JustynaKwapiszetal.

Hepcidinanditsroleinironhomeostasis

HEPCIDINANDITSROLEINIRONHOMEOSTASIS

JustynaKwapisz1,ArturSlomka1,EwaZekanowska1

1)DepartmentofPathophysiology,NicolausCopernicusUniversityinTorun,CollegiumMedicuminBydgoszcz,Poland

Correspondence
EwaZekanowska,DepartmentofPathophysiology,CollegiumMedicumUMK,SkodowskiejCurie9,85094Bydgoszcz,
Poland;email:zhemostazy@cm.umk.pl;phone.:(+4852)5853591;fax:(+4852)5853595.

Abstract
Hepcidin,asmallpeptidesecretedmainlybytheliver,playsacentralroleinironstatusregulation.Theexperiments
onhepcidinseemedverypromisingandgavenewlifetounderstandingironmetabolism.Manyauthorssuggestthat
hepcidinmeasurementcanbeusedasaclinicaltoolforthediagnosisandmanagementofawiderangeofironrelated
disorders.Thecurrentreviewpresentsdataconcerninghepcidin,especiallyitsbiology,mechanismofactionandits
roleinpathomechanismofmanydiseases.

Keywords:hepcidin,ironmetabolism

1.INTRODUCTION
Hepcidinwasfirstdiscoveredinhumanbloodultrafiltrateandurinesamplesasasmallbactericidalpeptide(defensin
and cathelicidin) and named liverexpressed antimicrobial peptide (LEAP1) [110]. The name hepcidin originates
from the place of synthesis in hepatocytes (hep) and its antimicrobial activity (cidin) [10]. The gene encoding
hepcidin (HAMP, 19q13) is expressed in the liver, heart, lungs, brain, spinal cord, intestine, stomach, pancreas,
adipocytes,skeletalmuscles,testisandmacrophages[25,8,10,11,12].Thehepcidingeneshavebeenalsofoundin
mice, pigs, birds and fish [3]. Hepcidin has antibacterial (Escherichia coli, Staphylococcus aureus, Staphylococcus
epidermidis, Streptococcus spp. group B) and antifungal activity (Candida albicans, Aspergillus niger, Aspergillus
fumigatus) [10]. This protein is a key regulator of iron level, it decreases the iron absorption from the duodenal
enterocytes,ironreleasefrommacrophagesanditstransportacrosstheplacenta[1,38,10,1316].Themainroleof
hepcidininironmetabolismwasconfirmedonanimalmodelsandinvitrostudies[13].Thesynthesisofhepcidinin
hepatocytescanberegulatedbyironoverload,inflammatorysignals,increasederythropoiesis,hypoxiaandanemia
[19,11,12,14,16,17].

2.THESTRUCTUREOFHEPCIDINGENEANDPEPTIDE
Thehumanhepcidingene(HAMP)islocatedonchromosome19q13.1[18].Itis2637basepairslongandcomposedof
threeexonsandtwointrons[19,20].ItcontainsbindingsitesforsuchregulatoryfactorsasHNF3,C/EBPandNFB
[19]. HAMP gene expression was detected mainly in the liver, but also in heart, brain, lung, prostate gland, tonsils,
salivary gland and trachea [21]. HAMP encodes a precursor of hepcidin preprohepcidin, which is 84 amino acids
proteincomprised24aaleaderpeptideattheNterminal,a35aaproregion,andtheCterminal20or25aamature
peptide.Preprohepcidiniscleavedto60aaprohepcidinwhichisfurtheraminoterminallyprocessedandgivesriseto
hepcidin.Therearethreeformsofhepcidin:25aa,22aaand20aapeptide.Allthreeformsaredetectableinurine,
butonlyhepcidin25andhepcidin20arepresentinhumanserum[18,22,23,24].Thestructureofhepcidin25,which
isamajorformofhepcidin,containseightcysteineresiduesconnectedbydisulfidebonds[21].Analysisofhepcidin
structure by NMR spectroscopy showed that this peptide forms a simple hairpin stabilized by four disulfide bonds
JustynaKwapiszetal. Hepcidinanditsroleinironhomeostasis

betweenthetwoantiparallelstrands.Unusualvicinaldisulfidebridgefoundattheturnofthehairpinprobablyplays
significantfunctionalrole[18,22,25].

3.MECHANISMSOFHEPCIDINACTION
Hepcidiniswellknownasironregulatoryhormone.Generally,itcausesadecreaseinserumiron.Themechanismof
hepcidinactivitydependsonhepcidininteractionswithferroportin.Ferroportinistheonlyknownmammaliancellular
iron exporter, which is expressed on the surface of reticuloendothelial macrophages, hepatocytes, duodenal
enterocytes and placenta cells. Hepcidin regulates posttranslationally ferroportin expression. Hepcidin binds to
ferroportinandcausesitsinternalizationanddegradationinendolysosomes,whatinturnblockstheirontransportvia
ferroportin.Whenironstoresareadequateorhigh,increasedhepcidinexpressioninhibitsintestinalironabsorption,
releaseofrecycledironfrommacrophagesanditstransportacrosstheplacenta.Ontheotherhand,whenironstores
are low, hepcidin production is suppressed. By modulating hepcidin expression, organism can control plasma iron
levelandmaintainironmetabolismhomeostasis[18,22,26,27,28].

4.FACTORSRESPONSIBLEFORTHEREGULATIONOFHEPCIDINSYNTHESIS
Hepcidin synthesis is modulated by different stimuli, which are divided into positive and negative regulators. The
molecularmechanismsofregulationofhepcidinexpressionarenotcompletelyunderstood.

POSITIVEREGULATORSOFHEPCIDINPRODUCTION

I NFLAMMATION
HepcidinisnotonlyironregulatoryhormonebutalsotypeIIacutephasereactant.Itmeansthatitssynthesiscanbe
induced by inflammatory cytokine IL6. Some studies show that inflammation and infection rapidly decrease serum
ironlevels,dietaryironabsorptionandironreleasefromREmacrophages[29,30].IL6actsviaitsreceptorandcauses
phosphorylationofsignaltransducerandactivatoroftranscription3(STAT3),dimerizationofphosphoSTAT3andits
translocationtonucleus,whereitinteractswithhepcidinpromoter.Whatisimportant,STAT3activationrequiresthe
presence of SMAD4 to affect the HAMP gene expression [6, 31]. This data confirms that hepcidin could be the
pathogenicmediatorofanemiaofchronicdiseases(ACD).

I NCREASEINIRONSTORES
Under normal conditions HAMP gene expression is regulated by BMP/SMAD and STAT3 pathways. Bone
morphogeneticproteins(BMPs),activatedbyelevatedcirculatingironlevel,bindandformcomplexwithtypeIandII
cell serine/threonine kinase BMP receptors in hepatocytes, that results in phosphorylation of SMAD proteins
receptors(RSMADs).PhosphoRSMADsformcomplexwithSMAD4,whichtranslocatesintothenucleusandactivates
thetranscriptionofHAMPgene[31,32].Membraneisoformofhemojuvelin(mHJV)asaBMPcoreceptoralsotakes
partinpositiveregulationofhepcidinexpressionviaBMP/SMADpathway.Anotherhepatocyteironsensorsactivating
hepcidinsynthesisarehemochromatosisprotein(HFE)andtransferrinreceptor2(TfR2).Furtherstudiesareneeded
todefinepreciselytheirroleinregulationofironstatus[31,32,33].

NEGATIVEREGULATORSOFHEPCIDINPRODUCTION

H YPOXIA , ANEMIA , INCREASEDERYTHROPOIESIS

Ithasbeenconfirmedthatanemiaandhypoxiabelongtoregulatorsofhepcidinexpression.Experimentsonmicehave
demonstratedthatanemiainducedbyphenylhydrazineorphlebotomiestriggeredaconsiderabledecreaseinhepcidin
mRNA[34].Hypoxiaandanemiaregulatetheerythrocytesproductionthrougherythropoietin(Epo)synthesis.Some
authorsclaimthatEpoisahormonedownmodulatinghepcidinmRNAexpression.IntheiropiniontheeffectofEpois
mediated via Epo receptor signaling and regulation of C/EBP. Moreover, observations on thallasemia patients
reported growth differentiation factor15 (GDF15) as a possible factor causing hepcidin downregulation [35, 36].
Obviously erythropoietic activity is a potential hepcidin synthesis suppressor, but specific erythropoiesisassociated
mediatorthatnegativelyregulateshepcidinproductionisstillunknown.

It is highly likely that in hypoxic conditions the hypoxia inducible factor/von HippelLindau (HIF/vHL) pathway can
inhibithepcidinexpressioninhepatocytes[31,33].Itisalsosuggestedthathypoxiaactivatesfurintoincreaseamount
ofsolublehemojuvelin(sHJV)byproteolyticcleavageofmembranehemojuvelin(mHJV)[31].
JustynaKwapiszetal. Hepcidinanditsroleinironhomeostasis

D ECREASEINIRONSTORES
In plasma hemojuvelin exists in soluble form (sHJV), which in response to low serum iron level binds toBMPs and
inhibitsBMP/SMADsignaling[31,33].Veryintriguingistheroleofmatriptase2(transmembraneserineprotease6,
TMPRSS6) in hepcidin production. Some authors indicate that matriptase2 is a negative regulator of hepcidin
expressionbecauseitiscleavingmembranehemojuvelin[37].

5.HEPCIDININTHEPATHOGENESISOFIRONDISORDERSANDOTHERDISEASES
Thestudiesonhepcidinprovideessentialinformationabouttheetiologyandpathomechanismsofironmetabolism
disordersandotherdiseases.

Hemochromatosis (HH), the most common form of genetic iron overload, is divided into two groups: iron overload
associatedwithdefectiveorsuppressedhepcidingeneandferroportindisorders[12,31].Thefirstgroupofdiseasesis
causedbymutationinfourgenes:HFE1,HJV,TfR2andHAMP.Patientswithdefectofthesegeneshavelowhepcidin
mRNA level in comparison with normal subjects [6, 10, 12, 13, 16, 31]. The mutation in HAMP causes rare form of
juvenile hemochromatosis(JH) type2B and leads to downregulationof hepcidin expression [10,13,31]. Moreover,
juvenilehemochromatosisisassociatedwithlowerhepcidinlevelthaninadultformsofhemochromatosis[31].

Hepcidin, as an acute phase protein is the key mediator of anemia observed in inflammatory disorders known as
anemiaofchronicdiseases(ACD)[3,7,8,10,16].PathogenesisofACDisassociatedwithdecreasedironabsorption
and impaired mobilization of iron stores [8]. The individuals with the anemia of inflammation, characterized by
disturbanceofironabsorption,hypoferremiaandhyperferritinemia,havehigherhepcidinlevelsthanhealthysubjects
[9,10,15,38].ThehigherconcentrationofserumhepcidininpatientswithACDthaninthehealthypeoplecanbe
explainedbytheIL6increase[1,2].Interestingly,therelationshipbetweentheIL6andhepcidinlevelwasobserved
inpatientswithacuteinflammatoryreactionandinhealthyvolunteersafterlipopolysaccharideinjection[1,10].The
increasedlevelofserumhepcidinisobservedinmanychronicinflammatorydiseasessuchas:chronickidneydiseases,
thallassemia,glucose6phosphatedehydrogenasedeficiency,sicklecelldisease(SCD),coronaryarterydisease(CAD)
and myelodysplasia [7, 9, 10, 14]. The recent studies have shown elevated hepcidin level during radiotherapy for
prostatecancerinindividualswithacuteproctitisandafterhematopoieticstemcelltransplantation(SCT)[1,8].

Theoverexpressionofhepcidinhasbeenshowninclinicalstudiesondysmetabolicironoverloadsyndrome(DIOS)[11,
12].InpatientswithDIOStheironabsorptionissignificantlydecreasedthanincontrolswithnormalironstatus[12].In
other liver diseases like obesity related to nonhereditary mild iron overloading hepatic disease (NHIOD), alcoholic
liver disorders and hepatitis C virus infection (HCV) enterocytes are resistant to circulating hepcidin, while
macrophages are more sensitive [11]. Serum hepcidin correlates positively with hepatic hepcidin mRNA level and
ferritinlevelinchronichepatitisC(CHC)[15].Hepcidinmaybeaprognosticandmonitoringtestofironoverloadin
patientswithNHIODandHCV.NormalizationofhepcidinconcentrationmaybealsoanindicatorofHCVeradication
[11].

6.HEPCIDINASAPOTENTIALDIAGNOSTICANDTHERAPEUTICTOOL
Thediscoveryofhepcidinin2000byKrauseetal.[21]andParketal.[20]notonlyopenedthewaytounderstandthe
ironmetabolismbutalsohelpedtoelucidatethepathomechanismsofmanydiseases.Thestudiesonhepcidinraise
thequestionoftheuseofhepcidinasadiagnosticandtherapeutictoolinmanydiseases.Hepcidinmeasurementcan
behelpfultestdistinguishinganemiaofchronicdiseases(ACD)fromirondeficiencyanemia(IDA),asitisknownthat
hepcidinproductionisinducedbyinflammation(ACD)andreducedinirondeficiencystates(IDA)[29,39].Oneofthe
greatest promises for the practical application of hepcidin assay is the utilization of hepcidin in diagnosis and
monitoring of hemochromatosis. What is more, the possible therapeutic value of hepcidin is investigated. The
development of synthetic hepcidin should be useful in the treatment of hemochromatosis and other ironloading
conditions[40].

In 2008 Ganzet al. [41] performed successful validationof a competitive enzymelinked immunoassay (CELISA) for
humanhepcidin.Thissimpleandrobustassaycanbeusedindetectingphysiologicandpathologicchangesinserumor
urinehepcidinlevels.Itisprobablethatthistestwillbewidelyavailableforuseinclinicalchemistrylaboratoriesinthe
nearfuture.

However,alotremainstobeuncoveredonthebiologyandfunctionofhepcidin.Itssignalingpathwaysareasyetto
bedelineated.Furtherstudiesareneededtodefinepreciselythehepcidinroleinironmetabolismhomeostasisandits
utilityinthediagnosisandtreatmentofirondisorders.
JustynaKwapiszetal. Hepcidinanditsroleinironhomeostasis

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