Abstracts Vol. 1 [International Conference on AIDS (11th: 1996: Vancouver, Canada)]

Track A: Basic Science

Tu.A.2027 - Tu.A.203 I

Results: Murine cell clones expressing human CD26 were found to bind specifically bovine
[I 251]ADA with a high affinity No specific binding of [I1251]ADA was observed to muroine
clones not expressing human CD26.These observations indicated that the murine cells
expressing human CD26 provide a highly suitable model, to investigate the potential binding
of HIV- I Tat to CD26. In contrast to previously published results however we could not
demonstrate a specific interaction between Tat and human CD26.The [I 51]ADA specific
binding to human CD26 was not affected by Tat, even at concentrations when Tat induced
cell death. Similarly the binding of several monoclonal antibodies to human CD26 was not
modified by the addition of Tat. More significantlyTat binding to different murine cell clones
(human CD26 negative or positive) was found not to be correlated with the expression of
human CD26. Finally, the toxic effect of Tat on the growth of different murine cell clones
was independent of human CD26 expression.
Conclusions: Taken together: these observations further confirm the specific binding of
ADA to human CD26, and point out that CD26 is not the target of HIV- I Tat protein. As
the toxic effect of Tat is not species specific, it is plausible to suggest thatTat interacts with
different cell surface antigens, in a non-specific manner, by virtue of its high content of basic
amino acids.
J. Blanco, Institut Pasteur, Unite VIC, 28 rue du Dr: Roux, Paris, France Tel [phone] Fax
33 I 406 130 12 email: jblanco@pasteucfr
Tu.A.2027
STABLE EXPRESSION OF INTEGRINS IN HIV-INFECTED CD4+ T LYMPHOCYTES.
Leblond V.**v* Legendre C.*, Gras G.T, Lafuma C.**, Dormont D.*. * Service de
Neurovirologie, IPSC, CEA and CRSSA, Fontenay aux Roses, France. **INSERM U296,
Creteil, France.
Issue: Integrins are a family of adhesion molecules which mediate both cell-cell and cellextracellular matrix (ECM) interactions.These interactions play a critical role in T cell recognition of foreign antigens and in T cell migration into solid organs. HIV infection may alter
integrin expression on CD4+ T cells, then modulate their recirculation and migration abilities. Such mechanism would thus be involved in AIDS pathogenesis.To asses this hypothesis,
we have investigated the effect of in vitro HIV infection on the expression of various integrin
chains expressed byT cells: CD29 and the four a chains CD49c (VLA-3), CD49d (VLA-4),
CD49e (VLA-5) and CD49f (VLA-6).
Methods: Peripheral blood lymphocytes (PBLs) were obtained by counterflow centrifugal
elutriation and activated with PHA-P for 3 days.They were then infected with different viral
strains and isolates which possess specific cell tropism and various replicative capacity and
cytopathicity in vitro (HIVI -LAI, HIVI-BaL, HIVI -DAS, HIVI-THI and HIV2-ROD). At different times after infection, flow cytometry analysis of integrin expression was performed: cells
were doubly stained with monoclonal antibody to CD4 (Phycoerythrin-labelled) on one
hand, and to CD29, CD49c, CD49d, CD49e or CD49f on the other hand (FITC-labelled).
Integrin-specific mean fluorescence intensity of gated CD4+ or CD4- cells was compared
to isotype-matched negative controls. Controls included two cultures of uninfected cells,
and mock-infected cells using heat-inactivated HIVI-LAI and HIVI-BaL.
Results: Integnrin expression was modulated by cell cultivation, regardless infectious virus was
used or not. In contrast, the level of each tested molecule expressed by either CD4+ or
CD4- lymphocytes remained the same in infected and control cultures, from the day of
infection to day 20 p.i., when all the CD4+ cells were depleted.The only effect specific of
HIV infection was that CD29bright CD4+ cells were depleted before CD29dim ones during peak viral expression.
Conclusion: Cell surface density of integrins is not affected by HIV infection, even during
viral peak expression and CD4+ cell depletion. Infected cells are thus likely to maintain their
migration potential unchanged up to virus-induced cell death.This result might be of interest when virus spread in the body is considered. On another hand, the rapid depletion of
CD29bright CD4+ cells in infected cultures is concordant with previous reports which suggested preferentiel expression of HIV in T cells of the memory phenotype.
V. Leblond, CE-FAR, DSV/DRM/SNV, BP6, F92265 Fontenay aux Roses, France.Telephone:
(33 I) 46 54 80 35 Fax: (33 I) 46 54 77 26 Email [email protected]
Tu.A.2028
EXCHANGES WITHIN AN IMMUNODOMINANT CTL-EPITOPE INFLUENCES THE
REPLICATION OF HIV-I
Bernd LeschonskyI, Elisabeth Peintner,Tanja Fitzon, Hans Wolf',Thomas Harrer2, Ralf
Wagner. Universitit Regensburg, Regensburg, Germany; 2 Universitat Erlangen, Erlangen,
Germany
Objective: To examine a possible coherence between the control of an HIV-infection
through the cell-mediated immune response of a long term nonprogressor and the functional preservation of highly conserved domains.
Methods: site-directed mutagenesis, recombinant HIV-proviral constructs, p24-capture assay
To prove this hypothesis we went back to a CTL-clone out of a long term nonprogressor
recognizing an immodominant, 9 aminoacid enclosing, HLA-B I4 restricted epitope within
the carboxyterminal part of the HIV- I p24-capsidprotein.This region, which not only exhibited the primary target for activated CD8+ CTLs but also confirmed to be highly conserved within all sequenced HIV-isolates, had been exchanged in each position both as conservative as possible and not conservative.Thereupon we inserted selected, functionally
essential flo-mutants into HIV-proviral constructs and studied their replication behaviour
(COS-7 cells) and finally their infectivity behaviour (MT-4 cells).
Results: In contrast to the conservative substitutions the non conservative exchange of
aminoacids arginin299 (essential for the anchoring of antigenic peptides on MHC-I molecules) as well as threonin303 (necessary for the recognition of the T-cell receptor) resulted
in a decreased level of virus release. Furthermore the non conservative altered virus
mutants turned out to be non infectious anymore which could be shown in a permissive
CDI+ cell line.The determination of the recognition rate of these epitope variants through
the original CTL-clone is under investigation.
Conclusion: It could be demonstrated, that two essential positions for the stimulation
of a CTL response within a selected epitope of a long term survivor are conservative

interchangeable without altering morphogenesis and infectivity Whereas the conservative
substitution of the same positions resulted in decreased levels of virus release as well as in
loss of infectivity
Bernd Leschonsky Institut fOr Medizinische Mikrobiologie und Hygiene, D-93053
RegensburgTel. [phone], Fax. +49-941 I -944-6402, email:
[email protected]
Tu.A.2029
TWO PARALLEL ROUTES OF THE COMPLEMENT-DEPENDENT ENHANCEMENT
OF THE HUMAN IMMUNODEFICIENCY VIRUS TYPE I INFECTION
Zoltin Prohaszka'--, Jtn Prohasz2,T0nde Hidvdgi, Ferenc D.Tegi2, Judit Szabe2, Eszter
Ujhelyi, NicoleThielens3, Manfred P Dierich4, Jolin Kiss2, Gerard Arlaud3, George FOss t.
National Institute of Haematology Blood Transfusion and Immunology Budapest, Hungary I
Institute of Microbiology, University Medical School, Debrecen, Hungary2; Institut de Biologie
Structurale, Grenoble, France3; Institute of Hygiene and Ludwig Boltzmann-Institut for AIDSForschung, Innsbruck, Austria4
Objective:To study the mechanism of the complement-dependent enhancement (CDE) of
human immunodeficiency virus (HIV) infection that may play a significant role in the progression of HIV-disease.
Methods: In vitro complement activating and complement-dependent HIVinfection enhancing abilities of three human anti-gp4 I monoclonal antibodies were tested. CDE was estimated using HIV- I IIIB and CR2 (CD2 I)-carrying MT-4 target cells. NHS, purified C I q, low
C I q-level (CI qD) and C2-deficient (C2D) human sera were applied as complement
source.
Results: All monoclonal enhancing antibodies mediated increased C Iq binding to solidphase gp4 I.The effect of 246-D was the most pronounced. All monoclonals had a marked
dose-dependent and strictly complement-dependent HIV-infection enhancing effect.
Mixtures of the monoclonals with purified C Iq also significantly increased HIV- I infection.
Pretreatment of the target cells with anti-CR2 antibodies only partially inhibited the
enhancing effect of the monoclonals plus normal human serum. Both C I qD and C2D
serum had a markedly lower enhancing effect than NHS.
Conclusion:These novel findings indicate that besides the well-known facilitation of entry of
HIV I by the interaction between virus-bound C3 fragments and CR2 present on the target cells, fixation of C Iq to intact virions also results in an enhanced productive HIV- I infection in the MT-4 cell cultures.
Z. Prohaiszka, 24 Daroczi str, POB 44 H-1518 Budapest, HungaryTel.: 361I 209 23 10 Fax:
361 209 23 10 E-mail: [email protected]
Tu.A.2030
EFFECT OF IL-13 ON HIV-I EXPRESSION IN LATENTLY INFECTED MONOCYTIC
CELL LINES
Rizzardi, G Paolo*, Shattock R**, Fain C*, Zecca B*, Dalgleish A**, Barcellini,W. *
University of Milan, Milan, Italy; ** University of London, London, UK
Objective:To investigate the effect of IL- 13 on HIV- I expression in some models of latent
HIV infection, and its co-operation with proinflammatory cytokines.
Methods: HIV- I infected promyelocytic OM I10. I and promonocytic UI cells were maintained in complete culture medium in log phase of growth, and then plated in 96-well flatbottomed plates at the concentration of 2 x 105 cells/mL, in the presence of different
cytokines. Membrane bound TNF-(t (mbTNF-a) surface expression were determined by
flow cytometry HIV- I expression was quantitated by reverse transcriptase assay and p24
antigen release.
Results:We had already demonstrated that IL-10 induces an up-regulation of HIV expression in UI cells through the endogenous production ofTNF-e and the up-regulation of
mbTNF-ae and TNFRI. Here, we found that different doses of IL 13 (10, 50, and 100 ng/mL)
failed to significantly increase HIV expression in OM 10. I cells. However we also found that
IL- 13 synergizes with TNF- a in enhancing HIV expression in OM-10. I and UI models of
latent infection. Likewise, when IL-I 3 was added to the cultures along with PMA, we
observed a clear additive effect on HIV expression. Furthermore, in 24 hr vitamin D3-treated OM 10. I cells, the IL-13-induced additive effect on HIV expression was still present when
IL- 13 was added along with TNF-oa, and more evident along with PMA.
Conclusions: IL- 13, like IL- 10, is known to inhibit proinflammatory cytokines in normal
human monocytes, and HIV expression in acutely infected monocyte-derived macrophages.
Interestingly, in a model of latent infection, these cytokines seem to play a more complex
role, being able to trigger HIV through mechanisms that involve TNF- a and the mbTNF-a
/TNFRI pathway
Gian Paolo Rizzardi, Ospedale Policlinico, via F. Sforza 35, Milano 20122 Italy Telephone: [phone] Fax: 02-551I9-2842 email: milancam@(imiucca.csi.unimi.it
Tu.A.203 I
EFFECTS OF AND METABOLITES ON HIV-I-RELATED CELL DEATH
Savarino Andrea, Pugliese A, Malavasi F, Pescarmcna GP University ofTurin, Italy
Objective. To assess whether the lack of CD38 and its catalytic activity may be responsible
for cell death from HIV- I in MT-4 cells. Results on some selected Trcell lines indicated that
ce lular death fiom HIV- I is inversely proportional to the expression of CD38, which catalyzes the hydrolysis of extricellulr NAD into nicotinamide and cyclic ADP ribose
(cADPR).
Methods. MT-4 cells were infected with I00 CCID50 of HIV 11B. Cell death was monitored daily by eosin-dye exclusion. 5mM nicotinamide was added in culture after incubation
with HIV I.
Results. A significantly increased number of hve cells were detected up to the 5th day after
infection utilizing 5mM nicotinamide. Furthermore, very preliminary results indicated that
live cells increased also utilizing 25pM cADPR in the culture medium and an association of
cADPR and nicotinamide at suboptimal doses.
Conclusions. These results support the hypothesis that the catalytic activity of CD8 plays
a protective role against cell death fiom HIV I.This may be one of the explanations for the
long survival of cells expressing surface CD38 (monocytes) after HIV- I infection.
Andrea Savarino, Dip. Scienze Medico-Chirurgiche C.so Svizzera I 64 - 10 I 149 Torino, ITALY
tel[phone],,O
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