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The Th1/Th2/Th17/Treg paradigm induced by stachydrine hydrochloride reduces uterine bleeding in RU486-induced abortion mice

Author(s): Li X, Wang B, Li Y, Wang L, Zhao X, et al.

Abstract

Ethnopharmacologicalrelevance:TheTh1/Th2/Th17/Tregparadigmplaysanimportantroleinachievingmaternal-fetalimmunotoleranceandparticipatesinRU486-inducedabortion.ExcessiveuterinebleedingisthemostcommonsideeffectofRU486-inducedabortion;however,itsetiopathogenesishasnotbeenfullyunderstood.Therefore,elucidatingthecorrelationbetweentheTh1/Th2/Th17/TregparadigmandthevolumeofuterinebleedingmayoffernoveltherapeutictargetforreducinguterinebleedinginRU486-inducedabortion.Leonurussibiricushasbeenusedinclinicstoreducepostpartumhemorrhagewithlowtoxicityandhighefficiency;however,theeffectiveconstituentsandtherapeuticmechanismhavenotbeendescribed.StachydrinehydrochlorideisthemainconstituentofL.sibiricus,thereforeL.sibiricusisregardedasacandidateforreducinguterinebleedinginRU486-inducedabortionmicebyregulatingtheTh1/Th2/Th17/Tregparadigm.

Aimofthestudy:ThepurposeofthisstudywastodeterminetheTh1/Th2/Th17/TregparadigminuterinebleedingofRU486-inducedabortionmiceandtoelucidatetheimmunopharmacologiceffectsofstachydrinehydrochlorideoninducingtheTh1/Th2/Th17/TregparadigminreducingtheuterinebleedingvolumeinRU486-inducedabortionmice.

Materialsandmethods:ToinvestigatetheTh1/Th2/Th17/TregparadigminuterinebleedingduringRU486-inducedabortionmice,pregnantBALB/cmiceweretreatedwithhigh-andlow-doseRU486(1.5mg/kgand0.9mg/kg,respectively),andtheserumprogesterone(P(4))proteinlevel,uterinebleedingvolume,andproportionsofTh1/Th2/Th17/Tregcellsinmiceatthematernal-fetalinterfaceweredetectedbyELISAassay,alkalinehematinphotometricassay,andflowcytometry,respectively.TodeterminetheregulatoryeffectofstachydrinehydrochlorideontheTh1/Th2/Th17/Tregparadigminvitro,splenocytesofnon-pregnantmicewereseparatedandtreatedwithP(4,)RU486,and/orstachydrinehydrochloride(10(-5)M,10(-4)M,and10(-3)M,respectively).TheproportionsofTh1/Th2/Th17/Tregcellswereanalyzedusingflowcytometry.ToevaluatetheeffectofstachydrinehydrochlorideinreducinguterinebleedingviaregulationoftheTh1/Th2/Th17/Tregparadigm,pregnantmiceweretreatedwithRU486(1.5mg/kg)and/orstachydrinehydrochloride(2.5mg/kg,5mg/kg,and10mg/kg).TheserumP(4)level,uterinebleedingvolume,andproportionsofTh1/Th2/Th17/Tregcellsatthemicematernal-fetalinterfaceweredetected.Moreover,theproteinlevelsofcytokines(IL-12andIL-6)andthecytokinesolublereceptorswereanalyzedbyELISAassay,andthemRNAexpressionoftranscriptionfactors(T-bet,GATA-3,RORγt,andFoxp3)weredetectedbyRT-PCRassay.

Result:Th1-andTh17-biasedimmunitywasobservedinRU486-inducedabortionmice.ThevolumeofuterinebleedingduringRU486-inducedabortionwasnegativelyrelatedtotheproportionsofTh1andTh17cells,aswellastheratiosofTh1:Th2cellsandTh17:Tregcells,andpositivelyrelatedtotheproportionsofTh2andTregcells.StachydrinehydrochloridepromotedtheproteinexpressionofIL-12andIL-6,aswellasthemRNAexpressionofT-betandRORγt,whileinhibitingthemRNAexpressionofGATA-3andFoxp3.Therefore,theTh1/Th2/Th17/TregparadigminRU486-inducedabortionmiceshiftedtoTh1andTh17afterstachydrinehydrochlorideadministration.ThevolumeofuterinebleedingduringRU486-inducedabortionwasreducedsignificantlyafterstachydrinehydrochlorideadministration.

Conclusion:TheTh1/Th2/Th17/TregparadigmiscloselyrelatedtothevolumeofuterinebleedinginRU486-inducedabortionmice.TheTh1/Th2/Th17/TregparadigminducedbystachydrinehydrochloridecontributedtothereductioninuterinebleedinginRU486-inducedabortionmice.

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