pn A,
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`64606/
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`he
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`Miltenyi Ex. 1030 Page 1
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`Miltenyi Ex. 1030 Page 1
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`Eachissue contains hundreds of pages of up-to-date research. The
`“Cutting Edge” section of the journal features the highest impact
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`Miltenyi Ex. 1030 Page 2
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`

`

`VOL. 171 | NO. 11 | December 1, 2003 | Pages 5649-6312
`
`eek che
`
`5653
`
`Early T Lineage Progenitors: New Insights, but Old Questions Remain
`Avinash Bhandoola, Arivazhagan Sambandam, DavidAllman, Anita Meraz, andBenjamin Schwarz
`
`[F
`
`
`5659 Cutting Edge: SDS-Stable Fas Microaggregates: An Early Eventof Fas Activation Occurring with
`Agonistic Anti-Fas Antibody but Not with Fas Ligand
`Patrick Legembre, Marie Beneteau, Sophie Daburon, Jean-Francois Moreau, andJean-Luc Taupin
`5663 Cutting Edge: IncreasedNK CellActivityin HIV-1-Exposed but Uninfected Vietnamese Intravascular:
`Drug Users
`DanielScott-Algara, Lien X. Truong, Pierre Versmisse, Annie David, Tram T. Luong, Ngai V. Nguyen, .
`loannis Theodorou, Francoise Barré-Sinoussi, and Gianfranco Pancino
`5668 Cutting Edge: Efficient MHC Class I Cross-Presentation during Early Vaccinia Infection Requires the
`Transfer of Proteasomal Intermediates between Antigen Donor and Presenting Cells
`Amparo Serna, Maria C. Ramirez, Anna Soukhanova, and Luis]. Sigal
`5673 Cutting Edge: Transplantation Tolerance throughEnhanced CTLA-4 Expression
`CharlotteAriyan, Paolo Salvalaggio, ScottFecteau, Songyan Deng, Linda Rogozinski, DidierMandelbrot, Arlene Sharpe,
`Mohamed H. Sayegh, Giacomo Basadonna, and DavidM. Rothstein
`
`5678 Cutting Edge: Self-Peptides Drive the Peripheral Expansion of CD4*CD25+* Regulatory T Cells
`Cristina Cozzo, Joseph Larkin II, andAndrewJ. Caton
`
`5683 Cutting Edge: TCR Contacts as Anchors: Effects on Affinity and HLA-DMStability
`Matthew W. Anderson'andJack Gorski
`
`
`—_—_——-—
`_
`we
`Onthecover: Side-view projection of leukocyte-endothelial interaction reconstructed from serial section confocal
`microscopy. Endothelial ICAM-1 (green)rapidly relocalized to newly formed microvilli-like membrane projectionsin
`responseto binding LFA-1 (red) on leukocytes. These ICAM-1-enriched projectionsencircled the leukocytes extending up
`their sides, and clustered LFA-1 underneath into linear tracks. Colocalized ICAM-I and LEA-1 appear yellow. Carman et
`al. Endothelial cells proactively form microvilli-like membrane projections upon intercellular adhesion molecule 1
`engagementofleukocyte LFA-1. J. Immunol. 171:6135.
`
`
`
`TheJournalofImmunology (ISSN 0022-1767)is published twice each month by The American Association ofImmunologists,
`9650 Rockville Pike, Bethesda, MD 20814-3998, Subscription rates: U.S.A. AND POSSESSIONS:personal, $330;
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`www.jimmunol.org for current rate information. Air Printed Matterrates are available on request from Customer Service, The
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`U.S.A.’Printed on acid-free recyclable paper. Postmaster: Send address changes to TheJournalofImmunology, Room L-2310,
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`\
`Miltenyi Ex. 1030 Page 3
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`
`
`
`
`
`
`
`
`
`5689
`
`5697
`
`5707
`
`5718
`
`5727
`
`5736
`
`5743
`
`5751
`
`5760
`
`5769
`
`5778
`
`5787
`
`5795
`
`5802
`
`5812
`
`Requirements for T Cell-Polarized Tubulation of Class IIT Compartments in Dendritic Cells
`Nicolas Bertho, Jan Cerny, You-Me Kim, Edda Fiebiger, Hidde Ploegh, and Marianne Boes
`
`CD4* T Cell-Associated Pathophysiology Critically Depends on CD18 Gene Dose Effects in a Murine
`Model ofPsoriasis
`Daniel Kess, Thorsten Peters, Jan Zamek, Claudia Wickenhauser, Samir Tawadros, Karin Loser, Georg Varga,
`Stephan Grabbe, Roswitha Nischt, Cord Sunderkotter, Werner Miiller, Thomas Krieg, and Karin Scharffetter-Kochanek
`
`CD40, but Not CD 154, Expression on B Cells Is Necessary for Optimal Primary B Cell Responses
`Byung O. Lee, Juan Moyron-Quiroz, Javier Rangel-Moreno, Kim L. Kusser, Louise Hartson, Frank Sprague,
`Frances E. Lund, and Troy D. Randall
`
`Tethering ofApoptotic Cells to Phagocytes through Binding of CD47 to Src Homology 2 Domain-
`Bearing Protein Tyrosine Phosphatase Substrate-1
`Kazutoshi Tada, Masato Tanaka, Rikinari Hanayama, Keiko Miwa, Azusa Shinohara, Akihiro Iwamatsu,
`and Shigekazu Nagata
`
`IL-2 Is Not Required for the Initiation of CD8 T Cell Cycling but Sustains Expansion
`Warren N. D'Souza and Leo Lefrangois
`
`AberrantExtracellular and Dendritic Cell (DC) Surface Expression of Heat Shock Protein (hsp)70 in the
`Rheumatoid Joint: Possible Mechanisms of hsp/DC-Mediated Cross-Priming
`Carla A. Martin, Steven E. Carsons, Robert Kowalewski, David Bernstein, Michael Valentino,
`and Frances Santiago-Schwarz
`
`PeroxisomeProliferator-Activated Receptor-y-Deficient Heterozygous Mice Develop an Exacerbated
`Neural Antigen-Induced Th1 Response and Experimental Allergic Encephalomyelitis
`John Jj. Bright, Chandramohan Natarajan, Gladson Muthian, Yaavov Barak, and RonaldM. Evans
`
`TNF Receptor-Associated Factor 6 Deficiency during Hemopoiesis Induces Th2-Polarized Inflammatory
`Disease
`Elise Chiffoleau, Takashi Kobayashi, Matthew C. Walsh, Carolyn G. King, Patrick T. Walsh, Wayne W. Hancock,
`Yongwon Choi, and Laurence A. Turka
`
`Human CD25* Regulatory T Cells Maintain Immune Tolerance to Nickel in Healthy, Nonallergic
`Individuals
`Andrea Cavani, Francesca Nasorri, Chiara Ottaviani, Silvia Sebastiani, Ornella De Pita, and Giampiero Girolomoni
`
`In Vitro Differentiation from Naive to Mature E-Selectin Binding CD4 T Cells: Acquisition of Skin-
`Homing Properties Occurs Independently of Cutaneous Lymphocyte Antigen Expression
`Ryo Takahashi, Yoshiko Mizukawa, Yoshimi Yamazaki, Kazuhito Hayakawa, Jun Hayakawa, Akihiko Kudo,
`and Tetsuo Shiohara
`
`Ultraviolet B Radiation-Induced Cell Death: Critical Role of Ultraviolet Dose in Inflammation and Lupus
`Autoantigen Redistribution
`Roberto Caricchio, Lenese McPhie, and Philip L. Cohen
`
`Human’ErbB-2 (Her-2) Transgenic Mice: A Model System for Testing Her-2 Based Vaccines
`Marie P. Piechocki, Ye-Shih Ho, Shari Pilon, and Wei-Zen Wei
`
`Increased Severity of Murine Lupusin Female Mice Is Due to Enhanced Expansionof Pathogenic T Cells
`ThomasJ. Lang, Phuong Nguyen, John C. Papadimitriou, and Charles S. Via
`
`Role ofAntiproliferative B Cell Translocation Gene-1 as an Apoptotic Sensitizer in Activation-Induced
`Cell Death of Brain Microglia
`Heasuk Lee, Sanghoon Cha, Myung-Shik Lee, GyeongJae Cho, Wan Sung Choi, and Kyoungho Suk
`
`MHCClass II-Peptide Complexes in Dendritic Cell Lipid MicrodomainsInitiate the CD4 Th1
`Phenotype
`Vanessa Buatois, Marjorie Baillet, Stéphane Bécart, Nuala Mooney, Lee Leserman, and Patrick Machy
`
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`5828
`
`The Duration of Signaling through CD40 Directs Biological Ability of Dendritic Cells to Induce
`Antitumor Immunity
`‘
`Satoshi Watanabe, Hiroshi Kagamu, Hirohisa Yoshizawa, Nanae Fujita, Hiroshi Tanaka, Junta Tanaka,
`and Fumitake Gejyo
`
`5837 Maturation of Dendritic Cell 2 Phenotype by a Helminth Glycan Usesa Toll-Like Receptor 4-Dependent
`Mechanism
`Paul G. Thomas, Michele R. Carter, Olga Atochina, Akram A. Da’Dara, Danuta Piskorska, Edward McGuire,
`and DonaldA. Harn
`
`5842 Dendritic Cell-Induced Activation ofAdaptive and Innate Antitumor Immunity
`Leon T. van den Broeke, Emily Daschbach, Elaine K. Thomas, Gerda Andringa, andJay A. Berzofsky
`
`5853 Distinct Effects of STATS Activation on CD4* and CD8* T Cell Homeostasis: Developmentof
`CD4*CD25* Regulatory T Cells versus CD8* Memory T Cells
`Matthew A. Burchill, Christine A. Goetz, Martin Prlic, JenniferJ. O’Neil, Ian R. Harmon, Steven J. Bensinger,
`Laurence A. Turka, Paul Brennan, Stephen C. Jameson, and MichaelA. Farrar
`
`5865
`
`Systemic Administration of IL-18 Promotes Diabetes Development in Young Nonobese Diabetic Mice
`Yoichi Oikawa, Akira Shimada, Akira Kasuga, Jiro Morimoto, Tadashi Osaki, Hideaki Tahara, Tatsushi Miyazaki,
`Fumi Tashiro, Eyji Yamato, Jun-ichi Miyazaki, and Takao Saruta
`
`5876 Ligation of CD27 on B Cells In Vivo during Primary Immunization Enhances Commitment to Memory
`B Cell Responses
`Vanitha S. Raman, Rama S. Akondy, Satyajit Rath, Vineeta Bal, andAnna George
`
`‘
`
`5882
`
`5890
`
`5901
`
`Fibronectin-Associated Fas Ligand Rapidly Induces Opposing and Time-DependentEffects on the
`Activation and Apoptosis of T Cells
`Alexandya Zanin-Zhorov, Rami Hershkoviz, Iris Hecht, Liora Cabalon, and Ofer Lider
`
`Tolerance through Indifference: Autoreactive B Cells to the Nuclear Antigen La Show No Evidence of
`Tolerance in a Transgenic Model
`Brett D. Aplin, Catherine L. Keech, Andrea L.de Kauwe, Thomas P. Gordon, Dana Cavill, andJames McCluskey
`
`IL-4 and IL-13 Induce SOCS-1 Gene Expression in A549 Cells by Three Functional STAT6-Binding
`Motifs Located Upstream of the Transcription Initiation Site
`Daniel Hebenstreit, Petra Luft, Angela Schmiedlechner, Gerhard Regl, Anna-Maria Frischauf, Fritz Aberger,
`Albert Duschl, andJutta Horejs-Hoeck
`
`5908 Vaccination with Plasmid DNA Activates Dendritic Cells via Toll-Like Receptor 9 (TLR9) but Functions
`in TLR9-Deficient Mice
`Barbara Spies, Hubertus Hochrein, Martin Vabulas, Katharina Huster, Dirk H. Busch, Frank Schmitz, Antje Heit,
`and Hermann Wagner
`
`5913 Control of NKT Cell Differentiation by Tissue-Specific Microenvironments
`Yang Yang, Aito Ueno, Min Bao, Zhongying Wang, Jin Seon Im, Steven Porcelli, andJi-Won Yoon
`
`5921
`
`CD19 Function in Early and Late B Cell Development. II. CD19 Facilitates the Pro-B/Pre-B Transition
`Dennis C. Otero and Robert C. Rickert
`
`5931
`
`CD47*/CD25* Regulatory Cells Inhibit Activation of Tumor-Primed CD4* T Cells with IFN-y-
`Dependent Antiangiogenic Activity, as well as Long-Lasting Tumor Immunity Elicited by Peptide
`Vaccination
`Noelia Casares, Laura Arribillaga, Pablo Sarobe, Javier Dotor, Ascensién Lopez-Diaz de Cerio, Ignacio Melero,
`Jestis Prieto, Francisco Borrds-Cuesta, andJuan J. Lasarte
`5940 Dendritic Cells Charged with Apoptotic Tumor Cells Induce Long-Lived Protective CD4* and CD8* T
`Cell Immunity against B16 Melanoma
`Romina S. Goldszmid, Juliana Idoyaga, Alicia I Bravo, Ralph Steinman, José Mordoh, and Rosa Wainstok
`
`\
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`Miltenyi Ex. 1030 Page 5
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`Promiscuity of MHC Class Ib-Restricted T Cell Responses
`Alexander Ploss, Gregoire Lauvau, Brian Contos, Kristen M. Kerksiek, Patrick D. Guirnalda, Ingrid Leiner,
`Laurel L. Lenz, MichaelJ. Bevan, and Erie G. Pamer
`
`Molecular Cloning and Immunologic Characterization of a Novel cDNA Codingfor Progesterone-
`Induced Blocking Factor
`Beata Polgar, Gyula Kispal, Margit Lachmann, Gabriella Paar, Eszter Nagy, Peter Csere, Eva Miko, Laszlo Szereday,
`Peter Varga, andJulia Szekeres-Bartho
`
`Simultaneous Prediction of Binding Capacity for Multiple Molecules of the HLA B44 Supertype
`John Sidney, Scott Southwood, Valerie Pasquetto, andAlessandro Sette .
`
`Altered Regulation of FeyRII on Aged Follicular Dendritic Cells Correlates with Immunoreceptor
`Tyrosine-Based- Inhibition Motif Signaling in B Cells and Reduced Germinal Center Formation
`YiikselAydar, Péter Balogh, John G. Tew, andAndras K. Szakal
`‘
`
`Function of Bruton’s Tyrosine Kinase during B Cell DevelopmentIs Partially IndependentofIts Catalytic
`Activity
`Sabine Middendorp, Gemma M. Dingjan, Alex Maas, Katarina Dahlenborg, and RudolfW. Hendriks
`
`OX40-Mediated Memory T Cell Generation Is TNF Receptor-Associated Factor 2 Dependent
`Rodney A. Prell, Dean E. Evans, Colin Thathofer, Tom Shi, Castle Funatake, andAndrew D, Weinberg
`
`Ikaros Family Membersfrom the Agnathan Myxine glutinosa and the Urochordate Oikopleura dioica:
`Emergence of an Essential Transcription Factor for Adaptive Immunity
`Pauline M. Cupit, John D. Hansen, Aaron S. McCarty, Greg White, Mariacristina Chioda, Fabio Spada,
`Stephen T. Smale, and Charles Cunningham
`‘
`
`Inhibition of NF-«B Activation and Its Target Genes by Heparin-Binding Epidermal Growth Factor-Like
`Growth Factor
`Veela B. Mehta and Gail E. Besner
`
`ia
`
`INFLAMMATION
`Sustained Nitric Oxide Delivery Delays Nitric Oxide-Dependent Apoptosis in Macrophages:
`Contribution to the Physiological Function ofActivated Macrophages
`Sonsoles Hortelano, Paqui G. Través, Miriam Zeini, Alberto M. Alvarez, and Lisardo Boscd
`
`5948
`
`5956
`
`5964
`
`5975
`
`5988
`
`5997
`
`6006
`
`6014
`
`6032
`
`6039
`
`6046
`
`6052
`
`6059
`
`6065
`
`HOSTDEFENSE
`
`6023
`Allogeneic T Cells Treated with Amotosalen Prevent Lethal Cytomegalovirus Disease without Producing
`Graft-versus-Host Disease Following Bone Marrow Transplantation
`John D. Roback, Mohammad S. Hossain, Levan Lezhava, John W. Gorechlad, Sabina A. Alexander, DavidL. Jaye,
`Stephen Mittelstaedt, Sohel Talib, John E. Hearst, Christopher D. Hillyer, and Edmund K. Waller
`
`Activation ofAntigen-Specific CD8 T Cells Results in MinimalKilling of Bystander Bacteria
`Jiu Jiang, Lauren A. Zenewicz, Lani R. San Mateo, Lisa L. Lau, and Hao Shen
`
`NK Cells Respond to Pulmonary Infection with Mycobacterium tuberculosis, but Play a Minimal Role in
`Protection
`Ana PaulaJunqueira-Kipnis, Andre Kipnis, Amanda Jamieson, Mercedes GonzalezJuarrero, Andreas Diefenbach,
`David H. Raulet, Joanne Turner, and Ian M. Orme
`
`Human C-Reactive Protein Does NotProtect against Acute Lipopolysaccharide Challenge in Mice
`Gideon M. Hirschfield, JeffHerbert, Melvyn C. Kahan, and Mark B. Pepys
`
`Cross-Talk in the Innate ImmuneSystem: Neutrophils Instruct Recruitmentand Activation of Dendritic
`Cells during Microbial Infection
`Soumaya Bennouna, Susan K. Bliss, TylerJ. Curiel, and Eric Y. Denkers
`
`IL-1R-Associated Kinase 4 Is Required for Lipopolysaccharide-Induced Activation ofAPC
`Nobutaka Suzuki, Shinobu Suzuki, Urs Eriksson, Hiromitsu Haya, Christine Mirtosis, Nien-Jung Chen, Teiji Wada,
`Denis Bouchard, Irene Hwang, Kiyoshi Takeda, Takashi Fujita, Sandy Der, JosefM. Penninger, Shizuo Akira,
`Takashi Saito, and Wen-Chen Yeh
`
`Miltenyi Ex. 1030 Page 6
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`6080
`
`6090
`
`Cyclooxygenase-2-Derived E Prostaglandins Down-Regulate Matrix Metalloproteinase-1 Expression in
`Fibroblast-Like Synoviocytes via Inhibition of Extracellular Signal-Regulated Kinase Activation
`Michael H.Pillinger, Pamela B. Rosenthal, Sonia N. Tolani, Beth Apsel, Victoria Dinsell, Jeffrey Greenberg,
`Edwin 8, L. Chan, Paul F. Gomez, and Steven B. Abramson
`
`FeyRIIIb Allele-Sensitive Release of a-Defensins: Anti-Neutrophil Cytoplasmic Antibody-Induced
`Release of Chemotaxins
`Sumiaki Tanaka, Jeffrey C. Edberg, Winn Chatham, Giorgio Fassina, and Robert P. Kimberly
`
`6097 Activation of Endothelial Extracellular Signal-Regulated Kinase Is Essential for Neutrophil
`Transmigration: Potential Involvementof a Soluble Neutrophil Factor in Endothelial Activation
`Brian N. Stein, Jennifer R. Gamble, Stuart M. Pitson, Mathew A. Vadas, and Yeesim Khew-Goodall
`
`6105 Mac-1, but Not LFA-1, UsesIntercellular Adhesion Molecule-1 to Mediate Slow Leukocyte Rolling in
`TNF-a-Induced Inflammation
`Jessica L. Dunne, Robert G. Collins, Arthur L. Beaudet, Christie M. Ballantyne, and Klaus Ley
`
`6112 Regulation and Phenotypeof an Innate Th1 Cell: Role of Cytokines and the p38 Kinase Pathway
`JeffreyJ. Yu, Catherine S. Tripp, andJohn H. Russell
`
`6119
`
`6128
`
`6135
`
`6145
`
` FeeRI Signaling of Mast Cells Activates Intracellular Production of Hydrogen Peroxide: Role in the
`Regulation of Calcium Signals
`Yoshihiro Suzuki, Tetsuro Yoshimaru, Takashi Matsui, Toshio Inoue, Osamu Niide, Satoshi Nunomura, and Chisei Ra
`
`Increased Acute Inflammation, Leukotriene B4-Induced Chemotaxis, and Signaling in Mice Deficient for
`G Protein-Coupled Receptor Kinase 6
`Annemieke Kavelaars, Anne Vroon, Roel P. Raatgever, Alan M. Fong, Richard T. Premont, Dhavalkumar D. Patel,
`Robert]. Lefkowitz, and CobiJ. Heijnen
`
`Endothelial Cells Proactively Form Microvilli-Like Membrane Projections upon Intercellular Adhesion
`Molecule 1 Engagementof Leukocyte LFA-1
`.
`Christopher V. Carman, Chang-DukJun, Azucena Salas, and Timothy A. Springer
`
`Toll-Like Receptor 2 Pathway Drives Streptococcal Cell Wall-Induced Joint Inflammation: Critical Role
`of Myeloid Differentiation Factor 88
`Leo A. B. Joosten, Marije I. Koenders, Ruben L. Smeets, Marleen Heuvelmans-Jacobs, Monique M. A. Helsen,
`Kiyoshi Takeda, Shizuo Akira, Erik Lubberts, Fons A. J. van de Loo, and Wim B. van den Berg
`
`6154 Apoptosis-Associated Speck-Like Protein Containing a Caspase Recruitment DomainIs a Regulator of
`Procaspase-1 Activation
`,
`.
`Christian Stehlik, Sug Hyung Lee, Andrea Dorfleutner, Angela Stassinopoulos, Junji Sagara, andJohn C. Reed
`
`6164
`
`Proinflammatory Cytokines Disrupt Epithelial Barrier Function by Apoptosis-Independent Mechanisms
`Matthias Bruewer, Andreas Luegering, Torsten Kucharzik, Charles A. Parkos, James L. Madara, Ann M. Hopkins,
`andAsma Nusrat
`
`6173
`
`Suppression of ImmuneInduction of Collagen-InducedArthritis in IL-17-Deficient Mice
`Susumu Nakae, Aya Nambu, Katsuko Sudo, and Yoichiro Iwakura
`
`6178 Alterations in Granule Matrix and Cell Surface of Focal Adhesion Kinase-Deficient Mast Cells
`Daniel Vial, Constance Oliver, Maria CéliaJamur, Maria Verénica Ddvila Pastor, Edvaldo da Silva Trindade,
`Eka Berenstein, Juan Zhang, and Reuben P. Siraganian
`
`6187
`
`Toll-Like Receptor-2, but Not Toll-Like Receptor-4, Is Essential for Development of Oviduct Pathology
`in Chlamydial Genital Tract Infection
`Toni Darville, Joshua M. O'Neill, Charles W. Andrews, Jr, Uma M. Nagarajan, Lynn Stahl, andDavidM. Ojcius
`6198 Aberrant Inflammation andLethality to Septic Peritonitis in Mice Lacking STAT3 in Macrophages and
`Neutrophils
`Akihiro Matsukawa, Kiyoshi Takeda, Shinji Kudo, Takako Maeda, Motoko Kagayama, and Shizuo Akira
`
`\
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`Miltenyi Ex. 1030 Page 7
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`IVE
`
`6206
`
`Inhibition of Thl- and Th2-Mediated Airway Inflammation by the Sphingosine 1-Phosphate Receptor
`Agonist FT¥720
`Elzbieta Sawicka, Claudia Zuany-Amorim, Corinne Manlius, Alexandre Trifilieff, Volker Brinkmann, DavidM. Kemeny,
`and Christoph Walker
`
`CLINICALIMMUNOLOGY
`
`6215
`Placental Cell Expression of HLA-G2IsoformsIs Limited to the Invasive Trophoblast Phenotype
`Pedro ]. Morales, Judith L. Pace, Jeralyn Sue Platt, Teresa A. Phillips, Kim Morgan, Asgi T. Fazleabas, andJoan S. Hunt
`
`ors
`
`
`
`OEenactsSEN
`
`_AQEUBRARYQe, «AEIBR
`
`Dy__grrOSAncttey
`
`
`6225.
`
`B-1 B Cells Mediate Required Early T Cell Recruitmentto Elicit Protein-Induced Delayed-Type
`Hypersensitivity
`Marian Szczepanik, Moe Akahira-Azuma, KrzysztofBryniarski, Ryohei F. Tsuji, Ivana Kawikova, Wlodzimierz Ptak,
`Claudia Kiener, Regis A. Campos, and Philip W. Askenase
`
`6236 Mechanismsof SpontaneousResolution versus Fibrosis in Granulomatous Experimental Autoimmune
`Thyroiditis
`Kemin Chen, Yongzhong Wei, Gordon C. Sharp, and Helen Braley-Mullen
`
`6244 The Plant Lectin Wheat Germ Agglutinin Inhibits the Binding of PemphigusFoliaceus Autoantibodies to
`Desmoglein 1 in a Majority of Patients and Prevents Pathomechanismsof PemphigusFoliaceus In Vitro
`and In Vivo
`Susana Ortiz-Urda, Adelheid Elbe-Biirger, JosefSmolle, Yvonne Marquart, Yakov Chudnovsky, Todd W. Ridky,
`Pamela Bernstein, Klaus Wolff, and Klemens Rappersberger
`6251 Activity and Safety of CTLA-4 Blockade Combined with Vaccines in Cynomolgus Macaques
`Tibor Keler, EdHalk, Laura Vitale, Tom O’Neill, Diann Blanset, Steven Lee, Mohan Srinivasan, Robert F. Graziano,
`Thomas Davis, Nils Lonberg, andAlan Korman
`6260 Chromatin Specificity ofAnti-Double-Stranded DNA Antibodies and a Role for Arg Residuesin the
`Third Complementarity-Determining Region of the Heavy Chain
`Amanda M. Guth, Xianghua Zhang, Diana Smith, Thiago Detanico, and Lawrence]. Wysocki
`
`,
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`‘
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`‘
`
`6267
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`6275
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`6283
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`Immune Modulation in Multiple Sclerosis Patients Treated with the Pregnancy HormoneEstriol
`Samantha S. Soldan, Ana IsabelAlvarez Retuerto, Nancy L. Sicotte, and Rhonda R. Voskubl
`
`Injection of Immature Dendritic Cells into Adjuvant-Treated Skin Obviates the Need for Ex Vivo
`Maturation
`Smita Nair, Catherine McLaughlin, Alon Weizer, Zhen Su, David Boczkowski, Jens Dannull, Johannes Vieweg,
`and Eli Gilboa
`
`Identification of Five New HLA-B3501-Restricted Epitopes Derived from Common Melanoma-
`Associated Antigens, Spontaneously Recognized by Tumor-Infiltrating Lymphocytes
`Houssem Benlalam, Boris Linard, Yannik Guilloux, Agnes Moreau-Aubry, Laurent Derré, Elisabeth Diez, Brigitte Dreno,
`FrancineJotereau, and Nathalie Labarriére
`
`6290 Differential Regulation of Peripheral CD4* T Cell Tolerance Induced by Deletion and TCR Revision
`MohamedAli, Michael Weinreich, Stephanie Balcaitis, CristineJ: Cooper, and PamelaJ. Fink
`
`6297 Corrections
`
`6298 Author Index
`
`6301 AAI Membership Application
`
`6302 AAI Trainee Membership Application
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`Miltenyi Ex. 1030 Page 8
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`Miltenyi Ex. 1030 Page 8
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`The Journal of Immunology
`
`IL-2 Is Not Required for the Initiation of CD8 T Cell Cycling
`but Sustains Expansion?
`
`Warren N. D’Souza and Leo Lefrancois”
`
`Basedprimarily onin vitro data, IL-2 is believed to be the key cytokineforinitiation of thecell cycle of activated T cells. However,
`the role of IL-2 remains unresolved for T cell responses in vivo. We examined whetherthe absence of IL-2-mediated signaling in
`CD8T cells affectedinitiation of proliferation. Ourresults conclusively demonstrated thatinitial division ofAg-specific CD8T cells
`following priming was IL-2 independent, regardless of‘the context in which Ag was presented.In contrast, the latter stage of the
`proliferative phase was IL-2-dependent, particularly in nonlymphoidtissues. Thus, activated CD8,T cellsinitially undergo IL-2-
`independentproliferation, but reach a critical juncture where the requirementfor IL-2 as a growth factor gains prominence. The
`Journal of Immunology, 2003, 171: 5727-5735.
`
`
`
`pon stimulation with cognate Ag, naive T cells are ac-
`activated CD8T cells is indispensable for the cell cycle progres-
`tivated and typically undergo a responsethat consists of
`sion of recently activated-T cells.
`three phases, namely, an expansion phase, a contraction
`In light of the central role that IL-2 is thought to play in gov-
`phase, and a memory phase. An augmented expansion phase ap-
`erning T cell responses,it is quite intriguing that not only is there
`pears to lead to an increase in the size of the memory pool gen-
`a lack of definitive data in support of IL-2 being the quintessential
`erated and, as a result, much research is aimed at potentiating the
`T cell growth factorin vivo, but evidencealso exists opposingthis
`priming of T cells. Efficient priming is thought to be achieved by
`notion. For example, mice with targeted disruptions of the IL-2 or
`optimizing costimulation and providing the appropriate cytokine
`IL-2R genes are not immunodeficient (as would have been pre-
`milieu to naive T cells during activation. The expansion phaseis
`dicted), but in fact display a severe lymphoproliferative disorder
`believed to be driven primarily by cytokines and IL-2 is one such
`(13-16). Also, since the induction of anergy is thought to be due
`prototypical cytokine that was originally identified based on its
`to a failure of T cells to prodice IL-2, the demonstration that
`potent growth-promotingability for T cells in vitro (1). IL-2 andits
`proliferation usually precedes anergy in vivo (17-19) argues that
`high-affinity receptor (IL-2Rafyc) were shown to be expressed
`the production of IL-2: is not required for cell cycle initiation.
`rapidly following activation in vitro and the absence ofeither IL-2
`Experimentation with mice that lack IL-2 or IL-2R components
`or IL-2R was thoughtto result in a “failure? of the immunere-
`has yielded variable results, with some studies indicating no sig-
`sponse (2, 3), Also, one of the most important costimulatory mol-
`nificant requirement for IL-2 in the expansion of CD8Tcells in
`ecules, CD28,is believed to function for the most part by increas-
`vivo (20-22) while other studies pointto an essential role for IL-2
`ing the production of IL-2 by T cells (4~7). Consequently, anergy
`(23, 24). We also observed a minimal requirement for IL-2 in
`or unresponsiveness that results from TCR occupancy in the ab-
`mounting an antiviral CD8 T cell response within secondary lym-
`sence of costimulatory signals is assumed to be dueto the inability
`phoid tissue (25).
`of the cells to produce IL-2 andthe addition of exogenousIL-2 is
`Thus, although it is widely accepted that IL-2 plays a pivotal
`capable of reversing the unresponsive phenotype(8, 9). It is well
`role in the early events following CD8Tcell activation,it is ap-
`documentedthat the generation of certain CD8 T cell responsesis
`parent that the requirement for IL-2 in the generation of CD8 T.cell
`dependent upon the presence of CD4Tcell help and oneof the
`responses in vivo remains equivocal. More importantly, it is un-
`postulated mechanismsofhelp is the provision of IL-2 to the CD8
`clear whether the requirement for IL-2 observed in some model
`T cells (10). Also, recent data implicate IL-2 production by den-
`systems is due to an alteration in the entry and kinetics of CD8 T
`dritic cells as an essential factor for the initiation of both CD4 and
`cell cycling (impaired priming) or dueto lack of sustained prolif-
`CD8 T cell responses (11). In addition, it is now apparent that
`eration. We sought to examinethis issue directly by visualizing
`regulatory T cells are key players in the immune system andthese
`early cell division subsequentto activation in vivo and determinin2
`Tk cells are thought to suppress T cell proliferation by inhibiting
`what effect the absence of IL-2 or IL-2R signaling had on the
`IL-2 production by responding T cells (12). Thus, current dogma
`initiation and rate of CD8 T cell cycling. To this end, we tracked
`avers that the expression of IL-2 and the high-affinity IL-2R by
`Ag-specific CD8 T cells throughoutthe proliferative phase in re-
`eee
`sponse to OVA whenexpressed either as a viral Ag, soluble Ag,
`tumor Ag,or self-Ag. Our results provided compelling evidence
`that the latter half of the proliferative phase was dependent on
`IL-2, but the early division of Ag-specific CD8T cells subsequent
`to priming with cognate Ag was, even in the absence of overt
`inflammation, IL-2 independent.
`
`Department of Medicine, Division of Immunology, University of Connecticut Health
`Center, Farmington, CT 06030
`
`Received for publication June 20, 2003. Accepted for publication September
`24, 2003.
`
`Thecosts of publication ofthis article were defrayed in part by the payment of page
`charges. This article must therefore be hereby marked advertisement in accordance
`with 18 U.S.C. Section 1734 solely to indicate this fact.
`'This work was supported by National Institutes of Health Grants DK57932,
`DK45260, and Al41576.
`
`? Address correspondence and reprint requests to Dr. Leo Lefrancois, Division of Immu-
`nology, University of Connecticut Health Center, MC1319, Room L3072, 263 Farming-
`ton Avenue, Farmington, CT 06030-1319. E-mail address: llefranc@neuron.uchc.edu
`
`Copyright © 2003 by The American Association of Immunologists,Inc.
`
`Materials and Methods
`Mice
`
`C57BL/6], C57BL/6-IL-2~’~, and C57BL/6-CD25~’~ mice were obtained
`from The Jackson Laboratory (Bar Harbor, ME). C57BL/6-Ly5.2 mice
`were obtained from Charles River Breeding Laboratories (Wilmington,
`\
`
`0022-1767/03/$02.00
`
`Miltenyi Ex. 1030 Page 9
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`Miltenyi Ex. 1030 Page 9
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`5728
`
`IL-2 DOES NOTINITIATE CD8 T CELL CYCLING
`
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`Results
`MA)through the National CancerInstitute. The OT-I mouse line (26) was
`generously provided by W. R. Heath (Walter and Eliza Hall Institute,
`IL-2 and IL-2Ra are expressed before cell division on activated
`Parkville, Australia) and F. Carbone (Monash Medical School, Prahran,
`antiviral CD8Tcells in vivo
`Victoria, Australia). We also generated GT-I-CD25~"“RAG~ mice. and
`S. Schoenberger (La Jolla Institute of Immunology, La Jolla, @A) kindly
`T cells are known to express IL-2 and the high-affinity IL-2R
`provided us with OT-LIL-2~‘"RAG~’" mice that were used in some ex-
`(which includes ,IL-2Ra) immediately following activation in
`periments. Unless otherwise indicated, the cells used in the experiments
`were obtained ‘from RAGT’+ OT-I mice (either IL-27’~, CD25~", oO
`vitro. However, since recent in vivo data suggest cell division may
`precede the expression of IL-2 and JL-2Ra (35), we analyzed the
`normal). The presence of the OT-I transgene and the RAG mutation was
`detected by assessing the frequency of Va2*VB5*CD8°*cells and B220*
`kinetics of expression of IL-2 and the IL-2Ra subunit (CD25) by
`cells, respectively, in PBL. 232-4 mice expressing cytoplasmic OVA under
`CD8T cells followingaviral infection. We used an adoptive trans-
`control of the intestinal fatty acid-binding protein promoter have been pre-
`fer system in which trackable OVA-specific Ly5.1 OT-I CD8 -T
`viously described (27). OT-I-mice were mated to IL-2*/~ mice. Offspring
`celis (26) were CFSE-labeled and transferred to B6 (Ly5.2*) mice
`were screened for the IL-2 mutation by, PCR. OT-LIL-2*’~ mice were
`that were either left unimmunizedor immunized a daylater with a
`intercrossed to obtain QTI-IL-2~’~ animals and OTI-CD25~/~ animals
`weré generated in a similar’manner.
`recombinant VSV- encoding OVA (VSV-OVA). Mice were sac-
`rificed at early time points postinfection and splenic donorcells
`Isolation of lymphocyte populations
`were analyzed for the expression of CFSE, cell surface CD25, and
`intracellular IL-2 directly ex vivo (without restimulation in vitro).
`Intraepithelial lymphocytes (IEL)* and laminapropria cells from the small
`At 12hpostinfection? OT-I cells had not yet divided but were
`intestine were isolated as previously described (28, 29). Spleens and lymph
`already activated, as indicated by an increase in forward light scat-
`nodes (LN),were removed andsingle-cell suspensions were prepared using
`a tissue homogenizer. The resulting preparation wasfiltered through Nitex
`ter (Fig. 1B, inset). All of the OT-I cells also expressed high
`levels
`nylon mesh (Tetko, Kansas City, MO) andthefiltrate was centrifuged to
`of CD25 (Fig. 1B, top panels) and approximately one-fifth of them
`pellet the cells. To obtain lymphocytes from lungs, anesthetized mice were
`produced TL-2 diréctly ex vivo (Fig. 1A). By as early as 58 h
`perfused with PBS containing 75 U/ml heparin until the tissue was cleared
`postinfection, we were unable to detect any ex vivo IL-2 produc-
`of blood, andthe organs were removed and cells were isolated as previ-
`tion, but the,cells were capable of producing the cytokine after a
`ously described (30).
`5-h in vitro restimulation with peptide (data not shown). At 58 h,
`CD25 wasstill expressed at high levels on the dividing cells (Fig.
`
`Flow cytometric analysis
`
`Lymphocytes were resuspended in PBS, 0.2% BSA,and 0.1% NaN,(PBS,
`BSA, NaN,)at a con