核酶对人肺腺癌细胞多药耐药的逆转

:100853,(mdr1)mdr1mRNA880,,mdr1(pHApr21neo/mdr12Rb)A549/RRT2PCRMTT,mdr1mRNAP(P2glycoprotein,Pgp)mdr1,mdr1mRNAPgp,,200mdr1mdr1mRNA,Pgp,;;;Reversionofmultidrugresistanceinvitrooflungadenocarcinomabymdr1ribozymeCHENLiangan,LIUYouning.DepartmentofRespiratoryMedicine,GeneralHospitalofPLA,Beijing100853,ChinaAbstractObjectiveToinvestigatethepossibilityofreversionofmultidrugresistant(MDR)inlungadenocarcinoma2resistantcelllineA549/Rbymdr1ribozyme.MethodsWithGUCat880siteofmdr1mRNAselectedastargetpoint,plasmidexpressingmdr1ribozyme(pHApr21neo/mdr12Rb)wastransducedtoA549/Rbylipofectin.Theexpressionofmdr1mRNAandPgp,cellularrhodamineaccumulationandsensitivitytodoxorubicinwereexaminedinribozymetransducedandnon2transducedcells.ResultsTransductionofmdr1ribozymetoA549/Rcellsledtodecreaseinmdr1mRNAandPgpexpression;increaseinrhodamineaccumulation.Thesensitivityoftheribozymetransducedcellstodoxorubicinwas2002foldashighasthatoftheparentaldrug2resistantcells.ConclusionMDRoflungadenocarcinoma2resistantcelllineA549/Rcanbeeffectivelyreversedwithmdr1ribozymebycleavageofmdr1mRNAandinhibitionofPgpexpression.SubjectwordsLungneoplasms;Adenocarcinoma;Ribozyme;Multidrugresistance(multidrugresistance,MDR)MDRP(P2glycoprotein,Pgp),,,(ribozyme)MDR,mRNA,mRNA,,MDRMDRPgp,PCRRT2PCR,MTTSigma,G418GibcoA549/R()mdr1pHApr21neo/mdr12RbScanlon11:pHApr21neo/mdr12RbDNA,DNARNA,PCR,RT2PCR21A549/R:6,1105A549/R,70%DNA5g,10l,6h(A)(B)72h,,13;80%,80g/mlG418,,(800g/ml),2d,G418400g/ml;3,98120005223ChinJOncol,May2000,Vol22,No.331mdr1:Pgp[1,2]41(MTT):3104/ml,96,200l,;,1640(ml0.00001,0.0001,0.001,0.01,0.1,1,10,100g)200l/,48h,MTT20l(10mg/ml),4h,200l/,570nm,ID50A549/RMDR11DNA:PCRA549/Rmdr1(1),mdr1M:;Pc:;Nc:;A:A549/R1A549/Rmdr1PCR21mRNA:RT2PCRA549/Rmdr12mRNA(2),mdr1M:;Pc:;Nc:;B:;A:Ribozyme2mdr1RT2PCR31Pgp:A549/RPgp,mdr1mdr1MDR11mdr1mRNA:2,AmRNAB,AmRNA85%21Pgp:AB(A5187%,B12124%),APgpB,Pgp31Pgp:,AB(A159195%,B70136%)Pgp,Pgp,APgp,,,B41:ABID500.001g/ml0.2g/ml,AB200,,Amdr1[325]MDR,1280Pgp,,,MDR:mdr1mdr1mRNAPgpMDR,,MDRPgp,,,,mdr1mRNAMDR,Cech[6],,[7,8]RNA,RNA,,[9]:(1)RNA;(2);(3)RNA,,RNA,,;(4),GUC;(5)09120005223ChinJOncol,May2000,Vol22,No.3RNA,,,,RNA,RNAGUC,mdr1mRNA37GUC,56,3321880GUCmdr1mRNA,ATP,mdr1mRNAPgp,[8],,mdr1mRNA80%,Pgp,ID50200,RNA,RNA,,pHBApr21neo/mdr12rb,mdr1mRNA,,mdr1mRNAPgp,,A549/R1LabroilleG,BellocF,BilhouNC,etal.CytometricstudyofintracellularPgpexpressionandreversalofdrugresistance.Cytometry,1998,32:86294.2PetrizJ,GarciaLJ.FlowcytometricanalysisofP2glycoproteinfunctionusingrhodaminel23.Leukemia,1997,11:112421130.3RoninsonIB,AbelsonHT,HousmanDE,etal.AmplificationofspecificDNAsequencecorrelateswithMDRinChinesehamstercell.Nature,1984,309:6262628.4GrosP,NeriahYB,CroopJM,etal.IsolationandexpressionofacomplementaryDNAthatconfersmultidrugresistance.Nature,1986,323:7282731.5SteinWD.Kineticsofthemultidrugtransporter(P2glycoprotein)anditsreversal.PhysiolRev,1997,77:5452590.6CechT.Thechemistryofself2splicingRNAandRNAenzymes.Science,1987,236:153221539.7WalterNC,BurkeJM.Thehairpinribozyme:structure,assemblyandcatalysis.CurrOpinChemBiol,1998,2:24230.8BranchAD,KlotmanPE.Optimizingribozymeforsomaticcellgenetherapy.ExpNephrol,1998,6:78283.9BouffardDY,OhkawaT,KijimaH.Oligonucleotidemodulationofmultidrugresistance.EurJCancer,1996,32A:101021018.(:1999206207),15:36,84/min,105/68mmHg,,0152cm,,,:41621012/L,7159109/L,01533,01278,01186,130g/L,5413109/LB,X:01202,,;12;,,,:2,018cm110cm,:157011,();(),,:,,,,,,,,;,13,,:mac387(+++),2AT(++),lysozyme(+),CD20(-),CD45RO(-):,,mac387KP212ACT,2AT,lysoymeT,(:2000202218)19120005223ChinJOncol,May2000,Vol22,No.3