Effects Of TLR4 And TLR2 On Cerebral Ischemia Reperfusion Injury In Rats

ObjectiveCerebral ischemia reperfusion injury is a pathophysiological process involving multiple mechanisms.Inflammation plays an important role in cerebral ischemia reperfusion injury.Toll-like receptor 4(TLR4)and Toll-like receptor 2(TLR2)are subtypes of Toll-like receptors(TLRs)family.They play an important role in innate immune and inflammatory responses and participate in the pathological process of ischemic cerebral injury.The purpose of this study is to explore the role and mechanism of TLR4 and TLR2 in cerebral ischemia reperfusion injury,and to provide a theoretical basis for the treatment of ischemic cerebrovascular disease.MethodsEffect of TLR4 on cerebral ischemia reperfusion injury in rats:Adult rats were randomly divided into 3 groups:sham group,MCAO group and MCAO + TAK group.Occlusion of the right middle cerebral artery was used to block cerebral ischemia in rats.MCAO group administrated saline by intraperitoneal injection at 1 h after MCAO,MCAO + TAK group administrated TLR4 antagonist TAK by intraperitoneal injection at 1 h after MCAO,reperfusion was performed 2 h after ischemia.The brain was removed on day 1,3,7,14 after surgery.Zealonga method was used to evaluate the nerve function in each group of rats before and after surgery.The brain tissue of some rats was removed freshly for Western blotting to detect the expression of TLR4 and its regulatory factor P-IKK α/β in the cerebral cortex of rats.The other rats were perfused through the heart aorta with normal saline and 4%paraformaldehyde,then make paraffin section,the pathological changes were observed by HE staining.Effect of TLR2 on cerebral ischemia reperfusion injury in rats:Adult rats were randomly divided into 3 groups:sham group,MCAO group and MCAO + PAM group.In addition to MCAO group administrated saline by intraperitoneal injection at 30 min after MCAO,MCAO + PAM group administrated TLR2 agonist PAM3CSK4 by intraperitoneal injection at 30 min after MCAO,the rest processes were the same as the study with TLR4.ResultsEffect of TLR4 on cerebral ischemia reperfusion injury in rats:(1)The score of Zealonga method sham group,MCAO group and MCAO +TAK group is 0,2.38±0.493 and 1.92±0.739 respectively.(2)HE staining showed that there were different degrees of pathological changes the MCAO group and the MCAO + TAK group in the cerebral cortex.The MCAO+ TAK group were severer than the MCAO group on day 1 after surgery,however,the MCAO group exhibited severer damage at the other time points.(3)The expression of TLR4 in the cerebral cortex:at any time points,the sham group were lower than that the MCAO group and the MCAO + TAK group.On day 1 and 14 after surgery,the MCAO group were lower than the MCAO +TAK group.While on day 3 and 7 after surgery,the MCAO group were higher than the MCAO+ TAK group.(4)The expression of P-IKK α/β in the cerebral cortex:on day 1 after surgery,the MCAO + TAK group were the higher than the MCAO group,at the other time points,the MCAO group were the highest and the sham group were lower.Effect of TLR2 on cerebral ischemia reperfusion injury in rats:(1)The score of Zealonga method sham group,MCAO group and MCAO +PAM group is 0,2.49±0.506 and 1.56±0.0.502 respectively.(2)HE staining showed that there were different degrees of pathological changes the MCAO group in the cerebral cortex,on day 1 after surgery,the MCAO group were severer than the MCAO +PAM group,at the other time points,the MCAO +PAM group were no significant pathological change.(3)The expression of TLR2 in the cerebral cortex:at any time points,MCAO group were higher than that the sham group and the MCAO+PAM group.(4)The expression of P-IKK α/β in the cerebral cortex:on day 1 to 7 after surgery,the MCAO group were higher than that the sham group and the MCAO+PAM group;on day 14 after surgery,the MCAO+PAM group were the highest and the sham group were the lowest,however,there was no statistical difference between the three groups.Conclusions(1)The score of Zealonga method showed that the sham group was normal and the MCAO group was the most serious.(2)The effects of TLR4 and TLR2 on cerebral ischemia reperfusion injury mechanism may be due to the effect of P-IKKα/βexpression in cerebral cortex.(3)On day 1 after surgery,TLR4 may alleviate ischemia reperfusion injury through MyD88 dependent pathway down regulated the expression of P-IKKα/β,and on day 3 to 14 after surgery,TLR4 may exacerbate ischemia reperfusion injury through MyD88 dependent pathway up regulated the expression of P-IKKα/β.(4)TLR2 may alleviate ischemia reperfusion injury through MyD88 dependent pathway down regulated the expression of P-IKKα/β.