Abstract Book Vol. 2 [International Conference on AIDS (14th: 2002: Barcelona, Spain)]

364 Abstracts ThPeA7119-ThPeA7122

XIV International AIDS Conference

ThPeA71 19 HIV-1 Tat protein modulates proteasome
composition and activity
A. Bottoni', A. Bonaccorsi2, E. Gallerani1, M. Fabris2, C. Fortini', P. Rimessi2,
B. Ensoli3, R. Gavioli1. Department of Biochemistry and Molecular Biology
University of Ferrara, Ferrara, Italy; 2Department of Experimental and Diagnostic
Medicine, University of Ferrara, Ferrara, Italy; 3Laboratory of Virology, Istituto
Superiore di Sanita, Rome, Italy
Background: Tat is a regulatory protein of HIV-1 produced very early after infection and essential for HIV gene expression and replication. Tat is also released
from infected cells in the extracellular milieu and taken-up by neighbour cells
and modulates cellular functions depending on the concentration and cell type.
Perturbation of the proteasome system by viral proteins is often a key event in
viral immuno-escape and oncogenesis. Biochemical studies on the specificities
of the proteasome revealed three distinct proteolytic components, which are involved in chymotryptic, tryptic and post-acidic hydrolyzing activities, respectively.
When cells are exposed to IFN-y, the three catalytic P-subunits are substituted
with LMP2, LMP7 and MECL-1, respectively. These subunits are also expressed
in constitutive manner in specific cell types such as dendritic cells and B cells
and their incorporation alters the activity of the proteasome and enhances the
production of certain peptides.
Methods: We analyzed proteasome composition and activity in EBV-infected
cells endogenously expressing Tat.
Results: We demonstrated that constitutive Tat expression in lymphoblastoid cell
lines (LCL) induces up-regulation of LMP7 and MECL1 subunits and downmodulation of LMP2. To investigate whether the differences in subunit composition
correlated with differences in enzymatic activity, we have analyzed the cleavage specificity of 26S proteasomes isolated from LCL-vector and LCL-Tat. We
tested chimotryptic, tryptic and post-acidic activities using specific substrates. All
three activities detected in proteasome preparation from LCL expressing Tat were
higher then those detected in vector cells. In addition, we demonstrated that LCL
expressing Tat present with different efficiency EBV-derived CTL epitopes.
Conclusions: Thus, Tat can modulate proteasome activity and antigen processing of heterologous antigens.
Presenting author: Gavioli Riccardo, Department of Biochemistry and Molecular Biology, University of Ferrara, Via Borsari, 46, 44100 Ferrara, Italy, Tel.: [phone], Fax: [phone], E-mail: r.gavioli @ unife.it
ThPeA7120 Thymical reconstitution of CD4+ T-lymphocytes in
treated HIV-infected children
R. Correa, M.A. Mufoz-Fernandez. Hospital General Univ Gregorio Marah6n,
Division of Immunology, Hospital General Universitario "Gregorio MaraF6n", C/
Dr Esquerdo 46, 28007 Madrid, Spain
Background: Children maintain the thymical functionality and thus, the capacity
of replacing the CD4 T-cells clones lost by the HIV-infection, making possible the
immune reconstitution. Since HIV has an inhibitory effect on thymus, a suppression of viral replication would be advisable for recovering the thymical function.
The increase in CD4 after HAART could be explained by a recovery of the thymical function, but also by a redistribution or peripheral expansion of pre-existent
naive cells. We have studied the effect of ART on the thymical production of naive
T-cells, to assess the source of the increased CD4 T-cells in infected children.
Methods: 6 vertically HIV-infected children (3 treated with combination therapy
and 3 on HAART) were studied longitudinally for 4 years. CD4 percentages, viral
load and thymic function were measured every 2-3 months. Thymic function was
determined quantifying TCR rearrangement excision circles (TRECs) in PBMCs
by Real-time Quantitative PCR.
Results: ART induced a decrease in viral load in the 3 children on combination
therapy, and a complete suppression to undetectable levels in the 3 children on
HAART. A marked increase in CD4 was observed in the 6 children, showing a
significant inverse correlation with viral load (p<0.05). The increase in CD4 was
accompanied in all 6 children by a concomitantly marked increase in TRECs levels, disclosing a highly significant correlation between both variables (p<0.001).
A redistribution or increase in the division rates of peripheral T-cells, was rejected
since they would have produced a dilution of naive T-cells from thymus, and therefore a decrease in TRECs levels. It seems clear that in all our ART-treated children
an early thymical production of naive T-cells accounted for most of restored CD4
T-cells.
Conclusions: In children, the decrease in viral load by effect of ART leads to an
increase in CD4 T-cells mainly due to a recovery in the thymical production of new
T-cells.
Presenting author: Ma Angeles Mufioz-Fern.ndez, Division of Immunology,
Hospital General Universitario "Gregorio Marafin", C/ Dr Esquerdo 46, 28007
Madrid, Spain, Tel.: [phone], Fax: [phone], E-mail: [email protected]

ThPeA7121 Trafficking of HIV-specific CTL to mucosal tissues
and cerebrospinal fluid
B.L. Shacklett', C.A. Cox', J.K. Sandberg1, A. Nilsson2, N.H. Stollman3
M.A. Jacobson3, R.W. Price2, D.F. Nixon1. 'Gladstone Institute of Virology and
Immunology, Gladstone Institute of Virology and Immunology, PO Box 419100,
San Francisco, CA, 94141-9100, United States; 2San Francisco General
Hospital, San Francisco, CA, United States; 3Center for AIDS Research,
University of California, San Francisco, San Francisco, CA, United States
Background: Although extensive studies have described HIV-specific CTL in peripheral blood, little is known of the role of CTL in viral clearance from tissue reservoirs. Gut-associated lymphoid tissue (GALT) and the central nervous system
(CNS) may be considered distinct immunological compartments from peripheral
blood, and T-cells trafficking to these tissues may differ in cytokine expression,
homing receptors, activation, specificity and maturation status.
Methods: Peripheral blood, rectal biopsy tissue and/or cerebrospinal fluid (CSF)
were obtained from 16 clinically stable HIV-infected individuals not on antiretroviral therapy. HIV-specific CTL were quantified by MHC class I tetramer staining. CD8+ T-cells were assessed by flow cytometry for expression of adhesion
molecules [integrins o4p1, ct L2, c437, oEP7], chemokine receptors [CXCR3,
CXCR4, CCR5, CCR7], maturation markers [CD45RO, CD45RA] and cytotoxic
effector molecules [perforin, granzyme A].
Results: Expression of CCR5 and CXCR3 was strikingly increased in CD8+ Tcells from CSF relative to peripheral blood (P<0.001). CSF T-cells also expressed
high levels of integrins o41pl and oaLP2. CD8+ T-cells in GALT expressed high
levels of CCR5, CD45RO and integrins o4P7 and xEt7, but low levels of perforin (P=0.002). In two patients from whom all three tissues were sampled, CTL
specific for the HLA-A*0201-restricted Gag epitope SLYNTVATL were present in
blood, GALT and CSF.
Conclusions: These results suggest that recruitment of HIV-specific CTL to tissue reservoirs is driven by expression of multiple chemokine receptors, adhesion
molecules and activation markers. These pathways may be shared by other inflammatory diseases.
Presenting author: Shacklett Barbara, Gladstone Institute of Virology and Immunology, PO Box 419100, San Francisco, CA, 94141-9100, United States, Tel.:
[phone], Fax: [phone], E-mail: [email protected]
ThPeA7122 Expanded HIV-1 antigen-specific T cells after
treatment for TB in HIV-infected patients
A.M. Masemola', H. Maila', S. Nyoka, RP. Mohube', G. Khoury', S. Roux2,
E. Magcuntsu2, G. Churchyard2, A.D. Grant3, C.M. Gray'. 'National Institute for
Communicable Diseases, National Institute for Communicable Diseases, Private
Bag X4, Sandringham, Johannesburg 2131, South Africa; 2Aurm Health
Research, Welkom, South Africa; 3London School of Tropical Medicine and
Hygiene, London, United Kingdom
Background: Tuberculosis (TB) is the most important HIV-1-related disease in
sub-Saharan Africa. We investigated the impact of TB treatment on viral load and
T cell changes in antiretroviral-naive dually infected HIV-1/TB patients. Our aim
was to measure whether there was improved immune competence with TB treatment.
Methods: 45 mineworkers with a first episode of TB were recruited at a hospital
in the Free State, South Africa. Control groups consisted of 17 HIV-1+ve patients
with acute bacterial pneumonia and 22 HIV-1+ve patients. Patients with TB or
pneumonia had samples taken at baseline, 2, 8 and 24 weeks for viral load, T cell
subsets and HIV antigen-specific T cell (IFNg ELISPOT) changes.
Results: HIV-1 RNA copies were significantly (p<0.05) higher at baseline in HIV+/TB+ patients (median, 29861 copies/ml) when compared with
HIV+/pneumonia (12213 copies/ml) or HIV-1+ only (12722 copies/ml). There
were no significant changes in viral load after treatment in groups with TB or
pneumonia. CD4 counts were unchanged between treatment groups and remained between 300-600 cells/ul. Numbers of naive (45RA+62L+) CD4+ and
CD8+ cells were significantly depressed in HIV+/TB+ patients and remained depressed with treatment. Conversely, the numbers of activated (CD38+DR+) CD8+
T cells were significantly elevated in HIV+/TB+ infected patients and remained elevated during treatment. 9/13 patients showed significant (p=0.019) increase in
HIV-1 antigen-specific cells after therapy: median of 418-1630 sfc/10e6 PBMC
for HIV/pneumonia and 165-1985 sfc/10e6 PBMC for HIV/'B. These changes did
not correlate with viral load or numbers of activated T cells.
Conclusions: At 24 weeks after HIV/TB or HIV/pneumonia treatment, the HIV1 burden was not significantly lowered and the number of chronically activated
T cells remained high. However, there was evidence for improved immune competence where increased frequencies of anti-Gag and -Nef-specific T cell responses were identified.
Presenting author: Agatha Masemola, National Institute for Communicable Diseases, Private Bag X4, Sandringham, Johannesburg 2131, South Africa, Tel.:

[phone], Fax: [phone], E-mail: [email protected]