内源性调节性T细胞在SAH后脑血管痉挛及脑损伤中的作用.ppt

内源性调节性T细胞在SAH后脑血管痉挛及脑损伤中的作用,汇报人：张金卉导师：孙保亮,研究的目的及意义,SAH后脑血管痉挛、炎症反应及神经功能预后的改善是一个非常复杂的病理、生理、临床过程。虽然进行了大量研究，为止其发生机制仍未完全明了。研究小鼠体内内源性调节性T细对SAH后脑血管痉挛、炎症反应及神经功能预后的改善仍将是今后一个时期的热点。随着研究的逐渐深入，将会对内源性调节性T细胞移植SAH的临床治疗及神经功能预后带来深远的影响。,国内外相关文献,TheKineticsofLymphocyteSubsetsandMacrophagesinSubarachnoidSpaceAfterSubarachnoidHemorrhageinRats,ithasbeensuggestedthathumoralimmunityplaysaroleinthepathogenesisofcerebralvasospasmaftersubarachnoidhemorrhage,therehasbeennoquantitativeassayforcellularimmunity.Westudiedthekineticsofimmunecellsinthesubarachnoidspaceaftersubarachnoidhemorrhageintherat.,Aserialresponseofimmunoreactivecells,whichresemblesthatofthechronicallergicreactionobservedinautoimmunediseasesordelayed-typehypersensitivity,existsinthesubarachnoidspaceaftersubarachnoidhemorrhage.Thepresentresultssuggestthattheinitialresponseincellularimmunity,whichisfollowedbyhumoralimmunityandeicosanoidreactions,playsaroleinelicitingthedevelopmentofcerebralvasospasm.,蛛网膜下腔出血后脑血管痉挛分子机制的研究进展,与NO代谢无关的CVS,Statin-InducedT-LymphocyteModulationandNeuroprotectionFollowingExperimentalSubarachnoidHemorrhage,Statinsinfluenceimmunesystemactivitiesthroughechanismsindependentoftheirlipidloweringproperties.Tcellscanbesubdividedbasedoncytokinesecretionpatternsintotwosubsets:T-helpercellstype1(Th1)andtype2(Th2).IndependentlaboratorystudieshaveshownstatinstobepotentinducersofaTh2switchinimmunecellresponseandbeneuroprotectiveinseveralmodelsofcentralnervoussystem(CNS)disease.Thisstudywasthefirsttoevaluatetheimmunemodulatingeffectsofstatinsinsubarachnoidhemorrhage(SAH).,ThepresentstudyelucidatedthepotentialroleofaTh2immuneswitchinstatinprovidedneuroprotectionfollowingSAH.,Matrixmetalloproteinase-9concentrationinthecerebralextracellularfluidofpatientsduringtheacutephaseofaneurysmalsubarachnoidhemorrhage,Elevatedcerebralinterstitialpro-MMP-9relatestoearlybraininjuryinaSAHpatients.Alargerprospectivestudyshouldbeperformedtoconfirmwhetherpatientswithprolongedelevationorasecondpeakofcerebralpro-MMP-9wouldbemorelikelytodevelopDCIandtoconfirmwhetherMMP-9,amediatorofneurovascularinjury,wouldbeworthtoconsiderasapredictorofvasospasm.,Trehalosetreatmentsuppressesinflammation,oxidativestress,andvasospasminducedbyexperimentalsubarachnoidhemorrhage,Subarachnoidhemorrhage(SAH)frequentlyresultsinseveralcomplications,includingcerebralvasospasm,associatedwithhighmortality.AlthoughcerebralvasospasmisamajorcauseofbraindamagesafterSAH,otherfactorssuchasinflammatoryresponsesandoxidativestressalsocontributetohighmortalityafterSAH.Trehaloseisanon-reducingdisaccharideinwhichtwoglucoseunitsarelinkedby,-1,1-glycosidicbond,andhasbeenshowntoinducetolerancetoavarietyofstressorsinnumerousorganisms.Inthepresentstudy,weinvestigatedtheeffectoftrehaloseoncerebralvasospasm,inflammatoryresponses,andoxidativestressinducedbybloodinvitroandinvivo.,trehalosehassuppressiveeffectsonseveralpathologicalventsafterSAH,includingvasospasm,inflammatoryresponses,andlipidperoxidation.TrehalosemaybeanewtherapeuticapproachfortreatmentofcomplicationsafterSAH.,propertieswithinthecerebrospinalfluidaftersubarachnoidhemorrhageinvivoandinvitro,Tofunctionallycharacterizepro-inflammatoryandvasoconstrictivepropertiesofcerebrospinalfluidafteraneurysmalsubarachnoidhemorrhage(SAH)invivoandinvitro.,WefunctionallycharacterizedinflammatoryandvasoactivepropertiesofpatientsCSFafterSAHinvivoandinvitro.Thispro-inflammatorymilieuinthesubarachnoidspacemightplayapivotalroleinthepathophysiologyofearlyanddelayedbraininjuryaswellasvasospasmdevelopmentfollowingSAH.,研究内容,每项研究内容均分4组1、Tregs删除组2、SAH模型组3、假手术组4、PBS腹腔注射组体重、窝别、喂养方法相同的小鼠随机分组每组1015只小鼠；每只小鼠编号后按计算机随机表分4组，双盲（研究者、负责资料收集和分析的人员也不了解分组情况）较好地避免了偏倚。,研究方法实验动物处理1、Tregs删除,2、SAH模型制作,3、假手术：遇到阻力即停止继续插入，并迅速退出线栓4、PBS腹腔注射：经腹腔注射与0.25mgCD25特异性抗体等体积的PBS。,一、内源性调节性T细胞对蛛网膜下腔出血（SAH）后脑血管痉挛的检测内容,检测内容,局部脑血流量（regionalcerebralbloodflow,rCBF）检测：颅骨开窗，在立体定向仪控制下，在SAH前及SAH后12小时内（或6小时）用激光多普勒血流计动态测量动物顶叶皮层rCBF。,基底动脉形态学检测：,循环内皮细胞检测：于SAH后3天、2周取静脉血标本，用因子相关抗原免疫荧光染色检测循环内皮细胞（circulatingendothelialcell,CEC），以反映血管内皮细胞的损伤。,于SAH后3天、2周，取动物脑底Willis动脉环及其相连的动脉制备匀浆液PCR、Westernblot等检测脑动脉匀浆液中一氧化氮合酶（nitricoxidesynthase,NOS）mRNA和蛋白含量硝酸酶还原法测定NO含量放射免疫分析法测定cGMP含量,二、内源性调节性T细胞移植SAH后神经功能预后的改善作用,感觉评分,运动评分,认知功能障碍评估,数据表示为动物