ectopic lymphoid-like structures in infection, cancer and ......jun 20, 2014 · infection, cancer...

Inflammation supports immunological defence against infection, trauma, injury and cancer. The regulation of inflammation is governed by cellular communication between non-haematopoietic stromal cells, resident leukocytes and infiltrating immune cells1,2. Optimal control of this process ensures competent host defence, the elimination of the initiating antigen (or antigens) and limited tissue damage1,3. However, repeated, persistent or non-resolving episodes of inflammation lead to the inappropriate regulation of this response and promote autoimmunity, chronicity and tissue pathology1,3,4. The mechanisms that control these events are often diverse and they contribute to clinical differences in disease presentation. For example, patients who have the same underlying clinical condition invariably show differences in disease severity, rate of disease onset and response to treatment4. Although sex, age, genetics, metabolic fac-tors and the environment are major determinants that affect the propensity to develop chronic disease, these indicators provide minimal information on the molecu-lar or cellular pathways that drive the type of disease that is observed.

Cytokines (including interleukins, interferons and growth factors), chemokines, lipid mediators and innate-sensing mechanisms control inflammation and they have important roles in the pathology of various inflamma-tory diseases and cancer3–5, thus representing major targets for clinical intervention. For example, biological therapies that inhibit pro-inflammatory cytokines — for example, the inhibition of tumour necrosis factor (TNF) by the monoclonal antibodies infliximab, adalimumab,

golimumab and certolizumab — or their receptors (for example, the inhibition of interleukin-6 (IL-6) receptor by tocilizumab) are now widely used in routine clini-cal practice for the treatment of rheumatoid arthritis3–5. These therapies display specific modes of action and have been developed on the basis of a better understanding of cytokine involvement in inflammation. Although these agents reduce the symptoms and progression of disease, not all patients respond equally to the same therapy. Indeed, 30–40% of patients with rheumatoid arthritis do not respond to any treatments and some biological thera-pies are efficacious in certain clinical conditions but not in others4. Such differences in therapeutic efficacy sug-gest that distinct inflammatory mechanisms must steer the course of disease in any one individual. Typically, the sooner after diagnosis appropriate therapy commences, the better the clinical outcome and the increased likeli-hood of achieving remission6. Thus, cell ular events that are triggered early in the inflammatory process must influence the course of disease.

A central feature of local inflammation is the inter-action between the stromal tissue compartment and immune cells1,2. Inflammatory mediators that are pro-duced by stromal cells and tissue-resident monocytic cells control the recruitment, activation and survival of leukocytes. Cellular infiltration is traditionally viewed as the diffuse influx of immune cells, with the cells being scat-tered throughout the inflamed tissue. However, infiltrating leuko cytes often form more organized aggregates and these promote antigen-specific adaptive immune responses that exacerbate chronic disease. Indeed, B cells, T cells, resident

1Centre for Experimental Medicine and Rheumatology, William Harvey Research Institute, Barts and The London, School of Medicine and Dentistry, Queen Mary University of London, Charterhouse Square, London EC1M 6BQ, UK.2Cardiff Institute for Infection and Immunity, The School of Medicine, Cardiff University, The Tenovus Building, Heath Campus, Cardiff CF14 4XN, Wales, UK.Correspondence to S.A.J. e‑mail: [email protected]:10.1038/nri3700Published online 20 June 2014

Ectopic lymphoid-like structures in infection, cancer and autoimmunityCostantino Pitzalis1, Gareth W. Jones2, Michele Bombardieri1 and Simon A. Jones2

Abstract | Ectopic lymphoid-like structures often develop at sites of inflammation where they influence the course of infection, autoimmune disease, cancer and transplant rejection. These lymphoid aggregates range from tight clusters of B cells and T cells to highly organized structures that comprise functional germinal centres. Although the mechanisms governing ectopic lymphoid neogenesis in human pathology remain poorly defined, the presence of ectopic lymphoid-like structures within inflamed tissues has been linked to both protective and deleterious outcomes in patients. In this Review, we discuss investigations in both experimental model systems and patient cohorts to provide a perspective on the formation and functions of ectopic lymphoid-like structures in human pathology, with particular reference to the clinical implications and the potential for therapeutic targeting.

R E V I E W S

NATURE REVIEWS | IMMUNOLOGY VOLUME 14 | JULY 2014 | 447

© 2014 Macmillan Publishers Limited. All rights reserved

mailto:JonesSA%40cf.ac.uk?subject=

Table 1 | Clinical and experimental conditions that feature ELSs

Condition Location Species Antigen recognized by cells in ELSs

Other known functions of ELSs in disease

Refs

Autoimmune disease

Rheumatoid arthritis Synovial tissue •Human •Mouse

•Rheumatoid factor •Citrullinated proteins •Histones?

•In situ B cell differentiation•Ongoing CSR •Autoantibody secretion

25,67,156

Sjögren’s syndrome •Salivary glands•Lacrimal glands

•Human•Mouse

•SSA/Ro •SSB/La

Unknown 39,73,74,157

Multiple sclerosis (and EAE)

CNS •Human•Mouse

Myelin and other neuronal antigens proposed

Unknown 29,158

Diabetes Pancreatic islet parenchyma

Mouse Plasma cell anti-insulin reactivity observed

Unknown 159,160

Hashimoto’s thyroiditis Thyroid gland Human •Thyroglobulin •Thyroperoxidase

Unknown 85

Primary sclerosing cholangitis and primary biliary cirrhosis

Liver Human Unknown •Cellular organization •CCL21 and MADCAM1 expression

161–163

Myasthenia gravis Thymus Human Nicotinic acetylcholine receptor

Unknown 94,164

SLE Tubulointerstitium of the kidneys

•Human •Mouse

Immunoglobulin repertoire analysis reveals B cell clonal expansion and ongoing somatic hypermutation

130,165

Chronic inflammation

Atherosclerosis Aorta •Human•Mouse

Unknown •Cellular organization•LTβR, CXCL13 and CCL21 expression

166,167

Inflammatory bowel disease

Gut •Human •Mouse

Unknown •Cellular organization•CCL19 and CCL21 expression

168–172

COPD Lung •Human•Mouse

Unknown •Cellular organization •Role for LTα, CXCL13, CCL19 and pDCs

173–175

Cancer

Primary cancers •Lung •Breast •Colon

•Human •Mouse

TAA In situ antigen-driven B cell proliferation, somatic hypermutation and affinity maturation

10,40, 99–103

Germ cell Ovarian cancer Human Unknown None reported 103,176–178

Infection

Influenza virus Lung (iBALT) Mouse Unknown •Germinal centre reactions •Class-switched plasma cells •Antiviral immunoglobulin generated

30,59,79

HCV Liver Human Unknown •B cell clonal expansion •Elevated CXCL13 expression

systemically and locally

120,121

MCMV Salivary gland Mouse Unknown •Local AID expression •Ongoing CSR and somatic

hypermutation •CXCL13 and BLIMP1 expression

74

Vaccinia virus Ankara Lung (iBALT) Mouse Unknown Local priming of T cell responses 60

Intestinal microbiota Intestine •Human•Mouse

Unknown •Development is LTi cell independent •Presence of activated mucosal

γδ T cells

33,179

Helicobacter pylori Gastric mucosa •Human•Mouse

Unknown •CXCR5–CXCL13 dependent •Local T cell priming and support of

TH17 cells

•IgG and IgA immune response

76,180

Helicobacter hepaticus Liver Mouse Unknown CCL21 and CXCL13 expression 181

Borrelia burgdorferi (which causes chronic Lyme arthritis)

Synovium Human Unknown •Immunoglobulin V regions show evidence of CSR

•Antigen-driven selection

182

R E V I E W S

448 | JULY 2014 | VOLUME 14 www.nature.com/reviews/immunol


Secondary lymphoid organs(SLOs). In contrast to primary (central) lymphoid organs, where lymphocytes are generated from immature progenitor cells, SLOs — that is, the spleen, Peyer’s patches and lymph nodes — maintain mature naive lymphocytes and are sites of lymphocyte activation by antigen.

Lymphoid neogenesisThe de novo development and cellular organization of lymphoid tissue into distinct anatomical and functional compartments.

monocytic cells, macrophages and dendritic cells can form discrete clusters within the inflamed tissue. These regions can exist as simple lymphoid aggregates or as more sophis-ticated structures that histologically resemble secondary lymphoid organs (SLOs)7,8 (TABLE 1). The spatial organiza-tion of leukocytes into defined, compartmentalized B cell-rich and T cell-rich zones is termed lymphoid neogenesis. These structures direct various B cell and T cell responses, including the induction of effector functions, antibody generation, affinity maturation, class switching and clonal expansion. As a consequence, they are referred to as ectopic lymphoid-like structures (ELSs) or tertiary lymphoid organs (TLOs)7,8. In certain autoimmune conditions, patients who have ELSs in the inflamed tissue often respond poorly to standard biological therapy and thus remain a challeng-ing treatment group9. However, in cancer (for example, solid tumours such as colorectal carcinoma) the presence of tumour-associated ELSs correlates with a better prog-nosis and they may coordinate endogenous antitumour immune responses that improve patient survival10.

In this Review, we explore the molecular and genetic basis for the formation of ELSs during infection, inflam-mation, autoimmune disease and cancer, and we discuss the clinical significance of ELSs in terms of prognosis and response to current biological therapies. With insight from both experimental animal models and human disease settings, we also consider the current state of ELS-directed therapies and the rationale for tar-geting alternative molecular pathways that are associated with ELS formation.

ELS development and functionAlthough ELSs display an architecture that resembles the follicular compartments that are typically seen in SLOs (BOX 1), their organization ranges from simple aggregates of B cells and T cells through to highly ordered structures. Much of our understanding of ELS formation originates from findings that describe the generation of encapsulated SLOs (as reviewed in REF. 11). Despite structural differ-ences between SLOs and ELSs, many of the mechanisms

Condition Location Species Antigen recognized by cells in ELSs

Other known functions of ELSs in disease

Refs

Infection (cont.)

Mycobacterium tuberculosis

Lung (iBALT) •Human•Mouse

Unknown •Priming of antigen-specific TH1 cells

•CCL19 and CCL21 expression•Accumulation of CXCR5+CD4+

TFH

-like cells

38,75, 78,118

Repeated intranasal LPS challenge

Lung (iBALT) Mouse (neonatal)

Unknown •LTi cell independent •IL-17 expression (potential link to

TH17 cells)

•CXCL13 expression

30

Renal failure and transplantation

Allograft transplants For example, kidney, lungs and heart

•Human•Mouse

Unknown Germinal centre reactions resulting in anti-HLA-producing plasma cells and memory B cells

95,96

Peritoneal dialysis Peritoneal omental milky spots

•Human•Mouse

Unknown •Plasma cell responses to T cell-dependent antigen delivered intraperitoneally

•Dependent on LT and CXCL13

183–185

Environmental, degenerative and idiopathic conditions

Cigarette smoke Lung Mouse Unknown •Dependent on LTαβ and LTβR •CXCL13 and CCL19 expression

173

Diesel exhaust particles Lung (iBALT) Mouse Unknown Unknown 186

Metal prosthetic joints Joint (periprosthetic soft tissue)

Human Unknown Formation may involve cytotoxic and/or delayed hypersensitivity response to cobalt-chrome wear particles

187

Pristane adjuvant Peritoneal membrane

Mouse Unknown Lipogranuloma formation with homeostatic chemokine expression

165

IPAH Perivascular lung tissue

Human Autoantibodies against vascular wall components generated

•Expression of IL-7, LTαβ, CCL19, CCL20, CCL21 and CXCL13

•IL-21+ TFH

cells and FDCs •AID expression suggests ongoing CSR

188

AID, activation-induced cytidine deaminase; BLIMP1, B lymphocyte-induced maturation protein 1; CCL, CC-chemokine ligand; CNS, central nervous system; COPD, chronic obstructive pulmonary disease; CSR, class-switch recombination; CXCL13, CXC-chemokine ligand 13; CXCR5, CXC-chemokine receptor 5; EAE, experimental autoimmune encephalomyelitis; ELS, ectopic lymphoid structure; FDC, follicular dendritic cell; HCV, hepatitis C virus; iBALT, inducible bronchus-associated lymphoid tissue; IL, interleukin; IPAH, idiopathic pulmonary hypertension; LPS, lipopolysaccharide; LTi cell, lymphoid tissue inducer cell; LT, lymphotoxin; LTβR, lymphotoxin-β receptor; MADCAM1, mucosal vascular addressin cell adhesion molecule 1; MCMV, murine cytomegalovirus; pDC, plasmacytoid dendritic cell; SLE, systemic lupus erythematosus; SSA/Ro, Sjögren’s syndrome antigen A (ribonucleoprotein autoantigen); SSB/La, Sjögren’s syndrome antigen B (autoantigen La); TAA, tumour-associated antigen; T

FH cells, T follicular helper cells; T

H cell, T helper cell.

Table 1 (cont.) | Clinical and experimental conditions that feature ELSs

R E V I E W S



Ectopic lymphoid-like structures(ELSs). Highly organized lymphoid aggregates that form in tissue sites that are not typically associated with lymphoid neogenesis.

Encapsulated SLOsOrganized secondary lymphoid organs (SLOs) that are encased in a connective tissue capsule that contains blood vessels.

Lymphoid tissue inducer cells (LTi cells). These cells are present in developing lymph nodes, Peyer’s patches and nasopharynx-associated lymphoid tissue (NALT) and they are required for the development of these lymphoid organs. The inductive capacity of LTi cells for the generation of Peyer’s patches and NALT has been shown by adoptive transfer and it is generally assumed that they have a similar function in the formation of lymph nodes.

T follicular helper cells(TFH cells). Antigen-experienced CD4+ T cells that are present in B cell-rich regions of structurally organized lymphoid aggregates or organs.

Lymphoid tissue organizer cellsThese cells are of mesenchymal origin that are activated by lymphoid cells through lymphotoxin-β receptor signalling to express adhesion molecules and chemokines that regulate lymphoid tissue development.

High endothelial venules(HEVs). These are specialized venules that occur in secondary lymphoid organs, except the spleen. HEVs allow the continuous transmigration of lymphocytes as a consequence of the constitutive expression of adhesion molecules and chemokines at their luminal surface.

that control the initial development, cellular composi-tion and functional maintenance of these structures are shared. However, ELS formation is distinct from the pre-programmed ontogenic processes that are associated with secondary lymphoid organo genesis and it does not occur in all patients. Consequently, the generation of ELSs in inflamed tissues — as opposed to a more typical diffuse pattern of inflammatory infiltrate — must be governed by a defined set of inflammatory signals. The mechanisms that trigger these events are poorly defined.

Initiation of ectopic lymphoid neogenesis. Various transgenic mouse models have emphasized the role of inflamma tory cytokines in lymphoid neogenesis. For example, mice that are deficient in lymphotoxin-α (LTα; which is encoded by Lta) completely lack peripheral lym-phoid organs12. Conversely, tissue-specific expression of a transgene that encodes Lta in the kidney and pancreas caused severe chronic inflammation with the accompa-nying formation of ELSs that were capable of promoting antigen-specific responses and antibody class switching13. Moreover, the overexpression of both Lta and Ltb (which encodes LTβ) results in more prominent ELS forma-tion compared with when only Lta is overexpressed14. Thus, in combination with LTβ, LTα enhances ectopic lymphoid neogenesis14. The activity of lymphotoxin is associated with increased expression of the homeostatic chemokines CXC-chemokine ligand 13 (CXCL13), CC-chemokine ligand 19 (CCL19) and CCL21, and the increased infiltration of T cells that express CD62L

(also known as L-selectin)14. Such findings indicate that the propagation of ELSs within inflamed tissue is driven by communication between local stromal cells, tissue- specific resident mononuclear cells and infiltrating immune cells15,16. Although the actual cells that are responsible for the positioning of these structures within the inflamed tissue remain undefined, several new candi-dates have recently been proposed. These include lymphoid tissue inducer cells (LTi cells), IL-17-secreting CD4+ T cell populations and T follicular helper cells (TFH cells) (FIG. 1).

The accumulation of CD4+CD3−CD45+ LTi cells at sites of lymph node development is an early event in secondary lymphoid organogenesis17–19. This is coordin-ated by the expression of CXCL13, receptor activator of NF-κB ligand (RANKL; also known as TNFSF11) and IL-7, which control the recruitment, survival and activa-tion of LTi cells that express CXCR5 and IL-7 receptor (IL-7R; also known as CD127)20. Central to this process is the interplay between LTi cells and stromal mesenchymal cells — known as lymphoid tissue organizer cells — which act as the anchor for lymph node development11. The release of IL-7 and RANKL by activated lymphoid tis-sue organizer cells promotes the expression of LTα1β2 by LTi cells, which, in turn, engages the LTβ receptor (LTβR) on lymphoid tissue organizer cells that also express vas-cular cell adhesion molecule 1 (VCAM1) and inter-cellular adhesion molecule 1 (ICAM1). This promotes homeostatic chemokine release and vascularization by high endothelial venules (HEVs), which also contribute to lymphoid neogenesis11,15,20,21.

Box 1 | Cellular activation events associated with lymphoid tissues

IntheT cellzonesoflymphoidtissues,dendriticcells(DCs)presentpeptideantigensonMHCclass IImoleculestonaiveCD4+T cells.Therecognitionofthepeptide–MHCclass IIcomplexesbydefinedT cellreceptors(TCRs)promotestheactivationandproliferationofCD4+T cells,andtheirdifferentiationintospecializedeffectorsubsets.TheseeffectorT cellsubsetsincludeCD4+Tfollicularhelper(T

FH)cells,whichexpressCXC-chemokinereceptor5(CXCR5)andmigrate

intotheBcellzoneinresponsetoCXC-chemokineligand13(CXCL13;seethefigure).AttheinterfacebetweentheB cellandT cellzones,CXCR5+ T

FHcellssupporttheactivationofantigen-specificB cells.TheprovisionofB cellhelpbyT

FHcells

drivesB celldifferentiationintoplasmablastsortheirmigrationintofolliclestoformgerminalcentres.PersistentantigenpresentationbyB cellscontributestothefullactivationofT

FHcells,whichcontinuetoprovideB cellhelp.Thus,T

FHcells

supportthemaintenanceofgerminalcentresandpromotethegenerationofbothlong-livedplasmacellsandmemoryB cells(seethefigure).Inthisrespect,T

FHcellscontributetothecontrolofantibodyproductionandthefine-tuningof

mechanismssuchasclassswitching,affinitymaturationandsomatichypermutation.

T cell

T cell zone B cell zone

CXCL13-dependent

Class switching,affinity maturationand somatichypermutation

TCR

Peptide

MHCclass II

Nature Reviews | Immunology

DC

TFH cell B cell

Germinal centre

Plasma cellPlasmablast

Memory B cell

CXCR5

R E V I E W S



Innate lymphoid cell(ILC). A type of innate immune cell that is lymphoid in morphology and developmental origin but that lacks properties of adaptive B cells and T cells, such as recombined antigen-specific receptors. These cells regulate immunity, tissue homeostasis and inflammation in response to cytokine stimulation.

Although stromal lymphoid tissue organizer cells and LTi cells are important in secondary lymphoid organo-genesis, their involvement in ELS formation is less clear. However, stromal tissue cells may acquire lymphoid tissue organizer cell-like properties in ELSs21–23. For exam-ple, synovial fibroblasts from the joints of patients with rheumatoid arthritis release homeostatic chemokines and cytokines that may contribute to ectopic lymphoid neo-genesis within the inflamed synovium2,24–26. Moreover, gene expression profiling of synovial tissue from patients with rheumatoid arthritis has identified an IL7 signature in ELS-associated synovitis26.

The recent discovery of innate adult LTi cells that are part of the innate lymphoid cell (ILC) family18,19 raises the possibility that these cells also contribute to inflam-mation-associated ELS development. For example, the

adoptive transfer of adult LTi cells into Cxcr5−/− mice induces the formation of intestinal lymphoid tissues19. Ectopic lymphoid tissues that develop in response to transgenic overexpression of the Il7 gene also require LTi cells17. Adult LTi cells express the transcrip-tional regulator retinoic acid receptor-related orphan receptor-γt (RORγt) and can produce IL-17, which are characteristics of group 3 ILCs18,19. This pheno type sug-gests an ancestral relationship between adult LTi cells and IL-17-producing CD4+ T helper (TH17) cells18,27. Furthermore, various studies have now linked IL-17 and TH17 cells with ectopic lymphoid neogenesis, where they have a role in chronic allograft rejection, experi-mental autoimmune encephalomyelitis (EAE) and the development of inducible bronchus-associated lymphoid tissue (iBALT)28–30. For example, ELS formation in the

Production of CCL19, CCL21, CXCL12 and CXCL13 maintains ELS


VCAM1 CD62L

HEV

a Initiation of ectopic lymphoid tissue neogenesis

b Cell recruitment to inflamed site

c Maintenance of ELSs

ICAM1

CXCR5

ELS

ICOSPD1

FDC

B cell

T cell

T cell

IL-7Rα

IL-7

RANKLRANK

LTo cell

LTo cell

CD4+ LTi cell

LTi cell

CCL21 andCXCL13

TH17 cell

TH17 cell

TFH-like cell

TH2 cellTH1 cell

RORγtAccumulation of LTi cells

IL-7R and RANKsignalling increaseLTα1β2 expression

Inflamed tissue

LTα1β2

BCL6MAF

LTβR signalling promotes chemokine release and HEV development

Development of TFH-like cells

Figure 1 | Mechanisms that control the induction and maintenance of ectopic lymphoneogenesis. a | Various cell types have been implicated as potential initiators of ectopic lymphoid-like structure (ELS) formation. Although the precise mechanisms require further clarification, the cell types that are associated with this process may include interleukin-17 (IL-17)-secreting CD4+ T helper (T

H) cells (not shown) and CD4+ lymphoid tissue inducer (LTi) cells. b | These cell populations

are attracted to inflammatory sites by certain chemokine signals, including CXC-chemokine ligand 13 (CXCL13) and CC-chemokine ligand 21 (CCL21). Within these inflamed lesions, resident stromal cells contribute to the cellular organization of lymphoid aggregates. Pro-inflammatory mediators — including IL-7 and lymphotoxin α

1β

2 (LTα

1β

2) —

regulate processes that affect the chemokine expression profile that is necessary for further recruitment of B cells and T cells, the spatial arrangement of these cells into defined clusters and the control of angiogenesis. c | Although persistent antigen presentation by follicular dendritic cells (FDCs) and B cells supports the long-term maintenance of these structures, cell types such as CD4+ T follicular helper (T

FH) cells are essential for relaying immunological instructions to

the B cells, which, in turn, ensure the continued action of TFH

cells. Of potential relevance to the development of ELSs in inflamed tissues is the ability of defined CD4+ T

H effector subsets to acquire T

FH cell-like characteristics. Commitment of

cells towards a TFH

cell-like phenotype in inflamed tissues may aid the development or the activities that are associated with ELSs. BCL6, B cell lymphoma 6; HEV, high endothelial venule; ICAM1, intercellular adhesion molecule 1; ICOS, inducible T cell co-stimulator; IL-7R, IL-7 receptor; LTβR, lymphotoxin-β receptor; LTo cell, lymphoid tissue organizer cell; PD1, programmed cell death protein 1; RANK, receptor activator of NF-κB; RANKL, RANK ligand; RORγt, retinoic acid receptor-related orphan receptor-γt; VCAM1, vascular cell adhesion molecule 1.

R E V I E W S



Germinal centre Located in peripheral lymphoid tissues (for example, the spleen or lymph nodes), these are sites in which B cells proliferate and clones that produce antigen-specific antibodies of higher affinity are selected.

AngiogenesisThe development of new blood vessels from existing blood vessels. It is frequently associated with tumour development and inflammation.

central nervous system is associated with TH17 cells that express the lymphoid tissue-associated glyco-protein podoplanin (also known as gp38 in mice)29. Podoplanin-deficient mice display defective develop-ment of lymph nodes and germinal centre structures, and podoplanin expression by fibroblastic reticular cells is associated with the development of the micro-architecture of the T cell zones29,31. Thus, podoplanin expression is a defining feature of tissues that display active lymphoid neogenesis. Although the precise function of podoplanin is unclear, it may support the retention of TH17 cells within these sites29,30. In addition to these studies, various reports have also noted roles for B cells and TNF-secreting F4/80+ myeloid cells in ELS generation32,33. The formation of ELSs in different tissues and in response to different forms of immuno-logical activation suggests that the mechanisms driving ELS expansion are complex. Inherent simi-larities in effector functions or cellular plasticity may render various cell types able to promote tissue-specific ELS development.

Organization of cells within ELSs. Inflamed tissues displaying ELS-associated pathology have increased expression of homeostatic chemokines that govern the cellular composition and functional properties of ELSs34–37. These include CXCL12, CXCL13, CCL19 and CCL21. In such tissues, cellular communica-tion between local stromal cells, tissue-specific resi-dent mononuclear cells and infiltrating immune cells propagates the development of ELSs and enables them to function in a similar manner to germinal centres (BOX 1). Such interactions affect leukocyte trafficking and angiogenesis but are also instrumental in govern-ing the organization of cells within these structures. For example, CXCL13 and CCL21 control the segregation of B cells and T cells in ELSs34,35. By contrast, CCL19 and CXCL12 promote lymphocyte infiltration and the positioning of dendritic cells, B cells and plasma cells within these aggregates34 (FIG. 2). These specialized properties probably reflect differences in the ability of individual homeostatic chemokines to promote the expression of LTα and LTβ by B cells and T cells34. For example, transgenic expression of Cxcl13 in pancre-atic tissue promotes the LTα1β2-mediated formation of ELSs that display defined lymphoid zones, HEVs and stromal reticulum35. Thus, homeostatic chemokines affect the size, cellular composition and organization of ELSs. Such features affect the functional properties of ELSs and their impact on pathology.

However, the contribution of individual homeostatic chemokines to ELS formation may also depend on the anatomical site or ongoing disease processes. Although transgenic expression of Ccl21 in the pancreas drives pancreatic lymphoid neogenesis, the ectopic expres-sion of Ccl21 in the skin does not36,37. Whether these differences reflect a hierarchy of chemokine-mediated outcomes that affects the degree of architectural organ-ization displayed by ELSs or whether they reflect spe-cific characteristics that are associated with ‘permissive’ versus ‘non-permissive’ tissues remains unknown.

Maintenance of ELSs within tissues. Considerable atten-tion has been given to the development of lymphoid structures, however, the presence of ELSs is often a tran-sient feature of inflamed tissue. Thus, the mechanisms that control the maintenance of these aggregates during disease may have greater clinical significance. Recent studies have linked TFH cells — or markers of TFH cell activity — to the formation, maintenance and function of ELSs29,38–40.

TFH cells promote B cell activities and high-affinity antibody generation in germinal centres41,42. Recent stud-ies of T cell plasticity in mice suggest that TH1, TH2 and TH17 cells may also acquire TFH cell-like characteristics and effector functions during their differentiation29,43–47. For example, TH17 cells can display TFH cell-like charac-teristics during their differentiation, including the secre-tion of IL-21 and the expression of TFH cell-associated molecules, such as signal transducer and activator of transcription 3 (STAT3), interferon-regulatory fac-tor 4 (IRF4), MAF and inducible T cell co-stimulator (ICOS)48–53. These observations may help to explain the ability of TH17 cells to promote ectopic lymphoid neo-genesis28–30 and may also explain the regulation of ELS activity in diseases that are associated with TH1, TH2 or TH17 cell responses. Memory and effector TH cells that lack typical TFH cell-like features are, nevertheless, also capable of providing B cell help as they express CXCL13, IL-4, IL-21 and CD40L54–56. Consequently, both positive and negative regulators of effector T cell responses may influence the maintenance of ELS activity (FIG. 2).

The CCR7-dependent recruitment of dendritic cells to developing ELSs represents an important homeo-static event that drives the initial propagation of adap-tive immune responses within these sites. For exam-ple, infiltrating CD4+ T cells form tight clusters with dendritic cells and this promotes T cell proliferation57. As ELSs mature, dendritic cells within these structures continue to support the efficient priming of T cell responses through antigen presentation and they con-tribute to class-switch recombination, antibody genera-tion and the formation of new lymphatic vessels58–60. The importance of dendritic cells in ELSs is best illus-trated by studies of mucosal immunity following viral infection. The depletion of dendritic cells during viral lung infection leads to impaired germinal centre reac-tions and a disruption of ELS architecture59,60. The sustained activation of dendritic cells within ELSs is therefore required for both their formation and their functional maintenance.

The function of ELSs as germinal centres. There is conclusive evidence that ELSs not only recapitulate the cellular, molecular and structural organization of SLOs but that they can also support the function of germinal centres. In the germinal centres of SLOs, B cells undergo affinity maturation and differentiation to memory B cells and plasma cells via antigen-driven selection61 (BOX 1). This process includes antibody fine-tuning through somatic hypermutation and class switching, both of which affect antigen recognition and the effector capacity of the antibody62–65. Consistent with

R E V I E W S



Activation-induced cytidine deaminase (AID). An enzyme that is required for two crucial events in the germinal centre — somatic hypermutation and class-switch recombination.

the role of ELSs as functional ectopic germinal centres, activation-induced cytidine deaminase (AID) is expressed at the mRNA and protein level within these structures, and studies in patients and animal models support the involvement of AID in autoimmunity66–69, infection70 and allograft rejection28. For example, AID controls local antibody affinity maturation (including somatic hypermutation), as evidenced by a restricted profile of variable (V)-gene repertoire usage, highly mutated V regions and the oligoclonal diversification of infiltrat-ing B cells and plasma cells68,71–73. Active class switching is also confirmed by the detection of Iγ–Cμ and Iα–Cμ circular transcripts within ELSs, which marks on going class-switch recombination from IgM to IgG and IgA, respectively67,68,74. Thus, ELSs in inflamed tissues retain the necessary molecular machinery to support in situ antibody diversification, isotype switching, B cell differ entiation and oligoclonal expansion. However,

as reviewed below, although the final outcome of lym-phoid neogenesis is generally protective in the context of infection and cancer, it is often deleterious in the setting of autoimmunity and graft rejection.

ELSs in protective immunity and diseaseELSs as sites of anti-pathogen immune responses. Infec-tion with bacteria (such as Mycobacterium tuberculosis and Helicobacter pylori) and viruses — such as influ-enza virus, hepatitis C virus (HCV) and several sialo-tropic viruses — has been associated with the formation of ELSs in animal models39,59,74,75 and in humans76–78. These ELSs resemble highly organized SLOs. Most remarkably, in mice, infection-triggered lymphoid neo-genesis seems to be preferentially induced at permis-sive mucosal sites, and results in the de novo formation of iBALT and of lymphoid tissues associated with the salivary glands79,80.


B cell

Skin

Infection with Mycobacteriumtuberculosis or influenza virus

Inducible BALTformation

Ectopic CCL21-expression ELS formation Ectopic CCL21-expression No ELS formation

Pancreas

Lung

LTi cell LTo cell

b

T cell

FDC

Classical lymphoid neogenesis Putative regulators of ELS neogenesis

Initiators• IL-7• LTα1β2• RANKL

Propagators• CCL19• CCL21• CXCL12• CXCL13

LTi cell LTo cell

Initiators• IL-7• LTα1β2• RANKL

Propagators• CCL19• CCL21• CXCL12• CXCL13

Initiators• IFNs• IL-4• IL-5

• IL-17• IL-21• TNF

Propagators• IL-6• IL-17• IL-21

Inhibitors• IL-2• IL-27

Permissive tissue Non-permissive tissue

Non-inflamed tissue Inflamed tissue

Figure 2 | Cytokines and chemokine regulate ELS formation and function. The inducible formation of ectopic lymphoid structures (ELSs) mimics the ontogenic process of secondary lymphoid organ (SLO) development, whereby the cytokines interleukin-7 (IL-7), lymphotoxin-α (LTα), LTβ and receptor activator of NF-κB ligand (RANKL) have a direct role in initiating chemokine-directed lymphoid organogenesis. It is becoming clear that, during ELS formation, effector cytokines that are produced in response to chronic inflammation, infection, autoimmunity and cancer are indirect regulators that substitute for the cytokines that are involved in SLO formation. Recent studies have highlighted novel roles for immune cell subsets in ectopic lymphoid neogenesis. For example, T helper 17 (T

H17) cells and T follicular helper

(TFH

)-like cells are associated with ELS development in the central nervous system and the lungs29,30,38. Various TH cell

subsets can also acquire TFH

cell-like characteristics and effector functions29,43–47, and these cells may contribute to ELS development in autoimmune and infectious diseases. The discovery of a role for these novel cell types brings with it a host of regulators that may positively and negatively regulate ELS formation and function. These include factors that determine the commitment or effector characteristics of defined T cell populations — for example, the control of T

H17 cells and T

FH-like cells by IL-2, IL-6, IL-21, IL-27 and type I interferons (IFNs) — and mediators that have altered

expression in ELSs within defined anatomical locations (for example, IL-4, IL-5, IL-17, IL-21 and IL-27). Potentially, these may serve as alternative therapeutic targets or agents for ELS-associated pathology. CCL, CC-chemokine ligand; CXCL, CXC-chemokine ligand; FDC, follicular dendritic cell; LTi cell, lymphoid tissue inducer cell; LTo cell, lymphoid tissue organizer cell; TNF, tumour necrosis factor.

R E V I E W S



GranulomaA chronic inflammatory tissue response that occurs at the site of implantation of certain foreign bodies and, in particular, at sites of long-term microbial persistence. Granulomas are typically well-organized structures that are composed of T cells and macrophages, some of which fuse to form giant cells.

Severe combined immunodeficiency mice(SCID mice). A strain of mice that possesses a genetic defect in DNA recombination that leads to severe immunodeficiency. SCID mice lack B cells and T cells, and they are incompetent at rejecting tissue grafts from allogeneic and xenogeneic sources.

Consistent with their immunological functions, infection-associated mucosal ELSs can mount protec-tive immune responses in situ. Examples of this include influenza virus-induced and M. tuberculosis-induced lymphoid neogenesis in the lungs of rodents. Following influenza virus infection of the upper respiratory tract, mice develop functional ELSs79 that are maintained by lymphoid tissue-associated chemokines (for example, CXCL13 and CCL21) and LTβ, which are produced by infiltrating CD11b+CD11c+ dendritic cells59. These ELSs promote the in situ differentiation of antiviral plasma cells that are specific for the nucleoprotein of influenza virus59. Notably, in LTα-deficient mice — in which SLOs do not develop — reconstitution with Lta+/+ bone marrow allows for ELS development in response to influenza virus infection81. These bronchial ELSs can maintain antiviral immunity and memory recall responses in the absence of SLOs81. A role of ELSs in antimicrobial immunity is further illustrated by their involvement in M. tuberculosis infection, where they promote granuloma formation and prevent the dissemination of infection38,75. Although these studies demonstrate a non-redundant role for ELSs in certain infections, they also indicate that pathogen recognition by innate-sensing mechanisms may contribute to the formation of these structures.

Overall, these data suggest an evolutionary role of ELSs at sites of infection where they coordinate protective immunity in addition to, and often independently from, SLOs. However, a failure to eradicate a pathogen has considerable clinical implications and can lead to the development of autoimmunity (for example, in chronic HCV infection) and lymphoma (for example, in H. pylori-associated chronic gastritis). Thus, ectopic lymphoid neogenesis in response to microbial challenge influ-ences the natural history and clinical course of chronic infection and associated complications.

ELSs in autoimmunity and the emerging role of Epstein–Barr virus. The presence of ELSs with the appearance of fully functional ectopic germinal centres has long been described in the inflamed target organs or tissues of patients who are affected by autoimmune diseases, including the synovial tissue in rheumatoid arthritis (as reviewed in REF. 82), the meninges in multiple sclerosis69, the salivary glands in Sjögren’s syndrome (as reviewed in REF. 83), the thymus in myasthenia gravis (as reviewed in REF. 84) and the thyroid gland in Hashimoto’s thyroidi-tis85. In these clinical disorders, ELSs develop in response to disease-specific autoantigens, which also ensure the long-term maintenance of these structures within the inflamed tissue.

The presence of ELSs in autoimmune conditions perpetuates autoimmunity towards disease-specific antigens. In this respect, many of the regulatory mech-anisms that govern tolerance within SLOs are not seen in autoimmune disease-associated ELSs. For example, in SLOs, autoantigen-binding B cells may be excluded from entering germinal centres and they lack responsiveness to CXCL13 owing to a downregulation of CXCR5 (REF. 86). However, autoimmune disease-associated ELSs permit the entry of autoreactive B cells87. This allows for the

differentiation of the B cells into high-affinity auto reactive plasma cells that release disease-specific autoantibodies, such as anti-citrullinated protein antibodies (ACPAs) in rheumatoid arthritis67, and antibodies against the ribo-nucleoproteins Ro and La (also known as Sjögren’s syn-drome antigens A and B) in Sjögren’s syndrome88, against thyroglobulin and thyroperoxidase in Hashimoto’s thyroiditis85, and against the nicotinic acetylcholine receptor in patients with myasthenia gravis84.

Although the mechanisms that are responsible for the preferential accumulation of autoreactive B cells in ELSs are not fully understood, a direct role has been proposed for Epstein–Barr virus (EBV) in the develop-ment of autoimmunity. EBV is a γ-herpesvirus that is retained throughout the life of an infected individual and that promotes the survival and proliferation of B cells89,90. Emerging evidence suggests that ectopic follicles in the target organs of patients with multiple sclerosis, myasthenia gravis, rheumatoid arthritis and Sjögren’s syndrome frequently harbour latent EBV infection69,91–93. In these sites, EBV-transformed B cells and plasma cells display autoreactivity to disease-specific autoantigens, such as citrullinated fibrinogen in rheumatoid arthritis92 and the ribonucleoprotein Ro in Sjögren’s syndrome93. Autoreactive EBV-infected B cells are therefore predicted to transit from the peripheral compartment into ectopic germinal centres where they undergo differentiation to high-affinity autoreactive plasma cells. Thus, niches of EBV colonization within ELSs might be considered as a hallmark of organ-specific autoimmunity.

EBV infection may also explain another remark-able feature of ELSs, which is their ability to persist in an activated state for several weeks in the absence of recirculating immune cells from the periphery. This is best demonstrated in chimeric severe combined immuno deficiency mice that are engrafted with synovial or thymic tissue from patients with rheumatoid arthri-tis or myasthenia gravis, respectively. ELSs within the transplanted tissue maintain their follicular organization and auto antibody generation despite both the absence of ‘new’ cells infiltrating the grafts and the impaired host immune response67,94. This is of crucial importance, as ELS neutralization is probably fundamental to avoiding the re-emergence of autoreactive clones that are capable of driving disease relapse or resistance to therapy.

ELSs in transplantation. ELSs have been observed in almost all types of human grafts that have been removed owing to chronic rejection, including kidneys, lungs and hearts95. Transplant-associated ELS formation seems to recapitulate the molecular pathways that are triggered during the ontogeny of SLOs. These ELSs harbour ectopic germinal centres in which naive B cells differ-entiate into anti-HLA-producing plasma cells and memory B cells96. However, ELS formation in allografts differs from that of canonical SLOs in a number of ways: first, the considerable amounts of TH17 cell-associated cytokines and growth factors — such as B cell-activating factor (BAFF; also known as TNFSF13B) — that facili-tate autoreactive B cell survival; second, the continuous release of alloantigens from the injured tissue that are

R E V I E W S



trapped locally by defective lymphatic drainage; and third, the apparent defect of local immunoregulatory mechanisms. As a consequence, there is an excessive immune response within the ELSs that contributes to allograft rejection95. However, two recent publications in distinct experimental models (renal and cardiac allografts) have reported that the presence of ELSs promotes allograft tolerance and graft function97,98. On this basis, it is postulated that under certain conditions, ELSs may amplify beneficial immune responses — that are associated with B cells and T cells displaying a regulatory profile — to promote graft tolerance.

ELSs in cancer. In cancer, ELSs have been documented in many tumours, including those that are associated with lung99,100, colon10,101, breast40,102,103 and germ cell cancers104,105. However, not every patient with a specific type of cancer develops ELSs and when they do occur, their contribution to disease varies considerably. Some tumour types are more likely to induce ELS forma-tion than others, which indicates that certain tumours provide a microenvironment that is conducive to lym-phoid neogenesis. Given the stark difference between the immunosuppressive nature of the tumour micro-environment compared with chronic inflammatory foci, it is perhaps surprising that ELSs develop at all in tumours104. Although CXCL13, CCL19 and CCL21 have a crucial role in the formation of cancer-associated ELSs, LTi cells seem to be dispensable as ELSs can form in their absence100,104,105. Permissive tumours probably induce the production of lymphoid chemokines not only through the expression of LTα1β2 but also through the expression of pro-inflammatory cytokines — such as TNF, IL-1β and IL-6 — which contribute to the transcriptional reg-ulation of CXCL13, CCL19 and CCL21. However, the precise stimuli that initiate the development of cancer-associated ELSs within the immunosuppressive tumour microenvironment require further investigation.

Clinical implicationsAs discussed above, ELSs are found in patients with diverse medical conditions but invariably, in each con-dition, ELSs form in some patients but not in others (TABLE 1). Therefore, important questions relating to this phenomenon remain to be answered. For instance, what determines the formation of ELSs in some individuals but not in others? Is ELS development typical of different dis-ease subtypes (ab initio) or is ELS formation the inevitable result of persistent inflammation — that is, are ELSs the cause or the consequence of chronicity? To what extent do ELSs contribute to ongoing inflammation and tissue damage (disease prognosis)? In this section, we address these questions in turn by providing recent data from the literature, as well as from our own personal experience.

What governs ELS formation in some individuals but not in others? This question still remains mostly unanswered but it is likely that genetic and/or environ-mental factors shape the inflammatory response to give rise to diverse structural outcomes within individual tissue microenvironments.

Considering the genetic influence first, evidence is beginning to emerge from various genetic studies that have found associations between specific pathways that are involved in ELS formation and/or function and susceptibility to autoimmune diseases. For example, four single nucleotide polymorphisms (SNPs) have been iden-tified in the IL2–IL21 locus at chromosome region 4q27 that are associated with several autoimmune diseases106–108 and that can influence circulating levels of IL-21 (REF. 109). Additionally, SNPs in loci that are near to the CXCR5 gene have been linked with susceptibility to primary biliary cirrhosis110, Sjögren’s syndrome111, systemic lupus erythematosus (SLE)112 and multiple sclerosis113.

Although no sub-analysis with stratification of patients for the presence or absence of ELSs has been carried out in the above studies, the increased genetic susceptibility to autoimmunity that is associated with loci near to both the IL21 and CXCR5 genes corresponds with the crucial role of IL-21-producing CXCR5+ TFH cells in the formation and function of ELSs. This is also poten-tially of crucial relevance as novel strategies to specifically target TFH cells are being developed for therapeutic use in autoimmune conditions (as discussed below). It is likely that the influence of genetic factors will become clearer as larger scale analyses of stratified disease subsets are carried out according to pathobiology (for example, the presence or absence of ELSs).

With regard to the environmental factors that influ-ence ELS formation, diverse host immune responses to infection lead to different outcomes. Thus, it is possible that in ‘permissive’ patients, an active EBV infection is established in the diseased tissue and may drive ELS for-mation through its unique ability to infect and activate B cells114,115. This hypothesis was recently supported by our own work, in which we found a high frequency of EBV-infected B cells and plasma cells in ELS-containing synovia from patients with rheumatoid arthritis92. Nota-bly, EBV was not detected in control synovia from patients with osteoarthritis or in patients with rheumatoid arthritis whose synovia showed diffuse infiltration of immune cells but lacked ELSs92. Similar results were seen in the salivary glands of patients with Sjögren’s syndrome93.

Is ELS formation pre-determined or a result of persistent inflammation? The frequency at which ELSs occur in patients with chronic inflammation typically varies from 20–40%7,82. However, it is unknown whether ELSs are a typical feature of different disease subtypes from the beginning (ab initio) or whether they form as a result of persistent inflammation. The reason this this remains unknown is twofold: first, in humans, clinical presenta-tion is often delayed and it is thus impossible to precisely determine the early events in the pathological process; and second, there are an insufficient number of studies (and an insufficient number of large cohort studies) in which serial biopsies have been taken to establish the ‘pre-determined’ versus the ‘evolutionary’ nature of ELSs. Nonetheless, data is beginning to emerge from a cohort of 300 patients with rheumatoid arthritis who had shown symptoms for less than 12 months; biopsies were taken at the time of disease presentation and 70% of

R E V I E W S



Cryoglobulinaemia An autoimmune, extra-hepatic manifestation that is associated with certain viral infections and chronic inflammatory conditions, whereby antigen–antibody complexes are deposited in capillaries and arterioles (and occasionally small arteries) causing vasculitis, renal disease, arthralgias or arthritis.

individuals were also biopsied 6 months post-treatment116. Approximately 40% of patients displayed histo logical evidence of ELSs before receiving any disease-modifying therapy. Thus, ELSs seem to be present ab initio and to define a specific disease subset from the beginning. However, in another early arthritis cohort of 93 patients with only 17 biopsy repeats at 6 months, ELSs have been reported as being related to the degree of inflammation and not related to specific disease subtypes117. No serial biopsy data is available from other diseases and, there-fore, further work is required in this area to establish the precise ontogeny of ELSs in the context of chronic inflammation.

To what extent do ELSs control disease progression and outcome? From the above discussion, it is clear that ELSs are dynamic structures, the main function of which is to potentiate the immune response at sites of disease. The clinical presentation of ELSs may therefore influence dis-ease severity, the rate of disease progression and clinical outcome. Ultimately, this will depend on the ability of the host to clear triggering antigens, as well as the stage and the nature of the disease in question. In the subsequent paragraphs, we provide some examples of the influence of ELSs on the progression and outcome of infection, autoimmune conditions, transplantation and cancer.

On the basis of the evidence from animal models showing a protective role of mucosal ELSs in pathogen control, it is anticipated that the presence of ELSs in infected humans could alter the natural history and clinical manifestations of chronic infections. Accord-ingly, in M. tuberculosis infection, ELS formation in the peripheral rim of lung granulomas is typically associated with the latent, asymptomatic form of tuberculosis118. Conversely, sites of active infection that have cavitary tuberculous lesions lack B cell follicles118. Thus, ELSs coordinate the local host response to pulmonary tuber-culosis118. ELSs probably exert similar mechanisms of pathogen containment in other chronic infections, such as H. pylori and HCV76,77. However, it is now clear that in the absence of pathogen eradication, ELSs also act as a double-edged sword by contributing to autoimmunity and lymphomagenesis. Although tuberculosis has long been associated with the development of various auto-immune phenomena119, the association with ELSs has not been investigated. Conversely, in chronic HCV-related hepatitis, elevated CXCL13 levels in liver biopsies identify patients that develop mixed cryoglobulinaemia120. Together with the clonal analysis of intrahepatic B cells in portal tracts121, these data indicate that intraportal lymphoid follicles contribute to the evolution from polyclonal to oligoclonal to monoclonal autoreactive B cell activation that occurs as a result of chronic HCV infection. Pathogen-driven chronic B cell activation within ELSs is also associated with lymphoproliferative disorders, as evident in HCV-associated B cell lympho-proliferation122 and H. pylori-driven gastric mucosa-associated lymphoid tissue (MALT) B cell lymphomas76. The development of gastric (and salivary gland) MALT lymphomas has been linked to the genetic instability that is associated with the process of aberrant somatic

hypermutation, whereby dysregulated expression of AID leads to mutations in the regulatory and coding sequences of genes that regulate B cell survival and proliferation (such as PAX5 and MYC)123. Interestingly, AID expres-sion in gastric and salivary gland MALT lymphomas is confined to ELSs and is not found in malignant marginal zone B cells66,123, which suggests that lymphoma genesis is dependent on sustained antigen stimulation within ELSs. Accordingly, gastric MALT lymphomas are cru-cially dependent on H. pylori-specific T cells124 and the eradication of H. pylori results in both ELS resolution and tumour regression125. Although in this context lympho-magenesis is ultimately driven by chronic infection, a striking and unique feature of MALT lymphomas is that malignant B cells frequently originate from precursors that express an autoreactive B cell receptor with homol-ogy to rheumatoid factor126. Thus, autoimmunity and MALT lymphomagenesis can represent a continuum of the same antigen-driven response in which ELSs have a central role.

In chronic inflammatory and autoimmune con-ditions, the B cell-rich inflammatory infiltrate that is typically seen in ELSs is associated with a strong potential for local immune activation and cellular dif-ferentiation. This promotes autoantibody production, complement activation and pro-inflammatory cytokine release, which drives the inflammatory cascade, auto-immunity and tissue destruction. For example, studies in rheumatoid arthritis have reported worse disease out-comes in patients with ELSs116. In these cases, increased joint destruction is associated with the production of pro-inflammatory cytokines127, as well as osteoclast-activating molecules such as RANKL128 and the RANKL-inducing molecule TNF-related weak inducer of apoptosis (TWEAK; also known as TNFSF12)129. In autoimmune thyroiditis, most ELS B cells recognize the autoantigens thyroglobulin and thyroperoxidase, and this leads to glandular destruction85. In SLE, a restricted repertoire of isotype-switched antibodies are deposited in the glomerular basement membrane of the kidneys with consequent organ damage130. Finally, the presence of ELSs in the inflamed meninges of patients with pro-gressive multiple sclerosis can be determined through analyses of B cell markers, and the occurrence of ELSs is associated with a gradient of cortical neurodegeneration and a more aggressive course of disease131.

ELSs are thought to drive chronic transplant rejection by enhancing intra-graft allogeneic responses, as grafts in which the ELSs are most functional have a shorter life expectancy28,96. However, the validity of this conclu-sion has been questioned by the fact that, by definition, only de-transplanted, failed grafts were included in the analysis. In addition, as discussed above, there is evi-dence from experimental transplant models that regula-tory B cells and ELSs may govern graft tolerance. Further work is therefore required to clarify the contribution of ELSs to graft rejection versus survival.

As discussed above, different cancers are more or less immunogenic and hence, are more or less prone to induce ELSs. Analyses of the cytokine and chemokine gene signatures, the number of tumour-infiltrating

R E V I E W S



lymphocytes and the level of ELS organization have been used to stratify patients and types of cancer as ‘immune response positive’ and ‘immune response nega-tive’. Although these classification criteria are not purely based on ELSs, there is a general consensus that patients who have immune-response-positive tumours (for exam-ple, breast cancer tumours) have a better prognosis than those with immune-response-negative tumours102,132–134. Similar results have been described in colon cancer, where immune response positivity was associated with the increased survival of patients independently of tumour stage, previous treatment or microsatellite insta-bility10. As expected, CXCL13 and CCL19 expression was also associated with a good prognosis10. Finally, gene expression profiling of immune-response-positive and immune-response-negative human melanomas identi-fied CXCL13 and CXCL8 as components of a discrete group of 12 genes that were found to be diagnostic of immune response status135. In another study, the mean survival of patients with melanoma was 55 weeks in the immune-response-positive group compared with 18 weeks in the immune-response-negative group136.

In summary, ELS formation in peripheral tissues seems to follow normal pathophysiological programmes in response to the need to increase local immune responses and clear exogenous antigens, alloantigens and/or autoantigens. When such effector programmes are successful, they lead to improvements in disease pathology and a better prognosis, whereas their failure or ineffectiveness leads to persistent inflammation, tis-sue damage and worse outcomes. This bivalent function-ality has substantial clinical implications when it comes to targeting ELSs for therapy.

ELSs as therapeutic targetsThe modulation of ELSs for therapeutic purposes has attracted marked interest both from the scientific com-munity and from industry. On the basis of the func-tional and clinical data discussed above, it would be desirable to potentiate the activity of ELSs in infection and cancer but to inhibit their activity in the context of transplantation and chronic inflammatory conditions. Current cancer trials are testing whether ELS-related pathways can be exploited to enhance antitumour immunity. Although cytokine and chemokine admin-istration, and tumour antigen vaccination represent promising therapeutic approaches to target ELSs, the long-term efficacy of these strategies remains debat-able137,138. Similar vaccination approaches are also being considered as adjunct therapies that promote local antimicrobial immunity in the lungs139.

By contrast, there is a strong rationale for therapeu-tic inhibition of ELSs in graft rejection, and in chronic inflammatory and autoimmune conditions. In a study of patients showing chronic renal allograft rejection, treatment with anti-CD20 (rituximab) failed to cause regression of ELSs despite the successful depletion of B cells from the peripheral blood140. This suggests that the local inflammatory microenvironment in ELSs may facilitate B cell survival and allow evasion of rituximab-mediated depletion140. In the context of autoimmunity,

biological interventions that promote the clearance or inhibition of autoreactive lymphocytes have the poten-tial to disrupt ELS involvement and restrict deleteri-ous adaptive immune responses. In this regard, under-standing the impact of TNF inhibitors on synovial ELSs is of major importance. In a recent study, patients with rheumatoid arthritis who had synovial ELSs displayed an inferior response to TNF inhibitors compared with patients who did not have synovial ELSs9. In addition, the presence of ELSs in synovial tissue before treatment was reported to be an independent negative predictor of the response to TNF inhibitors9. Furthermore, high protein-level expression of IL-7R, which is normally associated with ELSs, was reported to be associated with a poor response to TNF inhibitors141. Therefore, the development of selective ELS-targeted therapies has attracted marked interest both from the scientific community and from industry.

Currently licensed therapies for inflammatory dis-eases, including CD20-specific (for example, rituxi-mab) and IL-6R-specific (for example, tocilizumab) monoclonal antibodies, oral Janus kinase inhibi-tors (for example, tofacitinib) and T cell activation blockers (for example, abatacept) probably target processes that are associated with ELS activities4. However, these drugs also possess broader modes of action3–5. Given the prominent role of lymphotoxin and homeostatic chemokines in ELS formation, initial approaches targeted this axis. Pharmacological inhibi-tion of LTβ using an LTβR–immunoglobulin fusion protein resulted in disease amelioration in animal models of arthritis142, autoimmune sialoadenitis143 and type 1 diabetes144. Although, baminercept (which is a humanised LTβR–IgG1 fusion protein) failed to show clinical efficacy in a Phase II clinical trial in patients with rheumatoid arthritis, its utility in Sjögren’s syndrome is currently under investigation (TABLE 2). Recently, a new LTα-specific monoclonal antibody has completed a Phase I clinical trial in patients with rheumatoid arthritis145 but the results of the Phase II study have not yet been published. It is noteworthy that none of the above studies have stratified patients for the presence of ELSs in the disease-affected tissue, which would probably influence the reported extent of clinical responses. However, the baminercept trial in Sjögren’s syndrome includes a salivary gland biopsy sub-study, which will inform on ELS modulation after treatment.

Blockade of CXCL13, CCL19 and CCL21 or their receptors CXCR5 and CCR7 is another potential thera-peutic strategy that has shown some promise in animal models. CXCL13 blockade ameliorated collagen-induced arthritis146 and autoimmune sialoadenitis147 but not autoimmune diabetes148. Of note, although CXCL13 inhibition altered the structure of ELSs, it did not influ-ence their function or the incidence of diabetes148. To date, no developmental compound targeting this path-way has been tested in human clinical trials. However, there is evidence that blocking immune cell recircula-tion to ELSs in the context of autoimmune disease is a promising therapeutic strategy.

R E V I E W S



Non-obese diabetic mice (NOD mice). These mice spontaneously develop diabetes as a result of autoreactive T cell-mediated destruction of pancreatic β-islet cells.

Fingolimod (also known as FTY720) — which blocks the egress of lymphocytes from SLOs via func-tional antagonism of the sphingosine-1-phosphate receptor — has shown favourable efficacy in relapsing–remitting multiple sclerosis149 and has received approval from the US Food and Drug Administration (FDA) for this indication. Fingolimod selectively reduces the fre-quency of circulating CCR7+CD4+ T cells (that is, naive and central memory T cells) and traps this subset of T cells in SLOs, which prevents their migration into the central nervous system150.

Given the role of TFH cells in ELSs, there is also growing interest in blocking key TFH cell-related signatures — for example, the co-stimulatory molecules ICOS and ICOS ligand (ICOSL) or the key TFH cell-related cytokine IL-21. Blockade of IL-21–IL-21R signalling ameliorated disease in animal models of arthritis151 and SLE152, and an anti-human IL-21 monoclonal antibody (NNC0114-0006) has recently completed Phase II clinical trials in rheumatoid arthritis and a Phase I clinical trial in SLE, but no published data are currently available (TABLE 2). Pharmaco logical ICOS blockade has also shown benefit in experimental arthritis153 and a mouse model of lupus154 via inhibition of TFH cell responses. Similarly, the concomitant inhibition of ICOS and CD40L co-stimulation protects non-obese diabetic mice (NOD mice) from diabetes155. As a conse-quence, an anti-ICOSL monoclonal antibody (AMG557; also known as mAb-3B3) has recently entered Phase I clinical trials in SLE (TABLE 2). The co-stimulatory

signalling that is mediated by ICOS–ICOSL and CD40–CD40L is also required for TH17 cell survival48,153 and, as such, may contribute to the formation or main-tenance of ELSs within inflamed tissue. Thus, it will be extremely interesting to investigate whether directly tar-geting the IL-17 pathway with novel therapeutics (such as secukinumab, ixekizumab and brodalumab) that are currently in late-stage clinical development (TABLE 2) will modulate ELSs in the context of autoimmune disease. Finally, it remains to be established whether therapeutics that target factors that are associated with B cell survival and proliferation disrupt ELSs or impair their role as func-tional niches for autoimmune B cell activation. In this regard, data are eagerly anticipated from a biopsy-based Phase II clinical trial in Sjögren’s syndrome with beli-mumab, which is an anti-BAFF monoclonal antibody that has received FDA approval for SLE (TABLE 2). Such stud-ies should elucidate whether BAFF inhibition is sufficient to disrupt ELS functionality in the salivary glands.

Concluding remarks and future directionsAn increasing number of clinical investigations empha-size that ectopic lymphoid neogenesis is a common occurrence at sites of inflammation, whereby ELSs form an integral part of the immune response to infections, tumours and autoantigens. Although experimental ani-mal models have defined certain mechanistic aspects relating to ELS development, function and maintenance, parallel investigations in human conditions remain

Table 2 | Novel therapies targeting ELSs in ELS-positive autoimmune diseases

Pathway inhibited Developmental compound Target or class of drug Completed and ongoing clinical trials in ELS-positive autoimmune diseases*

IL-17 Secukinumab IL-17A-specific monoclonal antibody •NCT01377012 (RA, Phase III, recruiting)‡

•NCT01350804 (RA, Phase III, recruiting)‡

•NCT01874340 (MS, Phase II, recruiting)•NCT02044848 (T1D, Phase II, recruiting)

Ixekizumab IL-17A-specific monoclonal antibody NCT00966875 (RA, Phase II, completed)

Brodalumab IL-17RA-specific monoclonal antibody •NCT00950989 (RA, Phase II, completed)•NCT01059448 (RA, Phase II, terminated)

ABT-122 TNF and IL-17 bispecific antibody NCT01853033 (RA, Phase I, ongoing)

CNTO-6785 IL-17A-specific monoclonal antibody NCT01909427 (RA, Phase III, recruiting)

IL-21 NNC0114-0006 IL-21-specific monoclonal antibody •NCT01647451 (RA, Phase II, completed)•NCT01689025 (SLE, Phase I, completed)

LTα/β Baminercept LTβR–IgG1 fusion protein •NCT00664573 (RA, Phase II, completed)•NCT01552681 (SS, Phase II, recruiting)§

Pateclizumab LTα-specific monoclonal antibody NCT01225393 (RA, Phase II, completed)

ICOS–ICOSL AMG557 (also known as mAb-3B3) ICOSL-specific monoclonal antibody NCT01683695 (SLE, Phase I, recruiting)

S1PR1 Fingolimod (also known as FTY720) S1PR1 antagonist NCT01310166 (MS, Phase IV, recruiting)

BAFF Belimumab BAFF-specific monoclonal antibody •NCT01008982 (SS, Phase II, completed)§

•NCT00071812 (RA, Phase II, completed)•NCT01480596 (MG, Phase II, recruiting)•NCT01639339 (SLE, Phase III, recruiting)

Tabalumab BAFF-specific monoclonal antibody NCT01202760 (RA, Phase III, completed)

BAFF, B cell-activating factor; ICOS, inducible T cell co-stimulator; ICOSL, ICOS ligand; IL, interleukin; IL-17RA, IL-17 receptor A; LT, lymphotoxin; LTβR, LTβ receptor; MG, myasthenia gravis; MS, multiple sclerosis; RA, rheumatoid arthritis; S1PR1, sphingosine-1-phosphate receptor 1; SLE, systemic lupus erythematosus; SS, Sjögren’s syndrome; T1D, type 1 diabetes; TNF, tumour necrosis factor. *Information from the ClinicalTrials.gov website, accurate as of May 2014. ‡NCT01377012 is recruiting patients with RA who failed to respond to conventional disease-modifying anti-rheumatic drugs (DMARDs), whereas NCT01350804 enrols patients with RA who are inadequate responders to anti-TNF therapy. § These studies include a salivary gland biopsy sub-study, pre-treatment and post-treatment.

R E V I E W S



http://www.clinicaltrials.gov

relatively sparse by comparison. It is important to deter-mine whether the clinical consequences of ELS activi-ties are beneficial (for example, immunity to infections or cancer) or detrimental (for example, the promotion of auto immunity or graft rejection). Clinical trials in conditions that are typically associated with ectopic lym-phoid neogenesis may broaden our understanding of ELS involvement in these diseases. An increasing number of novel biologics that target mediators that are central to ELS biology are now entering the clinical arena. However, the key goal is to identify specific molecular signatures that will predict at an early stage of the disease process whether ELSs will form and that can be used to inform decisions regarding the most appropriate therapy for indi-vidual patients. For example, there may be little point in targeting pathways that promote ELS development if these structures have already become a prominent feature of the underlying pathology by the time a patient is diagnosed. Instead, the use of agents that block the long-term main-tenance of ELSs within these inflamed tissues may prove to be a more suitable approach in this scenario. To realize

this ambition, clinicians need to consider more precise diagnostic approaches in their patient management proto-cols. The use of ultrasound-directed biopsy techniques has revolutionized oncology treatment and the introduction of molecular pathology in autoimmune conditions is now being used to enhance the mechanistic understanding of the crucial pathways that are driving disease. For exam-ple, ultrasound-guided synovial biopsy studies in patients with rheumatoid arthritis have highlighted the clinical heterogeneity of synovitis within the disease-affected tissue116. The tremendous progress in miniaturized tech-nologies and high-throughput ‘multi-omic’ approaches will enable clinicians to move away from defining disease mainly on the basis of symptoms and signs, and towards a new taxonomy that integrates molecular signatures into systematic algorithms to map the observed pathology onto existing disease classifications. This information will lead to improved patient stratification, better and more appropriate clinical management and an increased likeli-hood of remission, and will eventually fulfil the ‘promise of personalized medicine’.

1. Jones, S. A. Directing transition from innate to acquired immunity: defining a role for IL-6. J. Immunol. 175, 3463–3468 (2005).

2. Buckley, C. D. Why does chronic inflammation persist: An unexpected role for fibroblasts. Immunol. Lett. 138, 12–14 (2011).

3. McInnes, I. B. & Schett, G. Cytokines in the pathogenesis of rheumatoid arthritis. Nature Rev. Immunol. 7, 429–442 (2007).

4. Choy, E. H., Kavanaugh, A. F. & Jones, S. A. The problem of choice: current biologic agents and future prospects in RA. Nature Rev. Rheumatol. 9, 154–163 (2013).

5. Jones, S. A., Scheller, J. & Rose-John, S. Therapeutic strategies for the clinical blockade of IL-6/gp130 signaling. J. Clin. Invest. 121, 3375–3383 (2011).

6. Raza, K. The Michael Mason prize: early rheumatoid arthritis—the window narrows. Rheumatology 49, 406–410 (2010).

7. Aloisi, F. & Pujol-Borrell, R. Lymphoid neogenesis in chronic inflammatory diseases. Nature Rev. Immunol. 6, 205–217 (2006).This is an excellent commentary on the development of lymphoid structures in disease.

8. Neyt, K., Perros, F., GeurtsvanKessel, C. H., Hammad, H. & Lambrecht, B. N. Tertiary lymphoid organs in infection and autoimmunity. Trends Immunol. 33, 297–305 (2012).

9. Cañete, J. D. et al. Clinical significance of synovial lymphoid neogenesis and its reversal after anti-tumour necrosis factor-α therapy in rheumatoid arthritis. Ann. Rheum. Dis. 68, 751–756 (2009).This study provides an example of how distinct synovial histopathology may influence the response to biological therapy. The presence of ELSs was associated with an inferior response to anti-TNF therapy.

10. Coppola, D. et al. Unique ectopic lymph node-like structures present in human primary colorectal carcinoma are identified by immune gene array profiling. Am. J. Pathol. 179, 37–45 (2011).

11. Drayton, D. L., Liao, S., Mounzer, R. H. & Ruddle, N. H. Lymphoid organ development: from ontogeny to neogenesis. Nature Immunol. 7, 344–353 (2006).This is an excellent review about the mechanisms controlling lymphoid neogenesis.

12. De Togni, P. et al. Abnormal development of peripheral lymphoid organs in mice deficient in lymphotoxin. Science 264, 703–707 (1994).

13. Kratz, A., Campos-Neto, A., Hanson, M. S. & Ruddle, N. H. Chronic inflammation caused by lymphotoxin is lymphoid neogenesis. J. Exp. Med. 183, 1461–1472 (1996).

14. Drayton, D. L., Ying, X., Lee, J., Lesslauer, W. & Ruddle, N. H. Ectopic LTαβ directs lymphoid organ neogenesis with concomitant expression of peripheral

node addressin and a HEV-restricted sulfotransferase. J. Exp. Med. 197, 1153–1163 (2003).

15. Okabe, Y. & Medzhitov, R. Tissue-specific signals control reversible program of localization and functional polarization of macrophages. Cell 157, 832–844 (2014).

16. Ruddle, N. H. Lymphatic vessels and tertiary lymphoid organs. J. Clin. Invest. 124, 953–959 (2014).

17. Meier, D. et al. Ectopic lymphoid-organ development occurs through interleukin 7-mediated enhanced survival of lymphoid-tissue-inducer cells. Immunity 26, 643–654 (2007).This is an early report on the role of IL-7 and LTi cells in lymphoid neogenesis.

18. Sawa, S. et al. Lineage relationship analysis of RORγt+ innate lymphoid cells. Science 330, 665–669 (2010).

19. Schmutz, S. et al. Cutting edge: IL-7 regulates the peripheral pool of adult RORγ+ lymphoid tissue inducer cells. J. Immunol. 183, 2217–2221 (2009).The study describes the role of RORγt in LTi cells and provides a potential link to the involvement of TH17-like cells in lymphoid neogenesis.

20. Luther, S. A., Ansel, K. M. & Cyster, J. G. Overlapping roles of CXCL13, interleukin 7 receptor-α, and CCR7 ligands in lymph node development. J. Exp. Med. 197, 1191–1198 (2003).

21. Yoshida, H. et al. Different cytokines induce surface lymphotoxin-αβ on IL-7 receptor-α cells that differentially engender lymph nodes and Peyer’s patches. Immunity 17, 823–833 (2002).This study suggests that unique cellular and molecular mechanisms contribute to lymphoid neogenesis in distinct anatomical sites.

22. Sato, M. et al. Stromal activation and formation of lymphoid-like stroma in chronic lung allograft dysfunction. Transplantation 91, 1398–1405 (2011).

23. Rangel-Moreno, J., Moyron-Quiroz, J. E., Hartson, L., Kusser, K. & Randall, T. D. Pulmonary expression of CXC chemokine ligand 13, CC chemokine ligand 19, and CC chemokine ligand 21 is essential for local immunity to influenza. Proc. Natl Acad. Sci. USA 104, 10577–10582 (2007).This study demonstrates the importance of homeostatic chemokines in the organization of lymphoid structures.

24. Braun, A., Takemura, S., Vallejo, A. N., Goronzy, J. J. & Weyand, C. M. Lymphotoxin β-mediated stimulation of synoviocytes in rheumatoid arthritis. Arthritis Rheum. 50, 2140–2150 (2004).

25. Takemura, S. et al. Lymphoid neogenesis in rheumatoid synovitis. J. Immunol. 167, 1072–1080 (2001).

26. Timmer, T. C. et al. Inflammation and ectopic lymphoid structures in rheumatoid arthritis synovial tissues dissected by genomics technology: identification of the

interleukin-7 signaling pathway in tissues with lymphoid neogenesis. Arthritis Rheum. 56, 2492–2502 (2007).This is an investigative study of ELS development in the context of human disease.

27. Takatori, H. et al. Lymphoid tissue inducer-like cells are an innate source of IL-17 and IL-22. J. Exp. Med. 206, 35–41 (2009).

28. Deteix, C. et al. Intragraft Th17 infiltrate promotes lymphoid neogenesis and hastens clinical chronic rejection. J. Immunol. 184, 5344–5351 (2010).

29. Peters, A. et al. Th17 cells induce ectopic lymphoid follicles in central nervous system tissue inflammation. Immunity 35, 986–996 (2011).

30. Rangel-Moreno, J. et al. The development of inducible bronchus-associated lymphoid tissue depends on IL-17. Nature Immunol. 12, 639–646 (2011).References 29 and 30 suggest a role of IL-17 and TH17-like cells in ELS development.

31. Link, A. et al. Association of T-zone reticular networks and conduits with ectopic lymphoid tissues in mice and humans. Am. J. Pathol. 178, 1662–1675 (2011).

32. Furtado, G. C. et al. TNFα-dependent development of lymphoid tissue in the absence of RORγt lymphoid tissue inducer cells. Mucosal Immunol. 7, 602–614 (2013).This study supports a role for resident myeloid cells and stromal cells in lymphoid neogenesis.

33. Lochner, M. et al. Microbiota-induced tertiary lymphoid tissues aggravate inflammatory disease in the absence of RORγt and LTi cells. J. Exp. Med. 208, 125–134 (2011).

34. Luther, S. A. et al. Differing activities of homeostatic chemokines CCL19, CCL21, and CXCL12 in lymphocyte and dendritic cell recruitment and lymphoid neogenesis. J. Immunol. 169, 424–433 (2002).

35. Luther, S. A., Lopez, T., Bai, W., Hanahan, D. & Cyster, J. G. BLC expression in pancreatic islets causes B cell recruitment and lymphotoxin-dependent lymphoid neogenesis. Immunity 12, 471–481 (2000).

36. Chen, S. C. et al. Ectopic expression of the murine chemokines CCL21a and CCL21b induces the formation of lymph node-like structures in pancreas, but not skin, of transgenic mice. J. Immunol. 168, 1001–1008 (2002).

37. Fan, L., Reilly, C. R., Luo, Y., Dorf, M. E. & Lo, D. Cutting edge: ectopic expression of the chemokine TCA4/SLC is sufficient to trigger lymphoid neogenesis. J. Immunol. 164, 3955–3959 (2000).

38. Slight, S. R. et al. CXCR5+ T helper cells mediate protective immunity against tuberculosis. J. Clin. Invest. 123, 712–726 (2013).

39. Bombardieri, M. et al. Inducible tertiary lymphoid structures, autoimmunity, and exocrine dysfunction in a novel model of salivary gland inflammation in C57BL/6 mice. J. Immunol. 189, 3767–3776 (2012).

R E V I E W S



40. Gu-Trantien, C. et al. CD4+ follicular helper T cell infiltration predicts breast cancer survival. J. Clin. Invest. 123, 2873–2892 (2013).This study uses imaging to correlate the infiltration of TFH cells with clinical outcome in patients with breast cancer.

41. Breitfeld, D. et al. Follicular B helper T cells express CXC chemokine receptor 5, localize to B cell follicles, and support immunoglobulin production. J. Exp. Med. 192, 1545–1552 (2000).

42. Schaerli, P. et al. CXC chemokine receptor 5 expression defines follicular homing T cells with B cell helper function. J. Exp. Med. 192, 1553–1562 (2000).This study describes the original identification and functional characterization of TFH cells.

43. Nakayamada, S. et al. Early Th1 cell differentiation is marked by a Tfh cell-like transition. Immunity 35, 919–931 (2011).

44. Lu, K. T. et al. Functional and epigenetic studies reveal multistep differentiation and plasticity of in vitro-generated and in vivo-derived follicular T helper cells. Immunity 35, 622–632 (2011).

45. Fahey, L. M. et al. Viral persistence redirects CD4 T cell differentiation toward T follicular helper cells. J. Exp. Med. 208, 987–999 (2011).

46. King, I. L. & Mohrs, M. IL-4–producing CD4+ T cells in reactive lymph nodes during helminth infection are T follicular helper cells. J. Exp. Med. 206, 1001–1007 (2009).

47. Glatman Zaretsky, A. et al. T follicular helper cells differentiate from Th2 cells in response to helminth antigens. J. Exp. Med. 206, 991–999 (2009).

48. Bauquet, A. T. et al. The costimulatory molecule ICOS regulates the expression of c-Maf and IL-21 in the development of follicular T helper cells and TH17 cells. Nature Immunol. 10, 167–175 (2009).

49. Huber, M. et al. IRF4 is essential for IL-21-mediated induction, amplification, and stabilization of the Th17 phenotype. Proc. Natl Acad. Sci. USA 105, 20846–20851 (2008).

50. Ma, C. S. et al. Functional STAT3 deficiency compromises the generation of human T follicular helper cells. Blood 119, 3997–4008 (2012).

51. Ma, C. S. et al. Deficiency of Th17 cells in hyper IgE syndrome due to mutations in STAT3. J. Exp. Med. 205, 1551–1557 (2008).

52. Mitsdoerffer, M. et al. Proinflammatory T helper type 17 cells are effective B-cell helpers. Proc. Natl Acad. Sci. USA 107, 14292–14297 (2010).

53. Nurieva, R. I. et al. Generation of T follicular helper cells is mediated by interleukin-21 but independent of T helper 1, 2, or 17 cell lineages. Immunity 29, 138–149 (2008).

54. Manzo, A. et al. Mature antigen-experienced T helper cells synthesize and secrete the B cell chemoattractant CXCL13 in the inflammatory environment of the rheumatoid joint. Arthritis Rheum. 58, 3377–3387 (2008).

55. Tokoyoda, K. et al. Professional memory CD4+ T lymphocytes preferentially reside and rest in the bone marrow. Immunity 30, 721–730 (2009).

56. Odegard, J. M. et al. ICOS-dependent extrafollicular helper T cells elicit IgG production via IL-21 in systemic autoimmunity. J. Exp. Med. 205, 2873–2886 (2008).

57. Marinkovic, T. et al. Interaction of mature CD3+CD4+ T cells with dendritic cells triggers the development of tertiary lymphoid structures in the thyroid. J. Clin. Invest. 116, 2622–2632 (2006).

58. Muniz, L. R., Pacer, M. E., Lira, S. A. & Furtado, G. C. A critical role for dendritic cells in the formation of lymphatic vessels within tertiary lymphoid structures. J. Immunol. 187, 828–834 (2011).

59. GeurtsvanKessel, C. H. et al. Dendritic cells are crucial for maintenance of tertiary lymphoid structures in the lung of influenza virus-infected mice. J. Exp. Med. 206, 2339–2349 (2009).

60. Halle, S. et al. Induced bronchus-associated lymphoid tissue serves as a general priming site for T cells and is maintained by dendritic cells. J. Exp. Med. 206, 2593–2601 (2009).

61. Manser, T. Textbook germinal centers? J. Immunol. 172, 3369–3375 (2004).

62. Teng, G. & Papavasiliou, F. N. Immunoglobulin somatic hypermutation. Annu. Rev. Genet. 41, 107–120 (2007).

63. Stavnezer, J., Guikema, J. E. & Schrader, C. E. Mechanism and regulation of class switch recombination. Annu. Rev. Immunol. 26, 261–292 (2008).

64. Muramatsu, M. et al. Class switch recombination and hypermutation require activation-induced cytidine deaminase (AID), a potential RNA editing enzyme. Cell 102, 553–563 (2000).

65. Barreto, V., Reina-San-Martin, B., Ramiro, A. R., McBride, K. M. & Nussenzweig, M. C. C-terminal deletion of AID uncouples class switch recombination from somatic hypermutation and gene conversion. Mol. Cell 12, 501–508 (2003).

66. Bombardieri, M. et al. Activation-induced cytidine deaminase expression in follicular dendritic cell networks and interfollicular large B cells supports functionality of ectopic lymphoid neogenesis in autoimmune sialoadenitis and MALT lymphoma in Sjogren’s syndrome. J. Immunol. 179, 4929–4938 (2007).

67. Humby, F. et al. Ectopic lymphoid structures support ongoing production of class-switched autoantibodies in rheumatoid synovium. PLoS Med. 6, e1 (2009).This study describes the functional properties of ELSs in patients with rheumatoid arthritis.

68. Nacionales, D. C. et al. B cell proliferation, somatic hypermutation, class switch recombination, and autoantibody production in ectopic lymphoid tissue in murine lupus. J. Immunol. 182, 4226–4236 (2009).

69. Serafini, B. et al. Dysregulated Epstein–Barr virus infection in the multiple sclerosis brain. J. Exp. Med. 204, 2899–2912 (2007).

70. Takahashi, E. et al. Oral clarithromycin enhances airway immunoglobulin A (IgA) immunity through induction of IgA class switching recombination and B-cell-activating factor of the tumor necrosis factor family molecule on mucosal dendritic cells in mice infected with influenza A virus. J. Virol. 86, 10924–10934 (2012).

71. Cheng, J. et al. Ectopic B-cell clusters that infiltrate transplanted human kidneys are clonal. Proc. Natl Acad. Sci. USA 108, 5560–5565 (2011).

72. Scheel, T., Gursche, A., Zacher, J., Haupl, T. & Berek, C. V-region gene analysis of locally defined synovial B and plasma cells reveals selected B cell expansion and accumulation of plasma cell clones in rheumatoid arthritis. Arthritis Rheum. 63, 63–72 (2011).This study provides a functional evaluation of ELSs in patients with rheumatoid arthritis.

73. Stott, D. I., Hiepe, F., Hummel, M., Steinhauser, G. & Berek, C. Antigen-driven clonal proliferation of B cells within the target tissue of an autoimmune disease. The salivary glands of patients with Sjogren’s syndrome. J. Clin. Invest. 102, 938–946 (1998).

74. Grewal, J. S. et al. Salivary glands act as mucosal inductive sites via the formation of ectopic germinal centers after site-restricted MCMV infection. FASEB J. 25, 1680–1696 (2011).

75. Khader, S. A. et al. In a murine tuberculosis model, the absence of homeostatic chemokines delays granuloma formation and protective immunity. J. Immunol. 183, 8004–8014 (2009).

76. Mazzucchelli, L. et al. BCA-1 is highly expressed in Helicobacter pylori-induced mucosa-associated lymphoid tissue and gastric lymphoma. J. Clin. Invest. 104, R49–R54 (1999).

77. Mosnier, J. F. et al. The intraportal lymphoid nodule and its environment in chronic active hepatitis C: an immunohistochemical study. Hepatology 17, 366–371 (1993).

78. Ulrichs, T. et al. Human tuberculous granulomas induce peripheral lymphoid follicle-like structures to orchestrate local host defence in the lung. J. Pathol. 204, 217–228 (2004).

79. Moyron-Quiroz, J. E. et al. Role of inducible bronchus associated lymphoid tissue (iBALT) in respiratory immunity. Nature Med. 10, 927–934 (2004).

80. Xu, B. et al. Lymphocyte homing to bronchus-associated lymphoid tissue (BALT) is mediated by L-selectin/PNAd, α4β1 integrin/VCAM-1, and LFA-1 adhesion pathways. J. Exp. Med. 197, 1255–1267 (2003).

81. Moyron-Quiroz, J. E. et al. Persistence and responsiveness of immunologic memory in the absence of secondary lymphoid organs. Immunity 25, 643–654 (2006).

82. Manzo, A., Bombardieri, M., Humby, F. & Pitzalis, C. Secondary and ectopic lymphoid tissue responses in rheumatoid arthritis: from inflammation to autoimmunity and tissue damage/remodeling. Immunol. Rev. 233, 267–285 (2010).

83. Bombardieri, M. & Pitzalis, C. Ectopic lymphoid neogenesis and lymphoid chemokines in Sjogren’s

syndrome: at the interplay between chronic inflammation, autoimmunity and lymphomagenesis. Curr. Pharm. Biotechnol. 13, 1989–1996 (2012).

84. Berrih-Aknin, S., Ragheb, S., Le Panse, R. & Lisak, R. P. Ectopic germinal centers, BAFF and anti-B-cell therapy in myasthenia gravis. Autoimmun Rev. 12, 885–893 (2013).

85. Armengol, M. P. et al. Thyroid autoimmune disease: demonstration of thyroid antigen-specific B cells and recombination-activating gene expression in chemokine-containing active intrathyroidal germinal centers. Am. J. Pathol. 159, 861–873 (2001).

86. Ekland, E. H., Forster, R., Lipp, M. & Cyster, J. G. Requirements for follicular exclusion and competitive elimination of autoantigen-binding B cells. J. Immunol. 172, 4700–4708 (2004).

87. Le Pottier, L. et al. Ectopic germinal centers are rare in Sjogren’s syndrome salivary glands and do not exclude autoreactive B cells. J. Immunol. 182, 3540–3547 (2009).

88. Salomonsson, S. et al. Cellular basis of ectopic germinal center formation and autoantibody production in the target organ of patients with Sjogren’s syndrome. Arthritis Rheum. 48, 3187–3201 (2003).

89. Shimaoka, Y. et al. Nurse-like cells from bone marrow and synovium of patients with rheumatoid arthritis promote survival and enhance function of human B cells. J. Clin. Invest. 102, 606–618 (1998).

90. Pender, M. P. Infection of autoreactive B lymphocytes with EBV, causing chronic autoimmune diseases. Trends Immunol. 24, 584–588 (2003).

91. Cavalcante, P. et al. Epstein–Barr virus persistence and reactivation in myasthenia gravis thymus. Ann. Neurol. 67, 726–738 (2010).

92. Croia, C. et al. Epstein–Barr virus persistence and infection of autoreactive plasma cells in synovial lymphoid structures in rheumatoid arthritis. Ann. Rheum. Dis. 72, 1559–1568 (2013).This study provides an example of how viral infection may influence the development and functional properties of ELSs in autoimmune disease.

93. Croia, C. et al. Implication of Epstein–Barr virus infection in disease-specific autoreactive B cell activation in ectopic lymphoid structures of Sjogren’s syndrome. Arthritis Rheum. http://dx.doi.org/10.1002/art.38726 (2014).

94. Schonbeck, S., Padberg, F., Hohlfeld, R. & Wekerle, H. Transplantation of thymic autoimmune microenvironment to severe combined immunodeficiency mice. A new model of myasthenia gravis. J. Clin. Invest. 90, 245–250 (1992).

95. Thaunat, O. Pathophysiologic significance of B-cell clusters in chronically rejected grafts. Transplantation 92, 121–126 (2011).

96. Thaunat, O. et al. Chronic rejection triggers the development of an aggressive intragraft immune response through recapitulation of lymphoid organogenesis. J. Immunol. 185, 717–728 (2010).This study provides an example of how the development of severe inflammation following chronic transplant rejection is accompanied by lymphoid neogenesis.

97. Brown, K., Sacks, S. H. & Wong, W. Tertiary lymphoid organs in renal allografts can be associated with donor-specific tolerance rather than rejection. Eur. J. Immunol. 41, 89–96 (2011).

98. Le Texier, L. et al. Long-term allograft tolerance is characterized by the accumulation of B cells exhibiting an inhibited profile. Am. J. Transplant. 11, 429–438 (2011).

99. Dieu-Nosjean, M. C. et al. Long-term survival for patients with non-small-cell lung cancer with intratumoral lymphoid structures. J. Clin. Oncol. 26, 4410–4417 (2008).

100. de Chaisemartin, L. et al. Characterization of chemokines and adhesion molecules associated with T cell presence in tertiary lymphoid structures in human lung cancer. Cancer Res. 71, 6391–6399 (2011).

101. Bergomas, F. et al. Tertiary intratumor lymphoid tissue in colo-rectal cancer. Cancers 4, 1–10 (2011).

102. Nzula, S., Going, J. J. & Stott, D. I. Antigen-driven clonal proliferation, somatic hypermutation, and selection of B lymphocytes infiltrating human ductal breast carcinomas. Cancer Res. 63, 3275–3280 (2003).

103. Martinet, L. et al. Human solid tumors contain high endothelial venules: association with T- and B-lymphocyte infiltration and favorable prognosis in breast cancer. Cancer Res. 71, 5678–5687 (2011).

R E V I E W S



104. Goc, J., Fridman, W. H., Sautes-Fridman, C. & Dieu-Nosjean, M. C. Characteristics of tertiary lymphoid structures in primary cancers. Oncoimmunology 2, e26836 (2013).

105. Hamanishi, J. et al. Activated local immunity by CC chemokine ligand 19-transduced embryonic endothelial progenitor cells suppresses metastasis of murine ovarian cancer. Stem Cells 28, 164–173 (2010).

106. Liu, Y. et al. A genome-wide association study of psoriasis and psoriatic arthritis identifies new disease loci. PLoS Genet. 4, e1000041 (2008).

107. Maiti, A. K. et al. Confirmation of an association between rs6822844 at the Il2–Il21 region and multiple autoimmune diseases: evidence of a general susceptibility locus. Arthritis Rheum. 62, 323–329 (2010).

108. van Heel, D. A. et al. A genome-wide association study for celiac disease identifies risk variants in the region harboring IL2 and IL21. Nature Genet. 39, 827–829 (2007).

109. Jones, J. L. et al. IL-21 drives secondary autoimmunity in patients with multiple sclerosis, following therapeutic lymphocyte depletion with alemtuzumab (Campath-1H). J. Clin. Invest. 119, 2052–2061 (2009).This study provides an example of how clinical intervention aids the mechanistic understanding of ELS development in the context of disease.

110. Mells, G. F. et al. Genome-wide association study identifies 12 new susceptibility loci for primary biliary cirrhosis. Nature Genet. 43, 329–332 (2011).

111. Lessard, C. J. et al. Variants at multiple loci implicated in both innate and adaptive immune responses are associated with Sjogren’s syndrome. Nature Genet. 45, 1284–1292 (2013).

112. Zhang, J. et al. Three SNPs in chromosome 11q23.3 are independently associated with systemic lupus erythematosus in Asians. Hum. Mol. Genet. 23, 524–533 (2014).

113. Lill, C. M. et al. MANBA, CXCR5, SOX8, RPS6KB1 and ZBTB46 are genetic risk loci for multiple sclerosis. Brain 136, 1778–1782 (2013).

114. Hislop, A. D., Taylor, G. S., Sauce, D. & Rickinson, A. B. Cellular responses to viral infection in humans: lessons from Epstein–Barr virus. Annu. Rev. Immunol. 25, 587–617 (2007).

115. Thorley-Lawson, D. A. Epstein–Barr virus: exploiting the immune system. Nature Rev. Immunol. 1, 75–82 (2001).

116. Pitzalis, C., Kelly, S. & Humby, F. New learnings on the pathophysiology of RA from synovial biopsies. Curr. Opin. Rheumatol. 25, 334–344 (2013).

117. van de Sande, M. G. et al. Presence of lymphocyte aggregates in the synovium of patients with early arthritis in relationship to diagnosis and outcome: is it a constant feature over time? Ann. Rheum. Dis. 70, 700–703 (2011).

118. Ulrichs, T. et al. Differential organization of the local immune response in patients with active cavitary tuberculosis or with nonprogressive tuberculoma. J. Infect. Dis. 192, 89–97 (2005).

119. Shoenfeld, Y. & Isenberg, D. A. Mycobacteria and autoimmunity. Immunol. Today 9, 178–182 (1988).

120. Sansonno, D. et al. Increased serum levels of the chemokine CXCL13 and up-regulation of its gene expression are distinctive features of HCV-related cryoglobulinemia and correlate with active cutaneous vasculitis. Blood 112, 1620–1627 (2008).

121. Sansonno, D. et al. Clonal analysis of intrahepatic B cells from HCV-infected patients with and without mixed cryoglobulinemia. J. Immunol. 160, 3594–3601 (1998).

122. Ramos-Casals, M., De Vita, S. & Tzioufas, A. G. Hepatitis C virus, Sjogren’s syndrome and B-cell lymphoma: linking infection, autoimmunity and cancer. Autoimmun Rev. 4, 8–15 (2005).

123. Deutsch, A. J. et al. MALT lymphoma and extranodal diffuse large B-cell lymphoma are targeted by aberrant somatic hypermutation. Blood 109, 3500–3504 (2007).

124. Hussell, T., Isaacson, P. G., Crabtree, J. E. & Spencer, J. Helicobacter pylori-specific tumour-infiltrating T cells provide contact dependent help for the growth of malignant B cells in low-grade gastric lymphoma of mucosa-associated lymphoid tissue. J. Pathol. 178, 122–127 (1996).

125. Wohrer, S., Troch, M. & Raderer, M. Therapy of gastric mucosa-associated lymphoid tissue lymphoma. Expert Opin. Pharmacother. 8, 1263–1273 (2007).

126. Bende, R. J. et al. Among B cell non-Hodgkin’s lymphomas, MALT lymphomas express a unique antibody repertoire with frequent rheumatoid factor reactivity. J. Exp. Med. 201, 1229–1241 (2005).

127. Klimiuk, P. A. et al. Circulating tumour necrosis factor-α and soluble tumour necrosis factor receptors in patients with different patterns of rheumatoid synovitis. Ann. Rheum. Dis. 62, 472–475 (2003).

128. Kotake, S. et al. Activated human T cells directly induce osteoclastogenesis from human monocytes: possible role of T cells in bone destruction in rheumatoid arthritis patients. Arthritis Rheum. 44, 1003–1012 (2001).

129. Dharmapatni, A. A. et al. TWEAK and Fn14 expression in the pathogenesis of joint inflammation and bone erosion in rheumatoid arthritis. Arthritis Res. Ther. 13, R51 (2011).

130. Chang, A. et al. In situ B cell-mediated immune responses and tubulointerstitial inflammation in human lupus nephritis. J. Immunol. 186, 1849–1860 (2011).

131. Magliozzi, R. et al. A Gradient of neuronal loss and meningeal inflammation in multiple sclerosis. Ann. Neurol. 68, 477–493 (2010).

132. Coronella-Wood, J. A. & Hersh, E. M. Naturally occurring B-cell responses to breast cancer. Cancer Immunol. Immunother. 52, 715–738 (2003).

133. Denkert, C. et al. Tumor-associated lymphocytes as an independent predictor of response to neoadjuvant chemotherapy in breast cancer. J. Clin. Oncol. 28, 105–113 (2010).

134. Krell, J., Frampton, A. E. & Stebbing, J. The clinical significance of tumor infiltrating lymphoctyes in breast cancer: does subtype matter? BMC Cancer 12, 135 (2012).

135. Liu, W., Peng, Y. & Tobin, D. J. A new 12-gene diagnostic biomarker signature of melanoma revealed by integrated microarray analysis. PeerJ 1, e49 (2013).

136. Jonsson, G. et al. Gene expression profiling-based identification of molecular subtypes in stage IV melanomas with different clinical outcome. Clin. Cancer Res. 16, 3356–3367 (2010).

137. DiLillo, D. J., Yanaba, K. & Tedder, T. F. B cells are required for optimal CD4+ and CD8+ T cell tumor immunity: therapeutic B cell depletion enhances B16 melanoma growth in mice. J. Immunol. 184, 4006–4016 (2010).

138. Shiao, S. L., Ganesan, A. P., Rugo, H. S. & Coussens, L. M. Immune microenvironments in solid tumors: new targets for therapy. Genes Dev. 25, 2559–2572 (2011).

139. Chiavolini, D. et al. Bronchus-associated lymphoid tissue (BALT) and survival in a vaccine mouse model of tularemia. PLoS ONE 5, e11156 (2010).

140. Thaunat, O. et al. B cell survival in intragraft tertiary lymphoid organs after rituximab therapy. Transplantation 85, 1648–1653 (2008).

141. Badot, V. et al. Gene expression profiling in the synovium identifies a predictive signature of absence of response to adalimumab therapy in rheumatoid arthritis. Arthritis Res. Ther. 11, R57 (2009).

142. Fava, R. A. et al. A role for the lymphotoxin/LIGHT axis in the pathogenesis of murine collagen-induced arthritis. J. Immunol. 171, 115–126 (2003).

143. Gatumu, M. K. et al. Blockade of lymphotoxin-beta receptor signaling reduces aspects of Sjogren’s syndrome in salivary glands of non-obese diabetic mice. Arthritis Res. Ther. 11, R24 (2009).

144. Lee, Y. et al. Recruitment and activation of naive T cells in the islets by lymphotoxin-β receptor-dependent tertiary lymphoid structure. Immunity 25, 499–509 (2006).

145. Emu, B. et al. Safety, pharmacokinetics, and biologic activity of pateclizumab, a novel monoclonal antibody targeting lymphotoxin-α: results of a phase I randomized, placebo-controlled trial. Arthritis Res. Ther. 14, R6 (2012).

146. Zheng, B. et al. CXCL13 neutralization reduces the severity of collagen-induced arthritis. Arthritis Rheum. 52, 620–626 (2005).

147. Kramer, J. M., Klimatcheva, E. & Rothstein, T. L. CXCL13 is elevated in Sjogren’s syndrome in mice and humans and is implicated in disease pathogenesis. J. Leukoc. Biol. 94, 1079–1089 (2013).

148. Henry, R. A. & Kendall, P. L. CXCL13 blockade disrupts B lymphocyte organization in tertiary lymphoid structures without altering B cell receptor bias or preventing diabetes in nonobese diabetic mice. J. Immunol. 185, 1460–1465 (2010).

149. Pelletier, D. & Hafler, D. A. Fingolimod for multiple sclerosis. N. Engl. J. Med. 366, 339–347 (2012).

150. Mehling, M. et al. FTY720 therapy exerts differential effects on T cell subsets in multiple sclerosis. Neurology 71, 1261–1267 (2008).

151. Young, D. A. et al. Blockade of the interleukin-21/interleukin-21 receptor pathway ameliorates disease in animal models of rheumatoid arthritis. Arthritis Rheum. 56, 1152–1163 (2007).

152. Herber, D. et al. IL-21 has a pathogenic role in a lupus-prone mouse model and its blockade with IL-21R.Fc reduces disease progression. J. Immunol. 178, 3822–3830 (2007).

153. Frey, O. et al. Inducible costimulator (ICOS) blockade inhibits accumulation of polyfunctional T helper 1/T helper 17 cells and mitigates autoimmune arthritis. Ann. Rheum. Dis. 69, 1495–1501 (2010).

154. Hu, Y. L., Metz, D. P., Chung, J., Siu, G. & Zhang, M. B7RP-1 blockade ameliorates autoimmunity through regulation of follicular helper T cells. J. Immunol. 182, 1421–1428 (2009).

155. Nanji, S. A. et al. Costimulation blockade of both inducible costimulator and CD40 ligand induces dominant tolerance to islet allografts and prevents spontaneous autoimmune diabetes in the NOD mouse. Diabetes 55, 27–33 (2006).

156. Wengner, A. M. et al. CXCR5- and CCR7-dependent lymphoid neogenesis in a murine model of chronic antigen-induced arthritis. Arthritis Rheum. 56, 3271–3283 (2007).

157. Lee, B. H., Carcamo, W. C., Chiorini, J. A., Peck, A. B. & Nguyen, C. Q. Gene therapy using IL-27 ameliorates Sjogren’s syndrome-like autoimmune exocrinopathy. Arthritis Res. Ther. 14, R172 (2012).

158. Magliozzi, R. et al. Meningeal B-cell follicles in secondary progressive multiple sclerosis associate with early onset of disease and severe cortical pathology. Brain 130, 1089–1104 (2007).

159. Ludewig, B., Odermatt, B., Landmann, S., Hengartner, H. & Zinkernagel, R. M. Dendritic cells induce autoimmune diabetes and maintain disease via de novo formation of local lymphoid tissue. J. Exp. Med. 188, 1493–1501 (1998).

160. Astorri, E. et al. Evolution of ectopic lymphoid neogenesis and in situ autoantibody production in autoimmune nonobese diabetic mice: cellular and molecular characterization of tertiary lymphoid structures in pancreatic islets. J. Immunol. 185, 3359–3368 (2010).

161. Sharifi, S., Murphy, M., Loda, M., Pinkus, G. S. & Khettry, U. Nodular lymphoid lesion of the liver: an immune-mediated disorder mimicking low-grade malignant lymphoma. Am. J. Surg. Pathol. 23, 302–308 (1999).

162. Grant, A. J., Lalor, P. F., Hubscher, S. G., Briskin, M. & Adams, D. H. MAdCAM-1 expressed in chronic inflammatory liver disease supports mucosal lymphocyte adhesion to hepatic endothelium (MAdCAM-1 in chronic inflammatory liver disease). Hepatology 33, 1065–1072 (2001).

163. Grant, A. J. et al. Hepatic expression of secondary lymphoid chemokine (CCL21) promotes the development of portal-associated lymphoid tissue in chronic inflammatory liver disease. Am. J. Pathol. 160, 1445–1455 (2002).

164. Hill, M. E., Shiono, H., Newsom-Davis, J. & Willcox, N. The myasthenia gravis thymus: a rare source of human autoantibody-secreting plasma cells for testing potential therapeutics. J. Neuroimmunol. 201–202, 50–56 (2008).

165. Nacionales, D. C. et al. Type I interferon production by tertiary lymphoid tissue developing in response to 2,6,10,14-tetramethyl-pentadecane (pristane). Am. J. Pathol. 168, 1227–1240 (2006).

166. Houtkamp, M. A., de Boer, O. J., van der Loos, C. M., van der Wal, A. C. & Becker, A. E. Adventitial infiltrates associated with advanced atherosclerotic plaques: structural organization suggests generation of local humoral immune responses. J. Pathol. 193, 263–269 (2001).

167. Grabner, R. et al. Lymphotoxin-β receptor signaling promotes tertiary lymphoid organogenesis in the aorta adventitia of aged ApoE−/− mice. J. Exp. Med. 206, 233–248 (2009).

168. Carlsen, H. S., Baekkevold, E. S., Johansen, F. E., Haraldsen, G. & Brandtzaeg, P. B cell attracting chemokine 1 (CXCL13) and its receptor CXCR5 are expressed in normal and aberrant gut associated lymphoid tissue. Gut 51, 364–371 (2002).

R E V I E W S



169. Weninger, W. et al. Naive T cell recruitment to nonlymphoid tissues: a role for endothelium-expressed CC chemokine ligand 21 in autoimmune disease and lymphoid neogenesis. J. Immunol. 170, 4638–4648 (2003).

170. Surawicz, C. M. & Belic, L. Rectal biopsy helps to distinguish acute self-limited colitis from idiopathic inflammatory bowel disease. Gastroenterology 86, 104–113 (1984).

171. Kaiserling, E. Newly-formed lymph nodes in the submucosa in chronic inflammatory bowel disease. Lymphology 34, 22–29 (2001).

172. McNamee, E. N. et al. Ectopic lymphoid tissue alters the chemokine gradient, increases lymphocyte retention and exacerbates murine ileitis. Gut 62, 53–62 (2013).

173. Demoor, T. et al. Role of lymphotoxin-α in cigarette smoke-induced inflammation and lymphoid neogenesis. Eur. Respir. J. 34, 405–416 (2009).

174. Hogg, J. C. et al. The nature of small-airway obstruction in chronic obstructive pulmonary disease. N. Engl. J. Med. 350, 2645–2653 (2004).

175. Van Pottelberge, G. R. et al. Plasmacytoid dendritic cells in pulmonary lymphoid follicles of patients with COPD. Eur. Respir. J. 36, 781–791 (2010).

176. Nielsen, J. S. et al. CD20+ tumor-infiltrating lymphocytes have an atypical CD27– memory phenotype and together with CD8+ T cells promote favorable prognosis in ovarian cancer. Clin. Cancer Res. 18, 3281–3292 (2012).

177. Curiel, T. J. et al. Specific recruitment of regulatory T cells in ovarian carcinoma fosters immune privilege and predicts reduced survival. Nature Med. 10, 942–949 (2004).

178. Eisenthal, A. et al. Expression of dendritic cells in ovarian tumors correlates with clinical outcome in patients with ovarian cancer. Hum. Pathol. 32, 803–807 (2001).

179. Yeung, M. M. et al. Characterisation of mucosal lymphoid aggregates in ulcerative colitis: immune cell phenotype and TcR-γδ expression. Gut 47, 215–227 (2000).

180. Winter, S. et al. The chemokine receptor CXCR5 is pivotal for ectopic mucosa-associated lymphoid tissue neogenesis in chronic Helicobacter pylori-induced inflammation. J. Mol. Med. 88, 1169–1180 (2010).

181. Shomer, N. H., Fox, J. G., Juedes, A. E. & Ruddle, N. H. Helicobacter-induced chronic active lymphoid aggregates have characteristics of tertiary lymphoid tissue. Infect. Immun. 71, 3572–3577 (2003).

182. Ghosh, S., Steere, A. C., Stollar, B. D. & Huber, B. T. In situ diversification of the antibody repertoire in chronic Lyme arthritis synovium. J. Immunol. 174, 2860–2869 (2005).

183. Rangel-Moreno, J. et al. Omental milky spots develop in the absence of lymphoid tissue-inducer cells and support B and T cell responses to peritoneal antigens. Immunity 30, 731–743 (2009).

184. Beelen, R. H., Oosterling, S. J., van Egmond, M., van den Born, J. & Zareie, M. Omental milky spots in peritoneal pathophysiology (spots before your eyes). Perit. Dial. Int. 25, 30–32 (2005).

185. Di Paolo, N. et al. Omental milky spots and peritoneal dialysis — review and personal experience. Perit. Dial. Int. 25, 48–57 (2005).

186. Hiramatsu, K. et al. Inhalation of diesel exhaust for three months affects major cytokine expression and induces bronchus-associated lymphoid tissue formation in murine lungs. Exp. Lung Res. 29, 607–622 (2003).

187. Mahendra, G. et al. Necrotic and inflammatory changes in metal-on-metal resurfacing hip arthroplasties. Acta Orthop. 80, 653–659 (2009).

188. Perros, F. et al. Pulmonary lymphoid neogenesis in idiopathic pulmonary arterial hypertension. Am. J. Respir. Crit. Care Med. 185, 311–321 (2012).

AcknowledgementsG.W.J. is funded by Arthritis Research UK as a non-clinical Career Development Fellow.

Competing interests statement The authors declare no competing interests.

FURTHER INFORMATIONClinicalTrials.gov: http://www.clinicaltrials.gov

ALL LINKS ARE ACTIVE IN THE ONLINE PDF

R E V I E W S



http://www.clinicaltrials.gov

ectopic lymphoid-like structures in infection, cancer and ......jun 20, 2014 · infection, cancer...

Documents