基于“肝脑相关”理论研究扶肝阳化痰开窍法对抑郁模型大鼠Wnt-β-catenin信号通路的影响

基于“肝脑相关”理论研究扶肝阳化痰开窍法对抑郁模型大鼠Wnt-β-catenin信号通路的影响摘要：

本研究以抑郁模型大鼠为研究对象，探究了扶肝阳化痰开窍法对该模型大鼠Wnt/β-catenin信号通路的影响以及其作用机制。结果显示，扶肝阳化痰开窍法能够显著提高抑郁模型大鼠的运动能力，增强其学习记忆能力，缓解其抑郁情绪，并且能够降低大鼠端粒酶活性和细胞凋亡水平。同时，扶肝阳化痰开窍法还能够显著抑制抑郁模型大鼠脑组织中Wnt/β-catenin信号通路的活化和表达水平。进一步研究表明，扶肝阳化痰开窍法对Wnt/β-catenin信号通路的调节作用可能与其抑制DNA甲基转移酶1表达及促进BDNF、TrkB表达有关。因此，本研究结果为扶肝阳化痰开窍法应用于抑郁治疗提供了理论依据。

关键词：扶肝阳化痰开窍法；抑郁；Wnt/β-catenin信号通路；DNA甲基转移酶1；BDNF；TrkB

Abstract:

ThepresentstudyaimedtoinvestigatetheeffectofthetraditionalChinesemedicineformula“fuyanghuatankaqiao”onWnt/β-cateninsignalingpathwayinadepressionratmodelandexploreitspotentialmechanisms.Theresultsshowedthatfuyanghuatankaqiaosignificantlyimprovedthemotorandlearningabilities,easedthedepressivesymptoms,anddecreasedtheterminaldeoxynucleotidyltransferasedUTPnickendlabeling(TUNEL)positivecellsinthebrainofdepressionrats.Furthermore,fuyanghuatankaqiaodownregulatedtheactivationandexpressionofWnt/β-cateninsignalingpathwayintheratbrain.TheregulatoryeffectoffuyanghuatankaqiaoonWnt/β-cateninsignalingpathwaymightbeassociatedwiththeinhibitionofDNAmethyltransferase1expressionandthepromotionofbrain-derivedneurotrophicfactor(BDNF)andtyrosinekinasereceptorB(TrkB)expression.Thesefindingsprovideatheoreticalbasisforusingfuyanghuatankaqiaointhetreatmentofdepression.

Keywords:fuyanghuatankaqiao;depression;Wnt/β-cateninsignalingpathway;DNAmethyltransferase1;BDNF;TrkDepressionisacommonmentaldisordercharacterizedbypersistentsadness,lossofinterestorpleasure,feelingsofworthlessnessorguilt,anddisturbedsleeporappetite.Theexactcauseofdepressionisnotfullyunderstood,butitisbelievedtoresultfromacombinationofgenetic,environmental,andpsychologicalfactors.TheWnt/β-cateninsignalingpathwayhasbeenimplicatedinthepathogenesisofdepression,asitregulatesneuraldevelopmentandplasticity,anditsdysregulationhasbeenassociatedwithmooddisorders.FuyanghuatankaqiaoisatraditionalChinesemedicinethathasbeenusedforthetreatmentofdepressionwithpromisingresults.Itiscomposedofseveralherbs,includingChaihu(RadixBupleuri),Huangqin(RadixScutellariae),andDanggui(RadixAngelicaeSinensis),whichhavebeenshowntohaveantidepressanteffectsinpreclinicalstudies.

ThepresentstudyaimedtoinvestigatetheregulatoryeffectoffuyanghuatankaqiaoontheWnt/β-cateninsignalingpathwayanditsunderlyingmechanisms.Theresultsshowedthatfuyanghuatankaqiaosignificantlyincreasedtheexpressionofβ-cateninandWnt3a,twokeycomponentsoftheWnt/β-cateninsignalingpathway,inadose-dependentmannerinculturedhippocampalneurons.Moreover,fuyanghuatankaqiaotreatmentinhibitedtheexpressionofDNAmethyltransferase1(DNMT1),akeyepigeneticregulatorofgeneexpression,whichhasbeenimplicatedinthepathogenesisofdepression.ThissuggeststhatfuyanghuatankaqiaomaypromotetheexpressionofWnt/β-cateninsignalingpathwaygenesbymodulatingtheepigeneticstatusofthesegenes.

Furthermore,fuyanghuatankaqiaotreatmentincreasedtheexpressionofbrain-derivedneurotrophicfactor(BDNF)anditsreceptortyrosinekinasereceptorB(TrkB),twoimportantneurotrophinsthathavebeenshowntoplayacriticalroleinthedevelopmentandmaintenanceofneuralcircuitsinvolvedinmoodregulation.BDNFandTrkBexpressionwereupregulatedinadose-dependentmannerbyfuyanghuatankaqiaotreatment,suggestingthatfuyanghuatankaqiaomayenhancetheexpressionoftheseproteinsbyactivatingtheWnt/β-cateninsignalingpathway.Thesefindingsprovideatheoreticalbasisforusingfuyanghuatankaqiaointhetreatmentofdepressionandsuggestthatitsantidepressanteffectsmaybemediated,atleastinpart,byitsmodulationoftheWnt/β-cateninsignalingpathway,DNMT1expression,andBDNF/TrkBexpressionInadditiontothepotentialinvolvementoftheWnt/β-cateninsignalingpathway,othermechanismsmayalsocontributetotheantidepressanteffectsoffuyanghuatankaqiao.Forinstance,previousstudieshaveshownthatfuyanghuatankaqiaocanstimulatetheactivityofthehypothalamic-pituitary-adrenal(HPA)axisandincreasethesecretionofglucocorticoids,whichareknowntoregulatemoodandstressresponses(Shimizuetal.,2012;Liuetal.,2017).Moreover,fuyanghuatankaqiaohasbeenreportedtopossessantioxidantandanti-inflammatoryproperties,whichmayalsocontributetoitsantidepressanteffects(Lietal.,2018;Zhangetal.,2019).Furthermore,recentstudieshavesuggestedthatfuyanghuatankaqiaomaypromoteneurogenesisandsynapticplasticityinthehippocampus,whicharecloselyrelatedtothepathophysiologyofdepression(Gaoetal.,2013;Wangetal.,2018b).

Despitethesepromisingfindings,severallimitationsshouldbeacknowledgedwheninterpretingtheresultsofthepresentstudy.First,thepresentstudyonlyusedananimalmodelofdepressioninducedbyCUMS,whichmaynotfullyreflectthecomplexityandheterogeneityofhumandepression.Futurestudiesshouldinvestigatetheeffectsoffuyanghuatankaqiaoonotheranimalmodelsofdepressionandonhumanpatientswithdepression.Second,thepresentstudyfocusedontheWnt/β-cateninsignalingpathwayanditsdownstreamtargets,butdidnotexamineothersignalingpathwaysthatmaybeinvolvedintheantidepressanteffectsoffuyanghuatankaqiao.Futurestudiesshouldinvestigatetheeffectsoffuyanghuatankaqiaoonothersignalingpathways,suchasthemonoaminergicandneurotrophicsignalingpathways,whichareknowntobedysregulatedindepression.Finally,thepresentstudydidnotinvestigatethelong-termeffectsoffuyanghuatankaqiaooritspotentialsideeffects.Futurestudiesshouldinvestigatethesafetyandefficacyoffuyanghuatankaqiaoinanimalmodelsandinclinicaltrials.

Inconclusion,thepresentstudyprovidesevidencethatfuyanghuatankaqiaoexertsantidepressanteffectsbymodulatingtheexpressionofDNMT1,BDNF,andTrkBproteinsviatheactivationoftheWnt/β-cateninsignalingpathwayinadose-dependentmanner.Thesefindingsprovideatheoreticalbasisfortheuseoffuyanghuatankaqiaointhetreatmentofdepressionandsuggestthatitstherapeuticeffectsmaybemediatedbymultiplemechanisms,includingtheregulationofepigeneticmodifications,neurotrophicfactorexpression,andsynapticplasticity.FurtherstudiesareneededtoelucidatetheunderlyingmolecularmechanismsandtoevaluatethesafetyandefficacyoffuyanghuatankaqiaointhetreatmentofdepressionInadditiontothepotentialmechanismsdiscussedabove,studieshavealsosuggestedthatfuyanghuatankaqiaomayexertitstherapeuticeffectsthroughthemodulationoftheimmunesystem.Theimmunesystemplaysacriticalroleinthepathogenesisofdepression,andithasbeenshownthatchronicinflammationandimmunedysregulationareassociatedwithanincreasedriskofdevelopingdepressivesymptoms(Berketal.,2013).

Animalstudieshaveshownthatfuyanghuatankaqiaocanregulatetheexpressionofcytokinesandchemokines,whichareimportantmediatorsoftheimmuneresponse.Forexample,onestudyfoundthattreatmentwithfuyanghuatankaqiaoreducedthelevelsofIL-1βandTNF-αinthehippocampusofratswithchronicunpredictablestress-induceddepression(Chenetal.,2017).AnotherstudyshowedthatfuyanghuatankaqiaotreatmentincreasedthelevelsofIL-10,ananti-inflammatorycytokine,inthespleenandserumofratswithdepressioninducedbychronicmildstress(Caietal.,2018).

Moreover,fuyanghuatankaqiaomayalsoregulatethegutmicrobiota,whichhasbeenimplicatedinthepathogenesisofdepression.Thegutmicrobiotahasbeenshowntointeractwiththecentralnervoussystemthroughthegut-brainaxis,anddisruptionsinthegutmicrobiotahavebeenassociatedwithdepressionandanxiety(FosterandMcVeyNeufeld,2013).Animalstudieshaveshownthatfuyanghuatankaqiaocanmodulatethecompositionofthegutmicrobiota,increasingthelevelsofbeneficialbacteriaandreducingthelevelsofharmfulbacteria(Liuetal.,2018;Liuetal.,2019).

Finally,fuyanghuatankaqiaomayalsoexertneuroprotectiveeffectsthroughtheinhibitionofoxidativestressandapoptosis.Animalstudieshaveshownthatfuyanghuatankaqiaocanreducethelevelsofreactiveoxygenspeciesandlipidperoxidationinthehippocampusofratswithchronicunpredictablestress-induceddepression(Chenetal.,2017).Anotherstudyshowedthatfuyanghuatankaqiaotreatmentreducedtheexpressionofcaspase-3andBax,twomarkersofapoptosis,intheprefrontalcortexofratswithchronicunpredictablemi