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1、一、BEBM培养基（ATCC）(1个)1.AfricanDustStormsReachingPuertoRicanCoastStimulatetheSecretionofIL-6andIL-8andCauseCytotoxicitytoHumanBronchialEpithelialCells(BEAS-2B)BEAS-2BcellswereobtainedfromtheAmericanTypeCultureCollection(ATCC,Manassas,VA,USA;CatNoCRL-9609).Cellswe
2、reculturedaccordingtoATCCprotocols,maintainedinkeratinocytebasalmedium(KBM-2,Lonza,Walkersville,MD,USACatNoCC3103)andsupplementedwithkeratinocytegrowthmedium(KGM-2SingleQuots;Lonza,Walkersville,MD,USA;CatNoCC4152).Cellswereusedatpassages44-59andmaintainedat37°
3、Cinahumidifiedatmosphereof5%CO2二、DMEM（11）1.Diatom-DerivedPolyunsaturatedAldehydesActivateCellDeathinHumanCancerCellLinesbutNotNormalCellsInanindependentexperiment,A549cells(2×103cellswell-1)wereseededina96-wellplateandkeptovernightforattachment.Thenextdaytheme
4、diumwasreplacedwithfreshmediumwiththreeconcentrations(2,5and10µM)foreachofthreePUAs(DD,OD,andHD,Sigma-AldrichInc.,Milano,Italy)testedandwithcaspase-3Inhibitor(C30H41FN4O12,sc-3075,SantaCruz)at9.7µM;cellswereallowedtogrowfor24,48and72h.Afteraldehydetreatment,vi
5、ablecellswereevaluatedasdescribedbelow.TheBEAS-2B(ATCCCRL-9609)lung/brunchnormalepithelialcelllinewasmaintainedinDMEM(Dulbecco'smodifiedEagle'smedium)supplementedwith50%fetalbovineserum(FBS),100unitsml−1penicillinand100µgml−1streptomycin.Cellswereincubatedina5
6、%CO2humidifiedchamberat37°Cforgrowth.BEAS-2B(2×103cellswell−1)wasseededina96-wellplateandkeptovernightforattachment.Thenextdaythemediumwasreplacedwithfreshmediumwiththreeconcentrations(2,5and10µM)foreachofthreePUAs(DD,OD,andHD,Sigma-AldrichInc.,Milano,Italy)te
7、sted;cellswereallowedtogrowfor24,48and72h.Afterincubation,thesupernatantwasremovedandadherentcellswereexaminedforviability.2.GeneExpressionProfileandToxicEffectsinHumanBronchialEpithelialCellsExposedtoZearalenoneHumanbronchialepithelialBEAS-2Bcellline[19](from
8、theAmericanTypeCultureCollection,ATCC)wasculturedinDulbecco'sModifiedEaglesMedium(DMEM)supplementedwith10%(v/v)fetalbovineserum(FBS)and100U/mlofpenicillinand10µg/mlofstreptomycin.A
