人补体受体1型功能域scr1-3基因的克隆表达及对急性脑缺血再灌注损伤的保护作用

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1、第二军医人学硕+学位论文Cloning，ExpressionofTheSCRl·3FunctionalDomainofHumanComplementReceptorType1anditsProtectiveEffectsonAcuteCerebralIschemiaandReperfusionInjury术AbstractIschemiccerebrovasculardiseaseandcerebralreperfusionInflammatoryinjuryhasbeenoneofthemajordiseaseswhichhaveputaseriousth

2、reattohumanlifeandhealthintheworldwide．ItsPathogenesisiscomplex，andresearcheshaveshownthattheinflammatoryeffectivefragmentsproducedbycomplementactivationarethemaininitialfactorswhichcausecerebralischemiaandreperfusioninflammatoryinjury．Inthethreecomplementactivationpathwayswhichare

3、knownasclassic，mannose—bindinglectin(MBL)andalternativepathwayrespectively，althoughtheyareactivatedatdifferentstartingpoints，theycoverageatC3andformC3／C5convertase．SoamoleculethatcancombinewithC4b，C3bandinhibittheactivityofC3／C5convertingenzyme，wouldblocktheover—generationofpro—inf

4、lammatorycomplementfragments，suchasC3a，C4a，C5aandC5b一9，eventuallypreventingtheoccurrenceanddevelopmentoftheCI／Rdamageattheinitialstage．TheSCR1—3functionaldomainofhumancomplementreceptortype1whichhasacapacityofbindingC4bandinhibitingtheactivityofC3／C5convertingenzyme，wouldreducethee

5、xcess—generationofC3a，C5aandC5b一9incontrolofexcessiveactivationofcomplementsystemattheearlystage，andpreventtheoccurrenceanddevelopmentofCI／RinjuryInthisstudy，thetotalRNAwaspreparedfromhumanperipheralblood，andthegeneencodingsequenceofCR1一SCR1—3wasobtainedbyRT-PCRamplification．Afterr

6、ecombinantexpressionplasmidofpPIC9k—-CR1·-SCR1—·3wasconstructedandconformedtobecorrect．ThenitwaselectrotransportedintoPichiapastorisforproteinexpressionFinally，thetargetproteinwasanalyzedbySDS—PAGEandWestern—Blot．NI—NTAagarose+Thisworkwassupportedbygrantsfrom：(1)NationalNaturalScie

7、nceFoundationofChina(30471723)(2)NaturalScienceFoundationofChongqing(2007BB5011)4第三军医大学硕士学位论文metal—chelate—affinitychromatographywasusedforproteinpurification．Forfurtherstudy,ComplementcontrolactivityoftargetproteinwasinvestigatedbySRBChaemolyticassayinvitroanditsprotectiveeffectso

8、nacutecerebralischemiarepe