Ontogeny and regulation of matrix metalloproteinase activity in the zebrafish embryo by in vitro and in vivo zymography

摘要

Remodelingoftheextracellularmatrix(ECM)duringdevelopment,angiogenesis,woundhealing,tumormetastasis,andothermorphogeneticprocessesdependsontheexquisitelyregulatedactivitiesofmatrixmetalloproteinases(MMPs).Yetverylittleisknownabouttheactivitypatternsoftheseproteasesinvivo.WehaveemployedfluorescentMMP-substrates,bothinvitroandinvivo,tocharacterizepatternsofMMPactivityinthezebrafishembryo.QualitativelysimilarpatternsofdegradationaredetectedusingnativeTypeIorTypeIVcollagensubstrates,suggestingthatmultipleMMPsarebeingregulatedconcomitantly.MMPactivityisobservedprimarilyinECM-richstructurespredictedtobeundergoingactiveremodeling,suchastheperichordalsheathandsomiteboundaries.PatternsofTypeIandTypeIVcollagenhydrolysisaresimilar,butnotidenticalinembryosofanygivenstage.ConventionalgelatinzymographyshowsMMPspresentinembryosasearlyas3-somites(11h)andourinvivoassaysdetectTypeIVcollagendegradationatsomiteboundariesasearlyas4-somites(11.5h).However,weareunabletodetectsignificantinvitroactivityusinghomogenatesmadefromembryospriortoPrim-16(31h).Mixedlysateassaysdemonstratethatthisistheresultofendogenousinhibitorspresentinearlyembryos,suggestingamodelofmatrixremodelingregulatedbyspatiallyheterogeneousMMPinhibition./p/div