Recent Advances in Vitamin K Metabolism

3rd International Conference on Nutrition and Food Science(Nutritional Science-2014)Track 1: Nutrition and Basic Science14:10-14:30, September 23, 2014Committee Room 1-2Palacio de Congresos de Valencia, Spain

Kobe Pharmaceutical University

Toshio Okano

Dept. Hygienic Sciences

Kobe Pharmaceutical Univ.

O

O

O

O

O

O

Plasma vitamin K concentrations of healthy Japanese women

phylloquinone (ng/mL) MK-4 (ng/mL)

1.74±1.29 (0.13 ～ 8.83) 0.10±0.19 (n.d. ～ 1.44)

Naturally occurring vitamin K

Plant origin（ Major dietary

source ）

PK (phylloquinone: vitamin K1 ）

Bacterial origin（ Minor dietary

source ）

MK-4 (menaquinone-4: vitamin K2)

MK-7 (menaquinone-7)

Tsugawa N, Okano T, et al, Am J Clin Nutr, 2006;83:380-386.

Tissue concentrations of animals and humansMK-4 ＞＞ phylloquinone

Amounts of dietary intake and plasma concentrations phylloquinone ＞＞ MK-4

The aim of our study is to examine the above issues in animals and

humans.

Is phylloquinone converted into MK-4 in the body ?

What is the physiological significance of this conversion and what functions dose MK-4 have, and can we

develop MK-4 analogues for clinical use ?

Suhara Y, Okano T, et al, J Med Chem, 2011;54:4269-4273Suhara Y, Okano T, et al, J Med Chem, 2011;54:4918-4922Suhara Y, Okano T, et al, J Med Chem, 2012;55;1553-1558

Tissue concentrations of PK and MK-4 in mice fed a conventional laboratory chow diet

0

5

10

1520

25

30

35

40PK

Con

c.（

pm

ol/g

）

PKPK epoxide

0200400600800

100012001400160018002000

MK

-4 C

on

c.（

pm

ol/g

）

MK-4MK-4 epoxide

×100

Aort

a

Thyro

id

gla

nd

Sple

en

Sto

mach Fat

Ute

rus

Pancr

eas

Ovary

Adre

nal

gla

nd

Sem

inal

vesi

cle

Test

is

Duodenum

Thym

us

Skin

Sm

all

inte

stin

e

Bone

Kid

ney

Cere

bru

mH

eart

Eye

Musc

le

Lung

Liv

er

Pla

sma

　

Adre

nal

gla

nd

Ovary

Sple

en

Fat

Eye

Sto

mach

Ute

rus

Thym

us

Heart

Duodenum

Thyro

id

gla

nd

Bone

Liv

er

Pancr

eas

Lung

Musc

leSem

inal

vesi

cle

Ski

nTest

is

Cere

bru

m

Kid

ney

Sm

all

inte

stin

e

aort

a

Pla

sma

　

Okano T. et al., JBC 2008;283:11270-11279

LC-MS and NMR

MK-4 epoxide

PK epoxide

PK

Time,min

Authentic vitamin Ks

PKPK

epoxide

Time,min

PK

MK-4

MK-4 epoxide

PK epoxide

MK-4 fraction

Inte

nsi

ty,c

ps

MK-4

MK-4 epoxide PK

PK epoxide

Identification of MK-4 from brain of mice by LC-MS/MS

MK-4

H-5,88.06 H-6,7

7.67

H-2’,6’,10’,14’5.03

H-1’3.45

CH3 （ C-2 ）2.17

012345678 （ ppm （

H-5,88.06

H-6,77.68

H-2’,6’,10’,14’5.08

H-1’3.35

Authentic MK-4

MK-4 fraction CH3 （ C-2 ）2.17

Identification of MK-4 from brain of mice by 1H NMR spectrometryO

O

1 2

345

6

7

8

2’ 6’ 10’ 14’

1’

1 w 24 hr

PK-d7

12 hrC57BL/6J mice

7-weeks oldn=200

Fast Sacrifice

0

Oral administration

Cerebra

Analyses by LC-MS/MS and 2H NMR

Isolation and purification of MK-4-d7

O

O 3

PK-d7

D

D

D

D

CD3

O

O 3

MK-4 -d7

D

D

D

D

CD3

Experimental design

Experiment

？

3.5e4

1.5e4

（ m/z ）

Inte

nsit

y（

cp

s）

100 200 300 400 5000

4.5e4

3.0e4

Inte

nsit

y（

cp

s）

0

0

Authentic MK-4-d7

MK-4-d7 fraction

O

O

CH3

CD3

D

D

D

D

m/z ： 194

194.3452.5

194.4 452.6

+ H

Identification of MK-4-d7 from brain of mice by LC-MS/MS

2H-5,88.10

2H-6,77.70

C2H3

2.14

012345678 （ ppm （

2H-6,77.68

2H-5,88.08

Authentic MK-4-d7

MK-4-d7fraction

C2H3

2.14

O

O

CD3

D

D

D

D

1 2

345

67

8MK-4-d7

Identification of MK-4-d7 from brain of mice by 2H NMR spectrometry

Single peroral administration (10 g/kg)

0

20

40

60

80

100

120

vehicle PK-d7 MK-4-d9

pm

ol/g

of

bra

in

MK-7-d7

MK-4MK-4 epoxide

MK-4-d7

MK-4-d7 epoxideEndogenous MK-4

Not only PK-d7 but also MK-4-d9 and MK-7-d7 are converted into MK-4-d7 and accumulate in brain of mice

O

O 3

D

D

D

D

CD3PK-d7

O

O 3

D

D

D

D

CD3

D D

MK-4-d9 MK-7-d7O

O 6

D

D

D

D

CD3

24 hr12 hr

Fast Sacrifice

0

C57BL/6J ♀mice8 weeks old

O

O 3

D

D

D

D

CD3 MK-4-d7

Phylloqunone is converted into MK-4 via integral side-chain removal

Unsaturation of side chain

O

O 3

O

O 3

O

O 3

O

O 3

D DD D

MK-4-d7CD3

D

D

D

D

CD3

D

D

D

D

CD3

D

D

D

D

CD3

D

D

D

D

MK-4-d9CD3

D

D

D

D

D D

CD3

D

D

D

D

D D

K1-d9

D DD D

CD3

D

D

D

D

CD3

D

D

D

D

O

O

O

OPhytyl side chain

Geranylgeranylside chain

33

3

PP

PO OO-

OP :

PO O-

O-

OP

PO OO-

OP :

PO O-

O-

OPK3

Exchange of side chain

O

O 3

O

O 3

K vitamins are converted into MK-4 and accumulate in

tissues

Where does this conversion take place?

Following four routes for the conversion of PK or K3 into MK-4,　　　１． Oral route　　　２． Enteral route　　　３． Intravenous route　　　４． Intra-cerebroventricular routewere examined in mice.

O

O 3

K1 -d7

D

D

D

D

CD3

O

O 3

MK-4-d7

D

D

D

D

CD3

1 w 24 hr

K1-d7, K3-d8

12 hrC57BL/6J mice

7 weeks old

Fast Sacrifice

0

Oral(1), enteral(2), intravenous(3) or intra-cerebroventricular(4) dose

Cerebra

LC-APCI//MS

Experimental design

O

O K3 -d8

D

D

D

D

CD3

D

A single dose of 10 μmol/kg BW for (1), (2), (3) At 0.1 μmol/Kg BW for (4)

Concentrations of MK-4 in cerebra of mice

0

50

100

150

200

Co

nce

ntr

atio

n（

pm

ol/g

）

Vehicle K3-d8PK-d7Vehicle K3-d8PK-d7

0

100

200

300

MK-4 MK-4 epo MK-4-d7 MK-4-d7 epo

Vehicle K3-d8PK-d7

0

50

100

150

200

250

Co

nce

ntr

atio

n（

pm

ol/g

）

Vehicle K3-d8PK-d70

50

100

150

200

250

Endogenous MK-4 Converted MK-4

oral entero

intravenous Intra-cerebroventricular

Okano T, et al, J Biol Chem, 2008; 283:11270-11279

Po K1-d7

0 1 62 3 4 5 (h)

sacrificeWistar rat

Four Sites Cannulation experiments

Bile duct Portal vein

Inferior vena cava

Thoracic Lymph duct

heart

liver

intestine

Bile (BD)

serum (IVC)

Lymph(TLD)

Serum (PV)

180.06 180.08 180.10 180.12 180.14 180.16

0

20

40

60

80

100

180.10172[M+H]+

181.16181.14181.12181.10181.08181.060

20

40

60

80

100

181.10525[M+H]+

Authentic MD-d8

MD fraction from lymph

173.10173.08173.06173.04173.02173.00

0

20

40

60

80

100

173.05982[M+H]+

Authentic MD

(m/z )

Rela

tive

Abun

danc

e (%

)O

O

DD

DD

CD3

O

O

DD

DD

CD3

D

O

O

(m/z )

(m/z )

Rela

tive

Abun

danc

e (%

)Re

lativ

e Ab

unda

nce

(%)

0

10

20

30

40

50

60

0 1 2 3 4 5 6

Con

cent

ratio

n (p

mol/g) bile

Serum from IVC

Serum from PV

Lymph

0

0.01

0.02

0.03

0.04

0.05

0 1 2 3 4 5 6Time after administration (h)

MK-4-d7

K1-d7

0

0.4

0.8

1.2

0 1 2 3 4 5 6Time after administration (h)

Con

centr

ati

on

(pm

ol/m

L) MD-d7 　

Con

cent

ratio

n (p

mol/g)

high resolution mass spectrometry (HR-MS)

bile

Serum from IVC

Serum from PV

Lymph

bile

Serum from IVC

Serum from PV

Lymph

Concentrations of K1-d7, MK-4-d7 and MD-d7

MD-d7 　

200

400

0 1 2 3 4 5 6

K 1 con

cent

ratio

n (p

mol

/mL)

600

0 1 2 3 4 5 6

MK-

4 co

ncen

trati

on(p

mol

/mL)

0

2

4

6

8

10

0

6

12

18

24

0 1 2 3 4 5 6

MD

con

cent

ratio

n(pm

ol/m

L)

Time after administration (h)Time after administration (h)Time after administration (h)

0

K1 MK-4 MD

Time course changes in serum concentrations of K1, MK-4 and MD in humans orally given K1 capsules (40

mg)

(n=5) (n=5) (n=5)

GGPPMK-4-d3

O

O

CD3

O

O

CD2

D

H

O

O

CD2

H

H

OH

:OH

CD3

H

O

O

CD2

HH3

O

O

CHD2

H3

PPOH3

PPOH3

O

O

CD3

H3

HO

H

H

OH

O

CD3

H

H3

OH

OH

CD3

H3

HO

H

H

HO

H

H

GGPP

MK-4-d2

Route 2

Route 1

Conversion pathway of MD-d3 to MK-4-d3 demonstrated by HR-MS and 1H NMR analyses (Route 2)

MD-d3

Hirota Y, Okano T, et al, J Biol Chem, 2013; 288:33071-33080

O

O

H3

K1

OH

OH

Prenylation enzyme H3

Geranylgeranyl pyrophosphate

O-P

O

O

O-

PO

O-

O

O

O

H3

MK-4 (K2)

Removal of side chain

H3

Side-chain cleavage enzyme

O

O

MD (K3)(quinone form))

Lymph and blood stream

(intestine)

Reductase (tissues)

℗-℗ ℗-℗ ：

Hirota Y, Okano T, et al, J Biol Chem, 2013; 288:33071-33080

MD(K3) is a catabolic product of oral phylloquinone (K1) in the intestine and a circulating precursor of tissue MK-4 (K2) in mammals

MD (K3)(hydroquinone form))

New paradigm of the metabolic activation of vitamin K in brain and bone

Menaquinone-4 （ MK-4 ）

O

O 3

Biological actions

active metabolite

O

O 3

O

O

Foods

O

OR n-1

R:

phylloquinone(PK)

menaquinones(MKn)

Menadione(MD)

GGCX, SXR/PXR, PKA/PKC

brain

bone

O

O 3

O

OMenaquinone-4 （ MK-4 ）

O

O 3

GGCX, SXR/PXR, PKA/PKC

Biological actions

foods

O

OR n-1

R:

？Enzyme(s)

active metabolite

Menadione(MD)

What is the enzyme(s) involved in MK-4 biosynthesis in mammals ?

phylloquinone(PK)

menaquinones(MKn)

Biosynthesis of menaquinones in Escherichia. coli

menB

nH

OH

OH

menG

menF

menEmenCmenD

COO-

OHO COO-

Chorismate

yfbB

O-Succinylbenzoyl-CoA 1,4-Dihydroxy-2-napthol-CoA

1,4-Dihydroxy-2-napthanoate Demethlmenaquinonehydroquinone（DMK-n）

Menaquinone-nhydroquinone（MK-n）

Menaquinone-n（MK-n）

O

-OOCOH

COO-

Isochorismate

O-

OH

OH O

OH

OH

S-CoA

O

COO-

S-CoA

O

O

CO2

nP2O7

3- P2O73-

n

OH

OH

H

O

O n

menA

Alignment of the amino acid sequence of Ubia(E.coli), COQ2(Homo-sapiens), Men A(E.coli) and UBIAD1(Homo-sapiens)

ubiA_[Escherichia_coli] 1 ------------------------------------------------------------ COQ2_[Homo_sapiens] 1 MLGSRAAGFARGLRAVALAWLPGWRGRSFALARAAGAPHGGDLQPPACPEPRGRQLSLSA _menA_[Escherichia_coli] 1 ------------------------------------------------------MTEQQI UBIAD1_[Homo_sapiens] 1 ----------------------------MAASQVLGEKINILSGETVKAGDRDPLGNDCP ubiA_[Escherichia_coli] 1 -MEWSLTQNKLLAFHRLMRTDKPIGALLLLWPTLWALWVATPG--VPQLWILAVFVAGVW COQ2_[Homo_sapiens] 61 AAVVDSAPRPLQPYLRLMRLDKPIGTWLLYLPCTWSIGLAAEPGCFPDWYMLSLFGTGAI _menA_[Escherichia_coli] 7 SRTQAWLESLRPKTLPLAFAAIIVGTALAWWQGHFDPLVALLALITAGLLQILSNLANDY UBIAD1_[Homo_sapiens] 33 EQDRLPQRSWRQKCASYVLALRPWSFSASLTPVALGSALAYRSHGVLDPRLLVGCAVAVL ubiA_[Escherichia_coli] 58 LMRAAGCVVNDYADRKFDGHVKRTANRPLPSGAVTEKEARALFVVLVLISFLLVLTLNTM COQ2_[Homo_sapiens] 121 LMRGAGCTINDMWDQDYDKKVTRTANRPIAAGDISTFQSFVFLGGQLTLALGVLLCLNYY _menA_[Escherichia_coli] 67 GDAVKGSDKPDRIGPLRGMQKGVITQQEMKRALIITVVLICLSGLALVAVACHTLADFVG UBIAD1_[Homo_sapiens] 93 AVHGAGNLVNTYYDFSKGIDHKKSDDRTLVDRILEPQDVVRFGVFLYTLGCVCAACLYYL ubiA_[Escherichia_coli] 118 T------ILLSIAALALAWVYPFMKR---YTHLPQVVLGAAFGWSIPMAFAAVS-ESVPL COQ2_[Homo_sapiens] 181 S------IALGAGSLLLVITYPLMKR---ISYWPQLALGLTFNWGALLGWSAIKGSCDPS _menA_[Escherichia_coli] 127 F------LILGGLSIIAAITYTVGNRPYGYIGLGDISVLVFFGWLSVMGSWYLQAHTLIP UBIAD1_[Homo_sapiens] 153 SPLKLEHLALIYFGGLSGSFLYTGGIGFKYVALGDLIILITFGPLAVMFAYAIQVGSLAI ubiA_[Escherichia_coli] 168 SCWLMFLANILWAVAYDTQYAMVDRDDDVKIGIKSTAILFGQY-DKLIIGILQIGVLALM COQ2_[Homo_sapiens] 232 VCLPLYFSGVMWTLIYDTIYAHQDKRDDVLIGLKSTALRFGEN-TKPWLSGFSVAMLGAL _menA_[Escherichia_coli] 181 ALILPATACGLLATAVLNINNLRDINSDRENGKNTLVVRLGEVNARRYHACLLMGSLVCL UBIAD1_[Homo_sapiens] 213 FPLVYAIPLALSTEAILHSNNTRDMESDREAGIVTLAILIGPT--FSYILYNTLLFLPYL ubiA_[Escherichia_coli] 227 AIIGELNGLGWGYYWSILVAGALFVYQQKLIANREREACFKAFMNNNYVGLVLFLGLAMS COQ2_[Homo_sapiens] 291 SLVGVNSGQTAPYYAALGAVGAHLTHQIYTLDIHRPEDCWNKFISNRTLGLIVFLGIVLG _menA_[Escherichia_coli] 241 ALFNLFSLHSLWGWLFLLAAPLLVKQARYVMREMDPVAMRPMLERTVKGALLTNLLFVLG UBIAD1_[Homo_sapiens] 271 VFSILATHCTISLALPLLTIPMAFSLERQFRSQAFNKLPQRTAKLNLLLGLFYVFGIILA ubiA_[Escherichia_coli] 287 -YWHF---------------- COQ2_[Homo_sapiens] 351 NLWKEKKTDKTKKGIENKIEN _menA_[Escherichia_coli] 301 IFLSQWAA------------- UBIAD1_[Homo_sapiens] 331 PAGSLPKI-------------

Search for the enzyme(s) responsible for the MK-4 synthesis

E.coliOH

OH

O-

O1,4-Dihydroxy-2-naphthoate

（ DHNA ）

nH

OH

OH

Demethylmenaquinone-nhydroquinone （ DMK-n ）

MenA

Menaquinone-n（ MK-n ）

MenG

H

O

On

nH

Menaquinone-n biosynthesis in E.coli

O

O 3

MK-4O

O

+3

P P

GGPPMD

UBIAD1 orCOQ2 ?

MenA human homologs

338 a.a

384 a.a

Chromosome 1 11.25 M

Chromosome 4 84.40 M

MenA

UbiA prenyltransferasedomain containing 1

（ UBIAD1 ）

CoenzymeQ2 homolog,prenyltransferase

（ COQ2 ）

Prenylation domain

Prenylation domain

311 a.a

Prenylation domain

Conc

entr

ation

(pm

ol/m

g pr

otei

n)

Conversion of K3-d8 or MK-4-d12 to MK-4-d7 in siControl-, siUBIAD1- or siCOQ2-transfected MG-63 cells

MD-d8 MD-d8MD-d8MD-d8

EtOH MD-d8 PK-d7 MK-4-d12

0

10

20

30

40

50

60

70

80

90

Conc

entr

ation

(p

mol

/mg

prot

ein) MK4-d7

MK4-d7 epoxide

EtOH MD-d8 PK-d7 MK-4-d12

Sf9-Control Sf9-UBIAD1 expression vector

Conversion of PK-d7 and MK-4-d12 into MK-4-d7 in Sf9 cells transfected

with siControl or pcDNA3.3-UBIAD1(UBIAD1 expression vector)

Values are means and s.e.m. (n=6). Three asterisks, P<0.001 versus control-infected Sf9 cells with the same compound treatment.

UB

IAD

1 ac

tivi

ty (

pm

ol/m

g p

rote

in/m

in-1)

0

5

10

15

20

25

30

35

40

Substrate: MD-d8

0.01 　　　 0.1 　　 1 　　 (M)

0

10

20

30

40

50

0 0.01 　　 0.1 　　 1 　　　 (M)

GGPP

Sf9-control

Sf9-UBIAD1

UB

IAD

1 a

cti

vit

y (

pm

ol/

mg

pro

tein

/min

-1)

Substrate: K3-d8 (1 M)

Sf9-control

Sf9-UBIAD1

MK-4 biosynthetic activity of UBIAD1 in microsomes prepared from

UBIAD1 baculovirus-infected Sf9 cells with geranylgeranyl

pyrophosphate (GGPP) and MD

UBIAD1-GFPTransmitted ER-Tracker Red Merge

50 m

50 m

UBIAD1-GFPTransmittedBODIPY-TR

ceramide Merge

Subcellular localization of UBIAD1 in MG-63 cells

MG-63 cells stably transfected with a UBIAD1–GFP expression vector were stained with ER-tracker Red or BODIPY-TR ceramide (red) and were detected by GFP fluorescence (green). Merged images of GFP fluorescence and by ER-marker or Golgi-marker fluorescence are shown at the right. The control construct (mock-GFP) showed a diffuse fluorescence throughout the cytoplasm.

Nakagawa K. et al., Nature 2010; 468:117-121.

相対

比 (

UB

IAD

1/

-act

in)

0.0

0.2

0.4

0.6

0.8

1.0

1.2

1.4

1.6

1.8

0

200

400

600

800

1000

1200

1400

1600

1800

2000

濃度

(pm

ol/g

組織

)

MK-4

MK-4 epoxide

0

100

200

300

400

500

600

700

K3-d

8か

らの

生成

量 (

pm

ol/g

組織

)MK-4-d7

MK-4-d7 epoxide

UBIAD1 mRNA MK-4-d7 biosynthetic activity

Amounts of MK-4 and its epoxide

UBIAD1 mRNA expression, MK-4 biosynthetic activity, concentrations of MK-4 and its epoxide

in tissues of female mice

Cer

ebru

mC

ereb

ellu

mH

eart

Live

rP

ancr

eas

Kid

ney

Ova

ryU

teru

s

Bon

e

Cer

ebru

mC

ereb

ellu

mH

eart

Live

rP

ancr

eas

Kid

ney

Ova

ryU

teru

s

Bon

e

Cer

ebru

mC

ereb

ellu

mH

eart

Live

rP

ancr

eas

Kid

ney

Ova

ryU

teru

s

Bon

e

PK

MK-4

K3

Prenylation

UBIAD1

Side-chain cleavage

GGPP

MK-4

R= H

R =

R=

O

O 3

3

OP

O

OH

OP

O

OH

HO

O

O

R

3

3

UBIAD1 is a novel biosynthetic enzyme for MK-4 that may have both side-chain cleavage and prenylation activities

Identification of UBIAD1 as a novel human menaquinone-4 biosynthetic enzyme

Kimie Nakagawa, Yoshihisa Hirota, Natsumi Sawada, Naohito Yuge, Masato Watanabe, Yuri Uchino, Naoko Okuda, Yuka Shimomura, Yoshitomo Suhara & Toshio Okano

Department of Hygienic Sciences, Kobe Pharmaceutical University,

Nature 2010; 468:117-121.

O

O

DD

D

CD3

DD

K1-d7 K3-d8

Vitamin K homologues

O

O

DD

D

CD3

D3

MK-4-d7

O

O

DD

D

CD3

D3

UBIAD1

Geranylgeranyl pyrophosphate(GGPP)

Mevalonic acid

Acetyl-CoA

Mevalonate pathway

HMG-CoA

Geranyl pyrophosphate(GPP)

Farnesyl pyrophosphate(FPP)

HMG-CoA reductase

FPP synthase

Statin

Bisphosphonate

PP

3

POO-

O

O O-

O-

O

OO

O

P O-

O-

O

P :P

MK-4 biosynthesis in tissues is decreased by the treatments with statins and bisphosphonates

MK-4 synthetic Enz. (UBIAD1)

phylloquinone

geranylgeranyl pyrophosphate

SXR/PXR signaling

Biological functions

phytol

PKA/PKC signaling

VKDP（ active for

m ）

MK-4 Vitamin K cycle

VKDP（ inactive for

m ）

GGCX CO2, O2

MK-4-(epoxide

form)

MK-4（ quinone form ）

MK-4（ hydroquinone

from ）

prenylation

Mevalonate pathway

cleavage

Possible interaction of MK-4 biosynthesis, vitamin K cycle and vitamin K action

Generation of Ubiad1 knockout mice

15001000

500300

(bp)

M.W.

Posi500 bp 100 bp

A

B

Targeting vector

3.5 kb 1.1 kb 6.7 kb

1.7 kb

: loxP

: FRT

Ubiad1exon2

Ubiad1exon1

mTOR

exon1Wild-type allele

Targeted allele

neoR

Cre recombinase

neoRDTA

F R

SalI ClaII

NheISpeI

NotISacII

KpnI

PCR primer

Ubiad1-neofloxed allele

ATG

(A) Schematic presentation of ubiad1 genome, targeting vector and disrupted Ubiad1 genome. (B) PCR genotyping of Ubiad1+/+, Ubiad1+/− and Ubiad1−/− embryos. PCR genotyping of tail DNA of Ubiad1+/+, Ubiad1+/− and Ubiad1−/− embryos. Lane 1, positive controls for Ubiad1+/− allele. Lane 2, PCR bands of Ubiad1+/− embryos. Lane 3, PCR bands of Ubiad1+/+ embryos. Lane 4, PCR bands of Ubiad1−/− embryos.

Nakagawa K, Okano T, et al, PLOS ONE 2014; 9: 1-12, e104078

Morphological examination of Ubiad1+/+, Ubiad1+/− and Ubiad1−/− embryos and weanling mice (postnatal day 1) from pregnant Ubiad1+/− mice orally administered CoQ10

N.D.

E

N.D.

**

Day 1

DUbiad1 Ubiad1 Ubiad1

GAPDH

UBIAD1

F Ubiad1 Ubiad1 Ubiad1

Ubiad1 Ubiad1 Ubiad1

E15.5

HEstaining

A

B

C

UBIAD1staining

Ubiad1-deficient mouse embryos failed to survive beyond embryonic day 7.5 exhibiting small-sized body and gasturation arrest !!!

Flox/+ Cre/+

+/++/+

Cre/+Flox+/+

+/+

Flox/+Cre/+

-/+Cre/+

×

flox/- flox/+cre/+

flox/+flox/-cre/+

Nestin-cre

Flox/+ Flox/+

+/+ Flox/Flox Flox/+ Flox/+

×

Generation of a neural cell specific Ubiad1-/- mouse

Impairment of brain function

Co-Workers

(Kobe Pharmaceutical University)

Kimie Nakagawa, Yoshihisa Hirota, Natsumi

Sawada, Yuri Uchino, Masato Watanabe,

Syusaku Kimura, Naoto Yuge, Naoko Okuda,

Yuka Shimomura, Makiko Yamane

　　

Dept. Hygienic Sciences

Kobe Pharmaceutical Univ.

(Shibaura Institute of Technology)

Yoshitomo Suhara 　　

Thank you for your attention!