Protective Effects And Mechanism Of Limb Remote Ischemia Preconditioning On Blood-spinal Cord Barrier After Spinal Cord Ischemia Reperfusion Injury In Rats

Objective: Clinically,surgical repair of thoracoabdominal aneurysm can lead to temporary or permanent spinal cord ischemia-reperfusion injury.A major pathological change in spinal cord I/R injury is BSCB disruption,which further leads to neurological deficit.Strategies protecting BSCB integrity can improve neurological function.RIPC is a method in which short periods of non-lethal ischemia followed by reperfusion of tissue or organ protect remote tissue or organ to against a subsequent more severe I/R injury.RIPC can induce spinal cord ischemia tolerance,but the exact mechanism is unclear yet.Remote ischemic postconditioning alleviating the disruption of blood-brain barrier(BBB)to induce cerebral ischemic tolerance has been reported.Although BSCB and BBB are similar in both structure and function,the probability of RIPC preserving the integrity of BSCB following spinal cord I/R injury still needs to be confirmed.Receptor-mediated endocannabinoid system has been investigated as a latent neuroprotection target and it can alleviate ischemic injury.Endocannabinoid system affording a protection to the BBB during I/R injury has been demonstrated.Besides,a newly research suggested that CB2 receptor agonist might regulate the BSCB permeability.Therefore,this study will examine the effects of RIPC on BSCB after SCIRI,and then detect the role of CB receptor to explore the mechamism.Methods: 1.Transient aortic occlusion produced by aneurysm clips placed at the aortic arch(between the left common carotid and left subclavian arteries)and on the left subclavian artery(at its origin)to establish the SCIRI model.2.RIPC: right femoral artery occlusion and reperfusion were induced prior to spinal cord ischemia-reperfusion.3.Animals underwent intraperitoneal administration with cannabinoid receptor antagonist before RIPC.4.Histopathologic evaluation and evaluation of hind limbs motor function by Tarlov scale.5.Evaluation of BSCB integrity according to EB microvascular extravasation.6.Assay of occludin,cr1,cr2 expression in the injured spinal cord tissue by western blot method.7.Assay of occludin,cr1,cr2 colocalization with vascular endothelial cells in the injured spinal cord tissue by double immunofluorescence labeling.Results: 1.RIPC attenuated the motor dysfunction,BSCB disruption and downregulation of occludin after I/R injury.2.At 4h after injury,comparing with RIPC group,EB microvascular extravasation of AM251 group increased,occludin expression reduced;while there was no difference between AM630 group and RIPC group.It indicated that at 4h after injury,CB1 receptor antagonist AM251 could partly reverse the ptotective effects of RIPC on BSCB.3.At 24 h after injury,comparing with RIPC group,EB microvascular extravasation of both AM251 and AM630 groups were increased,occludin expression were reduced,which indicated that at 24 h after injury,both CB1 and CB2 receptors antagonists could partly reverse the ptotective effects of RIPC on BSCB.4.At 4h after injury,the expression of cr1 in injured spinal cord tissue was significantly increased,and RIPC group was more than control group;while there was no difference for cr2 expression.At 24 h after injury,the expression of cr2 in injured spinal cord tissue was significantly increased,and RIPC group was more than control group;while there was no difference for cr1 expression.5.In sham group,there was intermittent expression for cr1 along the microvascular,while there was almost no expression for cr2.In control group,the expression of both cr1 and cr2 were increased,and positive particles were increased.In RIPC group,the expression of both cr1 and cr2 were relatively continuous,and positive particles were more than control group.Conclusions: 1.Limb RIPC could improve the neurological function score after SCIRI and reduce normal motor neurons damage by attenuating BSCB disruption and increasing occludin expression.2.At rapid phase after SCIRI,CB1 receptor mediated the ptotective effects of RIPC on BSCB;while at delay phase,both CB1 and CB2 receptors mediated the ptotective effects of RIPC on BSCB.3.The expression of cr1 was increased at rapid phase after SCIRI,and returned to baseline at delay phase;while the expression of cr2 was not changed at rapid phase,but significantly increased at delay phase.RIPC could increase cr1 and cr2 expression.