In vitro identification of nonalcoholic fatty liver diseaserelated protein hnRNPM-中国期刊网

摘要 AIM:Tostudytheformationofintracellularglyceraldehyde-derivedadvancedglycationendproducts(Glycer-AGEs)inthepresenceofhighconcentrationsoffructose.METHODS:CellsofthehumanhepatocytecelllineHep3Bwereincubatedwithorwithoutfructoseforfivedays,andthecorrespondingcelllysateswereseparatedbytwo-dimensionalgradientsodiumdodecylsulfate-polyacrylamidegelelectrophoresis.Glycer-AGEsweredetectedwiththeanti-Glycer-AGEsantibody.Furthermore,theidentificationoftheproteinsthataremodifiedbyglyceraldehydeinthepresenceofhighconcentrationsoffructosewasconductedusingmatrixassistedlaserdesorption/ionizationtime-of-flightmassspectrometry(MALDI-TOF-MS).TheproteinandmRNAlevelsweredeterminedbyWesternblottingandrealtimereversetranscriptionPCR,respectively.RESULTS:Theresultsofthetwo-dimensionalgradientsodiumdodecylsulfate-polyacrylamidegelelectrophoresisindicatedagreateramountofGlycerAGEsinthesampleexposedtohighconcentrationsoffructosethaninthecontrol.ThedetectedGlycerAGEsshowedisoelectricpointsintherangeof8.0-9.0andmolecularweightsintherangeof60-80kDa.TheheterogeneousnuclearribonucleoproteinM(hnRNPM),whichplaysanimportantroleinregulatinggeneexpressionbyprocessingheterogeneousnuclearRNAstoformmaturemRNAs,wasidentifiedasamodifiedproteinusingMALDI-TOF-MS.Increasingtheconcentrationoffructoseinthemediuminducedaconcentration-dependentincreaseinthegeneratedGlycer-AGEs.Furthermore,inanexperimentusingglyceraldehyde,whichisaprecursorofGlycer-AGEs,hnRNPMwasfoundtobemoreeasilyglycatedthantheotherproteins.CONCLUSION:Theresultssuggestthatglyceraldehyde-modifiedhnRNPMaltersgeneexpression.Thischangemaycauseadverseeffectsinhepatocytesandmayserveasatargetfortherapeuticintervention.

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In vitro identification of nonalcoholic fatty liver diseaserelated protein hnRNPM

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In vitro identification of nonalcoholic fatty liver diseaserelated protein hnRNPM

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