Pathogenic Mechanisms of HIV Transmission by Breastfeeding
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Transcript of Pathogenic Mechanisms of HIV Transmission by Breastfeeding
Pathogenic Mechanisms of HIV Transmission by Breastfeeding
Philippe Van de Perre
Montpellier University Hospital, University of Montpellier 1
Research Unit 4205
IAS 2007, Sydney
Mechanism(s) of breastfeeding transmission of HIV: the moving target
An evolving host A complex and biologically active source of infection
Cells/ml
Weeks of lactation
103
104
105
2 4 6 8 10 12AS Goldman et al. J Pediatr 1982
Macrophages
Lymphocytes
Compartmentalization of HIV-1 in breast milk
Blood Colostrum
Mother # 25
Mother # 35
Mother # 65
M2m1
M2m1
m6
m4
m3m4
M2
M1
M7
m7M8
M5
M9
Viral DNA
Viral DNA
Viral DNA
Viral RNA
Viral RNA
Viral RNA
M3
P Becquart et al; Virology 2002
Further evidence for HIV compartmentalization in breast milk
• Different distribution of HIV quasispecies in right versus left breast, and in some women in different fractions of milk: suggest local micro environmental selection pressure (P Becquart, Virology 2007)
• In half of NVP-treated women: different expression of resistance profiles between plasma and breast milk (E Lee, J Infect Dis 2005)
Portal of entry
Mucosal lesion
AlternativeReceptor(Gal Cer)
Fcreceptor
Transcytosis M cells
T LymphocytesMonocytes/macrophages
DendriticCells
Ep
ithel
ial c
ells
Free virus
InfectedCells
Lum
en
Su
b m
uco
sa
From M Neutra, 1999
From M Neutra, 1999
Polarised HIV-1 infected cell
Gal Cer
Transcytosis in an enterocyte
Macrophages, lymphocytes and dendritic cells in the lamina propria
M Bomsel, 1997
ApicalChamber
BasalChamber
Target cells
EpithelialCellsMembrane
Transcytosis of HIV-1 across human enterocytes
• Concept of viral synapse
• HIV-1 gp41 recognises a membrane agrin (heparan sulfate proteoglycan) that favour interaction with GalCer and mediate transcytosis through an integrin associated mechanism
A Alfsen, 2005
Oral inoculation of macaques with SIV
* Non traumatic inoculation of tonsils with cell-free
and cell-associated SIV
• No infection of the epithelial cells
* Infection of CD4+ T cells close to M cells in the tonsil crypts
C Stahl-Henning, 1999
Breastfeeding transmission of HIV-1: by free virions or by HIV-infected cells?
Cell-free and cell-associated HIV-1 are both responsible for breast milk transmission
I Koulinska, 2006
Cell-free virus Cell-associated virus indetermined
< 9 m post p 2 8 6
> 9 m post p 11 8 5
Total 13 16 11
p=0.03
HIV-1 Transmission
SpinPlasma
FicollHypaque
Unwanted cells are cross-linked-to red blood cells
Breast milk cells plus
red blood cells of healthy control
Blood
EnrichedCD4+ TcellsFicoll -Hypaque
Redbloodcellsandrosettedcells
Irradiatedcells
Resting
CD4+ T cells
Day 5
Day 10
Day 0
Anti -CD3antibodies
Anti -CD28antibodies
Activated
CD4+ T cells
Quantification of HIV-1 DNA
by real-time PCR
Enumeration of the HIV-1-Ag SCs
by ELISPOT assay
HIV-1antigens
Detection of p24 antigen in
supernatantsby ELISA
A
B
18
Proportion of latently infected cells able to enter viral cycle
Blood Breast milk
HIV-1 DNA copies 6.948 4.788per 106 T CD4+ cells (2.351-23.043) (2.590-47.294)
HIV-1 Ag secreting cells 45 (9-108)* 500 (205-934)*per 106 T CD4+ cells
* Wilcoxson rank sum test for paired samples, P < 0.01
% of HIV-1 infected T CD4+ cells 0,9 - 1,8% 10,4 -32,4%able to enter viral cycle
1 to 3 copies / infected cell
(P Becquart, 2006)
Macrophages and dendritic cells in breast milk
-Are morphologically distinct from their possible precursor PBM
- Spontaneously produce GM-CSF
- Express DC-SIGN gene and protein
- Differentiate into CD1+ dendritic cells after incubation with IL4
M Ichikawa, 2003
Breast Milk macrophages
Soluble factors in breast milk
Soluble factors and innate immune factors with potential anti-HIV activity?
• Lactoferrin (suggested in vitro) (MC Hamsen, 1995)
• Lewis factorX (binding to DC-SIGN) (MA Naarding, 2005)
• SLPI (suggested in vitro) (SM Wahl, 1997)
• Defensins ( L Kuhn, 2005; R Bosire, 2007))
• Lysozyme ?• Complement ? • Mucines ?• Prostaglandins ?• Interleukins?
“The anti-infective activity of human milk is potentially greater than the sum of its microbicidal components” (CE Isaacs, 2004)
Anti-HIV IgG, IgA and IgM
in breast milk (Kigali, Rwanda 1988-1991)
50%
100%
0% D 15 M 6 M 18
% HIV+ mothers with +WB
in milk sample
Time post partum
IgG
IgA
IgM
P Van de Perre, 1993
Perspectives
MacrophagesMacrophagesInfected Lymphocytes Infected Lymphocytes
-
Free Virus Free Virus
Systemic CompartmentSystemic Compartment Mammary CompartmentMammary Compartment
MacrophagesMacrophagesActivated Infected Activated Infected
LymphocytesLymphocytes
« Reservoir Cells »« Reservoir Cells »
Macrophages,Macrophages,Infected LymphocytesInfected Lymphocytes
« Reservoir Cells »« Reservoir Cells »
Mucosal compartmentMucosal compartment
Free VirusFree Virus
Soluble Soluble factors?factors???
??« Reservoir Cells »« Reservoir Cells »
Infection of T CD4+ quiescente CellsInfection of T CD4+ quiescente Cells
CTL?
-
Why is the majority of infants exposed to HIV
through breastfeeding escaping infection?
• Presence of HIV-specific MHC class I-restricted CD8+ CTLs in breast milk (S Sabbaj, 2002; BL Lohman, 2003)?
• HIV specific antibodies?
• Other immune factors ?
Daily ingestion of 322,000 free viral particles and about 25,000 infected cells (RD Semba, 1999; RW Nduati, 1995)
Surprise to come?
• 2903 detected peptides
• 143 peptides corresponding to 53 genes
Host defense/immune related, enzyme, structural, transport, DNA binding and signal transduction proteins. An additional 13% of the proteins are of unknown or unclassified function.
LC / MS MS
G Smolenski, et al.
Use of maternal and/or infants antiretrovirals during breastfeeding ?
• Maternal HAART during lactation:
But…
- Breast milk CD4+ T lymphocytes expressing a high level of CCR5 and CXCR4 are preserved despite HAART (AP Kourtis, 2007);
- Cell-associated HIV persists despite maternal HAART (R Shapiro, 2005)
Alternative interventions to be tested?
• Periexposure prophylaxis in breastfed infants ?
• Postnatal mucosal vaccine?
• Pasteurisation? Heat treatment?
• Microbicides?
• Freezing / thawing?