The Neuroprotection Of Dexmedetomidine On Autophagy And Apoptosis In Rats With Traumatic Brain Injury

Backgrounds:Traumatic brain injury(TBI)is a severe disease with high morbidity and mortality around the world.Over 10 million people worldwide subject with traumatic brain injury every year,many survivors evolved with catastrophic complications,including neurological deficit and motor dysfunctio.Although the mortality of TBI has significantly improved in the last few decades due to advances in Emergency and Critical Care Medicine,there are still no evidence-base effective treatment for improving functional recovery and preventing long-term neuropsychiatric complications after TBI.It was convinced that the damages of TBI related with primary injury and secondary injury,which caused by cerebral edema,neuroinflammation,apoptosis and breakdown of blood-brain barrier.Secondary brain injury lasts from days to months,even lasts for years,which ultimately leading to the death of neuronal cell,loss of motor and cognition function and neurodegeneration diseases,and even central nervous system tumor.It is crucial for research on TBI to focus on the mechanisms of secondary brain injury,and explore the neuroprotective agents for attenuating and eliminate the influence for secondary brain injury post-TBI.Dexmedetomidine(DEX)is an a2-adrenoceptor agonist.It has been used as an anesthetic agent in analgesia and critical care management.Recent studies have shown that DEX also has potential beneficial effects for the central nervous system diseases.It may have the neuroprotective effect against ischemic stroke,subarachnoid hemorrhage,intracranial hemorrhage and SCI.As many pathophysiological mechanisms involved in traumatic brain injury are similar to other neural system diseases;Thus,we assume that DEX may have the neuroprotective effect for TBI.This study intends to explore the neuroprotective effect and mechanism of DEX on traumatic brain injury.Part 1Neuroprotective effects of Dexmedetomidine pretreatment for traumatic brain injuryObjective:To investigate the neuroprotective effect of DEX for TBI rat.Methods:(1)Exploring the optimal dose of DEX:rats were divided into five groups:sham operation group(Sham),TBI+NS pretreated group(normal saline ip),TBI+DEX50 pretreated group(DEX50μg/kg,i.p),TBI+DEX100 pretreated pretreated group(DEX100μg/kg,ip),and TBI+DEX200 pretreated group(DEX200μg/kg,i.p).(2)The modified neurological severity score(mNSS)was used to examine the effects of MB on neurological deficits;The brain water content(BWC)was calculated as follows:(wet weight-dry weight)/wet weight x 100%;Apoptosis-related proteins of Caspase-3,Bax and Bcl-2 were investigated by western blot analysis.The optimal dose of DEX was determined according to the neurological function,brain water content and apoptosis-related proteins 24 h post-TBI.Results:(1)The brain water content and mNSS were significantly increased after TBI compared to Sham group(P<0.05).DEX with 100μg/kg significantly reduced neurological deficits and brain water content relative to NS group 24 h post-TBI(P<0.05).However,this effect was not found in the TBI+DEX50 group and TBI+DEX200 group.Therefore,the optimal dose used in the following studies was 100μg/kg(i.p).(2)The level of Caspase-3 and Bax were significantly elevated 24 h post-TBI;Compared with TBI+NS group,DEX(DEX 100μg/kg,i.p)pretreatment significantly attenuated the expression of Caspase-3 and Bax(P<0.05);DEX pretreatment also significantly increase the level of Bcl-2(P<0.05).Conclusions:DEX has neuroprotective effect against TBI including alleviated cerebral edema,decreased neurological functional deficits,inhibited apoptic neuron cell death caused by TBI.Part 2 Effect of Dexmedetomidine on the apoptosis and autophagy in cortex after traumatic brain injury Objective:To investigate the possible mechanisms of DEX for TBI rat through the autophagic and apoptotic neural death pathway.Methods:(1)Rats were divided into three groups:sham operation group(Sham),TBI+NS pretreated group(normal saline i.p),TBI+DEX pretreated group(DEX 100μg/kg,i.p).Base on the previous studies,we evaluated the neurological function 24h,72h and 7 day post-TBI;brain water content,brain lesion volume was respectively measured at 24h and 14 days post-TBI.(2)Autophagic flux and apoptosis were investigated at 24h post-TBI.(3)The modified neurological severity score(mNSS)was used to examine the effects of DEX on neurological deficits.The brain water content(BWC)was calculated as follows:(wet weight-dry weight)/wet weight × 100%.Western Blot was employed to evaluate LC3-II,p62,Caspase-3,Bax and Bcl-2,to determine the autophagic flux and apoptosis in peri-contusional cortex.We also evaluated the brain lesion volume using Hematoxylin staining and then analyzed using Image J.Results:(1)The TBI resulted in neurological functional deficits.Compared with the NS group,mNSS scores were significantly lower in the TBI+DEX group 24 h and 72 h post-TBI(P<0.05).However,there was no significantly difference between NS group and DEX group on mNSS scores 7 days post-TBI(P>0.05).(2)Based on the increased Caspase-3 and Bax,as well as decreased Bcl-2,we found that apoptosis was significantly activated post-TBI.Compared to the TBI+NS group,the levels of apoptosis-related proteins were reversed by DEX pretreatment at 24 h post-TBI(P<0.05).(3)The increased levels of LC3-II and p62 indicated the autophagic flux was impaired.We found the pretreatment of DEX restored the autophagic flux post-TBI.(4)We evaluated the cerebral lesion volume to confirm the neuroprotective effect of DEX in TBI.The lesion volume of TBI+DEX group was significantly smaller than that of the TBI+NS group 14 days post-TBI(P<0.05).Conclusions:(1)We found that DEX exerts a neuroprotective effect by alleviating cerebral edema,decreasing neurological functional deficits and reducing brain lesion volume.(2)The inhibition of apoptotic neuron cell death and repairment of the autophagic flux induced by DEX may play a potential neuroprotective role in traumatic brain injury.