Vascular Immunology UnitVascular Immunology Unit

Transition écologique du paludisme et implication thérapeutique

Transition écologique du paludisme et implication thérapeutique

Ronan Jambou MD, PhD

VascularImmunology

Unit

1. Who is ill Rural Malaria / Urban Malaria

2. « BBB alteration » hypothesis • Histamine and BBB • Effector role of platelets• Microparticules• Direct effect of Infected red blood cells

PLAN

1. Who is ill Rural Malaria / Urban Malaria

2. « BBB alteration » hypothesis • Effector role of platelets• Histamine and BBB• Microparticules

PLAN

Severe Malaria

NewbornfeverdehydrationconvulsionMetabolic disorders= impaired consciousness

ChildSevere anaemia +++High parasitaemia +++Cerebral pathology

AdultRespiratory distresscerebral malarialow parasiteamia= delay in treatment

29

15 17

53

6154 55

85 82

42

76

3737

47

74

Monde Afrique Asie Amériquelatine/Caraïbes

Régions plusavancées

1950 2000 2030 (projections)

Source : Nations Unies, Perspectives de la population dans le monde, Edition 2003 (scénario moyen), 2004.

From rural to urban: trends of the population From rural to urban: trends of the population

15°

14°

13°

16°N

Dakar

17°W 16° 15° 14° 13° 12°

SENEGAL

Area of Dakar

Seasonal transmission

4 M inhabitants = 1/3 Senegal

3.5% surface

What’s a seasonal transmission

Gouly coulyIncidence of Malaria in a small village Gouye Kouly 08/2004-08/2006

0

50

100

150

200

250

8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8

2004 2005 2006mois et années

nom

bre

é

piso

des épisodes non febriles

épisodes fébrilesaccés

In West Africa

Prevalence and risk of Malaria are highly variable

They summarize all the parameters of the transmission

risk is not the same everywhere and for everyone

Adaptation of strategies

WHO Africa report 2006WHO Africa report 2006

ITN implementationITN implementationChange in drug strategyChange in drug strategy

Expected decrease of Rural Malaria transmissionExpected decrease of Rural Malaria transmission

Swaziland WHO Report 2006Swaziland WHO Report 2006Kilifi _ Kenya Malaria J 2007Kilifi _ Kenya Malaria J 2007

Kilifi _ East Africa Mwangi et al JID 2005Kilifi _ East Africa Mwangi et al JID 2005

Changes in transmission modulate Malaria profil Changes in transmission modulate Malaria profil

53%32Saharevo, Madagascar: ≤1 b.i.a./human/year

50%25Pikine, Sénégal: ≤1 b.i.a./human/year

41%62Ndiop, Sénégal: 10-30 b.i.a./human/year

23%43Dielmo, Sénégal: 100-300 b.i.a./human/year

10%25Danané, Côte d'Ivoire: ≥ 300 b.i.a./human/year

% withinadluts

Total Nb of attacks 60 years old

53%32Saharevo, Madagascar: ≤1 b.i.a./human/year

50%25Pikine, Sénégal: ≤1 b.i.a./human/year

41%62Ndiop, Sénégal: 10-30 b.i.a./human/year

23%43Dielmo, Sénégal: 100-300 b.i.a./human/year

10%25Danané, Côte d'Ivoire: ≥ 300 b.i.a./human/year

% withinadluts

Total Nb of attacks 60 years old

Annu

al N

bof

mal

aria

atta

cks

0

1

2

3

4

5

6

0 5 10 15 20 25 30 35 40 45 50

Age (years)

60

Annu

al N

bof

mal

aria

atta

cks

0

1

2

3

4

5

6

0 5 10 15 20 25 30 35 40 45 50

Age (years)

60

West Africa / Madagascar (meta-analysis)West Africa / Madagascar (meta-analysis)

Major challenge for the next 10 years

Changes of risks Changes in control strategies

Prevalence of malaria in consultations

0

0,05

0,1

0,15

0,2

0,25

0,3

septembre octobre novembre decembre

00,10,20,30,40,50,6

septembre octobre novembre decembre

00,050,1

0,150,2

0,250,3

0,350,4

sept oct nov dec

part of class of age in the malaria cases

prevalence of severe malaria.

0

200

400

600

800

1000

1200

1400

M F M F M F M F M F0-1 an 1- 4ans 5-14 ans 15- 49 ans 50ans &+

consultations by age

0-1 y1- 4 y5-14 y

15- 49 y> 50 y

From children to adults From children to adults

« most of the cases occur among children under 5 years » : still true ?

« most of the cases occur among children under 5 years » : still true ?

Nb countries

Cases= or > 5y

Cases < 5y

ratio CountriesDeaths ratio >1

(>= 5y / <5) total 25 19.5 13.4 1.4

West Africa 13 9.5 4.9 1.88 9/4

Central Africa 6 3.7 3.2 1.12 3/0

East – southern 9 6.2 5.3 1.2 5/3

WHO African report 2006 : data 2005WHO African report 2006 : data 2005

57,8%31,7%35,3%patients consulting before 4 days after beginning of the symptoms

18,4 %38,5 %27,5 %patients treated before consultation

1043612762656Parasitemia (paras. / µL)

10,612,29,7Hemoglobin (mean g/L)

14 %12,5%14,7%Patients with temperature > 40°C (%)

0.81.51Sex ratio

13.7 (11)15.5 (13)14.5 (10.3)Mean age (SD)

1024234N° patients

Mild Mal. dispensary

Mild Mal Hospital

Cerebral malaria

CM is associated with delay in efficient treatment

Self treatment / DelaySelf treatment / Delay

+++triple+++++two

+-+oneNRImutant

SCNwild1085951codon

MM n= 28CM n= 16

MM n= 102CM n= 37

DHFR CRT exon 2

0%5%

10%15%20%25%30%35%40%45%

Wild 1 mutat. 2 mutat. 3 mutat.

0%

10%

20%30%

40%

50%

60%70%

80%

90%

CVIET CVIET/CVMNK CVMNK

CM

MM

CM is associated with drug resistance

0102030405060708090

100

Cerebral Malaria

Mild Mal (hosp)

Mild Mal. (dispens)

0

0,5

1

1,5

2

2,5

Percent of multi-infectionn° isolates / patients

0

5

10

15

20

25

Fann HPD Gued All0,660,680,70,720,740,760,780,80,82A

He

HPD n=59GUED n=129

-15 -5 5 15-15

-5

0

5

10

15

Axis 1

Axi

s 2

10

CM is associated to multi-infection and specific isolates

FstatFstat

PCAPCA

toxtox

pathogenpathogenpathogenhosthosthost

toxtox

Infectious pathologyInfectious pathology

ImmunopathologyImmunopathology

Which mechanisms ?

Adaptation of the treatmentHealthcare supply

Adaptation of the treatmentHealthcare supply

models models additive treatments additive treatments

toxtox

pathogenpathogenpathogenhosthosthost

toxtox

Infectious pathologyInfectious pathology

ImmunopathologyImmunopathology

Which mechanisms ?

Adaptation of the treatmentHealthcare supply

Adaptation of the treatmentHealthcare supply

additive treatments additive treatments

1. Who is ill Rural Malaria / Urban Malaria

2. « BBB alteration » hypothesis • Role of platelets and TNF• Histamine and BBB• Microparticules• Direct effect of IRBC

PLAN

sequestration

Schofield & Grau

5: 722-735, 2005

Hunt & Grau

24: 491-499, 2003

mechanisms?mechanisms?

Plasmodium?

NKT

inflammatory CTKs ↑ Mo

CD8

CD4

NKMφ

DC

?

Schofield & Grau

5: 722-735, 2005

Mo?

endotheliumendothelium

Which effectors

Which interaction between blood cells?

1. Who is ill Rural Malaria / Urban Malaria

2. « BBB alteration » hypothesis • Role of platelets and TNF• Histamine and BBB• Microparticules• Infected red blood cells

PLAN

TNF: a central mediator in immunopathology

adhesionmolecules

microvessel(brain, lung, ...)

TISSUE LESIONS :graft-versus-host

pulmonary fibrosis granuloma

arthritisDTH

TNF

anti-TNF antibody VASCULAR LESIONS :

septic shock (*)cerebral malaria

(*) Beutler & Cerami, 1985

Grau et al., Immunol Rev; 112: 49-70 (1989)Int Rev Exp Pathol 34: 159-171 (1993)

Cytokines and pathology: approaches

+S ?cytokinecytokine

mini-pumps

Δ pathology?S

Pathogen

R

S + ?antibodies,inhibitors

observationobservation

inductioninduction

inhibitioninhibition

brain endothelial cells

TNF plays a major role in sequestrationTNF plays a major role in sequestration

Lucas et al., Eur J Immunol 27: 1719, 1997Lou, Lucas & Grau Clin Microbiol Rev 14: 810, 2001

Stoelcker et al., Infect Immun 70: 5857, 2002

monocyte bindingICAM-1 upregulation

platelet binding

memTNF

TNFR2TNFR2

sol or

memLT

PRBC binding

Functional consequence of TNFR2 upregulation (Functional consequence of TNFR2 upregulation (mouse modelmouse model, PbA infection), PbA infection)

TNF-dependent pathology is also platelet-dependent

Pulmonary fibrosisPulmonary fibrosis

Cerebral malariaCerebral malaria

Shwartzmann reactionShwartzmann reaction

DTHDTH J Exp Med. 1991 Mar 1;173(3):673-9 J Exp Med. 1991 Mar 1;173(3):673-9

Nature. 1990 Mar 15;344(6263):245-7.Nature. 1990 Mar 15;344(6263):245-7.

Science. 1987 Sep 4;237(4819):1210-2.Science. 1987 Sep 4;237(4819):1210-2.

J Leukoc Biol. 1993 Jun;53(6):636-9.J Leukoc Biol. 1993 Jun;53(6):636-9.

Accumulation of platelets in a TNF-mediated pathology (LPS shock)

Platelets: the conventional viewPlatelets: the conventional view

PreventionPrevention of LPSof LPS--induced mortalityinduced mortalityby antiby anti--platelet antibodiesplatelet antibodies

LPSLPS survivors/total (%)survivors/total (%)

--++++++++++

10/10 (100)10/10 (100)4/41 (10)4/41 (10)

0/6 (0)0/6 (0)0/6 (0)0/6 (0)

10/12 (83%)10/12 (83%)7/9 (77%)7/9 (77%)

nonenonenonenonenormal mouse IgGnormal mouse IgGnormal rabbit IgGnormal rabbit IgGmouse antimouse anti--platelet mAbplatelet mAbrabbit antirabbit anti--platelet IgGplatelet IgG

pretreatmentpretreatment

Prevention of hemorrhagic necrosis by anti-platelet or anti-CAM mAbs

Prevention of hemorrhagic necrosis Prevention of hemorrhagic necrosis by antiby anti--platelet or antiplatelet or anti--CAM mAbsCAM mAbs

0 25 50 75 100 125

*

*

*

-- 24 h (id)24 h (id) -- 2 h (ip)2 h (ip) 0 h (id)0 h (id)

PBSPBS

LPSLPS

LPSLPS

LPSLPS

LPSLPS

LPSLPS

nonenone

nonenone

normal IgGnormal IgG

antianti--CD11aCD11a

antianti--CD54CD54

antianti--plateletplatelet

TNFTNF

PBSPBS

TNFTNF

TNFTNF

TNFTNF

TNFTNF

5151CrCr--erythrocytes (% cpm)erythrocytes (% cpm)

Platelet-EC interactions

microvessel

stroke

metastasis

angiogenesis

THROMBOSIS

INFLAMMATION

Mol. Pathol. 50: 175-185 (1997)

PRBC

Wassmer et al., J Infect Dis 189: 180-9, 2004

PLT

PLTPLT

PLT

ECEC

PRBC

In vitroIn vitro evidence for a role of platelets in evidence for a role of platelets in PRBCPRBC--EC bridgingEC bridging

Wassmer et al., J Immunol 176: 1180-1184, 2006

Other roles of platelets in malaria: clumpingOther roles of platelets in malaria: clumping

Pain et al., Proc Natl Acad Sci USA 98: 1805-10, 2001

clump

•• aggregation of PRBCaggregation of PRBC

•• formation of giant clumps formation of giant clumps

•• role in sequestration and in role in sequestration and in ““sludgingsludging””

1. Who is illRural Malaria / Urban Malaria

2. « BBB alteration » hypothesis• Effector role of platelets• Histamine and BBB• Microparticules• IRBC

PLAN

Major role in: Major Major rolerole in: in: ECECEC

Vascular permeability

DiabetesDengue feverAge degenerescenceARDSMyocardial infarctionTumour angiogenesisBrain injury

DiabetesDengue feverAge degenerescenceARDSMyocardial infarctionTumour angiogenesisBrain injury

VVO Junctionremodelling

pinocytosis

pathogenspathogenspathogens stimulistimulistimuli

Histamine induces vascular congestionGuido Majno (1961)

vein

arteriole

Histamine induces opening of intercellular junctionsJ cell Biol (1969)

Histamine induces opening of intercellular junctionsJ cell Biol (1969)

Histamine

HRF/TCTPbasophyls

cytokine network (IL4, IL2..) eicosanoid pathway

endothelial cells responseto inflammation

platelets / neutrophils

DC response to TH2 typeproduction of IgE

vascular permeability

Cerebral malaria ??Histamine Increases

during malaria

( HR1, HR2, HR3, HR4, HR5 )

IgE AntigenAllergy / helminths

Histamine can modulate endothelial and immune cells functions

15°

14°

13°

16°N

Dakar

17°W 16° 15° 14° 13° 12°

SENEGAL

Area of Dakar

Seasonal transmission

4 M inhabitants = 1/3 Senegal

3.5% territory

An arabiensis

Gouly Couly

Longitudinal study: Gouly couly Longitudinal study: Gouly couly

Nov. 04Baso, IgE

Follow up

EnrolmentJun 04

Jun 05Baso, IgE

july 04IgE

EndJun 06

Rainy season Rainy seasondry season dry seasondry season

Drug resistance study

Low

high

medium

0

20

40

60

80

100

9-19 20-39 40-80

0

2

4

6

8

10

Total IgE ug/L

9-19 20-39 40-80

0

0,5

1

1,5

july 04 Nov 04 July 05

IgE

ug/m

L

0

0,5

1

1,5

2

2,5

IgE

ug/m

L

05

1015202530

1 2 3

IgE

ug/m

L

% of subjects with > 0.4µg/l

0

5

10

15

20

25

30

35

40

45

9-19 20-39 40-80

% Ig

E_Pf

pos

itive

nov_04Jun_05

% of subjects with IgE-Pf

No difference according to age for

high IgE level

Stability of IgE level

Higher percent of Pf-IgE during dry

season

IgE responses : Gouly couly IgE responses : Gouly couly

IgE Deposition in brain microvessels and on parasitized erythrocytes from cerebral malaria patientsYoshimasa maeno, AJTMH 2000,

Immunoglobulin E (IgE) containing complexes induce IL-4 production in human basophils: effect on Th1/Th2 balance in malariaM.A. Nyakeriga, Acta Tropica 2002

IgE+anti-E

IgG+anti-G

IgG+anti-E

Pathology ? TH2 response ?

IgE increase in CM and can induce IL4 productionIgE increase in CM and can induce IL4 production

Basophiles responses : Gouly couly Basophiles responses : Gouly couly

standard antigens

flmpD farinaeD pteronyssinus

0102030405060708090

9-19y 20-39y 40-80y

Perc

ent r

espo

nder

s nov jun

Percent of responders maximum during rainy

season

Pf can induce response for 10% of villagers during rainy season

High response for salivary glandsHemozoin

Pf ghostSalivary glands

0

1020

30

40

5060

70

80

9-19y 20-39y 40-80y

nov jun nov jun nov jun

Perc

ent r

espo

nder

s

Mosquitoes can trigger basophiles

activation

Pf antigens

Mosquitos bites induce mast cells activationJI Demeure (2005)

Mosquitos bites induce mast cells activationJI Demeure (2005)

ECIS™: Electric Cell-substrate Impedance Sensing

Monitoring of cells monolayer impedance to measure junction permMonitoring of cells monolayer impedance to measure junction permeabilityeability

HBEC-D3

HBEC1.5 h

washingHistamine, TNF

Spreading: 5 days

TEER

24 h

IRBC, NRBC

Attachment and spreading of the cells

0

5000

10000

15000

20000

0 20 40 60 80 100

Time (hours)

Res

ista

nce

(ohm

s)

Histamine or platelet supernatant induce opening of junctions

0

2000

4000

6000

8000

10000

12000

14000

16000

18000

29 29.5 30 30.5 31 31.5 32

stimulationHistamine

Control medium

hours

Resistanceohm

LPS

PLT SN

0

2000

4000

6000

8000

10000

12000

14000

16000

18000

23 28 33 38 43

stimulationMedium change

Histamine

Control medium

hours

Resistanceohm

LPS

PLT SN

ratio

at T

0

-0,35

-0,3

-0,25

-0,2

-0,15

-0,1

-0,05

0

0,05

0,1

1 2 3 4 5 6 7 8 10 11 12 13 14 16 17 18 19 20 21 22 23 24 25

his100-anti_H1histamine100TNFhist100-antiH2control

H1 receptor mediates histamine effect on HBECs

Histamine 100 µM

Histamine induce calcium mobilization thru HR1Histamine induce calcium mobilization thru HR1

Histamine 1 µM

-1000

100200300400500

0 5 10 15 20 25Time (min)

Ca

(nM

)

Histamine 10 µM

Histamine

Time (min)

0.6

1.6

2.6

3.6

0 20 40 60Time (min)

Nor

mal

ised

Rat

io

Diphenylhydramine (20µM)

Histamine

Cimetidine (40µM)

0.6

1.6

2.6

3.6

4.6

0 20 40 60 80

Nor

mal

ised

Rat

io

Histamine

(Pre-incubation 40 min)

H1

H2

Histamine increases VCAM but not ICAM expression

0

2

4

6

8

10

12

14

16

18

control Histamine100 Histamine100

+ antiH1H2

Histamine_10 Histamine_1

Per

cent

of p

ositi

ve c

ells

Histamine 100µM, 10µM, 1µMCimetidine 20µM, diphenylhydramine 20µM,

39.128.4237.246.

5.

DPHCiticoline ControlDay post-PbA infection

ControlCiticholinediphenyhydramine

0 7 140

25

50

75

100

Day post-PbA infection

Perc

ent s

urvi

val

Anti_H1 increases survival of P berghei infected mice

Beghdadi et al 2008

Histamine release and HRFsHistamine release and HRFs

Human TCTP (IgE-dependent HRF)-Gene locus 13q12-q14

- induces histamine release by basophiles of atopic patients

- in vitro : increase reactivity of basophiles to other stimulus (IgE, ..etc)

- induce proliferation of B cells and activation of eosinophiles

TCTP- found in all eukaryotes , with two conserved motifs (microtubules binding domain)

- control cells proliferation, division, and apoptose => overexpression in cancer

Induction of idiopathic allergy ?

Plasmodium can induce histamine release : TCTP/HRF protein

Plasmodium can induce histamine release : TCTP/HRF protein

PfTCTP is expressed by late trophozoitesPF

TCTP

Tr

ansc

ript R

Q

2 8 14 20 3226 38 44 50 hours0,5

11,5

22,5

33,5

44,5

05

10152025303540

P0 P6 P12 P18 P30P24 P36 P42 P48

- synchronisation + sampling every 6h over 54h = « sampling time »

- use of thin smear and timing genes (P David) to define the « parasite time » (gene 8h= MAL8P1.4, gene 12h=PFI1735c- normalisation of mRNA on average of (N1= PFC0255c and N2=PFA0570), then at time with less mRNA (T44 = 1 for PfTCTP)

Parasite cycle time

Sampling time

gene T8hgene T12h

Tim

ing

gene

s

Pf TCTP- 38% identity, 53% similarity with hTCTP

- mimics hTCTP in vitro on histamine release

- 0.1 to 1µg/ml in serum from patients with malaria

- totally conserved in 350 P falciparum field isolates => target human cells ??

PfTCTPhTCTP

10 µg/ml

His

tam

ine

histamine release by basophils induced by rTCTP is IgE dependant(MacDonald et al 2001)

Plasmodium can induce histamine release Plasmodium can induce histamine release

100 µg/ml

Conclusion 2 Conclusion 2

20% of population with high IgE level = stable

Seasonal variation of basophiles activation

Impact of mosquitoes bites on histamine release

Histamine modulates endothelial cells thru H1R

Histamine induces rapid opening of intercellular junction responsive for edema

PfTCTP poorly active on HBEC ( ongoing .. Hypoxia) => HRF

Anti-histamine = new way to improve the treatment of CM ?

1. Who is ill Rural Malaria / Urban Malaria

2. « Parasite » hypothesis3. « BBB alteration » hypothesis

• Effector role of platelets• Histamine and BBB• Microparticules• IRBC

PLAN

Membrane vesiculationMembrane vesiculationMembrane vesiculation

From Zwaal et al.From Zwaal et al.

Phosphatidyl-serinePhosphatidyl-serine

Phosphatidyl-ethanolaminePhosphatidyl-ethanolamine

resting

HUVEC

EMP

250

0

50

100

150

200

3EM

P / 1

0ce

lls

0 10 100 1000TNF (ng/ml)

Release in culture

Combes et al., J. Clin. Invest. 104: 93, 1999

Electron microscopy+ TNF

TNF enhances MP production by endotheliumTNF enhances MP production by endothelium

Combes et al, JAMA 291: 2542-4, 2004

acute phasefollow up

025

50

75

100

125

150

175

EMP

/ µl p

lasm

a

60N 138 80 37 35 27 1348

p = 0.01p = 0.005

p < 0.0001

Malaria - + + + + + + +CM - - + + - - + +SMA - - - - + + + +

Dramatic increase of plasma endothelial microparticles in Malawian children with CM

Dramatic Dramatic increase increase of pof plasma endothelial lasma endothelial microparticlesmicroparticles in Malawian children with in Malawian children with CMCM

Citicoline can protect against CMCiticoline can protect against CM

Protection of mice injected with PBA

Injection 1g/kg day 4 to end

First trial in human (Dakar HPD 2007)

Qunine + CTC 1.5g/day

no significant improvement of mortality at this dose

0 7 14 210

25

50

75

100 Control

Citicoline

Artesiminin

Day post-PbA infection

%su

rviv

al CTC + Artesiminin

1. Who is ill Rural Malaria / Urban Malaria

2. « BBB alteration » hypothesis • Effector role of platelets• Histamine and BBB• Microparticules• IRBC

PLAN

NKT

TregMo

CD8

host cells

P. falciparum-infected red cells

brainendothelialcellmembranes

What happens during IRBC/endothelial cells contactsWhat happens during IRBC/endothelial cells contacts

AutoMACS® - purified PRBC

CoCo--cultures of parasitised red blood cells (PRBC) and cultures of parasitised red blood cells (PRBC) and human brain endothelial cells (HBEC) human brain endothelial cells (HBEC)

HBEC 1 h 30MicroscopyTEERQ PCRFlow cytometry

unbound cellremoval

0 / 1 h / 2 h / 4h

TNF

O/N

INCUBATION

washing

PRBC-PKH26 Calcein AM

HBEC D3 + IRBC (3Ci)HBEC D3 + IRBC (3Ci)1 h incubation1 h incubation

HBEC

PRBC

hemozoin

merozoite

knobs

HBEC D3 + IRBC (3Ci)HBEC D3 + IRBC (3Ci)Engulfing ?Engulfing ?

HBEC 1 h 30 confocal microscopy

PRBC-PKH26

washing

0 / 1 h / 2 h

TNF

ON

washing

HIS + anti human-IgG

AutoMACS® - purified PRBC

[HIS : pool of 10 hyper-immune sera from African adults with malaria]

30 min co30 min co--cultureculture 40 min co40 min co--cultureculturebefore washingbefore washing

Beginning of transferAdhesion

HBEC: D3 + IRBC-PKH26-calcein

Ration : 50 RBC/1 HBEC

HZ

PKH26

Mergecalcein

1 h 30 min1 h 30 min coco--culture culture (after washing)(after washing)

Diffusion of membrane compounds

HBEC: D3 + IRBC: 3Ci

HZ

4 h4 h coco--cultureculture

Diffusion of membraneand cytosoliccompounds

HBEC+ IRBCcalcein

PKH26

HBEC

HBEC + PRBC-PK26 HBEC + PRBC + HIS Merge

PRBC

5.5 h5.5 h coco--culture HBEC / parasitised RBCculture HBEC / parasitised RBC

Time (0.1 hour)

0

0,2

0,4

0,6

0,8

1

1,2

1,4

1 6 11 16 21 26 31 36 41 46 51 56 61 66 71 76 81 86

resi

stan

ce ra

tion

(T/T

inje

ctio

n)

NRBC + IRBC (vol/vol)

0

0,2

0,4

0,6

0,8

1

1,2

1,4

1 7 13 19 25 31 37 43 49 55 61 67 73 79 85 91 97 103

Time ( 0.1 hour)

resi

stan

ce (r

atio

T/T

0)

controlNRBCIRBCHistamine 100

IRBC but not NRBC induce opening of the junctions IRBC but not NRBC induce opening of the junctions

Stimulus Stimulus

Early endosomes

Cell activation

Phosphorylation +++Apoptosis ?Src activation ?

Opening of junctions

Recyclingpresentation Increase of VCAM

TLR , Coagulation

Modulation of genesexpression

T cellsantibodies

Ca ++

MP=0

Antigen presentation

Y YYIRBC

Adhesion of IRBC

Trogocytosis-like adhesion and transfer

start as soon as 30 min

Membrane transfer

HBEC

Conclusion 3 Conclusion 3

Structure of parasite/HBECInterface and

proteins (SNARE , Vamp)

Dakar

Cell activation (Src, Rac1)

Opening of junctions

Recyclingpresentation

T cellsantibodiesAntigen presentation

Y YY

Adhesion of IRBC

Structure of the interface proteins involved ?

Africa

Ongoing Ongoing

Field isolates

T and B cells

University SydneyV Combes

A Sanchez PerezF ElassaadMJ Jambou

GE. Grau

IMTSSA- Le PharoS PelleauD Parzy

Institut Pasteur de Dakar

D AldebertL Marrama

R PaulY Seck

ML VarelaF Diène-SarrIbrahima Dia

Hopitaux de DakarB Diop

JC Moreau

Institut Pasteur (PF5)F Nato

P Beguin