Most HIV DNA in PBMC is present in non-gut-homing, resting … · 10 4 10 5 23.6 0 10 2 10 3 10 4...
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Most HIV DNA in PBMC is present in non-gut-homing, resting memory CD4 T cells with ß7-CD38-CD127high phenotype Kersten Koelsch, Yin Xu, Michelle Bailey, Kristin McBride, Nabila Seddiki, Kazuo Suzuki, John Murray, David A Cooper, Anthony Kelleher and John Zaunders Centre for Applied Medical Research, St Vincent’s Hospital, National Centre in HIV Epidemiology and Clinical Research, UNSW Sydney, Australia
Transcript of Most HIV DNA in PBMC is present in non-gut-homing, resting … · 10 4 10 5 23.6 0 10 2 10 3 10 4...
Most HIV DNA in PBMC is present in
non-gut-homing, resting memory CD4 T cells
with ß7-CD38-CD127high phenotype
Kersten Koelsch, Yin Xu, Michelle Bailey, Kristin
McBride, Nabila Seddiki, Kazuo Suzuki, John Murray,
David A Cooper, Anthony Kelleher and John Zaunders
Centre for Applied Medical Research, St Vincent’s Hospital,
National Centre in HIV Epidemiology and Clinical Research, UNSW
Sydney, Australia
St Vincent’s Research Campus Sydney
Gut-associated lymphoid tissue (GALT) as a reservoir of HIV infection
√ CCR5+ CD4 T cells in GALT are a major target during primary HIV infection
√ GALT may be a site of ongoing replication during chronic infection and where CD4 memory cells become latently infected
√ Subset of memory CD4 with integrins αααα4+ß7+, gut-homing CD4 T cells that can migrate from blood into GALT and back into blood via draining lymphatics and thoracic duct
St Vincent’s Research Campus Sydney
Gut-homing CD4+ T cells
CD45RO
Integrin αα αα4 (CD49d)
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23.6
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19.1 8.79
675.16
Integrin ß1 (CD29)
Integrin ß7
Non-gut-homing
memory cells
αααα4+
memory cells
Gut-homing
memory cells
Gated on CD4 T cells
St Vincent’s Research Campus Sydney
Methods - Cryopreserved leukapheresis packs
√ 8 relatively early HIV+ subjects, from pre-HAART era,
(median CD4 cell count of 622 cells/µl)
√ CD4 cells enriched by negative selection
√ Stained for subsets of CD45RO+ memory CD4
√ Subsets purified by cell sorting on FACS Aria
√ HIV gag DNA measured by quantitative real-time PCR
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Cell sorting of gut-homing CD4+ T cells
CD45RO
Integrin ß7
Naive
cells
Non-gut-homing
memory cells
Gut-homing
memory cells
8%
39%
Leukapheresis donor
HIV DNA copies/500ng DNA
HIV DNA levels in integrin ß7+ vs ß7- memory cells
Relative amounts of HIV DNA
in ß7+ vs ß7- memory CD4 T cells % of HIV DNA in CD45RO+ CD4 T cells
Leukapheresis donor
Relative amounts of HIV DNA
in ß7+ vs ß7- memory CD4 T cells % of HIV DNA in CD45RO+ CD4 T cells
Donor
Tregs vs long-term memory CD4+ T cells
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103
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0
102
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105
0 102
103
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0
102
103
104
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Gated on CD4
CD45RO-ECD
CD38-PE-Cy7
55%7.5%
74%
CD127-Pacific BlueCD25-APCMemory CD4
Tregs
CD127 high
St Vincent’s Research Campus Sydney
Relative amounts of HIV DNA
in Tregs vs CD127 high memory CD4 T cells % of HIV DNA in CD45RO+ CD4 T cells
Leukapheresis donor
Activated CD38+ CD45RO+ CD4+ T cells
Healthy adult
control
gated on CD4 cells
Primary HIV
infection
CD45RO-ECD
CD38-PE-Cy7
0 102
103
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105
0
102
103
104
105
Leukapheresis
donor 2
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103
104
105
0
102
103
104
105
0 102
103
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0
102
103
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2.5% 19% 7%
Relative amounts of HIV DNA
in CD38+ vs CD38- memory CD4 T cells % of HIV DNA in CD45RO+ CD4 T cells
Leukapheresis donor
St Vincent’s Research Campus Sydney
Cell sorting of CD4+ cells in gut biopsiesCell sorting of CD4+ cells in gut biopsies
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Single cell suspension
from gut biopsies
CD45-PerCP
EpCAM-FITC
81 %
19%
6 %
9 %
CD4-PE
CD8-APC
Lymphocytes
Epithelial cells
CD4
St Vincent’s Research Campus Sydney
Conclusions
√ Isolated gut-homing memory CD4+ cells from PBMC and shown infection with HIV DNA
√ Isolated CD4 cells from gut biopsies and confirmed infection with HIV DNA
√ However, the majority of HIV DNA in PBMC was found in CD4+ T cells that were not gut-homing (ß7-)
√ And in resting CD4 memory cells (high IL-7R and CD38-)
√ Overall, the majority of HIV DNA infected CD4+ T cells in PBMC were unlikely to have been activated and infected in GALT
St Vincent’s Research Campus Sydney
Acknowledgements
SVH / NCHECR
Yin Xu
Michelle Bailey
Nabila Seddiki
Kersten Koelsch
Kristin McBride
Kazuo Suzuki
Michael Gao
Mark Danta
Linda Gelgor
Christoph Boesecke
David Cooper
Tony Kelleher
John Zaunders
HIV Trials
Kat Marks
Kate Merlin
Julie Yeung
Maria Piperias
Bertha Fsadni
UNSW
John Murray
Barbara Cameron
Suzy Teutsch
Andrew Lloyd
Funding
NHMRC Project Grant
NHMRC Program Grant
