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1、MethodsinMolecularBiologyMethodsinMolecularBiologyTMVOLUME142TTransformingransformingGrowthGrowthFactor-BetaFactor-BetaProtocolsProtocolsEditedbyEditedbyPhilipH.HowePhilipH.HoweHUMANAPRESSMeasuringTGF-}Growth11InVitroAssaysforMeasuringTGF-}GrowthStimulationandInhibitionMaryanneEdensand
2、EdwardB.Leof1.IntroductionTransforminggrowthfactors(TGFs)wereinitiallyisolatedfromthecondi-tionedmediumoftransformedcelllinesthroughtheirabilitytostimulateanchorage-dependentcellstoformcoloniesinsoftagar(1,2).Theabilitytoproliferateinananchorage-independentmannerisstilloneofthebestinvi
3、trocorrelateswithtumorigenicity.Subsequentstudiesdemonstratedthatthegrowth-promotingactivityintheconditionedmediumconsistedoftwouniquepeptides,TGF-
4、andTGF-}(3–5).Dependingontheindicatorcelllineused,soft-agarcolonygrowthcouldoccurwhenTGF-
5、andTGF-}(i.e.,NRKcells)orTGF-}alone(i.e.,AKR-2Bc
6、ells)wereaddedtotheserum-containingmedium(6,7).ThisreviewwillfocusonTGF-}andcellularsystemscapableofrespondinginvitrotoitsgrowthmodulatoryactivityindependentofaddi-tionalfactors.Transforminggrowthfactor-}isa25-kDahomodimericproteinrepresenta-tiveofafamilyofmoleculescapableofregulatingc
7、ellgrowthanddifferentia-tion(8–10).ThreemammalianTGF-}isoforms,TGF-}1,TGF-}2,andTGF-}3,havebeenisolated(11).Althoughthesemoleculeshavesimilarandoverlap-pingactivityinthemajorityofinvitroassays,theirrole(s)invivoappearstobequitedistinct(12).Thisdistinctionbecomesreadilyapparentwhentheph
8、e-notypesofTGF-}knockoutmicearecompared.Forinstance,whereasTGF-}1nullanimalsdevelopamultifocalinflammatoryresponseandwastingfollow-ingweaning,thelackofTGF-}2orTGF-}3resultsinavarietyofdevelop-mentaldefects(13,14).ThecellularresponsetoTGF-}isquitedistinct,whereasmesenchymalcellsare(inge
9、neral)growthstimulated(bothinvitroandinvivo),themajorityofFrom:MethodsinMolecularBiology,Vol.142:TransformingGrowthFactor-}ProtocolsEditedby:P.H.Howe©HumanaPressInc.,Totowa,NJ12EdensandLeofothercelltypes(i.e.,epithelial,hematopoietic)aregrowthinhibited.Itisunknownhowasinglegrowthfact
