The Tolllike receptor 2 ligand Zymosan inhibits HIV1 replication in human primary cells'

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The Toll-like receptor 2 ligand Zymosan inhibits
HIV-1 replication in human primary cells.
Haynna Kimie Pimenta-Inada Oswaldo Cruz
Institute Fiocruz Rio de Janeiro, RJ Brazil
XVII International AIDS Conference Mexico City,
August 2008
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Introduction

• Toll-Like Receptors (TLR) are type I membrane
  glycoproteins that recognize specific molecular
  patterns found in microbial pathogens, triggering
  inflammatory and responses
• TLR ligation initiates complex signaling
  cascades, leading to NF-?B activation and
  production of Inflammatory cytokines
• TLR ligands modulate dendritic cell, monocyte and
  lymphocyte function
• Zymosan is an insoluble carbohydrate from cell
  wall of the yeast Saccharomyces cerevisae, and is
  a ligand of the TLR-2
• After phagocytosed by macrophages, zymosan can
  induce a prolonged inflammatory response, leading
  to secretion of inflammatory mediators (e.g.
  TNF-alpha and IFN-gamma)
• LPS, a TLR-4 ligand, inhibits HIV-1 replication
  in macrophages (Verani et al, 1997)
• It is not clear whether ligation of TLR-2
  modulates HIV-1 replication

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Toll-like Receptors
(Sher et al, 2007)
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Toll-like Receptors
(Akira et al, 2006)
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Zymosan inhibits HIV-1 replication in PBMCs and
macrophages.
n 6 (12.5 and 25µg/mL), 3 (50 and 100µg/mL) or 2
(Pam3Cys)
n 4 (3, 6 and 12.5µg/mL) or 2 (0.75, 25 and
50µg/mL)
PBMCs and macrophages were infected with HIV-1
(R5-tropic isolate) and treated with Zymosan or
Pam3Cys. Viral replication was assessed after 7
(PBMCs) or 10 (macrophages) days. plt0,001
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Pre-treatment with Zymosan inhibits HIV-1
replication in PBMCs.
PBMCs were treated with 25?g/mL of Zymosan for
24h, cells were washed and, then, infected with
HIV-1 (R5-tropic isolate). Viral replication was
assessed after 7 days.
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Production of beta-chemokines in cells stimulated
with Zymosan.
n6
n7
PBMCs and macrophages were treated with 25?g/mL
of Zymosan for 24h and, then, RANTES, MIP-1? and
MIP-1? production was assessed by ELISA. plt0,05
plt0,01 plt0,001.
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Zymosan does not modulate IL-10 secretion in
macrophages
Macrophages either uninfected or infected with
HIV-1 (R5-tropic isolate) and treated with
Zymosan (10?g/mL) for 3h 6h, 9h or 10d. IL-10
production was assessed by ELISA. n4.
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Zymosan inhibits HIV-1 primary isolates
replication.
PBMCs were infected with HIV-1 primary isolates
95BRRJ10 (X4-tropic) and 95BRRJ21 (R5-tropic) and
treated with 25?g/mL of Zymosan. Viral
replication was assessed after 7 days.
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Zymosan does not inhibit proviral integration in
HIV-1-infected PBMCs
After acute infection
Pre-treatment 24h
HIV-1-infected PBMCs were treated or not with
Zymosan (100?g/mL) for 24 hrs and total proviral
DNA (LTR) and integrated proviral DNA (ALU/LTR)
was evaluated by PCR methods C positive
control C- negative control LTR HIV Long
terminal repeated ALU sequence MW Molecular
weight. n1
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Conclusions

• Zymosan inhibits viral replication in
  HIV-1-infected human PBMCs and macrophages after
  acute infection
• Zymosan inhibits R5- and X4-tropic primary
  isolates
• Pam3Cys, a synthetic ligand of TLR-2, also
  inhibits HIV-1 replication in PBMCs
• Treatment with Zymosan prior to HIV-1 infection
  also inhibits viral replication
• Zymosan up-modulates production of
  beta-chemokines in PBMCs and macrophages
• Zymosan does not inhibit pro-viral integration
• Further studies should be done to elucidate the
  mechanisms underlying the antiretroviral activity
  derived from TLR-2 ligation.

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Acknowledgments

• Oswaldo Cruz Institute/Fiocruz
• Haynna Kimie Pimenta Inada
• Lívia Gobbo Jucá
• Cláudio César Cirne-Santos
• Eduardo Garcia N. P. Régis
• Victor Barreto de Souza
• Jairo Ramos Temerozo
• Diego Queiroz Rodrigues
• Thiago Moreno Lopes e Souza
• Thyago Martins Costa B. Pereira
• Caroline Passaes
• Mariza Morgado
• Dumith Chequer Bou-Habib

Reagents NIH AIDS Reagent Program,
Hemotherapy Service (HUCCF - UFRJ)
Financial Support Papes/Fiocruz, CNPq, Faperj