Recent Advances in Vitamin K Metabolism 3 rd International Conference on Nutrition and Food Science...
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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!