Oral Abstract Session 01 - Global HIV Vaccine Enterprise

Topic 7: Innate Immunity
P07.15 LB
Apoptotic Microparticles Generated During Acute
HIV-1 Infection Inhibit Human Dendritic Cells Via CD44
D. Frleta 1 , C.E. Ochoa 1 , H.B. Kramer 2 , S.A. Khan 1 , A.R. Stacey 2 ,
P. Borrow 2 , B.M. Kessler 2 , B.F. Haynes 3 , N. Bhardwaj 1
1 New York University Langone Medical Center, New York, NY,
USA; 2 University of Oxford, Oxford, United Kingdom (Great
Britain); 3 Duke University Medical Center, Durham, NC, USA
Background: Acute human immunodeficiency virus type 1 (HIV-
1) infection results in dysregulated immunity which contributes
to poor control of viral infection. Dendritic cells (DCs) are key
regulators of both adaptive and innate immune responses
needed for controlling HIV-1 and we surmised that plasma
factors elicited during acute HIV-1 infection (AHI) may impede DC
function. Such inhibitory factors present in AHI plasma include
apoptotic microparticles (MPs), small membranous blebs derived
from dying cells.
Methods: Plasma samples over sequential time points were
obtained from AHI patients or healthy controls. Apoptotic MPs
were isolated from supernatant of UV-irradiated PBMCs, AHI
patient plasma or control plasma. Human DCs were treated with
MPs or 10% plasma (control or AHI) and subsequently stimulated
with various TLR agonists. DC activation was then assessed.
MP-specific receptors were isolated from the DC surface and
sequenced by mass spectrometry.
Results: AHI plasma inhibited TLR-stimulated DC cytokine
production. The inhibitory capacity of AHI plasma occurs at
time of viral ramp-up, whereas plasma at times before plasma
viremia is not inhibitory. We determined this inhibition was not
mediated by virus. Because apoptotic MPs are elevated in AHI
plasma, we treated DCs with experimental and AHI plasmaderived
MPs, both of which reduced DC activation. The inhibition
of DCs by AHI plasma or MPs blocked DC capacity to prime IFNgproducing
T helper 1 CD4+ T cells as well as NK cell activation.
Mass spectrometric analysis revealed CD44 a MP receptor, and
blocking CD44 on DCs relieves MP-mediated suppression. Direct
ligation of CD44 also inhibits DC activation. MP-CD44 interaction
activates Rac1, c-Abl, and AKt signaling.
Conclusion: Determining the factors in AHI which block DC
function will provide potential targets to stimulate HIV-specific
immunity. We are currently investigating molecular mechanisms
of CD44-mediated DC inhibition and downstream signaling
events that can be targeted to alleviate DC inhibition.
P07.16 LB
AIDS Vaccine 2012
Posters
HIV Triggers Immunoregulatory Dendritic Cells And
Regulatory T Cells Through The Non-Canonical NF-kB
Pathway
O. Manches 1 , M.V. Fernandez 1 , J. Plumas 2 , L. Chaperot 2 ,
N. Bhardwaj 1
1 NYU Cancer Institute, New York City, NY, USA; 2 Inserm, U823,
Immunobiologie et Immunotherapie des cancers, La Tronche,
France
Background: HIV stimulates plasmacytoid dendritic cell (pDC)
through TLR7 and induce the secretion of high levels of IFNα.
pDC stimulated by HIV also upregulate the expression of the
enzyme indoleamine 2,3 dioxygenase (IDO). IDO is critical for the
induction of regulatory T cells (Treg) by HIV-activated pDC. We
investigated the molecular mechanisms of IDO induction and its
consequences for Treg function.
Methods: The cells used were purified primary pDC and
the GEN pDC cell line. A combination of siRNA knock-down,
immunoprecipitation of TLR signaling pathway molecules, IDO
promoter engineering and chromatin immunoprecipitation was
used to determine the molecular mechanisms of IDO induction in
pDC. To analyze Treg function and interaction with conventional
DC (cDC), blocking antibodies to CTLA- 4 and CTLA-4-Ig were used.
Results: We demonstrate that HIV induces activation of the
non-canonical NF-kB pathway in pDC, and is essential for
IDO induction. TLR7 triggering induces recruitment of TRAF3
to the TLR-MyD88 complex, followed by release of NIK and
phosphorylation of IKKα. Activation of the non-canonical NFκB
pathway culminates in p52/RelB nuclear translocation and
binding to the IDO promoter.
Furthermore, IDO-expressing pDC trigger the generation of
Treg, which dampen cDC activation through CTLA-4. CTLA-4
also induces IDO expression in cDC in a NIK-dependent fashion,
allowing cDC to induce Treg from naïve CD4+ T cells.
Conclusion: The non-canonical NF-kB pathway plays a central
role in regulating IDO expression in pDC and cDC upon HIV
infection, and may be a potential target for regulating Treg
activity in chronic or acute HIV infection.
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POSTERS