血红素加氧酶-1基因转染在大鼠脑缺血再灌注损伤中的作用

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1、Protectiveeffectsofhemeoxygenase一1genetransfectiononcerebralischemiareperfusioninjuryinratsAbstract：bjective：Toobservetheeffectsoncerebralischemia-reperfusion(CIR)injuryaftertransfectionofhemeoxygenase-1geneinratfordiscussingthetherapyprospectofregulatingthehemeoxygenasegenetostrokeinpractice．M

2、ethods：AdenovirusVectorProduction：H0一lgenewasclonedbyusingthereverse-transcriptionpolymerasechainreaction(RT-PCR)methodfromthetotalRNAextractedfi'omratspleencells．AccordingtopublishedGeneBanksequenceofratHO一1(J02722)theprimersequencewasdesigned．AdenoviralvectorscarryingaratHO-1genewereconstruct

3、edbyAdenovirus-bacterialrecombinationsystem(AdEasy)．TheviraltitersofAd．Ho．1andAd．GFPwere2．5×1011pfu／mland2．0x1011pfu／m1．MaleSpragueDawley(SD)rats(n=78)wererandomlydividedintofourgroups：shamgroup(Sri)，vehiclegroup(v)，emptyadenovirusvectorgroup(Ad)andAd—HO—Itransfectiongroup(18eachgroup)．RatsofVA

4、dandAd—HO一1groupswererespectivelyinjectedwith20Ixlsaline，20Ixlsalinecontaining1mofemptyvectoradenovirusor1山ofrecombinantHO—ladenovirusthroughtherightventricleforthreedaysbeforeischemia．InjectionsiteWasreferencetoPaxinosandWatsonratbrainstereotacticatlas．Then,·cerebralischemia-reperfusionwasesta

5、blishedbymiddlecerebralarteryocclusion(MCAO)accordingtoamethodZea--Longadescribed．Underhalothane4physiologicallevels(36．5_37．5oC)usingathermalblanket．Regionalcerebralbloodflow(rCBF)wasmeasuredusingalaserDopplerflowmeterprobe．Neurologicaldeficitswereevaluatedat24hoursafterischemiausingthescoring

6、systemreportedbyGarcia．Thenthebraintissuewasremovedandwassectionedintoslices，infarctvolumewasevaluatedwith2,3，5一triphenyltet·razoliumhydrochloride(TTC)staining．Infarctvolumeswereexpressedaspercentageofthecontralateralhemisphere．Twohoursafterischemia，braintissuewashomogenized．Thehomogenateswereu

7、singtodetectneuronalcaspase一3activity．24hafterischemia,braintissuesectionswereusingtodetectapoptoticcellsbyTUNELstain，GFPexpressionWaSobservedbyfluorescencemicroscope，andcalculatedthetransfectionrate．TheexpressionsofHO一1inbraintis