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Matrix Gla-protein: The calcification inhibitorinneed of vitaminK
Leon J. Schurgers,Ellen C. M. Cranenburg, CeesVermeer
VitaKand Cardiovascular ResearchInstitute (CARIM),Maastricht University,Maastricht, TheNetherlands
Summary
Among theproteins involved in vascular calcium metabolism,
the vitamin K-dependentmatrix Gla-protein (MGP) plays a
dominant role.Althoughonamolecular levelits mechanism of
action is not completely understood,it is generallyacceptedthat
MGPisapotent inhibitor of arterial calcification.Its pivotal im-
portance forvascular health is demonstratedbythe factthat
thereseemstobenoeffectivealternativemechanism forcalcifi-
cation inhibition in thevasculature.Anoptimal vitamin Kintake
is therefore important to maintain theriskand rate of calcifi-
cationaslow as possible.With the aid of conformation-specific
Keywords
Matrix Gla-protein, vitamin K, calcification, cardiovascular dis-
ease,oral anticoagulants
antibodies MGPspecies in bothtissueand the circulationhave
been detectedinthe healthypopulation, and significantdiffer-
enceswerefoundinpatientswith cardiovascular disease (CVD).
Using ELISA-basedassays, uncarboxylatedMGP (ucMGP) was
demonstrated to be apromising biomarker forcardiovascular
calcification detection.Theseassays mayhavepotential valuefor
identifyingpatientsaswellasapparentlyhealthysubjects at high
risk forCVD and/or cardiovascular calcification and formoni-
toringthe treatment of CVDand vascular calcification.
ThrombHaemost 2008; 100: 593–603
Theme Issue Article
Correspondence to:
Dr.L.J.Schurgers
VitaK, MaastrichtUniversity
Universiteitssingel 50
6200 MD Maastricht, TheNetherlands
Te l.: +31433881680, Fax: +3143388 4159
E-mail: l.schurgers@bioch.unimaas.nl
Received February14, 2008
Accepted after minor revision July 2, 2008
Prepublished onlineSeptember 5, 2008
doi:10.1160/TH08-02-0087
Background
Matrix Gla-protein (MGP) is asmall secretoryprotein thatcan
undergo twotypes of posttranslational modification: γ-gluta-
mate carboxylation and serine phosphorylation. The protein was
first described in 1983byPrice et al. whopurified it from the
bovine bone matrix (1).The authorsconcludedthat this approxi-
mately 14 kD protein contains five unusual amino-acids desig-
natedasγ-carboxyglutamate(abbreviated as Gla), and therefore
the protein wasdesignatedasmatrixGla-protein (Fig. 1).Soon
after its discovery in bone, MGP synthesis in cartilage,lung,
heart,kidney, arteries and calcified atherosclerotic plaqueswas
confirmed (1–3). Thematureprotein consistsof84amino-acids
and has atheoreticalPIof9.7. Of the nine glutamate residues
onlyfivecan be γ-carboxylated in avitamin K-dependent reac-
tion, and three of its five serine residuescan be phosphorylated
into phosphoserine (abbreviated as Pser). TheMGP gene is lo-
cated on chromosome12(p13.1-p12.3),consistsoffour exons
and threelarge introns and has alength of 3.9 kb.Itcontains
metal responsive elementsand presents putative binding sites for
AP1 and AP2 and cAMP-dependent transcription factors. At
physiological levels, vitamin D3 increased MGPtranscription in
VSMC whereas retinoic aciddownregulates its expression (4).
The best studied posttranslational modulation of MGP is
gamma-glutamatecarboxylation. Gla-residues areformed in a
unique posttranslational modification carriedout by the enzyme
gamma-glutamatecarboxylase (5).The onlyunequivocal role of
vitamin Kistoprovide the energy to drivethe carboxylase reac-
tion. The Gla-residues formed arenegativelycharged and pro-
teins in whichtheyare found are denominatedasGla-proteins.A
common characteristic of allknown membersofthis protein
familyisthat the Gla-residues areabsolutelyrequired forprotein
activity (6). In all Gla-proteins the affinity for gamma-glutamate
carboxylase is determinedbyapro-sequence located immedi-
atelyatthe N-terminal siteofthe protein. In most Gla-proteins
the pro-sequence is cleaved offduring maturation; MGP is the
exception in this respect, since the matureprotein contains an in-
ternal pro-peptide whichmay contribute to its unique properties.
Phosphorylation, the otherposttranslational modification in
MGP,may take placeatserine residuesinpositions 3, 6and 9
(Fig. 1). Price et al. showedthat the motifinMGP recognizedfor
serine phosphorylation is the tandemlyrepeated Ser-X-Glu se-
quence (4). Phosphorylation is carried outbythe Golgi casein ki-
nase (4, 7). The function of serine phosphorylation is not pre-
cisely known, butrecent datasuggest that it plays arole in regu-
lating the secretion of proteins into the extracellularenviron-
©2008 Schattauer GmbH,Stuttgart
593
Newvitamin K-dependentproteins
Schurgers et al.Matrix Gla-proteinand vascular calcification
594
ment. Wajih et al.showedthat phosphorylated MGP exits vascu-
lar smooth muscle cells (VSMCs) viathe secretorypathway,
whereas the non-phosphorylated MGPappears in the cytosol,
and is thus not secreted (7).
Thefat-soluble vitamins Aand Dmay modulate MGPex-
pression. Retinoicacidisaregulator of chondrocyte maturation
and mineralization (8).Its effect on mRNAexpression levels of
MGPiscelltype-dependent: in fibroblasts, chondrocytes, os-
teoblasts, and type II pneumocytes, retinoicacidupregulates
MGP mRNA expression (9, 10) whereas in kidney cells and
VSMCs, retinoic aciddownregulates MGP expression (11, 12).
1,25(OH)D3wasshown to increase MGP expression in vitro in
VSMCs(11). In models in vitro (13),animal models (14) and hu-
mans (15),extremelyhigh vitamin Dintakes maycausevascular
calcification, most likelydue to an effect on calcium-metab-
olism.
Functions of MGP
Although theprecisemolecular mechanisms of MGP function
arenot known, accumulating data demonstrate its major role in
the inhibition of soft-tissue calcification. The first clues for a
Gla-protein being involved in the inhibition of tissue calcifi-
cation camefrom rats treated with the vitamin K-antagonist war-
farin(16). These animals developedmassive cartilage calcifi-
cation, notablyinthe epiphyses and facial bones,leading to im-
paired growth, maxillonasal hypoplasia and reduction in the
length of the nasalbones (17).Itwas only after the identification
of MGPincartilage thatitwas recognized thatthe cartilage cal-
cification wasbroughtabout by loss of MGPfunction (18).After
its discovery, it wasthought for manyyears thatthe importance
of MGP wasrestrictedtobone and cartilage metabolism. By tar-
geteddeletion of the MGPgene in miceitbecameclear,however,
thatits main function is the inhibition of medialcalcification of
the arteries: MGP-deficient animals alldiedwithin six to eight
weeks after birthdue to calcification of the elasticlamellaeinthe
tunica media, resulting in rupture of the large arteries (19).The
arterial calcification in the MGP null mice resulted from the pre-
cipitation of calcium-phosphate in aratio similartohydroxyapa-
tite, thus mimicking bone mineralisation. Using histochemical
techniques, the authorsdemonstrated thatthe arterial calcifi-
cation wasassociatedwith the differentiation of VSMCs into
chondrocyte-likecells. Amechanism explaining the strong cal-
cification inhibitoryactivity of MGPwas putforwardbyPrice,
whosuggestedthat MGP binds tightly to the crystalnucleithus
preventing further growth (20). Inhibition of the differentiation
of VSMCs into chondocyte- and osteoblast-likecells maybea
secondfunction of MGP for whichfurther supportwas provided
in MGP-deficient micebydemonstrating aloss of smooth
muscle markers and upregulatedexpression of the bone-specific
transcription factor cbf1a/Runx2 and the osteogenicprotein os-
teopontin (21). Theability of MGP to keep VSMCs in the con-
tractile phenotype maybeaccomplishedbybindingtothe bone
morphogenetic protein-2 (BMP-2) (22, 23). BMP-2 is amember
of the transforming growth factor-beta(TGF-beta) superfamily,
and is an osteogenicgrowthfactor.BMP-2 hasbeen shown to be
expressed in human atherosclerotic lesions (24).Wallin et al.
demonstrated thatonlythe carboxylated formofMGP binds to
BMP-2(22); moreover,Bostrom et al. presenteddatasuggesting
that MGP blocks the osteo-inductive propertiesofBMP-2 (25).
This inhibitoryfunction is further supported by work of Shana-
hanetal., whoshowedthat MGP expression is lowerinthe media
of arteries from diabetic patients with Mönckeberg's sclerosis
thaninnormal vessels (26).Via its C-terminalregion, MGPcan
also bind to the extra-cellularglycoprotein vitronectin, whichis
present in the extracellularmatrixofthe arteries (27).The C-ter-
minal partofMGP is hydrophobic and does not contain Gla- or
Pser-residues, whichare all present in the morehydrophilic
Figure1:Matrix Gla-pro-
tein (MGP) is asmall84aa
vitaminK-dependent pro-
tein. Althoughits 14 kD size,
it can undergotwo posttrans-
lationmodifications: at posi-
tion 3, 6, and 9the serine resi-
dues can be phosphorylatedby
aGolgi-caseinkinase,and at
positions2,37, 41, 47 and 52
theglutamate residues can be
γ-carboxylated(117).
Newvitamin K-dependentproteins
Schurgers et al.Matrix Gla-proteinand vascular calcification
595
N-terminaland mid-section of the molecule.Itmay be hypothes-
izedthat MGP’sbindingtovitronectin results in aconcentration
of calcification-inhibitoryactivity in the milieusurrounding the
elasticfibers, therebyprotecting themfrom mineralization.
Formation of matrix vesicles(MV) and apoptotic bodies
(AB)isthought to precede and/or initiate arterial calcification.
VCSMs undergoing apoptosis provide negatively charged mem-
brane particleswhich –ifnot phagocytosed properly–playarole
in the initiation of calcification (28).The physiological function
of these extracellularmembrane particlesistoserve as the initial
nidusofcalcification in cartilage.Also in the vesselwall, both
MV and AB are relativelycommon, notably in atherosclerotic
plaques(29, 30), arterial injury(31) and Mönckeberg's sclerosis
(32, 33).When VSMCsare growninculture theycan form multi-
cellularnodules, containing ahigh number of AB.MGP ex-
pression is highest in this phase,suggesting an association be-
tweenMGP and apoptosis.Reynolds et al. showedincellculture
systems that VSMC derived MV and AB both containedMGP
whichisthought to limit the rate of calcification (34).
The specific knock-in expression of MGP in VSMCsof
MGP-deficient micecompletelyrescued the calcification phe-
notype (35). In the same article the authors also expressed MGP
in the liverofMGP-deficient mice, resulting in high levels of
circulating MGP. However, the elevatedsystemic levels of MGP
hadnoeffect on inhibition of arterial calcification implying that
MGPinhibitscalcification by acting locally withinits tissue of
synthesis, not systemically.Inhumans, mutations in the gene en-
coding for MGP –predicting anon-functional protein –cause
the Keutel syndrome (36),arare disordercharacterizedbyab-
normal cartilage calcification and peripheral pulmonary steno-
sis (37).Post mortem examination of ayoung Keutel patient also
revealed extensive arterial calcification (38).
Arterialcalcification
Until adecade ago,calcification of arteries wasthought to be a
passive,clinically irrelevant process, resulting from ahigh cal-
cium xphosphate product, inflammation, lipidaccumulation or
diabetes. However, during recent years it hasbecomeincreas-
inglyclear thatvascular calcification is an activeprocessand an
important, independent pathology that is stronglyassociated
with increased risk of cardiovascularmorbidity and mortality
(39–41).Clinically,vascular calcification causes stiffening of
the vascularwall, whichmay result in decreased arterial com-
pliance, development of left ventricularhypertrophyand de-
creased coronaryperfusion leading to an increased risk of fatal
complications (42,43). Calcification is common in the elderly
population, and in patients suffering from diseases such as
chronic kidneydisease(CKD), diabetes, aorticstenosis, and
atherosclerosis (44).Therefore,alot of efforts have beendi-
rected towards retarding or reversing the development of calcifi-
cation in the vasculature.Inanimal models it has beenshown that
arterial calcification is reversible(45–48), demonstrating that
also the regression processisanactivelyregulated process. In
humans, attempts to use lipidlowering drugs(statins) to stabilize
or regresscalcification have so farfailedtoshowasignificant ef-
fect (49, 50).
CKDpatients have the highestincidence of arterial calcifi-
cation, and cardiovascularmortality is 20-fold higher than in the
apparentlyhealthypopulation (51, 52). Moreover,moderate to
severe vascularcalcifications are foundin60–80% of patients on
hemodialysis (53, 54). Recently, it wasshown that vitamin
K-status in CKD patients is low(55). Circulating vitamin Klev-
elswere measured and reportedthat some 30% of the haemo-
dialysis patients had sub-clinicalvitamin K-deficiency. The au-
thors discussed the possibility of givingthese patients extravit-
amin Ktoreduce the risk for cardiovascular events(55). Addi-
tionally, the need forvitamin Kinpatients might be muchhigher
thaninthe generalpopulation. Anticoagulation therapywith vit-
amin Kantagonists, whichisregularly prescribed in these pa-
tients, will exacerbatethe lowvitamin K-status in these patients.
Togetherwith the additionalimmunohistochemical evidenceof
high levels of uncarboxylated MGP (ucMGP) present in calci-
fied areas (47, 56,57), these data aresuggestive for high vitamin
Kintakeasanoveltreatment option for cardiovascular calcifi-
cation (see alsobelow). Thefirst clinical studiesindialysis pa-
tients are in progress.
Factors affectingMGP activity
VitaminK/warfarin
It has been knownfor along timethat womenreceiving anti-
coagulant therapywith vitamin Kantagonists (coumarin deriva-
tives) during the first trimesterofpregnancyare at risk of de-
livering children with asyndrome characterized by nasalhypo-
plasia, depression of the nasalbridgeand punctuate calcifi-
cations in the axialskeleton, proximalfemursand calcanei (58).
This syndrome is known as warfarin embryopathy(fetal warfarin
syndrome), and the abnormalitieswere first believedtobe
caused by haemorrhages in the developingfetal cartilageswith
subsequent calcification of these areas (58–60). However, it was
soon recognized thatthis wasunlikely, sinceclotting factors
were knowntobeabsent during the first trimesterofpregnancy
(58,61). Similarities betweenthe facial and skeletal abnormal-
itiesseen in warfarin embryopathyand the fetal phenytoin (hy-
dantoin) syndrome suggested that prenatalvitamin K-deficiency
mayunderlie these abnormalities (58, 62). This wasconfirmed
by Pauli et al. whodescribed acongenital deficiencyofthe
enzyme vitamin K-epoxide reductase (VKOR, needed forrecyc-
ling of vitamin K),causing prenatal vitamin Kdeficiency, and
resulting in asimilarphenotype (63, 64). In later years it wasre-
ported thatalso dietary vitamin K-deficiencyresults in com-
parable calcification abnormalities(65–69), whichwere remark-
ably similar to the bone and cartilage defects observedinwarfa-
rin-treated rats(16). The same animal model provided the first
evidencethat impairment of MGP function resultsinvascular
calcification (20). It wasfound that within twoweeks of warfa-
rintreatment,the elastinfibres in the tunicamediawere signifi-
cantly calcified. Further evidencefor the pivotalrole of Gla resi-
dues for MGP function wasprovided by Murshedetal. whoused
the MGP null mice in whichMGP cannot be carboxylated,since
the glutamate residues in the Gla-domain were mutatedinto as-
partate; in this wayitwas demonstrated thatonlycarboxylated
MGP (cMGP) exhibitsanti-mineralization properties(35). Vit-
amin Kantagonists arefrequently used to prevent thrombosis in
Newvitamin K-dependentproteins
Schurgers et al.Matrix Gla-proteinand vascular calcification
596
patients at increased risk for thrombosis (70). Treatment periods
rangefrom several weeks to many years,evenoften life-long
(71).After demonstration in animalmodels that vitamin Kan-
tagonists induce vascularcalcification, studiesintwo indepen-
dent populations revealed thatindeedtreatment with coumarin
derivativesinducesexcessive calcification of the coronaryar-
teries and the aortic heartvalve (72,73). Schurgersetal. com-
pared valvularcalcification in patients receiving oral anticoagu-
lant treatment for aperiod of between 16 and 35 monthswith pa-
tients not on oral anticoagulation (72). Histopathological evalu-
ation of the valves fromthe patients whohad receivedoral anti-
coagulation showedpartialortotal valvedestruction induced by
amorphouscalcified deposits. Quantification of the calcium
contents of the aortic valves showedastatistically significant
differencebetween valves fromthose whohad neverreceived
oralanticoagulant treatment and those whohad receivedthis
treatment. These data were confirmed by Koos et al., whoused
multislice spiralcomputed tomography(CT) to quantitate the
extent of aortic calcification in patients on long term oralantico-
agulant treatment (73).Itwas foundthat these patients had in-
creased coronarycalcification compared to patients without
anticoagulation treatment (Agatston score1,561 and 738, re-
spectively). Thepresentpolicyisopposite, however: large
numbersofcardiovasculardisease(CVD) patients receive anti-
coagulant therapywith vitamin Kantagonists, whichincreases
their calcification tendency. The data described above suggest
that–ifpossible –otherforms of anticoagulation (specific pro-
thrombin or factor-X inhibitors) should be employed,preferably
in combination with high vitamin Kintake. This treatment could
activate MGP, and the intriguingquestion remains whether it de-
creases CVDinparallel. Together, these data demonstrateahi-
thertounrecognizedadverse side-effect of coumarinderivatives
whichshould be consideredwhen designing optimal anti-throm-
botic treatments for patients.
Vitamin Kcomprises afamilyincluding vitamin K1(phyllo-
quinone) and vitamin K2(menaquinones). The mechanism of
vitamin K1is believedtobemost important for activation of he-
patic clotting factorswhereasK
2also is importantfor proteins
synthesizedinextra-hepatic tissues such as the vasculature.
Moreover, there is nowscientific evidencethat K2vitamins have
additionalproperties, includingapoptosisand cell-cycle arrest
and anticancerproperties(74, 75), inhibition of the synthesis of
prostaglandin E2 (PGE2) (76),osteoclastapoptosis(77), and
binding to the SXR in the osteoblast, resulting in induction of os-
teoblastic markers (78). Recently,itwas shown thatvitamin K2
could also down regulate osteoprotegrin and increase DT-dia-
phorase, implicating thatvitamin K2is an anti-calcification
componentinthe vesselwall(79). Foramore extensive review
on the gene regulatoryfunctions of vitamin K2,see Shearer ar-
ticle,this volume beginning on page 530.
Thequestion of whether high vitamin K-intake is protective
againstarterial calcification wasfirst addressedinapopulation-
basedstudy amongparticipants of the Rotterdam Study.Itwas
demonstrated thatdietary vitamin K2intake(and notK
1
)was in-
verselycorrelated with cardiovascularcalcification and cardio-
vasculardeath(6). Elderly people in the highest tertileofvitamin
K2intake had about 50% reduction in both aortic calcification
and cardiovascularmortality and 25% decreasedall-causemor-
tality.Inaclinical intervention studyinwhich 78 womenbe-
tween 55 and 65 years of age receivedeithervitamin K(1mg/
day) or placebo for threeyears,vascular characteristics were as-
sessed (elasticity and distensibility) (80).Insubjects in the
placebo group vascularelasticity haddecreased by 10–13%,
whichisconsistent with the normal decrease during the time-
period of threeyears;inthe vitamin Kgroup, however, vascular
characteristics hadremained unchanged,suggesting that the pro-
cess of vascularaging canberetarded by increased vitamin Kin-
take.
The concept of calcification inhibition by high vitamin Kin-
takewas confirmed in experimental animals (81).Inthis model
the efficacyofvitamins K1and K2in preventing arterial calcifi-
cation wascompared. It wasfound that K2completely inhibited
tissue calcification, whereas asimilarorevenaneight-fold
higher dose of K1had no measurable effect.Tofullyunderstand
this model it is important to knowthat vitamin K1canbecon-
verted into MK-4 viaMK4-O,but that warfarin blocks the con-
version of MK4-O into reduced MK4, whichisthe active cofac-
tor (82). Buitenhuisetal. showedthat K2vitamins,especially the
long chainK
2vitamins such as menaquinone-7, have lowerK
M
valuesfor the enzyme γ-glutamyl carboxylase,demonstrating
thattheyare the preferred cofactor forvascular carboxylase (83).
Additionally, recently Wallin et al.showedthat specificallyK
2
actsasananti-calcification component in the vesselwallbyin-
creasing the gene expression with a4.8-fold higher specific ac-
tivity of DT-diaphorase, an enzyme of the vitamin K-cycle(79).
The same animal model wasusedtostudy the effect of high vit-
amin Kintakeonpotential regression of arterial calcification
(47). Whereas rats receiving the standard chow(control) had
very lowaortic calcium during the entire experiment, asix-week
warfarin treatment ledtothe accrual of calcium salts up to
12-fold above the baseline values. During the followingsix-
week period warfarin treatment wasstopped and the animals re-
ceivedeitherstandard choworstandard foodfortified with a
high dose of vitamin K1or K2.Itwas shown thatonce calcium
deposits had formed,the accrual of calcium salts in the vascula-
ture increased linearlyafter the warfarin diet had been replaced
by the normaldose of vitamin K. At high doses of either K1or K2,
however, the processofcalcification wasnot onlystopped,but a
significant fraction (some 40%)ofthe previouslyformed cal-
cium salts had beenremoved within six weeks.This effect was
found both in the aorta and in the coronaryarteries.Using immu-
nohistochemistryitwas demonstrated thatparalleltothe regres-
sion of aortic calcium content, cMGP had increased and ucMGP
had decreased,suggesting arole of activatedMGP in theregres-
sion of calcified plaques.The fact thatvitamins K1and K2had a
similar effect in this experiment maybeexplained by the very
high dosagesused, and by the fact thatinthe absence of warfarin
up to 25% of the vitamin K1maybeconverted into K2(84). Only
by performing dose-response studies the efficacies of both vit-
amins maybecompared in this model.
Carboxylase /VKOR
Mutations in the γ-glutamate carboxylase gene result in bleeding
disorders due to the inability to activate sufficient vitamin K-de-
pendent clotting proteins (85,86). Since onlyone gene encodes
for the enzyme, also Gla-proteins produced in extra-hepatic tis-
Newvitamin K-dependentproteins
Schurgers et al.Matrix Gla-proteinand vascular calcification
597
suesare affected by this mutation. Morerecently it wasshown
that patients with the γ-glutamate carboxylase mutation not only
presentedwith haemostatic disorders,but also with soft tissue
calcifications,asisseen in patients with apseudoxanthoma elas-
ticum (PXE)-likephenotype (87).Pseudoxanthoma elasticum is
an autosomal recessive multi-systemdisordercharacterizedby
dystrophic mineralization of soft tissues, including skin, eyes,
and arterial blood vessels (88).Whereasclassic PXE is caused by
mutations in the ABCC6 gene (ATP-binding cassette subfamily
Cmember 6),patients with the PXE-likesyndrome harboured
known γ-glutamate carboxylase mutations in six out of sevenpa-
tients analyzed.The involvement of MGP in classic PXE wasre-
cently demonstrated by twogroups,showing that fibroblasts
from PXE patients almost exclusivelyproduce the inactive
ucMGP, whichisnot able to block or inhibitcalcification (89,
90). Theverylow cMGP production in pathological fibroblasts
comparedtocontrols suggests these cells have adeficient vit-
amin Kmetabolism whichmay playanimportant roleinthe ec-
topic calcification in PXE.
The vitamin K-epoxide reductase (VKOR) enzyme is acru-
cial enzyme in vitamin Kmetabolism and ensures the re-utiliz-
ation of vitamin Kafter it hasbeen oxidized in the carboxylase
reaction. Because of this recycling, humanvitamin Krequire-
ment is extremelylow (91). On amolecular levelVKORreduces
vitamin K-epoxide in twosteps: first to the quinone,and subse-
quentlytovitamin Khydroquinone (KH2), whichisthe active co-
factor for γ-glutamate carboxylase.VKORisalso the target for
warfarin and related coumarinderivatives, whichblockthe re-
cycling of vitamin Ktherebydecreasing the vitamin K-status.
Both vitamin K-epoxide and vitamin Kquinone needtobind to
the VKOR before being reduced. Wallin et al.showedthat the
enzyme DT-diaphorase in VSMCsis100-fold less active thanin
the liver. The cytoplasmic DT-diaphoraseiscapable of reducing
vitamin Kquinones to their hydroquinone cofactors, and serves
as arescue enzyme in casethe VKOR is blocked by coumarin
(92,93). Therefore, coumarin treatment has adetrimentaleffect
in the arterial vesselwall, by blocking vitamin K-metabolism
leading to impairedMGP.Moreover,the vitamin Kbindingsite
in VKOR is thought to be closetothe coumarin-binding siteand
recentlyitwas shown thatthe presenceofvariousVKORC1 ha-
plotypes correlates with arterial vasculardisease(94).
Besides being acofactor in the vitamin K-dependent car-
boxylation, KH2also possesses antioxidant activity (95, 96).
This is consistent with its high sensitivity to free radicals, which
mayoxidize (and thus inactivate)KH
2before it cantakepartin
the carboxylation reaction. Especiallyinthe atherosclerotic
plaque,high levels of oxidized LDLare found,which maythus
contribute to alocalvitamin Kdeficiency.
MGPasbiomarker
As discussed above,MGP is one of thestrongestinhibitors of ar-
terial calcification, its function depending on the presenceofvit-
amin K. MGPisalocal inhibitor of vascularcalcification and it
hasbeen demonstrated thatcirculating MGPhas no biological
function (35). However, circulating MGPmay reflect calcifi-
cation processesand inhibition of those processesinthe vascular
wall. Belowwewill discuss the presence of MGP in vasculartis-
sue and in the circulation, and the potentialofcirculating MGP
as abiomarker for cardiovascular calcification.
MGPinvasculartissue
Immunohistochemical studieshaveshown that in healthyvessels
MGP is synthesizedatrelativelylow rate (2, 57,97), most likely
because the needfor calcification inhibition is low. However,
Shanahanetal. showedthat in arteries of diabetic patients lower
levels of MGPprotein were present thaninnormal vessels, sug-
gesting thatlow MGP levels might predispose forcalcification
(26). High MGPlevelshavebeen detected in arteries with cal-
cification (2, 57,97). This mayoriginatefrom increased MGP
synthesis, whichhas been reportedinboth medial and intimal ar-
terial calcification (2, 57, 97), or increased subsequent adsorp-
tion to the calcium salt crystals.
With the development of conformation-specificantibodies,
enabling the detection of active, carboxylated and inactive,un-
carboxylated MGP (cMGP and ucMGP, respectively), it became
clear thatspecificallythe ucMGPconformation accumulatesin
atherosclerotic and calcified arteries (56,57, 90). The cMGP
conformation wasnearlyabsent in these arteries. These con-
formation-specific antibodies have proventheir va