Treadmill Training Reduces Cerebral Ischemia-reperfusion Injury By Inhibiting Ferroptosis

Cerebral ischemia is a common nervous system disease,which has become the leading cause of adult physical disability in the world.However,the mechanism by which exercise training attenuates cerebral ischemia/reperfusion(I / R)injury,especially in the regulation of iron level in neuronal damage,has not been systematically studied.The purpose of this study is to reveal that treadmill training can inhibit ferroptosis after ischemic stroke in rats.Therefore,in the first part of this study,treadmill training was used to treat rats with brain I / R injury and observe its curative effect and ferroptosis related mechanism.In the second part,erastin(an ferroptosis activator)was used to induce the down-regulation of the expression level of cystine / glutamate reverse transporter(SLC7A11),and treadmill training was used to intervene rats with brain I / R,to observe whether erastin blocks the neuroprotective effect of treadmill training.This study can further reveal the molecular mechanism of brain protection of rehabilitation training and provide evidence of clinical rehabilitation as effective training.Part Ⅰ.The neuroprotective effect of treadmill training on rats with cerebral I / R injury and the mechanism of ferroptosis PurposeCerebral ischemia is a global disease that seriously threatens human health.Treadmill training has been widely used to treat cerebral ischemia.Treadmill training has a significant therapeutic effect on cerebral ischemia,but its mechanism related to ferroptosis is not clear.This study observed the protective effect of treadmill training on neurological function in rats with brain I / R injury,and the changes of lipid peroxidation,iron ion level and ferroptosis related protein level in cortical penumbra of rats,so as to explore the neuroprotective effect and mechanism of treadmill training.MethodThe internationally recognized longa method was used to establish the middle cerebral artery occlusion(MCAO)model in rats.One day after the successful construction of MCAO model,they were divided into sham operation group,MCAO group and MCAO +treadmill training group.Modified neurological severity scores(mNSS)and hanging wire test were performed on day 1,7 and 14 to evaluate motor ability and reflex function.Ferroptosis related index kit was used to detect the content of iron ions and lipid peroxidation products in the cortical penumbra of rats.The molecular and morphological features were detected by immunofluorescence and Western blot.ResultmNSS test showed that the motor function,reflex and balance abilities in the I/R injury rats after treadmill intervention were significantly improved.Treadmill training decreased the level of lipid peroxides and iron ions in the cerebral cortex of ischemic rats.We found that the protein levels of ferroptosis-related proteins including nuclear transcription factor E2 related factor 2(Nrf2),SLC7A11 and glutathione peroxidase 4(GPX4)were decreased in rats after cerebral I / R injury,while treadmill training increased the expression of these ferroptosis related proteins.ConclusionThis study provides the first evidence that treadmill training can inhibit ferroptosis to protect the cerebral cortex from I / R injury.This study shows that treadmill trainning may be a valuable training method for the treatment of cerebral ischemia.Part Ⅱ.Effects of treadmill training and ferroptosis activator on neurological dysfunction and ferroptosis after cerebral ischemia injury PurposeTo observe the effects of injection of erastin and treadmill training on neurological dysfunction in rats after brain I / R injury,as well as the effects of lipid peroxidation,iron ion level and ferroptosis related protein expression in cortical penumbra,and to explore whether injection of erastin blocks SLC7A11 / GPx4 pathway and reverses the neuroprotective effect of treadmill training by inhibiting ferroptosis.MethodOne day after the MCAO model was successfully constructed,it was divided into sham operation group,MCAO group,MCAO + treadmill training group,MCAO +treadmill training + erastin group.Erastin was injected continuously for 20 days after the MCAO model was established to induce ferroptosis.Modified neurological severity scores(MNSs)and hanging wire test were performed on day 1,27 and 34 to evaluate motor ability and reflex function.The content of iron and related lipid peroxidation products in the cortex of rats was detected by iron peroxidation kit.The molecular and morphological features were detected by immunofluorescence and Western blot.ResultmNSS test showed that the neurological deficit score in the I / R injury rats after treadmill training intervention was significantly reduced.Hanging wire test showed that the grip strength in the I / R injury rats after treadmill training intervention was significantly improved.However,these neuroprotective effect was reversed by erastin.Treadmill training decreased the levels of lipid peroxide and iron ion in the cerebral cortex of ischemic rats,but the injury of ferroptosis after cerebral ischemia was aggravated after injection of erastin.Treadmill training can significantly increase the protein expression levels of SLC7A11 and GPX4,and the use of erastin,an inhibitor of SLC7A11,reduces the expression levels of SLC7A11 and GPX4.We demonstrated that erastin-induced downregulation of SLC7A11 reversed the neuroprotective effect of treadmill training.ConclusionThis study provides the first evidence suggesting that treadmill training suppresses ferroptosis by activating the SLC7A11 / GPX4 pathway,thereby against cerebral I / R injury.