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1、EXPRESSIONOFNEUTRALGLYCOSPHINGOLIPIDSINTHEBRAINANDSPLEENOFMICELACKINGTNFRECEPTOR1AnitaMarkotić1*,AnaMarušić21DepartmentofBiochemistry,SplitUniversitySchoolofMedicine,Split,Croatia2InstituteforBrainResearchandDepartmentofAnatomy,ZagrebUniversitySchoolofMedicine,Zagreb,Croatia*Correspondingauthor
2、AnitaMarkotićSplitUniversitySchoolofMedicineŠoltanska221000SplitCROATIATelephone:385.21.557938Telefax:385.21.557625E-mail:markotic@bsb.mefst.hrRunningtitle:NeutralglycosphingolipidsinTNFR1knockoutmiceABSTRACTWefounddifferentexpressionofneutralglycosphingolipids(GSLs)inthebrainandspleenofmicelac
3、kingthegeneforthetumornecrosisfactor-areceptorp55(TNFR1).NeutralGSLsoftheganglio-,globo-,andneolacto-seriesweredeterminedinthetissuesofhomozygous(TNFR1-/-)andheterozygous(TNFR1+/-)animalsbyhighperfomancethinlayerchromatography(HPTLC)overlayimmunostainingwithspecificantibodies.Thespleenofhomozyg
4、ousTNFR1knockoutmicelackedglucosylceramidesubstitutedwithpalmiticacid,GlcCer(C16)andshowedseverereductionintheexpressionofGlcCer(C16).Inaddition,gangliotetraosylceramide,Gg4Cer,substitutedwithpalmiticacid,Gg4Cer(C24),andglobotetraosylceramide,Gb4Cer,weredown-regulatedinTNFR1-/-spleenincompariso
5、nwithheterozygouscontrol.Thebrainofbothgroupsofanimals(TNFR1-/-andTNFR1+/-)didnotexpressdetectablelevelsofGg4Cer,Gb5CerandGb4Cer,butthebrainofTNFR1knockoutmiceexpressedabundantglobotriaosylceramide,Gb3Cer,comparedtonoexpressionincontrolheterozygousmice.nLcCer(C24)wasslightlyhigherexpressedinthe
6、brainofTNFR1-/-micecomparedtocontrolanimals.ThisstudyprovidesinvivoevidencethatTNFsignalingviatheTNFR1isinvolvedintheacquisitionofadistinctGSLassemblyinthebrainandspleen.DecreasedtrendofpalmiticacidsubstitutioninsomeofGSLsfractionspointsatpossibleroleofpalmitoylationinthesignaltransductionpathw
7、ayofTNFR1.Keywords:neutralglycosphingolipids,HPTLCimmunostaining,geneknockoutmice,TNFreceptor1,brain,spleenINTRODUCTIONTNFR1playsanimportantroleinthemorphogenesisofthelymphoidorgansandgenerationofimmuneresponsetoantigens(1,2),aswe
