腺相关病毒介导PSA基因转染DC的实验研究.pptVIP

腺相关病毒介导PSA基因

转染DC的实验研究;AdaptiveandInnateImmuneSystems;Ag/MHCClassII;DendriticcellsexpressavarietyofcellsurfacemoleculeswhichplaykeyrolesintheirinteractionswithTcells.TheMHCmoleculespresentantigenicfragments(peptides)toTcellsinaformthatcanberecognizedinahighlyspecificfashionbytheappropriateTcellreceptor,whichisuniqueforeveryTcellandeveryantigen.Thecostimulatorymolecules(CD80andCD86)provideasecond-signalthatamplifiestheantigenrecognitionprocess.AccessorymoleculeslikeICAMsandLFAsenhancethephysicalinteractionbetweenTcellsandDCs.;前列腺癌细胞大量表达PSA;Loadingdendriticcellswith

theantigen;DisadvantagesofProtein/peptideloading

T1/2ofproteins/peptidesisveryshort.Allarerapidlylost.

2)MostprotocolsforgeneratingcytotoxicTlymphocytes（CTL）take3-5weeks.

;Retroviruses(Retrov)(8kb,RNA,enveloped)

1)TransgeneinactivationbyDNAmethylation．

2)Causescancer:

proviralchromosomalintegrationactivatesproto-oncogene．

3)Retrovirusvectorsareeasilydestroyed,easilyinactivated.

;Adenoviruses(Ad)(35kb,DNA,

non-enveloped)

-arehighlyimmunogenic,causinginflammationandcelldeath，thatisthenon-therapeuticimmuneresponse

-cannotinfectnon-dividingcells（Ｇ０）

;Adeno-AssociatedVirus(AAV)(5kb,DNA,non-enveloped)

non-pathogenic

lowimmuneresponse

chromosomalintegration

longtermexpressioninvivo

infectdividingandnon-dividingcells;;Ag/MHCClassII;ProtocolofDendriticCellsGeneration;rAAVinfection;day3

day5

day7;;;;;;;;51Cr释放法检测CTL对靶细胞杀伤的抗原特异性;51Cr释放法检测CTL对靶细胞杀伤的MHCI类限制性;研究结论

成功构建了AAV/PSA质粒，其能够高效转染DC；

AAV/PSA基因成功整合入DC染色体内；

DC内成功表达PSA蛋白；;

AAV/PSA转染DC后，仅1周即可诱导得到大量、高活性的CTL；

所获得CTL对PSA阳性的前列腺癌细胞具有特异、高效的细胞毒杀伤效应。;Thankyou!