-
CELLULAR ASPECTS OFHIV INFECTION
Edited by
ANDREA COSSARIZZASection of General PathologyDepartment of Biomedical SciencesUniversity of Modena and Reggio Emilia School of MedicineModena, Italy
DAVID KAPLANDepartment of PathologyCase Western Reserve University School of MedicineCleveland, Ohio
A JOHN WILEY & SONS, INC., PUBLICATION
Innodata0471459151.jpg
-
CELLULAR ASPECTS OFHIV INFECTION
-
CYTOMETRIC CELLULAR ANALYSIS
Series Editors
J. Paul Robinson George F. BabcockPurdue University Cytometry
LaboratoriesDepartment of Surgery
Purdue UniversityUniversity of Cincinnati College of
MedicineWest Lafayette, Indiana Cincinnati, Ohio
Phagocyte Function: A Guide for Research and Clinical EvaluationJ. Paul Robinson and George F. Babcock, Volume Editors
ImmunophenotypingCarleton C. Stewart and Janet K. A. Nicholson, Volume Editors
Emerging Tools for Single Cell Analysis: Advances in Optical MeasurementTechnologiesGary Durack and J. Paul Robinson, Volume Editors
Cellular Aspects of HIV InfectionAndrea Cossarizza and David Kaplan, Volume Editors
-
CELLULAR ASPECTS OFHIV INFECTION
Edited by
ANDREA COSSARIZZASection of General PathologyDepartment of Biomedical SciencesUniversity of Modena and Reggio Emilia School of MedicineModena, Italy
DAVID KAPLANDepartment of PathologyCase Western Reserve University School of MedicineCleveland, Ohio
A JOHN WILEY & SONS, INC., PUBLICATION
-
Designations used by companies to distinguish their products are often claimed as trademarks. In
all instances where John Wiley & Sons, Inc., is aware of a claim, the product names appear in
initial capital or all capital letters. Readers, however, should contact the appropriate
companies for more complete information regarding trademarks and registration.
All rights reserved.
No part of this publication may be reproduced, stored in a retrieval system or transmitted in any
form or by any means, electronic or mechanical, including uploading, downloading, printing,
decompiling, recording or otherwise, except as permitted under Sections 107 or 108 of the 1976
United States Copyright Act, without the prior written permission of the Publisher. Requests to
the Publisher for permission should be addressed to the Permissions Department, John Wiley &
Sons, Inc., 605 Third Avenue, New York, NY 10158-0012, (212) 850-6011, fax (212) 850-6008,
E-Mail: PERMREQ @ WILEY.COM.
This publication is designed to provide accurate and authoritative information in regard to the
subject matter covered. It is sold with the understanding that the publisher is not engaged in
rendering professional services. If professional advice or other expert assistance is required, the
services of a competent professional person should be sought.
This title is also available in print as ISBN 0-471-38666-9.
For more information about Wiley products, visit our web site at www.Wiley.com.
Copyright � 2002 by Wiley-Liss, Inc.
http://www.Wiley.com
-
CONTENTS
Preface ix
Contributors xi
Part I MOLECULES 1
1 HIV and Molecular Biology of the Virus-Host Interplay 3Massimo Clementi
2 Telomere Length, CD28C T Cells, and HIV Disease Pathogenesis 13Rita B. E¨ros
3 Immune Dysregulation and T-Cell Activation Antigens inHIV Infection 33Mara Biasin, Fulvia Colombo, Stefania Piconi, and Mario Clerici
4 Quantification of HIV/SIV Coreceptor Expression 53Benhur Lee and Robert W. Doms
Part II CELLS 67
5 B Cells in the Line of Sight of HIV-1 69Yolande Richard, Eric A. LefeÁvre, Roman Krzysiek, Christophe
Legendre, Dominique Dormont, Pierre Galanaud, and Gabriel Gras
6 Cytotoxic T-Cell (CTL) Function in HIV Infection 103M. L. Garba and J. A. Frelinger
7 Analysis of the a /b T-Cell Receptor Repertoire in HIV Infection 127Hugo Soudenys
v
-
8 Gamma-Delta (gd) T Cells and HIV-1 Infection 147Roxana E. Rojas and W. Henry Boom
9 Natural Killer Cells in HIV Infection and Role in the Pathogenesisof AIDS 183Benjamin Bonavida
10 Alveolar Macrophages 207Jianmin Zhang and Henry Koziel
11 Dendritic Cells Ferry HIV-1 from Periphery into Lymphoid Tissues 229Teunis B. H. Geijtenbeek and Yvette van Kooyk
Part III PROCESSES 249
12 Homeostasis and Restoration of the Immune System in HAART-Treated HIV-Infected Patients: Implications of Apoptosis 251Marie-Lise Gougeon, Herve LeCoeur, Luzia Maria de Oliveira
Pinto, and Eric Ledru
13 Mitochondria Functionality During HIV Infection 269Andrea Cossarizza, Marcello Pinti, Milena Nasi, Maria Garcia
Fernandez, Laura Moretti, Cristina Mussini, and Leonarda Troiano
14 Multiple Roles of Cytokines in HIV Infection, Replication, andTherapy 293Massimo Alfano and Guido Poli
Part IV TECHNOLOGIES 313
15 The Use of Peptide/MHC Tetramers to Visualize, Track, andCharacterize Class I-Restricted Anti-HIV T-Cell Response 315Clive M. Gray and Thomas C. Merigan
16 Tagging of HIV with Green Fluorescent Protein 333Nadya I. Tarasova
17 Flow Cytometric Analysis of Cells from Persons with HIV-1 Diseaseby Enzymatic Amplification Staining 351David Kaplan
vi Contents
-
18 Antigen-Specific Cytokine Responses in HIV Disease 371Vernon C. Maino, Holden T. Maecker, and Louis J. Picker
Part V ORGANISMS 383
19 Chimeric Models of SCID Mice Transplanted with Human Cells:The Hu-PBL-SCID Mouse and Its Use in AIDS Research 385S. M. Santini, C. Lapenta, M. Logozzi, S. Parlato, M. Spada,
T. Di Pucchio, S. Fais, and F. Belardelli
20 Immune Reconstitution of the CD4 T-Cell Compartment inHIV Infection 399Guislaine Carcelain, Taisheng Li, Marc Renaud, Patrice DebreÂ,
and Brigitte Autran
21 New Perspectives in Antiretroviral Therapy of HIV Infection 423Stefano Vella
Index 439
viiContents
-
Preface
Infection with human immunode®ciency virus (HIV) has produced one of themost dramatic epidemics of the twentieth century. It has spread worldwide,leaving no region of the world una¨ected. Before e¨ective therapies were de-veloped, infection with HIV meant an inexorable decline in health until deathwas a welcome relief. Now that the capability to decrease viral replicationhas been achieved, those who can a¨ord this expensive treatment survive bykeeping the infection dormant, not by eliminating the virus altogether. Un-fortunately, the antiviral reagents available come with serious side e¨ects,and resistance to these agents develops readily for a consistent percentage ofpatients. At the same time that therapies with speci®c antiviral agents havedecreased morbidity and mortality, they have resulted in a relaxation of ap-propriate public and private health measures, which threatens a recrudescenceof epidemic infection. Of course, many if not most infected persons world-wide cannot even a¨ord treatment and perish quickly without any speci®cintervention.
Many biomedical scientists have investigated HIV and the disease syndromethat it produces in infected persons. These investigators have contributedgreatly to our understanding of the mechanisms that the virus uses to replicate,to infect new hosts, and to cause disease. These mechanisms have been de-scribed in molecular, cellular, organismal, and social terms.
At the cellular level, investigators ®rst identi®ed the cells that are infected byHIV or that act as reservoirs for the virus. Then the crucial mechanisms of theimmune response, including the importance of HIV-speci®c cytotoxic cells andhumoral responses, the way in which cells die after the infection, the death ofinnocent bystanders, and the role of costimulatory molecules and coreceptorswere described. These studies at the cellular level have relied on many di¨erenttechnologies, one of the most important being ¯ow cytometry.
Flow cytometry is a powerful technique for the analysis of multiple param-eters of single cells. It is capable of assessing six to ten parameters on 10,000cells in less than a minute. Moreover, cells with speci®ed characteristics can besorted live and cultured for additional investigation. Flow cytometry has beenused since the beginning of the HIV era as a key approach to study the cellular
ix
-
level in HIV infection. Millions of analyses have been performed on samplesfrom persons infected with HIV. These analyses have allowed us to follow thecourse of the infection, to observe the complex response of the immune sys-tem to the virus, and to help in deciding how to treat infected patients, and tounderstand the patients' cellular response to the therapy.
This book includes chapters by renowned experts on various aspects of HIVinvestigations. The main aim of this book is to present these descriptions andanalyses with particular attention to the role that ¯ow cytometric techniqueshave played in shaping our current conceptualizations. The book is divided into®ve partsÐmolecules, cells, pathophysiological processes, technologies, andorganisms. Each chapter emphasizes an intelligent, concise synthesis of thetopic without an attempt to provide an exhaustive review.
The book is intended for experts in the ®eld of HIV studies, including im-munologists, virologists, and clinicians, as well as for other researchers whoare primarily interested in the use of ¯ow cytometric techniques in biomedicalinvestigations.
Andrea Cossarizza and David Kaplan
x Preface
-
CONTRIBUTORS
Massimo AlfanoAIDS Immunopathogenesis UnitSan Ra¨aele Scienti®c InstituteMilano, Italy
Brigitte AutranLaboratoire d'Immunologie
Cellulaire et TissulaireHoÃpital PitieÂ-SalpeÃtrieÁreParis, France
Filippo BelardelliLaboratory of VirologyIstituto Superiore di SanitaÁ
Roma, Italy
Mara BiasinCattedra di ImmunologiaUniversitaÁ di MilanoMilano, Italy
W. Henry BoomDivision of Infectious DiseasesCase Western Reserve UniversityCleveland, Ohio
Benjamin BonavidaDepartment of Microbiology,
Immunology, and MolecularGenetics
UCLA School of MedicineUniversity of CaliforniaLos Angeles, California
Guislaine CarcelainLaboratoire d'Immunologie
Cellulaire et TissulaireHoÃpital PitieÂ-SalpeÃtrieÁreParis, France
Massimo ClementiDepartment of Biomedical SciencesUniversity of TriesteTrieste, Italy
Mario ClericiCattedra di ImmunologiaUniversitaÁ di MilanoMilano, Italy
Fulvia ColomboCattedra di ImmunologiaUniversitaÁ di MilanoMilano, Italy
Andrea CossarizzaDepartment of Biomedical SciencesUniversity of Modena and Reggio
Emilia School of MedicineModena, Italy
Patrice DebreÂ
Laboratoire d'ImmunologieCellulaire et Tissulaire
HoÃpital PitieÂ-SalpeÃtrieÁreParis, France
xi
-
Robert W. DomsUniversity of Pennsylvania Medical
CenterDepartment of Pathology and
Laboratory MedicinePhiladelphia, Pennsylvania
Dominique DormontINSERM U131Institut Paris-Sud sur les CytokinesClamart, France
Rita B. EffrosDepartment of Pathology and
Laboratory MedicineUCLA Medical CenterLos Angeles, California
Stephano FaisLaboratory of ImmunologyInstituto Superiore di SanitaÁ
Roma, Italy
J. A. FrelingerUniversity of North CarolinaChapel Hill, North Carolina
Pierre GalanaudINSERM U131Institut Paris-Sud sur les CytokinesClamart, France
M. L. GarbaUniversity of North CarolinaChapel Hill, North Carolina
Maria Garcia FernandezDepartment of Human PhysiologyUniversity of MalagaMalaga, Spain
Teunis B. H. GeijtenbeekDepartment of Tumor ImmunologyUniversity Medical Center St.
RadboudNijmegen, The Netherlands
Marie-Lise GougeonUnite d'Oncologie Virale and
CNRS URA 1930Department SIDA et RetrovirusInstitut PasteurParis, France
Gabriel GrasService de NeurovirologieCEA/CRSSAInstitut Paris-Sud sur les CytokinesFontenay aux Roses, France
Clive M. GrayCenter for AIDS Research at
StanfordDivision of Infectious Diseases and
Geographic MedicineStanford University Medical CenterPalo Alto, California
David KaplanDepartment of PathologyCase Western Reserve UniversityCleveland, Ohio
Henry KozielAssistant Professor of MedicineHarvard Medical SchoolBoston, Massachusetts
Roman KrzysiekINSERM U131Institut Paris-Sud sur les CytokinesClamart, France
C. LapentaLaboratory of VirologyIstituto Superiore di SanitaÁ
Roma, Italy
Herve LecoeurUnite d'Oncologie Virale and
CNRS URA 1930
xii Contributors
-
Department SIDA et RetrovirusInstitut PasteurParis, France
Eric LedruUnite d'Oncologie Virale and
CNRS URA 1930Department SIDA et RetrovirusInstitut PasteurParis, France
Benhur LeeUniversity of Pennsylvania Medical
CenterDepartment of Pathology and
Laboratory MedicinePhiladelphia, Pennsylvania
Eric A. LefeÁvreINSERM U131Institut Paris-Sud sur les CytokinesClamart, France
Christophe LegendreService de NeurovirologieCEA/CRSSAInstitut Paris-Sud sur les CytokinesFontenay aux Roses, France
Taisheng LiLaboratoire d'Immunologie
Cellulaire et TissulaireHoÃpital PitieÂ-SalpeÃtrieÁreParis, France
M. LogozziLaboratory of VirologyIstituto Superiore di SanitaÁ
Roma, Italy
Holden T. MaeckerBD BiosciencesSan Jose, California
Vernon C. MainoBD BiosciencesSan Jose, California
Thomas C. MeriganCenter for AIDS Research at
StanfordDivision of Infectious Diseases and
Geographic MedicineStanford University Medical CenterPalo Alto, California
Laura MorettiDepartment of Biomedical SciencesUniversity of Modena and Reggio
Emilia School of MedicineModena, Italy
Cristina MussiniInfectious Diseases ClinicsUniversity of Modena and Reggio
Emilia School of MedicineModena, Italy
Milena NasiDepartment of Biomedical SciencesUniversity of Modena and Reggio
Emilia School of MedicineModena, Italy
Luzia Maria de Oliveira PintoUnite d'Oncologie Virale and
CNRS URA 1930Department SIDA et RetrovirusInstitut PasteurParis, France
S. ParlatoLaboratory of VirologyIstituto Superiore di SanitaÁ
Roma, Italy
Louis J. PickerUniversity of Oregon Health
Sciences CenterBeaverton, Oregon
xiiiContributors
-
Stefania PiconiDivisione di Malattie InfettiveOspedale L. SaccoMilano, Italy
Marcello PintiDepartment of Biomedical SciencesUniversity of Modena and Reggio
Emilia School of MedicineModena, Italy
Guido PoliAIDS Immunopathogenesis UnitSan Ra¨aele Scienti®c InstituteMilano, Italy
T. Di PucchioLaboratory of VirologyIstituto Superiore di SanitaÁ
Roma, Italy
Marc RenaudLaboratoire d'Immunologie
Cellulaire et TissulaireHoÃpital PitieÂ-SalpeÃtrieÁreParis, France
Yolande RichardINSERM U131Istitut Paris-Sud sur les CytokinesClamart, France
Roxana E. RojasDivision of Infectious DiseasesCase Western Reserve UniversityCleveland, Ohio
S. M. SantiniLaboratory of VirologyIstituto Superiore di SanitaÁ
Roma, Italy
M. SpadaLaboratory of VirologyIstituto Superiore di SanitaÁ
Roma, Italy
Hugo SoudeynsUnite d'immunopathologie viraleCentre de recherche de l'Hopital
Sainte-JustineDepartments de microbiologie and
immunologie et de peÂdiatrieFaculte de meÂdicineUniversite de MontreÂal
Nadya I. TarasovaNational Cancer InstituteFrederick Cancer Research and
Development CenterFrederick, Maryland
Leonarda TroianoDepartment of Biomedical SciencesUniversity of Modena and Reggio
Emilia School of MedicineModena, Italy
Yvette Van KooykDepartment of Tumor ImmunologyUniversity Medical Center St.
RadboudNijmegen, The Netherlands
Stefano VellaIstituto Superiore di SanitaÁ
Roma, Italy
Jianmin ZhangResearch FellowHarvard Medical SchoolCambridge, Massachusetts
xiv Contributors
-
P A R T I
MOLECULES
-
1HIV and Molecular Biology of theVirus-Host Interplay
Massimo ClementiDepartment of Biomedical Sciences, University of Trieste, Trieste, Italy
INTRODUCTION
The precise understanding of the molecular mechanisms in each step of thehuman immunode®ciency virus (HIV) life cycle has provided an essential basisfor designing antiviral compounds and strategies aimed at blocking viral repli-cation and preventing or delaying disease progression. As in other retroviralinfections, the replication cycle of HIV can be described as proceeding in twophases. The ®rst phase includes entry of the virion into the cell cytoplasm,synthesis of double-stranded DNA (provirus) using the single-stranded genomeRNA as a template, transfer of the proviral DNA to the nucleus, and inter-gration of the DNA into the host genome. The second phase includes synthesisof new copies of the viral genome, expression of viral genes, virion assembly byencapsidation of the genome by precursors of the HIV structural proteins,budding, and ®nal processing of the viral proteins. Whereas the former phaseis mediated by proteins that are present within the virion and occurs in theabsence of viral gene expression, the latter, leading to production of infectiousvirions, is a complex process requiring the interplay of viral and cellular factors.
The precise understanding of the molecular mechanisms of HIV replicationand the use of new technologies in virology has lead to exciting discoveries onmany aspects of the biology of this virus. In particular, a growing body of newdata on the HIV replication mechanisms together with the results from molec-
3
-
ular studies carried out directly in vivo have allowed researchers to address thevirus-host relationships, including the pathogenic role of this virus in diseaseprogression.
In this chapter, two virologic aspects that are regulated by the complexmechanisms of the HIV-host interplay and have crucial pathogenic implicationsare discussed: the dynamics of HIV replication and the intrahost evolution ofthe HIV population.
HIV GENOME AND CONTROL OF VIRUS EXPRESSION
The HIV genome encodes for precursor polypeptides of structural and func-tional virion proteins, regulatory proteins, and other proteins that are dispens-able for replication and are called accessory proteins (Table 1.1). As for other
T A B L E 1.1. Genes of HIV, Proteins, and Function
HIV
Gene Protein Function
Essential for
Replication
gag Pr55gag Polyprotein precursor for matrix protein
(p17), capsid protein (p24), nucleocapsid
protein p9, and p7
Yes
pol Pr160gag-pol Polyprotein precursor for virion enzymes
reverse transcriptase (RT)/RNAse-H (p51),
protease (PR) (p10), and integrase (IN)
(p32)
Yes
env gp160 Polyprotein precursor for envelope
glycoproteins gp120 and gp41 (receptor
binding and membrane fusion, respectively)
Yes
tat p14 Transcriptional transactivator (initiation and
elongation of viral transcripts)
Yes
rev p19 Regulates viral gene expression at post-
transcriptional levels (regulates splicing and
transport of viral RNAs from the nucleus
to the cytoplasm)
Yes
nef p27 Downregulates CD4 receptor, enhances virion
infectivity, in¯uences T-cell activation
No
vif p23 Viral infectivity factor (infectivity reduced
in vif-minus mutants)
No
vpr p15 Virion protein (associated with the
nucleocapsid) implicated in regulation of
viral and cellular gene expression
No
vpu p16 In¯uences virus release No
4 HIV and Virus-Host Interplay
-
retroviruses, the genomic HIV RNA is synthesized and processed by the cellu-lar mRNA handling machinery starting from the proviral HIV DNA. For thisreason, the viral genome contains a cap structure at the 5 0 end and a poly-A tailat the 3 0 end. Moreover, the diploid lentiviral genome has the additional featureof being rich in A residues (on average 38±39%) (Myers and Pavlakis, 1992). Asa direct consequence, the HIV codon usage di¨ers dramatically from that ofcellular genes (Berkhout and van Hemert, 1994; Kypr et al., 1989).
Control of HIV RNA synthesis is complex and requires the presence ofseveral cellular proteins as well as of viral transactivators and cis-acting viralelements. Indeed, retroviral long terminal repeats (LTRs) are divided into do-mains (designated U3, R, and U5) that have distinct functions in transcriptioneither in regulating basal levels or inducing high levels of HIV gene expression.The U3 domain of HIV contains basal promoter elements, including the TATAbox for initiation by RNA polymerase II and the site for binding the cellulartranscription factor SP1. Immediately upstream of the core promoter, the viruscontains one or more copies of a 10-bp sequence recognized by the enhancerfactor nuclear factor (NF)-kB. However, whereas in simple retroviruses regu-lation of viral transcription is passive (i.e., regulated by cellular factors), in HIVinfection, this process is more complex and products of the HIV genome arerequired to achieve high levels of expression. Initiation of HIV RNA occurs atthe U3/R level (cap site) of the 5 0 LTR, and the viral transactivator Tat func-tions through a cis-acting sequence (designated Tat-responsive element, TAR)an RNA encoded by a region located in R (+ 19 to + 43). R-U5 is the leadersequence of the full-length and spliced viral transcript, whereas the 3 0 ends ofmRNAs are de®ned by the R/U5 border in the 3 0 LTR. Finally, the accessorygenes of HIV (vif, vpr, vpu, and nef ) (Table 1.1) are generally de®ned as dis-pensable for viral replication based on studies in tissue culture systems. On theother hand, accessory genes are expressed in vivo and increasing data indicatethat they play important roles in the virus-host interplay.
MOLECULAR CORRELATES AND DYNAMICS OF HIV ACTIVITYIN VIVO
The relevant data on mechanisms of HIV replication have been coupled withthe results from in vivo studies, thus obtaining a precise understanding of thevirus-host relationships. Indeed, natural history and pathogenicity studies havesupplied a pro®le of HIV activity during the di¨erent phases of this infection,have contributed to a better understanding of virus-host interactions, haveallowed the application of mathematical models to evaluate the intrahost HIVdynamics, and, ®nally, have provided a theoretical basis for therapeutic anti-viral intervention.
In vivo, systemic HIV activity is a formal entity that consists of a sum ofdynamic processes, including productive infection of target cells, release ofvirions outside the infected cell and eventually in the blood compartment, and
5Molecular Correlates and Dynamics of HIV Activity in vivo
-
de novo infection of permissive cells. The virus variables in¯uencing the levelof systemic HIV activity and cell-free virus dynamics include degree of viralexpression and host cell range, whereas the host variables include the speci®c(humoral and cytotoxic) immune response and polymorphism of genes codingfor cell receptors of HIV.
The vast majority of quantitative studies carried out in vivo have highlightedthe role of cell-free viremia as a reliable index of mean viral activity in HIVinfection. Indeed, viremia-based studies have provided clear evidence thatchanges in HIV load during the di¨erent phases of this infection can be e½-ciently evaluated by measuring cell-free virus in plasma samples (Bagnarelliet al., 1994; Perelson et al., 1996), and that substantial increases in viral loadparallel or even predict (Mellors et al., 1996) the disease progression. These®ndings have greatly contributed in the last few years to a clearer understand-ing of the virologic correlates of disease progression, to driving new attemptsat understanding the pathogenic potential of HIV, and to designing e¨ectiveantiretroviral strategies. Although recent research has highlighted the diagnos-tic role of other quantitative parameters, including viral transcription patternand provirus copy numbers, and although in some cases virus compartmen-talization may in¯uence the exact correspondence between cell-free plasmaviremia and systemic viral activity, the analysis of viral genome molecules inplasma samples is still a major molecular correlate of systemic viral activity atthe level of the whole body in many human viral infections.
The evaluation of patients undergoing potent antiviral treatments has allowedthe dynamics of cell-free virus in plasma to be addressed in vivo (Ho et al.,1995; Perelson et al., 1996; Wei et al., 1995). Importantly, these studies havedocumented the dynamics of cell-free virions in plasma (half-life being approx-imately 5.7 h) and the turnover of infected cells. Furthermore, the sensitivityand speci®city performances of most quantitative molecular methods haveprovided in the last few years a simple approach to the evaluation of genetranscription in vivo and in vitro. In HIV infection, consistent evidence has in-dicated that progression of disease is driven by an increase in viral load eval-uated as cell-free plasma virus. To address whether this increase is contributedby the dysregulation of the molecular mechanisms governing virus gene ex-pression at the transcriptional or post-transcriptional levels, several quantitativevirologic parameters (including provirus transcriptional activity and splicingpattern) have been analyzed in subjects with nonprogressive HIV infection andcompared with those of matching groups of progressor patients. It was ob-served not only that high levels of unspliced (US) and multiply spliced (MS)viral transcripts in peripheral blood mononuclear cells (PBMCs) correlate withthe decrease in CD4 T cells (Bagnarelli et al., 1996; Furtado et al., 1995) fol-lowing the general trend of systemic HIV-1 activity, but also that MS mRNAlevels in PBMCs are closely associated with the number of productively infectedcells (Bagnarelli et al., 1996), because the half-life of this class of transcriptsafter administration of a potent protease inhibitor is very consistent with that ofproductively infected cells. The transcriptional pattern observed during in vitro
6 HIV and Virus-Host Interplay
-
infections of T-cell lines, primary PBMCs, and monocytes/macrophages sup-ports these ®ndings.
INTER- AND INTRASUBJECT HIV VARIABILITY
Comparative analysis of the sequences of the HIV env gene from a greatnumber of viral isolates has revealed a pattern of ®ve hypervariable regions(designated V1 to V5) interspersed with more conserved sequences in the gp120.This sequence variation consists of mutations (resulting in amino acid sub-stitutions), insertions, and deletions (Leigh-Brown, 1991). Among HIV isolatesfrom geographically di¨erent locations, gp120 amino acid sequences may di-verge up to 20±25%, whereas other regions of the genome are relatively con-served. More recently, molecular epidemiology surveys based on env sequencesof numerous HIV isolates have revealed at least nine distinct HIV subtypes (orclades) in the acquired immunode®ciency syndrome (AIDS) pandemic (Myers,1994; Myers et al., 1994) (intersubject HIV variability).
Subsequent analysis has revealed that both linear and conformational de-terminants in¯uence the functional and antigenic structure of the gp120; this isa crucial pathogenic issue, inasmuch as all neutralizing antibodies are directedagainst env-encoded domains in HIV-infected hosts. Indeed, infections withretroviruses are also characterized by di¨erent (from moderate to high) levels ofintrahost viral genetic variation. This viral variability is dependent upon muta-tion, recombination, degree of viral replication, and the host's selective pressure(Dougherty and Temin, 1988; Hu and Temin, 1990; Pathak and Temin, 1990a,b; Temin, 1993). In HIV infection, the viral population is represented by re-lated, nonidentical genetic variants (Goodenow et al., 1989; Hahn et al., 1996;Meyerans et al., 1989; Pedroza Martins et al., 1992), designated quasispecies.The error-prone nature of the HIV reverse transcriptase (RT) and the absenceof a 3 0-exonuclease proofreading activity determine in vitro about 3 × 10ÿ5
mutations per nucleotide per replication cycle (Yu and Goodman, 1992).Although the mutation rate observed in vivo is lower than that predicted fromthe ®delity of puri®ed RT (because a number of newly generated variants areunable to replicate or are cleared by the host's immune system) (Mansky andTemin, 1995), the viral replication dynamics (Ho et al., 1995) and the host'sselective forces determine a continuous process of intrahost HIV evolution(Bagnarelli et al., 1999; Holmes et al., 1992; McNearney et al., 1992; Wolinskyet al., 1996). Overall, the data currently suggest that viral genetic variability isthe molecular counterpart of a continuous dynamic interplay between viral(i.e., HIV-1 replication dynamics and generation of variants by mutation andrecombination) and host factors (i.e., selective pressure). In this context, intra-host evolution of HIV-1 populations may be compatible with a Darwinianmodel system, as recently suggested (Bagnarelli et al., 1999; Ganeshan et al.,1997; Wolinsky et al., 1996).
The complete elucidation of the mechanisms driving intrahost HIV-1 evolu-
7Inter- and Intrasubject HIV Variability
-
tion is of crucial importance for understanding the natural history of this in-fection and developing e¨ective anti-HIV strategies. In particular, the envelopeglycoproteins of HIV-1 interact with receptors of the target cells and mediatethe process of virus entry. This process is complex, including binding of theviral gp120 to CD4, conformational changes of the viral glycoprotein, andsubsequent use of a coreceptor before gp41-mediated fusion of the viral enve-lope and the cellular membrane (Kwong et al., 2000; Rizzuto and Sodroski,2000; Rizzuto et al., 1998; Wu et al., 1996; Wyatt et al., 1995; Zhang et al.,1999) (Table 1.2). The evolutionary changes characterizing the HIV-1 popula-tion during the natural history of infection strongly in¯uence crucial regionsof the viral env gene (Bagnarelli et al., 1999; Menzo et al., 1998; Salvatori etal., 1997; Scarlatti et al., 1997; Shankarappa et al., 1998, 1999; Wolinsky et al.,1996). Because di¨erent variable domains of the HIV-1 gp120 play a key role indriving the early steps of the viral infection cycle, including coreceptor usage(Isaka et al., 1999; Sato et al., 1999; Verrier et al., 1999; Xiao et al., 1998) andCD4 independence (LaBranche et al., 1999), careful analysis of the intrahostevolution of the HIV-1 env gene is strategic for addressing the relevant featuresof the virus-host relationships (Yamaguchi and Gojobori, 1997; Yamaguchi-Kabata and Gojobori, 2000). In addition, HIV entry is at present an attractivetarget for new classes of antiretroviral compounds (Sodroski, 1999); at present,these compounds include inhibitors of HIV binding to CCR5 and CXCR4coreceptors and fusion inhibitors (Murakami et al., 1999; Ono et al., 1997;Sakaida et al., 1998; Torre et al., 2000).
The V3 sequence is a variable domain in the HIV gp120 and contains 35amino acids arranged in a loop. This domain plays a crucial role in drivingimportant biological properties of the virus, including cell tropism. Generally,mutations in the V3 loop do not a¨ect the ability of gp120 to interact with theCD4 receptor, although several studies have unambiguously indicated that V3sequences play an important role in two correlated biological features withpathogenic implications, that is, syncytium formation (Willey et al., 1994) and
T A B L E 1.2. HIV Cell Receptors and Their Natural Ligands
CD4 CCR5 CXCR4 CCR3 CCR2b BOB BONZO
HIV
NSI ÿ SI ()
Natural ligands
MHC MIP-1a SDF-1 RANTES MCP-1 ? ?
Cl. II MIP-1b MCP-3 MCP-2
RANTES EOTAXIN MCP-3
MCP-4
NSI, nonsyncytium inducing; SI, syncytium inducing.
8 HIV and Virus-Host Interplay
-
coreceptor usage (Isaka et al., 1999). Importantly, analysis of chimeric viruseshas revealed that changes in the V3 loop can convert a nonsyncytium inducing(NSI), slowly replicating virus into a syncytium inducing (SI), rapidly repli-cating virus (Shioda et al., 1992).
CONCLUSION
Expanded analysis of the molecular biology of HIV has been the key to under-standing the mechanisms by which this virus persists in the host and causesAIDS, and to developing e¨ective antiretroviral strategies. Application of pow-erful molecular biology tools has allowed researchers to obtain fundamentalresults on many aspects of HIV biology in vitro (i.e., in cell-free and tissueculture systems) and in vivo (i.e., directly in samples from the susceptible host).Importantly, knowledge of the molecular mechanisms in each step of the viruslife cycle has provided an essential basis for discovering new antiviral com-pounds. Otherwise, a ®rm understanding of the relevant features of both theHIV turnover in vivo and the intrahost HIV evolution is crucial for developinge¨ective anti-HIV strategies. Indeed, the HIV biology poses several challengesto the development of these strategies. In particular, sequence variation result-ing from errors of the viral RT and recombination renders HIV an elusive tar-get for both antiviral compounds and vaccines. In this context, novel diagnos-tic molecular tools to control development of viral resistance to the di¨erentclasses of antivirals and new e¨ective therapeutic approaches, including geneticand immunologic strategies, could be the key to inhibiting HIV replication inthe future.
REFERENCES
Bagnarelli P, Valenza A, Menzo S, Manzin A, Scalise G, Varaldo PE, Clementi M. 1994. Dynamics
of molecular parameters of human immunode®ciency virus type 1 activity in vivo. J Virol 68:
2495±2502.
Bagnarelli P, Valenza A, Menzo S, Sampaolesi R, Varaldo PE, Butini L, Montroni M, Perno CF,
Aquaro S, Mathez D, Leibowitch J, Balotta C, Clementi M. 1996. Dynamics and modulation of
human immunode®ciency virus type 1 transcripts in vitro and in vivo. J Virol 70: 7603±7613.
Bagnarelli P, Mazzola F, Menzo S, Montroni M, Butini L, Clementi M. 1999. Host-speci®c mod-
ulation of the selective constraints driving human immunode®ciency virus type 1 env gene evo-
lution. J Virol 73: 3764±3777.
Berkhout B, van Hemert, FJ. 1994. The unusual nucleotide content of the HIV RNA genome re-
sults in a biased amino acid composition of HIV proteins. Nucleic Acid Res 9: 1705±1711.
Dougherty JP, Temin HM. 1988. Determination of the rate of base-pair substitution and insertion
mutations in retrovirus replication. J Virol 62: 2817±2822.
Furtado MR, Kingsley LA, Wolinsky SM. 1995. Changes in the viral mRNA expression pattern
correlate with a rapid rate of CD4 T cell number decline in human immunode®ciency virustype 1-infected individuals. J Virol 69: 2092±2100.
9References
-
Ganeshan S, Dickover RE, Korber BT, Bryson YJ, Wolinsky SM. 1997. Human immunode®ciency
virus type 1 genetic evolution in children with di¨erent rates of development of disease. J Virol
71: 663±677.
Goodenow M, Huet T, Saurin W, Kwok S, Sninsky J, Wain-Hobson S. 1989. HIV-1 isolates are
rapidly evolving quasispecies: evidence for viral mixtures and preferred nucleotide substitutions.
J Acquired Immune De®c Syndr 2: 344±352.
Hahn BH, Shaw GM, Taylor ME, Red®eld RR, Markham PD, Salahuddin SZ, Wong-Staal F,
Gallo RC, Parks ES, Parks WP. 1996. Genetic variation in HTLV III/LAV over time in patients
with AIDS or at risk for AIDS. Science 232: 1548±1553.
Ho DD, Neuman AU, Perelson AS, Chen W, Leonard JM, Markowitz M. 1995. Rapid turnover of
plasma virions and CD4 lymphocytes in HIV-1 infection. Nature 373: 123±126.
Holmes EC, Zhang LQ, Simmonds P, Ludlam CA, Leigh-Brown AJ. 1992. Convergent and diver-
gent sequence evolution in the surface glycoprotein of human immunode®ciency virus type 1
within a single infected patient. Proc Natl Acad Sci USA 89: 4835±4839.
Hu WS, Temin HM. 1990. Genetic consequences of packaging two RNA genomes in one retroviral
particle: pseudodiploidy and a high rate of genetic recombination. Proc Natl Acad Sci USA 87:
1556±1560.
Isaka Y, Sato A, Miki S, Kawauchi S, Sakaida H, Hori T, Uchiyama T, Adachi A, Hayami M,
Fujiwara T, Yoshie O. 1999. Small amino acid changes in the V3 loop of human immuno-
de®ciency virus type 2 determines the coreceptor usage for CXCR4 and CCR5. Virology 264:
237±243.
Kwong PD, Wyatt R, Sattentau QJ, Sodroski J, Hendrickson WA. 2000. Oligomeric modeling and
electrostatic analysis of the gp120 envelope glycoprotein of human immunode®ciency virus. J
Virol 74: 1961±1972.
Kypr J, Mrazek J, Reich J. 1989. Nucleotide composition bias and CpG dinucletide content in the
genomes of HIV and HTLV 1/2. Biochim Biopys Acta 1009: 280.
LaBranche CC, Ho¨man TL, Romano J, Haggarty BS, Edwards TG, Matthews TJ, Doms
RW, Hoxie JA. 1999. Determinants of CD4 independence for a human immunode®ciency
virus type 1 variant map outside regions required for coreceptor speci®city. J Virol 73: 10310±
10319.
Leigh-Brown AJ. 1991. Sequence variability of human immunode®ciency viruses: pattern and pro-
cess in viral evolution. AIDS 5S: 35±42.
Mansky LM, Temin HM. 1995. Lower in vivo mutation rate of human immunode®ciency type 1
than that predicted from the ®delity of puri®ed reverse transcriptase. J Virol 69: 5087±5094.
McNearney T, Hornickowa Z, Markham R, Birdwell A, Arens M, Saah A, Ratner L. 1992. Rela-
tionship of human immunode®ciency virus type 1 sequence heterogeneity to stage of disease.
Proc Natl Acad Sci USA 89: 10247±10251.
Mellors JW, Rinaldo CR, Gupta P, White RM, Todd JA, Kingsley LA. 1996. Prognosis in HIV
infection predicted by the quality of virus in plasma. Science 272: 1167±1170.
Menzo S, Sampaolesi R, Vicenzi E, Santagostino E, Liuzzi G, Chirianni A, Piazza M, Cohen OJ,
Bagnarelli P, Clementi M. 1998. Rare mutations in a domain crucial for V3-loop structure pre-
vail in replicating HIV from long-term non-progressors. AIDS 12: 985±997.
Meyerans A, Cheynier R, Albert J, Seth M, Kwok S, Sninsky J, Morfeldt-Manson L, Asjo B,
Wain-Hobson S. 1989. Temporal ¯uctuations in HIV quasispecies in vivo are not re¯ected by
sequential HIV isolations. Cell 58: 901±910.
Murakami T, Zhang TY, Koyanagi Y, Tanaka Y, Kim J, Suzuki Y, Minoguchi S, Tamamura H,
Waki M, Matsumoto A, Fujii N, Shida H, Hoxie JA, Peiper SC, Yamamoto N. 1999. Inhibi-
tory mechanism of the CXCR4 antagonist T22 against human immunode®ciency virus type 1
infection. J Virol 73: 7489±7496.
Myers G. 1994. HIV: between past and future. AIDS Res Hum Retroviruses 10: 1317±1324.
10 HIV and Virus-Host Interplay
-
Myers G, Pavlakis GN. 1992. Evolutionary potential of complex retroviruses. In: Levy JA (ed). The
Retroviridae. New York, Plenum Press, pp 51±105.
Myers G, Kolber B, Wain-Hobson S, Jeang KT, Henderson LE, Pavlakis GN. 1994. Human
Retroviruses and AIDS. A Compilation and Analysis of Nucleic Acid and Amino Acid Sequences.
Los Alamos, NM, Los Alamos National Laboratory.
Ono M, Wada Y, Wu Y, Nemori R, Jinbo Y, Wang H, Lo KM, Yamaguchi N, Brunkhorst B,
Otomo H, Wesolowski J, Way JC, Itoh I, Gillies S, Chen LB. 1997. FP-21399 blocks HIV en-
velope protein-mediated membrane fusion and concentrates in lymph nodes. Nat Biotechnol 15:
343±348.
Pathak VK, Temin HM. 1990a. Broad spectrum of in vivo forward mutations, hypermutations,
and mutational hotspots in a retroviral shuttle vector after a single replication cycle: deletions
and deletions with insertions. Proc Natl Acad Sci USA 87: 6024±6028.
Pathak VK, Temin HM. 1990b. Broad spectrum of in vivo forward mutations, hypermutations, and
mutational hotspots in a retroviral shuttle vector after a single replication cycle: substitutions,
frameshifts, and hypermutations. Proc Natl Acad Sci USA 87: 6019±6023.
Pedroza Martins L, Chenciner N, Wain-Hobson S. 1992. Complex intrapatient sequence variation
in the V1 and V2 hypervariable regions of the HIV-1 gp120 envelope sequence. Virology 191:
837±845.
Perelson AS, Neumann AU, Markowitz M, Leonard JM, Ho DD. 1996. HIV dynamics in vivo:
virion clearance rate, infected cell life span, and viral generation time. Science 271: 1582±
1586.
Rizzuto C, Sodroski J. 2000. Fine de®nition of a conserved CCR5-binding region on the human
immunode®ciency virus type 1 glycoprotein 120 [In Process Citation]. AIDS Res Hum Retro-
viruses 16: 741±749.
Rizzuto CD, Wyatt R, Hernandez-Ramos N, Sun Y, Kwong PD, Hendrickson WA, Sodroski J.
1998. A conserved HIV gp120 glycoprotein structure involved in chemokine receptor binding
[see comments]. Science 280: 1949±1953.
Sakaida H, Hori T, Yonezawa A, Sato A, Isaka Y, Yoshie O, Hattori T, Uchiyama T. 1998.
T-tropic human immunode®ciency virus type 1 (HIV-1)-derived V3 loop peptides directly bind
to CXCR-4 and inhibit T-tropic HIV-1 infection. J Virol 72: 9763±9770.
Salvatori F, Masiero S, Giaquinto C, Wade CM, Brown AJ, Chieco-Bianchi L, De Rossi A. 1997.
Evolution of human immunode®ciency virus type 1 in perinatally infected infants with rapid and
slow progression to disease. J Virol 71: 4694±4706.
Sato H, Kato K, Takebe Y. 1999. Functional complementation of the envelope hypervariable V3
loop of human immunode®ciency virus type 1 subtype B by the subtype E V3 loop. Virology
257: 491±501.
Scarlatti G, Tresoldi E, Bjorndal A, Fredriksson R, Colognesi C, Deng HK, Malnati MS, Plebani
A, Siccardi AG, Littman DR, Fenyo EM, Lusso P. 1997. In vivo evolution of HIV-1 co-receptor
usage and sensitivity to chemokine-mediated suppression. Nat Med 3: 1259±1265.
Shankarappa R, Gupta P, Learn Jr GH, Rodrigo AG, Rinaldo Jr CR, Gorry MC, Mullins JI, Nara
PL, Ehrlich GD. 1998. Evolution of human immunode®ciency virus type 1 envelope sequences
in infected individuals with di¨ering disease progression pro®les. Virology 241: 251±259.
Shankarappa R, Margolick JB, Gange SJ, Rodrigo AG, Upchurch D, Farzadegan H, Gupta P,
Rinaldo CR, Learn GH, He X, Huang XL, Mullins JI. 1999. Consistent viral evolutionary
changes associated with the progression of human immunode®ciency virus type 1 infection. J
Virol 73: 10489±10502.
Shioda T, Levy JA, Cheng-Mayer C. 1992. Small amino acid changes in the V3 hypervariable re-
gion of the gp120 can a¨ect the T-cell line and macrophage tropism of human immunode®ciency
virus type 1. Proc Natl Acad Sci USA 89: 9434±9438.
Sodroski JG. 1999. HIV-1 entry inhibitors in the side pocket. Cell 99: 243±246.
11References
-
Temin HM. 1993. Retrovirus variation and reverse transcription: abnormal strand transfers results
in retrovirus genetic variation. Proc Natl Acad Sci USA 280: 1949±1953.
Torre VS, Marozsan AJ, Albright JL, Collins KR, Hartley O, O¨ord RE, Quinones-Mateu ME,
Arts EJ. 2000. Variable sensitivity of CCR5-tropic human immunode®ciency virus type 1 iso-
lates to inhibition by RANTES analogs. J Virol 74: 4868±4876.
Verrier F, Borman AM, Brand D, Girard M. 1999. Role of the HIV type 1 glycoprotein 120 V3
loop in determining coreceptor usage. AIDS Res Hum Retroviruses 15: 731±743.
Wei X, Gosh SK, Taylor ME, Johnson VA, Emini EA, Deutsch P, Lifson JD, Bonhoefer S, Nowak
MA, Hahn BH, Saag MS, Shaw GM. 1995. Viral dynamics in human immunode®ciency virus
type 1 infection. Nature 373: 117±122.
Willey RL, Theodore TS, Martin MA. 1994. Amino acid substitutions in the human immuno-
de®ciency virus type 1 gp120 V3 loop that change viral tropism also alter physical and functionsl
properties of the virion envelope. J Virol 68: 4409±4419.
Wolinsky SM, Korber BTM, Neumann AU, Daniels M, Kunstman KJ, Whetsell AJ, Furtado MR,
Cao Y, Ho DD, Safrit JT, Koup RA. 1996. Adaptive evolution of human immunode®ciency
virus-type 1 during the ntural course of infection. Science 272: 537±542.
Wu L, Gerard NP, Wyatt R, Choe H, Parolin C, Ru½ng N, Borsetti A, Cardoso AA, Desjardin E,
Newman W, Gerard C, Sodroski J. 1996. CD4-induced interaction of primary HIV-1 gp120
glycoproteins with the chemokine receptor CCR-5 [see comments]. Nature 384: 179±183.
Wyatt R, Moore J, Accola M, Desjardin E, Robinson J, Sodroski J. 1995. Involvement of the V1/
V2 variable loop structure in the exposure of human immunode®ciency virus type 1 gp120
epitopes induced by receptor binding. J Virol 69: 5723±5733.
Xiao L, Owen SM, Goldman I, Lal AA, deJong JJ, Goudsmit J, Lal RB. 1998. CCR5 coreceptor
usage of non-syncytium-inducing primary HIV-1 is independent of phylogenetically distinct
global HIV-1 isolates: delineation of consensus motif in the V3 domain that predicts CCR-5
usage. Virology 240: 83±92.
Yamaguchi Y, Gojobori T. 1997. Evolutionary mechanisms and population dynamics of the third
variable envelope region of HIV within single hosts. Proc Natl Acad Sci USA 94: 1264±1269.
Yamaguchi-Kabata Y, Gojobori T. 2000. Reevaluation of amino acid variability of the human
immunode®ciency virus type 1 gp120 envelope glycoprotein and prediction of new discontinuous
epitopes. J Virol 74: 4335±4350.
Yu H, Goodman MF. 1992. Comparison of HIV-1 and avian myeloblastosis virus reverse tran-
scriptase ®delity on RNA and DNA templates. J Biol Chem 267: 10888±10896.
Zhang W, Canziani G, Plugariu C, Wyatt R, Sodroski J, Sweet R, Kwong P, Hendrickson W,
Chaiken I. 1999. Conformational changes of gp120 in epitopes near the CCR5 binding site are
induced by CD4 and a CD4 miniprotein mimetic. Biochemistry 38: 9405±9416.
12 HIV and Virus-Host Interplay
-
2Telomere Length, CD28ÿ T Cells, and HIVDisease Pathogenesis
Rita B. EffrosDepartment of Pathology and Laboratory Medicine, UCLA Medical Center,Los Angeles, California, USA
INTRODUCTION
In human immunode®ciency virus (HIV) disease, as in all viral infections, CD8T cells constitute a critical component of the protective immune response(Borrow et al., 1994; Brodie et al., 1999; Koup et al., 1994). Loss of CD8 T-cellactivity coincides with the progression to acquired immunode®ciency syndrome(AIDS), and studies on long-term nonprogressors have underscored the impor-tance of cytotoxic T lymphocyte (CTL) function (Cao et al., 1995; Goulder etal., 1997; Harrer et al., 1996). One of the intriguing alterations in the peripheralT-cell pool of individuals infected with HIV is the progressive accumulationwithin the CD8 T-cell subset of a population of cells that lack expression of theCD28 costimulatory molecule. Indeed, in some HIV-infected persons, >65% ofthe CD8 T cells are CD28ÿ. A more complete characterization of this unusualcell population, therefore, is essential for understanding disease pathogenesis aswell as for the development of appropriate strategies for treatment. BecauseCD28ÿ T cells are poorly proliferative, do not contribute to production ofsoluble antiviral suppressive factors, and also show alterations in apoptosis andin cell-cell adhesion, the presence of large proportions of such cells will un-doubtedly have a profound in¯uence on the immune control over HIV infec-
This chapter is dedicated to my friend and colleague Janis V. Giorgi.
13
-
tion. Although there had been much speculation by AIDS researchers on theorigin of the CD28ÿ T cells, elucidation of the nature of this expanded popu-lation of cells in HIV disease has emerged from research in a totally di¨erentarena of scienti®c investigation, namely basic cell biology studies on the processof replicative senescence. This chapter will review the ®ndings that have led tothe unexpected convergence of these two seemingly unrelated ®elds.
REPLICATIVE SENESCENCE
Normal human somatic cells have an intrinsic natural barrier to unlimited celldivision. Following a fairly predictable number of cell divisions in culture,most, if not all, mitotically competent human cells reach an irreversible state ofgrowth arrest known as replicative senescence, a process ®rst identi®ed byHay¯ick in human fetal ®broblasts (Hay¯ick, 1965). Replicative senescence is astrict characteristic of human cells, and has, in fact, been proposed to constitutea tumor suppressive mechanism (Smith and Pereira-Smith, 1996). Interestingly,experimental cell fusion studies have demonstrated that the property of senes-cence is genetically dominant over immortality in a variety of human cell types,and spontaneous transformation of human cells in vitro rarely, if ever, occurs(Smith and Pereira-Smith, 1996). By contrast, most rodent cells have a highpropensity to bypass senescence and transform spontaneously in culture (Cam-pisi et al., 1996). The divergent behavior of human and mouse cells with respectto spontaneous immortalization in vitro suggests that conclusions regardingreplicative properties, telomeres, and telomerase drawn from murine studiesmay not be applicable to human cells.
The characteristics of replicative senescence, or the so-called Hay¯ick Limit,have been explored in a variety of human cell types for more than 30 years, butonly relatively recently has this model been applied to the immune system.Ironically, the Hay¯ick Limit may be particularly deleterious for immune cells,inasmuch as the ability to undergo rapid clonal expansion is absolutely essentialto their function.
During the past decade, human T cells have been extensively characterizedin cell culture models with respect to replicative senescence. A number of large-scale studies have shown that following multiple rounds of antigen, mitogen, oractivatory antibody-driven proliferation, T cells reach a state of growth arrestthat cannot be reversed by further exposure to antigen, growth factors, or anyother established T-cell stimuli (E¨ros and Pawelec, 1997). The occurrence ofreplicative senescence has been documented for both clonal and bulk culturesof CD4 and CD8 T cells (Adibzadeh et al., 1995; Grubeck-Loebenstein etal., 1994; McCarron et al., 1987). It has also been shown that the replicativepotential of memory CD4 T cells is reduced compared with naõÈve CD4 T cellsfrom the same individual, a ®nding that is consistent with the notion thatmemory cells are the progeny of antigen-stimulated naõÈve T cells (Weng et al.,1995). It is important to emphasize that although cell cycle arrest is the most
14 Telomere Length, CD28ÿ T Cells, and HIV Pathogenesis