Study On The Protective Effect Of AT1R Inhibitor On Cerebrovascular Autoregulation After Traumatic Brain Injury By Down-Regulating Notch Signaling

Background Traumatic brain injury(TBI)occurs when the head is hit directly or indirectly by an object or by blast waves,or when an object pierces the skull and enters the brain parenchyma.The mortality and morbidity of TBI are higher,which can make the burden of society and family heavier.TBI affects practically all tissue and cell types directly or indirectly involved in the regulation of CBF,leading to the disorder of Cerebrovascular autoregulation(CA),causing secondary injury.The primary injury of brain tissue caused by TBI is difficult to reverse,the secondary brain injury(in part)due to vascular/microvascular alterations,and dysregulation of cerebral blood flow(CBF)initiated by TBI is potentially preventable.Therefore,it can provide a new theoretical basis for the treatment of traumatic brain injury by studying the mechanism of cerebrovascular autoregulation after TBI.Objectives In this study,we aim to using functional magnetic resonance imaging(fMRI)to evaluate the cerebrovascular autoregulation of the whole brain and regions of interest in TBI rats.Then to explore the role and mechanism of AT1R and downstream signaling pathways in cerebrovascular autoregulation after TBI,and to provide a new theoretical basis for the treatment of TBI.Methods(1)Using functional magnetic resonance imaging(fMRI)to evaluate cerebrovascular autoregulation function in TBI rats with different injury degrees.using feeney free-fall blow device principle to prepare traumatic brain injury animal models:40 male SD rats(250-300g)were randomly divided into 4 groups:control group(Sham,n = 10),Mild TBI group(Mild,n = 10),Moderate TBI group(Moderate,n =10),Severe TBI group(Severe,n = 10).24 hours after TBI perform neurological severity score and water maze,then block the abdominal aorta in an instant to make the blood pressure transient change,using functional magnetic resonance imaging detection of cerebrovascular autoregulation function,evaluate the cerebrovascular autoregulation of the whole brain and regions of interests in TBI rats,then test for histologic evaluation(HE staining,NISSL staining,TUNEL).(2)Explore the protective effect and mechanism of Notch signaling pathway inhibitor DAPT on cerebrovascular autoregulation with Severe TBI rats,60 male SD rats(250-300g)were randomly divided into 6 groups:Mild TBI group(Mild,n = 10),Moderate TBI group(Moderate,n = 10),Severe TBI group(Severe,n = 10),Severe TBI + vehicle group(Severe + vehicle,n = 10),Severe TBI + DAPT group(Severe + DAPT,n = 10).?The protein and gene expression of the Notch and downstream signal Hes1 and Hes5,apoptosis and oxidative stress indicators were evaluated.24 hours after TBI,test protein and gene expression of Notch and downstream signal Hes1,Hes5,apoptosis index:the Bcl-2,Bax,Caspase 3,Caspase-9,and oxidative stress indicators:the MDA and SOD around the damaged area of rats.? To evaluate the protective effect and mechanism of Notch signal inhibitor DAPT on cerebrovascular autoregulation in severe TBI rats.Selection of severe TBI in rats with Notch inhibitors for treatment,severe TBI and severe TBI + vehicle as control group,24 hours after TBI,perform neurological severity score and water maze,using functional magnetic resonance imaging detection of cerebrovascular autoregulation function,evaluate the cerebrovascular autoregulation of the whole brain and regions of interests in TBI rats,then test for histologic evaluation(HE staining,NISSL staining,TUNEL).then test apoptosis index:the Bcl-2,Bax,Caspase 3,Caspase-9,and oxidative stress indicators:the MDA and SOD around the damaged area of rats.(3)Investigate the protective effect and mechanism of AT1R inhibitors on cerebrovascular autoregulation with Severe TBI rats.30 male SD rats(250-300g)were randomly divided into 2 groups:Severe TBI + vehicle group(Severe + vehicle,n = 10),Severe TBI + Losartan group(Severe + Losartan,n = 10).24 hours after TBI perform neurological severity score and water maze,using functional magnetic resonance imaging detection of cerebrovascular autoregulation function,evaluate the cerebrovascular autoregulation of the whole brain and regions of interest in TBI rats,test protein and gene expression of Notch and downstream signal Hesl,Hess,apoptosis index:the Bcl-2,Bax,Caspase 3,Caspase-9,and oxidative stress indicators:the MDA 'and SOD around the damaged area of rats.Results(1)24 hours after TBI,severe TBI in rats after abdominal aorta remove blocking cerebral blood flow is still in a downward trend in 16 minutes,show cerebrovascular autoregulating function is impaired,the normal group return to the base level of the cerebral blood flow after releasing obstruction,mild TBI group and moderate TBI group respectively in 8 or 12 minutes return to the basic level of the cerebral blood flow,prompt good cerebrovascular autoregulation function.As the degree of damage increased,the expression of Notch and downstream signal Hes1 and Hes5 gradually increased,and the differences were statistically significant(p<0.05).Apoptosis factors Bax,Caspase 3,Caspase-9 gradually rise,antiapoptotic factor the Bcl-2 gradually decreased,oxidative stress index:MDA increased SOD decreased,expression between groups with statistical significance(p<0.05).Compared with TBI group and TBI+ vehicle group,the brain blood flow in the severe TBI group was increased by 16 minutes after the removal of the Notch inhibitor DAPT,suggesting an improvement of the cerebrovascular autoregulation.In addition,the expression of Notch and downstream signal Hesl and Hes5 in DAPT treatment group decreased,and the differences among the groups were statistically significant(p<0.05).Apoptotic factor Bax,caspase-3,caspase-9 decreased,anti-apoptotic factor bcl-2 increased,MDA decreased,SOD increased,and the difference between the groups was statistically significant(p<0.05).The NSS score of DAPT treatment group and the water maze escape latency were all improved,and the differences in each group were statistically significant(p<0.05).(2)After the treatment of AT1R inhibitor Losartan,in severe TBI group the cerebral blood flow began to rise at 16 minutes after the abdominal aorta releasing occlusion,suggesting that the cerebrovascular autoregulation function was restored.After the treatment of Losartan,Notch and downstream signal factor Hesl and Hes5 decreased,and the differences were statistically significant(p<0.05).In addition,Bax,caspase-3 and caspase-9 were decreased in Losartan treatment group,and the anti-apoptotic factor bcl-2 increased,MDA decreased,SOD increased,and the differences between the groups were statistically significant(p<0.05).The NSS score in Losartan treatment group and the water maze escape latency were all improved,and the differences in each group were statistically significant(p<0.05).Conclusion(1)The cerebrovascular autoregulation of whole brain and region of interests are impaired in severe TBI rats in 24 hours after TBI(2)AT1R inhibitors Losartan can inhibit Notch and downstream signal to reduce apoptosis and oxidative stress,improve the cerebrovascular autoregulation function of severe TBI rat.