Prdx6-si RNA group with 8μl Prdx6-si RNA by lateral ventricle injection,Prdx6-si RNA+MJ33 group adding to MJ33(0.5μmol/kg) by the caudal vein injection.Scramble group with out-of-order si RNA.the remaining two groups with positioning only after skull drilling without injecting;(3)After 24 h of lateral ventricle injection,we built the model of transient middle cerebral artery occlusion(MCAO) to induce the left cerebral cortex ischemic/reperfusion in SD rats, of which Sham group with only surgery incision, without inserting suture;(4)After 24 h of cerebral ischemia/reperfusion injury,nerve function score,cerebral water content and cerebral infarction volume in each groups were determined.moreover,with WB and real-time q PCR,we detected on Prdx6 and by ELISA to detect i PLA2 activity in each groups;(5)With HE and methylene blue staining,neurons were detected in the areas of the cerebral ischemia/reperfusion injury;(6)B detecting SOD activity and MDA content to evaluate oxidative activity, with WB, real- time q PCR to detect caspase3, and using TUNLE staining method to explore relating apoptosis;(7)Through WB to detect the expression level of NF-?B,i NOS and COX2 on relating TLR2/4 signaling pathways,and with ELISA to detect theirs corresponding content of IL-1beta, IL-17 and IL-23.Results(1)Nerve function score, water content and cerebral infarction volume on left cerebral ischemic:compared to MCAO group,these significantly increase in Prdx6-si RNA group and Prdx6-si RNA+MJ33 group,Prdx6 si RNA+MJ33 group compared with Prdx6-si RNA group,these decrease,all change are statistically significant;(2)The expression of Prdx6 and i PLA2 activity:compared to MCAO group,these significantly decrease in Prdx6-si RNA group and Prdx6-si RNA +MJ33 group,Prdx6-si RNA+MJ33 group compared with Prdx6-si RNA group,these decrease more,all change are statistically significant;(3)HE and methylene blue staining showed brain ischemic neuronal damage:compared to MCAO group,these significantly increase in Prdx6-si RNA and Prdx6-si RNA+MJ33 group,Prdx6 si RNA+MJ33 group compared with Prdx6-si RNA group,these decrease, all change are statistically significant;(4) SOD activity and MDA content:compared to MCAO group,in Prdx6-si RNA group and Prdx6-si RNA+MJ33 group SOD activity significantly decrease,MDA content significantly increase,Prdx6-si RNA + MJ33 group compared with Prdx6-si RNA group,SOD activity increase, MDA content decrease,and these changes are statistically significant;(5) TUNLE dyeing and caspase3 expression:compared to MCAO group,these significantly increase in Prdx6-si RNA group and Prdx6-si RNA + MJ33 group,Prdx6 si RNA + MJ33 group compared with Prdx6-si RNA group,these decrease,all change are statistically significant;(6) The expression of inflammatory cytokines of TLR2/4 related pathways in post-ischemic inflammation: compared to MCAO group,these significantly increase in Prdx6-si RNA group and Prdx6-si RNA + MJ33 group, Prdx6 si RNA + MJ33 group compared with Prdx6-si RNA group,these decrease, all change are statistically significant.Conclusion:Prdx6 in cerebral ischemia reperfusion injury plays a protective role,the role involves its anti-inflammatory,antioxidant and antiapoptotic mechanisms;however,its related i PLA2 activity can weaken these protection effect.