反義RNA抑制人視網(wǎng)膜色素上皮細(xì)胞VEGF分泌的實(shí)驗(yàn)研究.pdf

1、復(fù)旦大學(xué)博士學(xué)位論文反義RNA抑制人視網(wǎng)膜色素上皮細(xì)胞VEGF分泌的實(shí)驗(yàn)研究姓名：樊瑩申請學(xué)位級(jí)別：博士專業(yè)：眼科學(xué)指導(dǎo)教師：張皙2001.5.1AntisenseRNAInhibittheSecretionofVascularEndothelialGrowthFactorinHumanRetinalPigmentEpithelialCellsAbstractRetinalneovascuIarizalioniSthemostcommo

2、ncauseofvisuaIlossintheworldtodayButunfortunately，forthisfataIdisorder，currentlyavailabletherapeuticproceduresarenotoptimalAnewkindofnOndestructivemannerwouldhavesignificanttherapeuticimplicationsUntilthesedays，investiga

3、tionhaveprovidedthatvascularendothelialgrowthfactor(VEGF)appearstobeamajormediatorofischemia—inducedocularneOVascuIarIzatlOnFrOmaclinicaIstandpointinhibitionofthesecretionofVEGFmaybeapotentiaItherapeutictargetfortheneOva

4、scularlzatlOnForthispurpose，weprobedintothefeasibilityofusinganantisenseRNAsequenceofhumanVEGFtosuppressthesecretionofthisproteinintargetcellsHVEGFl65andhVEGFl21cDNAswereamplifiedbyreverse—transcriptionPCR(RT—PCR)frOmfet

5、alhumankidneytissueandtheninsertedintotheexpressionplasmidpCR31separatelytoconstructtherecombinedplasmidsthatencodingVEGFl21orVEGFl65cDNAinanantisenseorientationThehumanretinalpigmentepithelialcells(RPE)andhumanumbilicve

6、inendotheliaIcells(HUVEC)werelransfectedwiththeseplasmidsmediatedbyliposomeSelectthestablecelIlinesThetransfectedcellswererepresentedjustastheirparentalcellsexceptforthedifferenceinsomeorganellesassociatedwiththeproteins

7、ecretionfunctionThegrowthratesofthetransfectedHUVECcellswerechangedwiththelevelofitssecretionVEGFproteinIdentifiedthepotentialfunctionoftheserecombinantsininhibitingthesecretionofVEGFintargetcellsanddownregulatingthegrow