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4RESULTS ANDDISCUSSIONTofacilitatethecomprehensionofthefindingsregardingthedietprovidedtothemaleoffspringtheresultswillbegroupedintotwostudies– StudyIEffectofFructoseconsumption versus(vs.) Tagatose.Thisanalysis willallow todetermineif theobservedeffectsarefructosespecific–StudyIIEffectofFructoseconsumptionalonevsFructoseplusCholesterolInaddition theeffectofmaternal feedingduringgestation(waterorfructose)onthecholesterolmetabolismoftheiroffspringwillbeanalysedwithineachgroup41StudyIEffectofFructoseconsumptionvsTagatose41aPlasma totalcholesteroland triglycerideslevelsTheresultsof the two-way ANOVAarepresentedin Table2Regardingtotalcholesterolconcentrationthechangesobservedseem tobedue to thedifferentdietsreceivedby themaleoffspringalthough thedifferenceisnot statistically significant(p=0097)Ontheotherhandboththematernaltreatment(M)andthediet(D)seem toaffect triglycerideslevelsbut thesedifferencesdidnotreachstatisticalsignificance(p=0054andp=0094respectively)ThusFigure10showsthedataafteraposterioriBonferronicorrectionThe totalcholesterolconcentrationin theoffspringofcontrolmothersshowsstatisticallysignificantdifferencesbetweenthecontrolandfructosegroupswhencomparedtothetagatosegroupThis wouldindicate that tagatoseandnot fructoseproducesamarkedincreasein totalcholesterollevelsHowever, whenstudying thedescendantsof fructose supplementedmothers wecanobserve thatthereareno variationsbetween thegroups which suggests that thisTable1SequenceoftheprimersusedforthecDNAamplification33cbSYBRGreenIReal-TimePCRProtocolTheCFX96TouchReal-TimePCRDetectionSystem(BioRadUSA)wasusedtoperformthisassay.Foreachsamplethefollowingvolumeswereadded1μLforwardprimer(10μM)1μLreverseprimer(10μM)10μLSYBRgreenPremixExTaq(TakaraBioIncJapan)6μLdeionisedwaterand2μLsampleTheinformationregarding theprimersusedin thepresent workcanbe foundinTable1Anagarosegelelectrophoresiswasperformedaftereachreal time-PCRexperiment to verify that theampliconobtainedmatchedtheexpectedresult(Figures8and9)34StatisticalanalysisTheassayswerecarriedoutwithsamplesfrom7animalspergroupexceptforgroupControl-Controlwhichhad8animalsResultswereexpressedasmeans±standarderror(SE)andtheywereanalysedby two-way ANOVA(Analysisof Variance)If thedifferences were statistically significant(p<005) they wereanalysedthroughaposterioriBonferronicorrectiontoidentifythevariancesignificanceThereweretwofactorsanalysed– Theeffectof fructoseintakeduringgestationby themothers(M)onthemaleoffspringStatisticallysignificantdifferencesbetweenControlMothers vsFructoseMothers willbeindicatedwithahash(#)andincase thedifferenceisalmost statisticallysignificantitwillbeindicatedwithanampersand(&)–Theeffectofthedietsreceivedbythemaleoffspringat3monthsofage(D) Thedifferencesbetweentreatmentswithinthesamemotherwillbeindicatedwithanasterisk(*)The valueofetasquared (2)determinestheproportionofthevariancethatisattributedtothemother(M)thediet(D)ortheinteractionbetweenbothparameters(M*D)All the statisticaldeterminations wereperformed with theprogramSPSS264ANALESRANFwwwanalesranfcomLaingestamaternade fructosamodulaelmetabolismodelcolesterolenrespuestaaunadietaoccidentalenladescendenciaPalomaLópezLaizElenaFausteetalAnRealAcadFarmVol88Nº3(2022)·pp257-275GenForwardPrimerReversePrimerAmpliconOriginLDLR5-CACCCAAGTTCACCTGCGCTTGCC-35-AGGACACTGTCACCGACTCCACTG-3291bpClontechMTTP5-TCTGTGGTACCGCGAGTCTA-35-GGGTACTGGGAGAACTGCAC-3165bpPrimer3SR-B15-GTTCCGTGAAGATGCAGCTGAG-35-AACCACAGCAATGGCAGGACTAC-397bp(49)Figure8AgarosegelelectrophoresisofRealTime-PCRofMTTP.Figure9AgarosegelelectrophoresisofRealTime-PCRofLDLR