949 1765 2480 3833 5372 5987743 4124 5438 7370

3386 5110 7441

capsid

P1 P2 P3

non-capsid

poly (A)60 ± n

VPg-pU

initiation oftranslation

5’termination oftranslation

polyprotein(247)

●P1(97) P2(65) P3(84)

●VP0(37) VP3(26) VP1(34)

2A2BC(48) 3AB(12) 3CD(72)

●VP4(7) VP2(30) 2C(38) VPg(2) 3Cpro(20) 3Dpol(52)

3C’(36.4) 3D’(35.6)2B

3’

IRES

digestive tract

Replication of polio virus and disseminationReplication of polio virus and dissemination

Oral infection

barrier （tonsils, Peyer’s patch）

blood stream

barrier(BBB)

central nervous system

motor nerve cell

Skeletal muscle

axon pathway

Research on Research on pathogenicitypathogenicity

1. Mechanisms for determining species-specificity

2. Mechanisms of dissemination

3. Mechanisms for determining tissue-specificity

4. Ability to cause damage to the target cell

Research on Research on pathogenicitypathogenicity

1. Mechanisms for determining species-specificity

2. Mechanisms of dissemination

3. Mechanisms for determining tissue-specificity

4. Ability to cause damage to the target cell

Role of poliovirus receptor during polio virus infectionRole of poliovirus receptor during polio virus infection

human, monkey cells

inside cell

outside cell

polio virus

Can infect Cannot infect

extraction

polio virus gene

transfection

mouse cell

Cannot infect

transfectionCan infect

outside cell

polio virus receptor

inside cell

exon 1 2 3 4 5 6 7 8

HC 3

HC 510 Kb

ATG TGA

PVR αPVR β

PVR γ

BamHI

Hindlll

Structure of Poliovirus Receptor （PVR; CD155）

C

N

SS Ｖ

SS Ｃ2

SS Ｃ2

Domain 1

Domain 2

Domain 3

Cell membrane

Cytoplasm

Principles of viral tropismPrinciples of viral tropism

１． Receptor-dependent tropism２． Protease-dependent tropism３． IRES-dependent tropism４． Natural immunity-dependent tropism５． Others

Research on pathogenicity

1. Mechanisms determining species-specificity

2. Mechanisms of dissemination

3. Mechanisms determining tissue-specificity

4. Ability to cause damage to the target cell

digestive tract

Replication of polio virus and transmission in the bodyReplication of polio virus and transmission in the body

Oral infection

barrier （tonsils, Peyer’s patch）

blood stream

barrier(BBB)

central nervous system

motor nerve cell

Skeletal muscle

axon pathway

NeurotropicNeurotropic Polio virus InfectionPolio virus Infection

hPVR ExpressingMouse (Tg21)

TissueDistribution

BBBPermeation

CNSToxicity

Mahoney

Sabin 1 ?

?

?

? Strong

Very Weak

In BBB percolation of PV in the mouse, In BBB percolation of PV in the mouse, proactive import occurs in the brain proactive import occurs in the brain independent of CD155.independent of CD155.

Speed of PV accumulation in the blood of a mouse cerebrumSpeed of PV accumulation in the blood of a mouse cerebrum

speed of accumulation ( µl / min / g tissue )

virus and control substances mouse with CD155 expression

mouse without CD155 expression

PV ( Mahoney )

Negative control ( Albumin )

Positive control( Cationized rat serum albumin )

0.164 0.123

― 0.001

0.123 ( Rat )

( Yang et al.,1997, Virology 229 , 421- 428 )

Percolation examination using thePercolation examination using theBBB BBB in vitroin vitro modelmodel

mouse cerebrovascularendothelial cell

＋

fluorescent dye

( Alexa Fluor )

fluorescent dyed PV ( 126.55 µg / ml )

Periodic measurement of fluorescence after PV addition

TimeCle

aran

ce

volu

me

( µl )

PS

( PS : Permeability-surface area product )

Clearance volume

( µl )

amount of PV moved to basolateral surface ( µg )

Primary concentration of PV at the luminal surface ( µg / µl )

＝＝

purified PV

PV percolation in the BBB PV percolation in the BBB in vitro in vitro model is extremely highmodel is extremely high

05

10152025303540

0 10 20 30 40

PV （ mw 9800K ）

Dextran （ mw 2000K ）

Transferrin （ mw 80K ）

Dextran （ mw 70K ）

0

5

10

15

20

25

30

0 10 20 30 40Time ( min )

PS = 0.7953

PS = 0.3103

PS = 1.0877

PS = 0.2112

Cle

aran

ce v

olum

e ( µ

l )

Cle

aran

ce v

olum

e ( µ

l )

Time ( min )

Examination of PV intake into a mouse Examination of PV intake into a mouse cerebrovascularcerebrovascular--endothelial cellendothelial cell

TransferrinDextranDextran

（（ mw 3K mw 3K ）） raft

Cholera Toxin

TransferrinTransferrinreceptorreceptor

DextranDextran（（ mw mw ＞＞ 15K 15K ））

fluorescent dyed PV

path between intercellular gap

??

Pathway into Pathway into endosomeendosome

??????

PVPV ++ DextranDextran （（mw. 3K mw. 3K ））

PVPV ++ Cholera ToxinCholera Toxin

_

Dextran( mw15K - 20K )

667μg / ml 1mg / ml

_ m.o.i. 5000Purified PV

EndocytosisEndocytosis of of transferrintransferrin ( ( TfTf ) is inhibited by PV ) is inhibited by PV

20 µm

Alexa 488 - Tf10μg / ml

( MBEC4 cell )

PV percolation in BBB PV percolation in BBB in vitro in vitro model is lowered by model is lowered by TfTf

0

5

10

15

20

25

30

35

0 5 10 15 20

Time (min)

Cle

aran

ce v

olum

e (µ

l)

PV

Tf + PV

FITC-Dextran(2000K)

PS = 1.89

PS = 1.02

digestive tract

Replication of polio virus and transmission in the bodyReplication of polio virus and transmission in the body

Oral infection

barrier （tonsils, Peyer’s patch）

blood stream

barrier(BBB)

central nervous system

motor nerve cell

Skeletal muscle

axon pathway

Sampling time PV antigen

1.5h － － ＋

3h ＋ ＋ ＋

6h ＋ ＋ ＋

12h － ± ±

Polio virus antigen inside an Polio virus antigen inside an ischiadicischiadic nerve axon nerve axon 2cm away from administration2cm away from administration

Tg +PV

Merge α-bungarotoxin PV

20μm

PV localized at neuromuscular junctions in PV localized at neuromuscular junctions in TgTg micemice

Neuromuscular junction of a Neuromuscular junction of a TgTg mousemouse1.5 hrs after PV injection1.5 hrs after PV injection

Immunity electronmicroscope(detects PV )

Transmissionelectronmicroscope

100nm

100nm100nm

Principles of viral tropismPercolation examination using the� BBB in vitro modelPV percolation in the BBB in vitro model is extremely highExamination of PV intake into a mouse cerebrovascular-endothelial cellPV percolation in BBB in vitro model is lowered by Tf