Regulatory B Cells: Methods and Protocols

289 Pages·2014·6.008 MB·English

by  Gaetano Vitale, Francesca Mion (eds.)

Methods in
Molecular Biology 1190
Gaetano Vitale
Francesca Mion Editors
Regulatory
B Cells
Methods and Protocols
M M B
ETHODS IN OLECULAR IOLOGY
Series Editor
John M. Walker
School of Life Sciences
University of Hertfordshire
Hat fi eld, Hertfordshire, AL10 9AB, UK
For further volumes:
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Regulatory B Cells
Methods and Protocols
Edited by
Gaetano V itale and Francesca Mion

Department of Medical and Biological Sciences, University of Udine, Udine, Italy
Editors
Gaetano Vitale Francesca Mion
Department of Medical and Biological Sciences Department of Medical and Biological Sciences
University of Udine University of Udine
Udine, Italy Udine, Italy
ISSN 1064-3745 ISSN 1940-6029 (electronic)
ISBN 978-1-4939-1160-8 ISBN 978-1-4939-1161-5 (eBook)
DOI 10.1007/978-1-4939-1161-5
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Pre face
The regulation of the immune responses is a rapidly growing and fascinating experimental
fi eld that in the last two decades has attracted the interest of an increasing number of labo-
ratories. An emerging concept in the fi eld of immune regulation is that immune homeostasis
is maintained as a result of the cooperation of many cell types that have both effector and
regulatory functions. Both innate and adaptive immune recognition mechanisms contribute
to the optimal induction of multiple cell types that interact to produce effector and regula-
tory functions. Immune regulation is obtained by tolerance versus self and harmless antigens
and by control of immune response amplifi cation. Tolerance prevents lymphocyte activation
whereas control restrains the activity of effector cells. Tolerance and control operate by
means of cell-intrinsic and cell-extrinsic mechanisms. In particular, cell-extrinsic immune
regulation depends upon the activity of several types of immune-suppressive cells often
indicated as regulatory cells.
B cells are new entrees in the fi eld of immune regulation despite evidence of their role
in immune suppression dating back in the 1970s. Nevertheless, numerous recent studies
have established their essential role in the control of immune-mediated infl ammation and
tolerance. The regulatory function of B cells has been related to their ability to produce the
anti-infl ammatory cytokine IL-10. IL-10 secretion is not the only regulatory mechanism
operated by B cells: regulatory B cells (Breg) are able to induce regulatory T cells and other
suppressive cell types both by contact-mediated interactions and by the production of
secreted factors. Many populations of B cells with different phenotypic markers and devel-
opmental origin have been recognized to exert a regulatory suppressive function in several
immunological settings, including infections, infl ammation, and autoimmunity.
The investigation of the mechanisms of immune regulation requires many techniques
applied to different cell types. Also, there is an emerging interest in the regulatory role of B
cells in many different pathological settings, both in the mouse and in humans. The aim of
this book is to present to immunologists as well as cell and molecular biologists interested in
B cell biology the current experimental setups and methodologies useful for the identifi cation
and characterization of B cells with suppressive functions and for the study of their biological
and immunological properties. The book should be of interest to many labs that focus on the
study of the regulatory function of B cells as well as to those labs that would like to study the
possible function of this cell type in the pathophysiology of a given disease.
The book is organized into four parts: the fi rst part contains the basic methods for the
isolation and immunophenotypical characterization of specifi c B cell subsets from mouse
and human tissues; the second part presents methods useful for the investigation of the
mechanisms of immune suppression operated by B cells; the third part presents several
experimental approaches for the ex vivo generation/expansion of IL-10-producing B cells;
and fi nally, the fourth part presents procedures useful for the study of the immune suppres-
sive function of B cells in specifi c pathological settings.
Udine, Italy Gaetano V itale
Francesca M ion
v

Contents
Preface. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . v
Contributors. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . i x
PART I PURIFICATION AND PHENOTYPIC CHARACTERIZATION
OF B CELL SUBSETS
1 Purification and Immunophenotypic Characterization
of Murine MZ and T2-MZP Cells. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3
M. M anuela R osado, M arco S carsella, S imona C ascioli,
Ezio Giorda, and Rita C arsetti
2 P urification and Immune Phenotyping of B-1 Cells from Body
Cavities of Mice . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 7
Vanessa Y enson and Nicole B aumgarth
3 P urification and Immunophenotypic Characterization
of Murine B10 B Cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 5
Chao Hong and X iao-Ming Gao
4 Purification and Immunophenotypic Characterization
of Human B Cells with Regulatory Functions. . . . . . . . . . . . . . . . . . . . . . . . . 45
Adèle de M asson, H élène Le B uanec, and J ean-David B ouaziz
PART II MECHANISMS OF IMMUNE SUPPRESSION BY B CELLS
5 IL-10 Detection in Murine B Cells: Pros and Cons
of the Different Techniques. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 55
Xiang L in, Jun D eng, and Liwei Lu
6 T GF-β Detection and Measurement in Murine B Cells:
Pros and Cons of the Different Techniques. . . . . . . . . . . . . . . . . . . . . . . . . . . 7 1
Yoshiyuki M ishima, S hunji I shihara, J onathan J. H ansen,
and Y oshikazu Kinoshita
7 Characterization and Activity of Fas Ligand Producing CD5+ B Cells . . . . . . . 8 1
Steven K. Lundy and Matthew W . K linker
PART III METHODS FOR THE EX VIVO GENERATION/EXPANSION
OF IL-10- PRODUCING B CELLS
8 Utilization of a Lentiviral System for the Generation of B Cells
with Regulatory Properties . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 05
Elisabeth C alderón-Gómez and Simon F illatreau
9 E x Vivo Generation of Murine IL-10-Producing B Cells by Fusokines. . . . . . . 1 15
Aurélie T ormo, J iusheng D eng, E douard Al-Chami,
Sonia Ziouani, and Moutih R afei
vii
viii Contents
10 Toll-Like Receptor Ligation for the Induction of Regulatory B Cells. . . . . . . . 1 27
Luciën E . P. M . v an der Vlugt, S imone H aeberlein,
Wouter de Graaf, T yrill E. D . M artha, and H ermelijn H. Smits
11 T he Generation of Regulatory B Cells by Helminth Parasites . . . . . . . . . . . . . 1 43
Adnan R. Khan, S ylvie A mu, S ean P . S aunders, and Padraic G . Fallon
12 U se of Cocultures for the Study of Cellular Interactions
Influencing B Cell Regulatory Functions. . . . . . . . . . . . . . . . . . . . . . . . . . . . . 163
Francesca Mion, F ederica B envenuti, C arlo P ucillo,
and Gaetano V itale
PART IV STUDY OF REGULATORY B CELLS IN PATHOLOGICAL SETTINGS
13 R egulatory B Cells in Experimental Mouse Models of Arthritis. . . . . . . . . . . . 1 83
Elizabeth C . Rosser and Claudia M auri
14 Regulatory B Cells in Mouse Models of Systemic Lupus
Erythematosus (SLE) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 95
Takashi M atsushita
15 Regulatory B Cells in Allergic Airways Disease and Asthma. . . . . . . . . . . . . . . 2 07
Prabitha Natarajan, L inda A. G uernsey, and C raig M. Schramm
16 Regulatory B Cells in Mouse Models of Intestinal Inflammation. . . . . . . . . . . 2 27
Atsushi N ishida, C indy W . L au, E miko M izoguchi,
and A tsushi M izoguchi
17 R egulatory B Cells in Experimental Autoimmune
Encephalomyelitis (EAE) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 43
Avijit Ray and Sreemanti B asu
18 Regulatory B Cells, Helminths, and Multiple Sclerosis. . . . . . . . . . . . . . . . . . . 2 57
Jorge Correale and Tomas Rivero E quiza
19 G eneration and Identification of Tumor-Evoked Regulatory B Cells . . . . . . . . 2 71
Arya B iragyn, C atalina Lee-Chang, and M onica Bodogai
Index . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 91
Contributors
EDOUARD AL-CHAMI • The Department of Pharmacology , Université de Montréal ,
Montreal , Q C , Canada
SYLVIE A MU • Institute of Molecular Medicine, St James’s Hospital , D ublin, I reland
SREEMANTI B ASU • BloodCenter of Wisconsin, Blood Research Institute , Milwaukee, WI , U SA
NICOLE B AUMGARTH • Center for Comparative Medicine, Schools of Medicine
and Veterinary Medicine, University of California , Davis, CA , U SA ; D epartment of
Pathology, Microbiology, and Immunology, School of Veterinary Medicine ,
University of California , Davis, CA , U SA
FEDERICA BENVENUTI • International Centre for Genetic Engineering and Biotechnology ,
Trieste, I taly
ARYA B IRAGYN • Immunoregulation Section, N ational Institute on Aging, National
Institutes of Health , B altimore, MD, USA; Laboratory of Molecular Biology and
Immunology, Biomedical Research Center, National Institute on Aging, National
Institutes of Health, Baltimore, MD, USA
MONICA BODOGAI • Immunoregulation Section, Laboratory of Molecular Biology
and Immunology , National Institute on Aging, National Institutes of Health , B altimore ,
MD, USA
JEAN-DAVID BOUAZIZ • INSERM U976 , Paris, F rance ; U niversité Paris Diderot, Sorbonne
Paris Cité , Paris, France ; L aboratory of Immunology, Dermatology & Oncology,
UMR-S 976 , P aris, France ; Service de Dermatologie, Hôpital Saint Louis , Paris, F rance
HÉLÈNE LE BUANEC • INSERM U976 , P aris , F rance ; U niversité Paris Diderot, Sorbonne
Paris Cité , Paris, France ; L aboratory of Immunology, Dermatology & Oncology,
UMR-S 976 , P aris, France ; Service de Dermatologie, Hôpital Saint Louis , Paris, F rance
ELISABETH CALDERÓN-GÓMEZ • Institut d’Investigacions Biomèdiques August Pi i Sunyer
(IDIBAPS) , B arcelona, S pain
RITA CARSETTI • Immunology Unit, Research Center Ospedale Pediatrico Bambino Gesù,
IRCSS, Rome, Italy; Clinical Laboratories, Research Center Ospedale Pediatrico
Bambino Gesù, IRCSS, Rome Italy
SIMONA CASCIOLI • Immunology Unit, Research Center Ospedale Pediatrico Bambino Gesù,
IRCSS , R ome , I taly
JORGE CORREALE • Department of Neurology, Institute for Neurological Research
Dr. Raúl Carrea, FLENI , B uenos Aires, Argentina
JUN DENG • Department of Pathology, T he University of Hong Kong , H ong Kong, Hong Kong
JIUSHENG DENG • Department of Hematology and Medical Oncology, Winship Cancer
Institute, Emory University , Atlanta , G A , USA
TOMAS R IVERO EQUIZA • Department of Neurology, I nstitute for Neurological Research
Dr. Raúl Carrea, FLENI , B uenos Aires, Argentina
PADRAIC G . F ALLON • School of Medicine, Trinity Biomedical Sciences Institute,
Trinity College Dublin , Dublin, I reland ; I nstitute of Molecular Medicine,
St James’s Hospital , Dublin, I reland; National Children’s Research Centre,
Our Lady’s Children’s Hospital, Crumlin, Dublin, Ireland
ix