基因多态性与冠心病

基因多态性与冠心病第1页/共52页CoronaryHeartDiseaseisstilltheNo.1killerintheworld.第2页/共52页GeneticsEnvironmentalLifestyleThepreventionofCHDisbasedonthecontrolofseveralfactorsassociatedwithadiseaseorclinicalconditionandsuspectedtoplayapathogeneticalrole,definedasriskfactors.TheriskfactorsofCHDincluded:第3页/共52页AgeClottingfactorsSexFibrinolyticfactors

HypertensionHyperhomocysteinaemia

SmokingInflammation

factorsDiabetesEndotheliumfactorHyperlipidaemia

NutritionfactorObesity

Post-menopausalstatus

GeneticfactorsEmergedCHDRiskFactors第4页/共52页Butonlyin5%ofhereditaryCHD,thegenebackgroundwasclear.Intheothers,eachgeneticfactorplayedaminorroleinoccurrenceanddevelopmentofthedisease.Raremutations(e.g.,intheLDLRandAPOEgenes)mayhaveamajoreffect,whereasgenesbelongingtonormalpolymorphismhaveonlyamoderateeffect.Butevengeneswithonlyaslighteffectcanbeclinicallyimportantincombinationwithothergenes.Theimportanceofpolymorphismanalyseswillincreasesignificantlyinthenearfuture.第5页/共52页What’snormalpolymorphism?Theoccurrenceinapopulation(oramongpopulations)ofseveralphenotypicformsassociatedwithallelesofonegeneorhomologsofonechromosome.Theoccurrencetogetherinthesamepopulationofmorethanonealleleorgeneticmarkeratthesamelocuswiththeleastfrequentalleleormarkeroccurringmorefrequentlythancanbeaccountedforbymutationalone.第6页/共52页PolymorphismandMutationTheyarebothsinglenucleotidepoly-morphism，SNP.polymorphism－－normalphenotypemutation －－diseasepolymorphism－－moremutation－－less第7页/共52页Cardiovasculardiseaseiscomplexasaconsequenceofpleiotropy.Theseincludedenvironmentalandgeneticsfactors.Genepolymorphismplayedanimportantroleintheoccurrenceanddevelopmentofcardiovasculardisease.Anditcanbeappliedontheprediction,diagnosis,treatmentandprognosis.第8页/共52页1.ThegenepolymorphismsasindependentriskpredictorsAnHphIpolymorphismintheE-selectingeneisassociatedwithprematurecoronaryarterydisease.ApoEgenepolymorphismisrelatedtocoronaryheartdisease.E23KpolymorphisminKCNJ11genehasrelationshipswithcoronaryheartdisease.第9页/共52页EverygenevariantsthatcontributetoCHDliketinyweightsinbalance.第10页/共52页E-selectinbelongstoafamilyofstructurallyrelated"selectin'moleculesincludingE-,P-andL-selectinandparticipatesintheendothelial-leukocyteadhesion.ExperimentsusingE-andP-selectin-double-knockoutmicesuggestthatE-andP-selectintogetherplayanimportantroleinbothearlyandadvancedstagesoftheatheroscleroticlesiondevelopment.SeveralpolymorphismsintheE-selectingenehavebeenidentifiedasnewriskfactorsfortheearlyatherosclerosis.1.1TheG98/TpolymorphisminE-selectingeneandCHD第11页/共52页ThetransversionofG98TmutationabolishestheHphIrecognitionsite.NT92TTGGGTGAAAAG103NT92TTGGGTTAAAAG103HphIHphI第12页/共52页bp332194138Kb1.350.630.310.190.12ThePCRproductwasdigestedbyHphIandseparatedon2%agarosegelelectrophoresis.第13页/共52页PCRamplificationofthegenomicDNA,subcloningandDNAsequencingwerecarriedout.9898GGgenotype:TTgenotype:第14页/共52页

TableFrequencyoftheE-selectinG98TmutationintheangiographicallydocumentedprematureCADandcontrols(Theoriginalpopulation:allmalesaged50yr.old,allfemalesaged60yr.old;thesubset:Allmalesaged45yr.old,allfemalesaged55yr.old)a:Incontrol,32males,39females;inCAD,51males,42females.b:Chi-squarestatisticalanalysiswasdoneusingSigmaStat(ver.2.0,SPSSInc.,Chicago,IL).c:Incontrol,21males,29females;inCAD,28males,23females.d:Include18heterozygotes(GT)andtwohomozygotes(TT).

NG-allele(%)T-allele(%)TotalallelesThepopulationa

Control71128(90.14)14(10.93)142CAD93160(86.02)26(13.98)186Pb NSThesubsetc:Control5090(90.00)10(10.00)100CAD5180(78.43)22d(21.57)102Pb <0.05第15页/共52页

variableOddsRatio(95%,CI)P

G98T3.58(1.20-10.67)=0.022S128R4.11(1.24-13.56)=0.020TC0.99(0.97-1.07)NSTG1.00(0.99-1.01)NSLDLB1.02(0.99-1.05)=0.071Smoke,Y/N5.87(1.84-18.75)=0.003

MultivariateLogisticRegressionAnalysis第16页/共52页TheATP-sensitivepotassiumchannel(KATP)werecomplexesoftwosubunits,aregulatorysulfonylureareceptor(SUR)andanATP-sensitiveandpore-forminginwardlyrectifyingK+channel(Kir6.X).TheKir6.XsubunitsincludingKir6.1andKir6.2hadtwotransmembranedomainsandformthepore,conferringchannelsensitivitytoATPandothercellmetabolitessuchasADP.TheGtoAmutationintheKir6.2,theATP-sensitivepotassiumchannelsubunit,resultedaGlutamate(E)toLysine(K)substitutionatcodon23,andtheAallelewasshowntohavearelationshipwithhighrisktotype2Diabetesinpreviousstudy.1.2TheE23KpolymorphisminKCNJ11geneandCHD第17页/共52页第18页/共52页

Group

Genotype(%)

Allele(%)

GGGA+AA

GACHD(n=119)Controls(n=101)χ2PvalueOR(95%CI)50(42.0)69(58.0)29(28.7)72(71.3)4.2020.0401.799(1.023~3.163)*151(63.4)87(36.6)115(56.9)87(43.1)1.9400.1641.313(0.895~1.927)***:

ThefrequencyofGGgenotypewascomparedwiththatofGA+AAgenotype;**:ThefrequencyofGallelewascomparedwithAallele.Table.GenotypeandAlleleFrequenciesoftheE23KPolymorphismofKir6.2GeneinCHDPatientsandControls第19页/共52页1.3

TheApolipoprotein(Apo)EgenepolymorphismandCHDApolipoproteinEgeneislocatedonchromosome19q13.2.ApoEplaysacriticalroleintheformationofverylowdensitylipporotein(VLDL)andchylomicrons.Genetically,ApoEispolymorphicisoformsofproteins,E2,E3,E4respectively.TheApoEallelesmodulatetheriskforCHD,cerebralaheroscerosisandAlzheimer’sdisease.

第20页/共52页Therewerethreeallelesandsixgenotypes.NH2112158COOH25’TGCTGC3’CysCys35’TGCCGC3’CysArg45’CGCCGC3’ArgArgHhaI

HhaIHhaI第21页/共52页ApoEPAGE/RFLPpatternMε4/4ε3/3ε2/2ε3/2ε4/3ε4/2第22页/共52页Thereisasignificantrelationshipbetween“+”alleleoflowdensitylipoproteingene,ε4andhightotalcholesterol,LDLcholesterollevels.ThecholesterollevelsofindividualswithLDL-RAvaⅡ(-/-)andNcoI(-/-)genotypewerelowerthanthosewithLDL-RAvaⅡ(+/+)andNcoI(+/+)genotypes.1.4TheApolipoprotein(Apo)EandlowdensitylipoproteingenepolymorphismandCHD第23页/共52页TheresultspresentedheredonotsuggestapositiveassociationbetweenHindIII,TaqI,MspI,and3076A/CpolymorphismsinFNgeneandCHD.Wedidn'tfindanyrelationshipbetweenfourpolymorphisminfibronectingeneandCHD.Butanondifferencedoesnotimplyanoneffect.ButwefoundpFNlevelsincirculationweredecreasedsignificantlyinpatientswithCHD.1.5FibronectingeneandCHD第24页/共52页Figure.MspI(a),TaqI(b),(3076A/Cc),andHindIII(d)digestionsofthePCRproductshowingdifferentgenotypes.Thesizeofeachfragmentisindicatedonthegel.M1：A100-bpDNAladder,MBIFermentas.M2：A50-bpDNAladder,MBIFermentas.P:PCRproducts第25页/共52页TableGenotypeandAlleleFrequenciesofthePolymorphismsoftheFNGeneinCHDPatientsandControls

GenotypeAllelePvalueMspIinintron26PhenotypeCCCDDDCD0.527CHD(n=109)62(56.9)47(43.1)0(0)171(78.4)47(21.6)Controls(n=123)75(61.0)48(39.0)0(0)3076A/Cinexon20PhenotypeAAACCCAC0.95CHD(n=109)96(88.0)12(11.0)1(1.0)204(93.6)14(6.4)Controls(n=123)108(87.8)15(12.2)0(0)第26页/共52页FigThesequencereports.a:genotypeAAfor3076A/C;b:genotypeCCfor3076A/C(Eco81I:CC↓TNA↑GG);c:genotypeCCforMspI﹡(MspI:C↓CG↑G)﹡allelewasdesignatedasC(CCAG),D(CCGG)fortheMspIpolymorphism第27页/共52页1.6Interleukin-1βgeneandCHDInterleukin-1β(IL-1β)belongstoafamilyofcytokinesincludingIL-1,IL-4,IL-8,IL-10,andIL-13.IL-1βplaysadominantroleinseveralimmunereactionsIL-1βalsowasfoundtoassociatetoatheroscleroticeventsinvitroandinvivo.第28页/共52页PolymorphismanalyzeCCCTTTPCR第29页/共52页ComparisonofdistributionofIL-1β3954C/TgenotypesbetweenCHDgroupandcontrolgroupgroupnGenetype(%)AllelefrequencyCCCTTTCTcontrol1300.9310.0690.0000.9650.035CHD780.897

0.0900.0130.9420.058(P>0.05)第30页/共52页TheconcentrationsofC-reactiveproteinindifferentgenotypegroupsGenetypenCRPmedianInter-quartilerangeCC1212.05*0.67-3.64*CT93.981.69-6.68*(P<0.05)第31页/共52页TheTaqⅠpolymorphismofIL-1βwasassociatedwiththeconcentrationsofCRPinnormalpeopleTheTaqⅠpolymorphismofIL-1βwasnotassociatedwithwithCHD第32页/共52页CRP,Thefirstacute-phaseproteintobedescribed,itsplasmaconcentrationincreasesduringinflammatorystates.Recently,CRPmighthaveanimportantroleinthepathogenesisandpredictionofCHD.1.7CRPgeneandCHD第33页/共52页PolymorphismanalysisFigure1-1Determinationofthe+1444C/TpolymorphismintheCRPgenebyPCR-RFLP.LaneM：DNAMarker;LaneA：PCR

product;LaneB：homozygousCC;LaneC：heterozygous

CT;第34页/共52页PolymorphismanalysisFigureThechromatogramofhomozygousCC第35页/共52页Comparisonofdistributionof+1444C/TgenotypesandallelesbetweenCHDgroupandcontrolgroupGroupsnGenotypeDistribution(%)AlleleDistribution(%)CCCT+TTCTCHD128114(89.1)14(10.9)242(94.5)14(5.5)Control119107(89.9)12(10.1)226(95.0)12(5.0)20.0480.045P0.8270.832OR(95%CI)0.913(0.404¬2.063)*0.918(0.416¬2.027)△

*:genotypeCCvs

CT+TT；△：alleleCvsT第36页/共52页TheconcentrationsofC-reactiveproteinindifferentgroupsGroupsnmedianInter-quartilerangeCHD1281.210.63～2.58CC1141.210.67～2.48CT+TT141.520.51～3.73Control1190.770.59～1.22CC1070.750.59～1.15CT+TT121.420.66～3.05第37页/共52页The+1444C/TpolymorphismofCRPwasassociatedwiththebasalconcentrationsofCRPinnormalpeople.The+1444C/TpolymorphismofCRPwasnotassociatedwith

CHD.第38页/共52页1.8ConclusionforthestudyonpolymorphismsaspredictorsofCHDWefoundthepolymorphismsofE-selectin,ApoEgeneandKCNJ11genewererelatedtoCHDdisease.ThepolymorphismsinLDL-RgeneandApoEgeneeffectedtheleveloflipid.ThepolymorphismsofIL-1βandCRPgenesinfluencedtheCRPbaseline.Wedidn'tfindanyassociationbetweenpolymorphisminfibronectingeneandCHD.第39页/共52页Patientswithanacutemyocardialinfarctionareofhighrisktodevelopischemia–inducedventriculararrhythmias,leadingtosuddencardiacdeathinaboutonethirdofallAMIpatients.Theindividualsusceptibilitytoischemia-inducedarrhythmiasmaybemodifiedbypolymorphismsingenesencodingionchannels.A.JeronstudiedtheKir6.2gene.OpeningoftheKATPchannelduringischemiaresultsinactionpotentialshorteninginvariousstudiesandmaythereforeinfluencetheoutcomeofAMIpatients.Howevertheydidn’tfindanysignificantinfluenceofKir6.2genepolymorphismontheriskofSCDinpatientswithCHD.ButtheyidentifiedtwonovelmissensemutationsinahighlyconservedregionoftheKir6.2gene.2.Genepolymorphismandprognosis第40页/共52页From:Johnson

J.A.TRENDSinGenetics2003

Vol.19No.11:660-6663.Genepolymorphismandpharmacogenomics第41页/共52页Thenewpharmacogeneticsusespowerfulexperimentalanddata-handlingtechniquesinDNAanalysistodiscoverandassembleacomprehensivelistofthevariationswithinthehumangenome–specifically,SNPs–andthendefinescomplexgeneticprofilesoftheseSNPsthatpredicttheuseofneworexistingtherapeuticagentswithmaximalefficacyandminimaltoxicity.第42页/共52页Agenetictestforcertainsinglenucleotidepolymorphisms(SNP)willpredictyouthat:Youshouldsufferasevereadversereactiontoit.Youareexpectedtoshownanexcellentresponsetoadifferentmedicationwithlittlechanceofsideeffects.Thisisthepromiseofpharmacogenetics—

Theoptimizationofdrugtherapybasedontheindividualgeneticprofile.第43页/共52页TheHMG-CoAreductaseistherate-limitedenzymeinthebiosynthesisofcholesterol.

Statins,theHMG-CoAreductaseinhibitors,arewidelydesignedtoreducedenovocholesterolbiosynthesis.TheefficacyandtoxicityofStatinsaredifferentindifferentindividuals.ForExample:第44页/共52页Recently,investigationintherelationshipofstatinseffectsandgenepolymorphismsrelatedtothelipoproteinmetabolismhasbeendeveloped.ThesegenesincludedtheapolipoproteinEgene,hepaticlipasegene,lipoproteinlipasegeneandCETPgene.第4