Download - Let's go mucosal: communication on slippery ground

Let’s go mucosal: communication onslippery groundPer Brandtzaeg1 and Reinhard Pabst2

1Laboratory for Immunohistochemistry and Immunopathology (LIIPAT), Institute of Pathology, Rikshospitalet University Hospital,

N-0027 Oslo, Norway2Department of Functional and Applied Anatomy, Medical School of Hannover, D-30625 Hannover, Germany

Mucosal immunology is increasingly gaining attention

as an area of great potential for the development of

vaccines and immunotherapy. However, some immu-

nologists confuse this field by neglecting recommended

and well-defined terminology. Although nomenclature

might seem unimportant to progressive scientists, it is a

crucial means of communication and is essential for

those being introduced to a new research topic. The

mucosal immune system is even more complex than its

systemic counterpart, both in terms of effectors and

anatomy. It is instructive to divide the various tissue

compartments involved in mucosal immunity into

inductive sites and effector sites according to their

main function. Inductive sites comprise lymphoid tissue,

in which the triggering of naıve immune cells and the

generation of memory–effector cells take place.

The vast majority of exogenous challenges confronting thebody, including microbes and soluble antigens, makecontact with mucosal surfaces rather than the skin. Tomaintain homeostasis in the extensive and vulnerablemucosae, they are protected by specialized anti-inflam-matory immune defenses such as the formation and exportof secretory IgA (SIgA) antibodies and induction oftolerance against innocuous soluble substances, as wellas commensal bacteria. Another characteristic of themucosal immune system is its ability to perform activeantigen sampling from mucosal surfaces, whereas induc-tion of systemic immunity depends on antigenic supply tothe lymph nodes (LNs) and spleen via afferent lymph andperipheral blood, respectively [1]. To carry out all of itsfunctions, the mucosal immune system relies on extra-ordinary complex and dynamic mechanisms involvingmany structurally different tissue compartments(Figure 1). It would be conducive to the communicationamong researchers interested in this field to agree on auniform and functionally justifiable nomenclature whichaccords with classical anatomy terminology.

Mucosally induced tolerance has been studied for 100years, particularly in relation to the gut (‘oral tolerance’),but the first glimpses of a solid mechanistic understandinghave only been obtained quite recently [2,3]. Conversely,the molecular specialization of the SIgA system wasdescribed as far back as in 1965 [2,4], and a common

Corresponding author: Per Brandtzaeg ([email protected]).Available online 22 September 2004

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epithelial transport model for dimers (and larger poly-mers) of IgA and pentamers of IgM – which later inprinciple turned out to be true –was proposed in 1974 [5–7].The epithelial glycoprotein designated as the secretorycomponent (SC) by the WHO in 1972 (previously called‘transport piece’) was later shown to be responsible for theexport of J chain-containing Ig polymers [7,8], and thefunctional name polymeric Ig receptor (pIgR) is nowcommonly used for the transmembrane form of SC. Thecleaved ectodomain of pIgR is incorporated into SIgA andsecretory IgM (SIgM) as so-called bound SC, which isstructurally similar to free SC derived from unoccupiedreceptor. This terminology (Table 1) was approved by theSociety for Mucosal Immunology in 1997 and also by theInternational Union of Immunological Societies–WorldHealth Organization (IUIS–WHO) Nomenclature Com-mittee [9]. However, many scientists and editors areapparently unaware of this international recommen-dation, and continue to use lower-case ‘s’ as a prefix forthe SIgs. It is also often neglected to specify SC as beingeither bound or free in distinction from membrane SC.Further confusion arises from the fact that no attempthas been made to standardize the terminology appli-cable to the various tissue compartments involved inmucosal immunity.

This article aims to improve communication about theanatomy of the mucosal immune system, and recommendsa nomenclature that harmonizes with its compartmenta-lized functional specialization as well as the classicalconcepts of histology.

Mucosal immunity is elicited in inductive lymphoid

tissue

The mucosal immune system can principally be dividedinto inductive sites, where antigens sampled frommucosalsurfaces stimulate cognate naıve T and B lymphocytes,and effector sites, where the effector cells after extravasa-tion and differentiation get to work, for instance tocontribute to the formation of SIgA antibodies. Inductivesites for mucosal immunity consist of organized mucosa-associated lymphoid tissue (MALT) as well as local andregional draining LNs, whereas the effector sites consist ofdistinctly different histological compartments, includingthe lamina propria (LP) of various mucosae, the stroma ofexocrine glands and surface epithelia (Figures 1–3).Peyer’s patches (PPs) in the distal small intestine of

Opinion TRENDS in Immunology Vol.25 No.11 November 2004

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M M M

TRENDS in Immunology

M cell

SIgA

Mucosal inductive site

Organized MALT

Endothelialgatekeeperfunction

Mucosal effector sites

HEV

α/β

α/βDC

Local lymph node

Peripheral blood

Peyer’s patches

Appendix

Isolated lymphoid follicles

Waldeyer’s ring (NALT)

APC

Antigen Antigen SIgM (IgG)

Naïvecells

Lymphvessels

FDC

pIgR(SC)

CD8

CD4

APC

IgM+J IgA+J

IgA+J

γ/δ α/β

IgG(J)

Figure 1. Depiction of the human mucosal immune system. Inductive sites for mucosal T and B cells are constituted by regional mucosa-associated lymphoid tissue (MALT)

with their B-cell follicles andM cell (M)-containing follicle-associated epithelium throughwhich exogenous antigens are actively transported to reach antigen-presenting cells

(APCs), including dendritic cells (DCs), macrophages, B cells and follicular dendritic cells (FDCs). In addition, quiescent intra- or subepithelial DCsmight capture antigens at an

effector site (exemplified by nasal mucosa in the centre of the figure) and migrate via draining lymph to local and regional lymph nodes, where they become active APCs,

which stimulate T cells for productive or downregulatory (suppressive) immune responses. Naıve B and T cells enter MALT (and lymph nodes) via high endothelial venules

(HEVs). After being primed to become memory–effector B and T cells, they migrate from MALT and lymph nodes to the peripheral blood for subsequent extravasation at

mucosal effector site (exemplified by the gut mucosa on the right-hand side of the figure). This process is directed by the local profile of vascular adhesion molecules and

chemokines, the endothelial cells thus exerting a local ‘gatekeeper’ function for mucosal immunity. The gut lamina propria contains B lymphocytes, J-chain-expressing IgA

and IgM plasma cells, IgG plasma cells with a variable J-chain level (J), and CD4C T cells. Additional features are the generation of secretory IgA (SIgA) and secretory IgM

(SIgM) via polymeric Ig receptor (pIgR; called also membrane secretory component; SC)-mediated epithelial transport, as well as paracellular leakage of smaller amounts

(broken arrow) of both locally produced and serum-derived IgG antibodies into the lumen. Note that IgG cannot interact with J chain to form a binding site for pIgR. The

distribution of intraepithelial lymphocytes (mainly T-cell receptor a/bCCD8C and some g/dC T cells) is also depicted. Insert (lower left corner) shows details of anM cell and its

‘pocket’ containing various cell types.

FAE

GC

GC

Crypt

(a) (c) (d)

(b)

CD20CD3M-cell area

IgAIgG CD4CD8

M-cell area

Figure 2. (a,b) Three-color immunofluorescence staining of B cells (CD20, green), T cells (CD3, red) and epithelium (cytokeratin, blue) in cryosections of human Peyer’s patch.

(b) Details from theM-cell areas framed on the left in the follicle-associated epithelium (a) covering a B-cell follicle. (c) Two-color immunofluorescence staining for IgA (green)

and IgG (red) in section from normal human large bowel mucosa. Crypt epithelium shows selective transport of IgA, and only a few scattered IgG-producing cells are seen in

the lamina propria, together with numerous IgA plasma cells. (d) Three-color immunofluorescence staining for CD4C (red) and CD8C (green) T cells in normal human

duodenal mucosa. The epithelium of the villi is blue (cytokeratin). Note that most of the elements with weak CD4 expression seen in the background are either macrophages

or dendritic cells. Panel (a) is reproduced from Brandtzaeg et al. [30]. The other immunofluorescence pictures are original images from the Brandtzaeg laboratory.

Opinion TRENDS in Immunology Vol.25 No.11 November 2004 571

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Table 1. Recommended nomenclature for the mucosal immune

system

Preferred

abbreviations

Explanations

Immune-function

proteinsa

SIgA (or S-IgA) Secretory IgA

SIgM (or S-IgM) Secretory IgM

PIgA Polymeric IgA

Refers mainly to dimers but includes also larger

polymers of J-chain-containing IgA

J chain Joining chain

SC Secretory component

Exists in three forms: membrane SC; bound SC;

and free SC

PIgR Polymeric Ig receptor

The same as membrane SC

Immune-cell

compartmentsb

LP Lamina propria

Refers mainly to the connective tissue stroma

of gut mucosa above the submucosa and

muscularis mucosae but can also be used in

relation to other mucosae

FAE Follicle-associated epithelium

Contains variable number of M cells (see table

in Figure 4)

MALT Mucosa-associated lymphoid tissue

The principal inductive sites for mucosal

immune responses, subdivided according to

anatomical location (see below)

GALT Gut-associated lymphoid tissue

PP Peyer’s patch

ILF Isolated (solitary) lymphoid follicle

PPs and ILFs constitute the major part of GALT,

but the appendix is also included

NALT Nose- or nasopharynx-associated lymphoid

tissue

In humans, NALT consists of the lymphoid

tissue of Waldeyer’s pharyngeal ring, including

the adenoids (nasopharyngeal tonsil) and

palatine tonsils. In addition, human nasal

mucosa might contain ILFs. Rodents lack

tonsils but do have paired NALT structures

dorsally in the floor of the nasal cavity

SALT/DALT Salivary gland- or duct-associated lymphoid

tissue

Identified in non-human primates but not

humans

CALT Conjunctiva-associated lymphoid tissue

LDALT Lacrimal drainage-associated lymphoid tissue

TALT Eustachian tube-associated lymphoid tissue

LALT Larynx-associated lymphoid tissue

BALT Bronchus-associated lymphoid tissue

Not present in normal lungs of adult humans

MLN Mesenteric lymph node

CLN Cervical lymph nodeaApproved by the International Union of Immunological Societies–World Health

Organization Subcommittee on IgA nomenclature [9].bSee also Figure 4.


humans, rodents and rabbits (Figure 4) are typical MALTstructures believed to be a main source of conventional(B2) surface (s)IgA-expressing primed (memory–effector)mucosal B cells [10,11]. It should be noted, however, thatalthough the LP is considered to be an effector site, it isalso important for expansion and terminal differentiationof B cells (see below). Likewise, expansion of memory–effector T cells can apparently take place in the surfaceepithelium. There is considerable ‘crosstalk’ both withinand between these two effector compartments (Figure 3).


The term MALT was first coined to emphasize the factthat solitary organized mucosa-associated B-cell folliclesand larger lymphoid aggregates have common featuresand are the origin of trafficking to mucosal effector sites[12]. MALT is subdivided according to anatomical regions(Table 1), and the occurrence of such lymphoid structuresdepends on species, age and tissue state (normal orchronically inflamed). Thus, bronchus-associated lym-phoid tissue (BALT) is not present in normal lungs ofadults, and occurs only in 40% of healthy adolescents andchildren [13]. Figure 1 does therefore not include BALTamong the listed MALT structures.

The MALT concept was originally based on mouserepopulation experiments with B cells obtained fromBALT and PPs, as well as the surrounding LP. Notably,all MALT structures resemble LNs – with variable T-cellzones intervening between the B-cell follicles – andcontain a variety of antigen-presenting cells (APCs).However, MALT lacks afferent lymphatics because allsuch lymphoid structures actively sample exogenousantigens directly from the mucosal surfaces through acharacteristic follicle-associated epithelium (FAE) contain-ing ‘microfold’ or ‘membrane’ (M) cells (Figures 1,2a,2b).These specialized thin epithelial cells transfer effectivelysoluble, and especially particulate, antigens such asmicroorganisms from the lumen [14].

As schematically depicted in Figure 4, gut-associatedlymphoid tissue (GALT) comprises PPs, the appendixand isolated lymphoid follicles (ILFs), which are allconsidered to be inductive sites for mucosal B and T cells(Figure 1). The occurrence of other GALT-like elements,such as lymphocyte-filled villi (LFVs) and cryptopatches(CPs), is species dependent (Figure 4), and thesestructures are clearly not generating B cells. In fact,no immunological function has been identified for theextremely rare T-cell-dominated LFVs [15]; andalthough CPs were originally thought to be the originof intraepithelial lymphocytes (IELs) in mice [16], recentdata have questioned this notion and documented thethymus dependency of IELs even in this species [17].

Early animal studies demonstrated that not only PPs,but also the mesenteric LNs (MLNs) are enrichedprecursor sources for intestinal IgA plasma cells (PCs)[12,18,19]. Differentiation of sIgAC B cells takes placeduring their dispersion to the effector sites [20,21]. Thus,the fraction of B cells with cytoplasmic IgAwas only 2% inPPs but increased to 50% in MLNs, 75% in thoracic ductlymph and 90% in the intestinal LP because of terminalmaturation to PCs [22]. Such seminal findings gave rise tothe term ‘IgA cell cycle’ [23]. In humans, a clonalrelationship between sIgAC B cells in PPs and IgA-producing PCs in the LP provides strong support for thisnotion [24]. Later studies demonstrated that B cells ofother Ig classes and Tcells induced in PPs also exhibit gut-seeking properties.

HumanPPs occurmainly in the distal ileumand contain,by definition, between 5 and 200 aggregated lymphoidfollicles [25]. Human PP anlagen, composed of CD4C

dendritic cells (DCs), can be seen at 11 weeks of gestation,and discrete T- andB-cell areas occur at 19weeks.However,no germinal centers appear until shortly after birth,



Unstirred layerof mucus

Tight junction

Basal lamina

Goblet cell

Paneth cell

Entero-endocrinecell

Enterocytes

Intraepitheliallymphocyte(mainly CD8+)

Intraepitheliallymphocyte

Plasma cells(IgA, IgM, IgG)

Lymph vessel(lymphatic)

Cytokines

Dendritic cell

B lymphocytes

Mucosalmast cell

Nerves

Eosinophilicgranulocyte

Venule

T lymphocytes(Th1, Th2)

Chemokines

Macrophages

Neutrophilicgranulocyte

Lamina propria Epithelium

Figure 3. Depiction of cellular elements within the two effector compartments of the gut mucosa (lamina propria and epithelium). Indicated are the surface mucus, the

monolayered epithelium with enterocytes and other cell types, the basal lamina (basement membrane) and the diffusely distributed cellular components, as well as

mediators, within the lamina propria. IgA-producing plasma cells are normally remarkably dominating (Figure 2c). The different cell types are in reality much closer to each

other and can modulate the microenvironment by secreted chemokines (e.g. derived from the epithelium) and cytokines (e.g. derived from mast cells), as well as by the

interaction with juxtaposed nerve fibers. Note that dendritic cells can penetrate the basal lamina and epithelial tight junctions to actively sample antigens from the mucosal

surface (see Figure 1), while maintaining barrier integrity by expressing tight junction proteins [49]. Modified from Pabst and Rothkotter [64].


reflecting dependency on antigenic stimulation, which alsoinduces some follicular hyperplasia [26]. Thus, the numberof macroscopically visible human PPs increases from some50 at the beginning of the last trimester to 100 at birthand 250 in the midteens, then diminishes to becomew100 between 70 and 95 years of age [25].

In addition, the human gut harbors at least 30 000ILFs, increasing in density distally [27]. The normal smallintestine contains, on average, only one follicle per 269villi in the jejunum but one per 28 villi in the ileum [15]; inthe normal large bowel, the density of ILFs, as seen intissue sections, increases from 0.02 per mm muscularismucosae in the ascending colon to 0.06 per mm in therectosigmoid [28]. Small-intestinal ILFs have recentlybeen characterized immunologically in mice, showingfeatures compatible with the induction of local IgAresponses [29]. Interestingly, the cellular–molecularmechanisms involved in the organogenesis of variousinductive sites for mucosal immunity show both simi-larities and differences (Box 1).

Prevailing confusion about MALT terminology

Although the functional distinction between inductive andeffector sites is not absolute (see below), the cues forextravasation, migration and accumulation of naıveversus memory–effector B and T cells within theseimmune compartments are different [30,31]. It is indeed


confusing when authors, even in major journals andtextbooks, refer to mucosal effector compartments andtheir cells (e.g. LP lymphocytes and IELs) as part ofMALT. This is in direct conflict with the classicaldefinition of lymphoid tissue, as stated in authoritativetexts such as Terminologia Anatomica: InternationalAnatomical Terminology [32]. Gray’s Anatomy – in anattempt at clarification – is also confusing the issue [33];this text introduces the term ‘organized MALT’ (O-MALT)for the proper MALT structures, whereas ‘diffuse MALT’(D-MALT) refers to ‘the disseminated population oflymphocytes within the LP and epithelial base’. Althoughwe are aware of the fact that some mucosal immunologistshave used these terms, we strongly recommend that theybe abandoned.

Furthermore, despite the fact that mucosal immuneresponses are amplified in mucosa-draining local orregional LNs (Figure 1), they should not be included inthe MALT terminology. Thus, although being part of theintestinal immune system,MLNs should not be referred toas GALT structures. This distinction is important,emphasizing the fact that MALT is sampling antigensdirectly from the lumen viaM cells [14]. Likewise, it wouldbe appropriate to refer to head-and-neck-draining LNs aspart of the mucosal immune system but to replace theacronym CONALT (for ‘cranial-, oral-, and nasal-associ-ated lymphoid tissue’ [34]) with CONALNs, or simplycervical LNs (CLNs).



Continous ilealPeyer’s patch

Liver

Portal vein

Appendix

Peyer’s patch (PP)

M cells perenterocytes

RatPig

Human

VimentinUlex europaeusagglutinin (UEA-I)Cytokeratin 18Cytokeratin 8

M-cell markers

?

?~ 50%

~ 10%?

~ 3%

M cell

To peripheralblood

Portallymph node

Lymph vessel(lymphatic)

MLN

ILF

Lympho-glandularcomplex

Only pig

Largeintestine

Smallintestine

DogPig

Ruminant

Mouse Rabbit

Onlymouse

Onlyrat andhuman

Afferentlymphatic

Efferentlymphatic

Intestinallymph duct

To thoracicduct

IELs

LP

LFV

CP

ILF

Figure 4. Delineation of lymphoid-cell distribution in various compartments of the gut wall, with some species differences indicated. Lymphocytes can leave the gut wall via

draining lymphatics afferent to mesenteric lymph nodes, or via portal blood reaching the liver, where important regulation of immunity apparently takes place, particularly

the induction of tolerance [65]. Commonly used abbreviations are shown for various aggregates of lymphoid cells – for example, Peyer’s patches (PPs). The frequency of

M cells in the follicle-associated epithelium of PPs is highly variable among different species, and amarker for this specialized epithelial cell has not been identified in humans

[66]. Note also that, in contrast to the antigen-dependent priming of B cells that takes place in the PPs of most species, the continuous ileal PP present in ruminants, pigs and

dogs appears to be a primary lymphoid organ responsible for antigen-independent B-cell development, similarly to the Bursa of Fabricius in chicken (not shown). This PP can

be up to 2 m long and constitutes 80–90% of the intestinal lymphoid tissue. Abbreviations: CP, cryptopatch; IELs, intraepithelial lymphocytes; ILF, isolated lymphoid follicle;

LFV, lymphocyte-filled villus; LP, lamina propria; MLN, mesenteric lymph node. Modified from Pabst and Rothkotter [64].


Induction of IgA class switch distinguishes MALT from

lamina propria

Among the MALT structures, GALT in particularshows remarkable generation of B cells with a high levelof IgA and J-chain expression [30]. In this process, naıveB cells subjected to activation usually first change theirB-cell receptor composition from sIgDCIgMC to becomesIgDKIgMC memory–effector cells, and then switchdirectly or sequentially (via IgG) to IgA. Activation-induced cytidine deaminase (AID) has an essential rolein this process [11]. AID knockout mice, which showdefective class-switch recombination (CSR), have beenused to study the switching potential of intestinal B cellssampled outside of PPs [35]. As in IgA-deficient humans,AID-deficient mice were found to harbor numerousIgM-producing gut PCs, giving rise to abundant SIgM.When sIgMC intestinal B cells from such mice were


transformed with retrovirus to overexpress AID, theydisplayed a strong T-cell-independent propensity for IgAswitch after in vitro stimulation with lipopolysaccharide,transforming growth factor-b and interleukin-5. Thesame tendency was observed for similarly harvestedsIgMC B cells from normal mice in conjunction with AIDupregulation, whereas this phenotype obtained from PPsshowed a lower IgA-switch efficiency under identicalconditions [35].

During CSR, the DNA between the switch sites islooped out and excised, thereby deleting the Cm locus,which is followed either directly or sequentially by loss ofother constant heavy-chain (CH) genes [11]. After directIgA switch, the Ia-Cm circle transcripts (aCTs) derivedfrom the excised recombinant DNA are gradually lostthrough dilution from progeny cells during proliferation;readily detectable aCTs are therefore considered to be a


Box 1. Disparate organogenesis of inductive sites for

mucosal immunity

Experiments in knockout mice have helped in the understanding, at

least in part, of how inductive lymphoid tissue develops and

interesting differences appear. The development of Peyer’s patches

and mesenteric lymph nodes begins in fetal life, and their organ-

ization depends on multiple interactions between mesenchymal

organizer cells and hematopoietic inducer cells. A crucial differ-

entiating event is the expression of lymphotoxin (LT)a1b2 on

CD3KCD4CIL-7RC inducer cells after stimulation of the receptor

activator of NF-kB–tumor necrosis factor (TNF)-associated factor

family 6 signaling pathway through interleukin-7 (IL-7)R, and

subsequent interaction with LTbRC stromal cells [67]. In addition,

there are several feedback loops involving chemokines and adhesion

molecules, such as CXCR5–CXCL13 and CXCR5-induced a4b1–

vascular cell adhesion molecule-1 interactions [68].

By contrast, isolated lymphoid follicles develop only after birth, in

response to microbial stimulation, but they nevertheless require

LTa1b2–LTbR interactions and TNF-RI function [29,69]. Surprisingly,

unlike the gut-associated lymphoid tissue structures, nasopharynx-

associated lymphoid tissue (NALT) organogenesis does not depend

on the LTa1b2–LTbR signaling pathway, although NALT anlagen

apparently requires the presence of CD3KCD4C inducer cells [47].

Nevertheless, NALT occurs in retinoic acid-related orphan receptor-g

(RORg)-deficient mice that lack CD3KCD4CIL-7RC cells [70]. However,

fully developed NALT is apparently dependent on the LT-LTbR

pathway [47,70].

Unfortunately, in these genetically manipulated mice, no exten-

sive studies of mucosal effector sites, such as the gut lamina propria

and surface epithelium, have been performed. In future knockout

studies, the effect of different chemokines, cytokines and adhesion

molecules, as well as their receptors, on the cellular composition of

these effector sites should be included.


marker of recent CSR, being useful for identifyinganatomical compartments where switching takes place[11]. Interestingly, aCTs were detectable in murineintestinal sIgAC B cells located outside of PPs in AIDknockout mice, which was taken to show that LP IgAC

PCs differentiate in situ from sIgMC B cells [35].It was subsequently reported by the same authors that

AID-deficient mice show a dramatic hyperplasia of ILFs inresponse to intestinal bacterial overgrowth [36]. Thus, thefinding referred to above could actually have reflected adifference in IgA-switching capacity between ILFs andPPs rather than formation of aCTs truly outside ofGALT, because these studies were based on cell suspen-sions dispersed from gut segments, which necessarilycontained ILFs. In a subsequent review, the same authorsapparently were aware of this problem and ‘redefined’ theLP by making the following statement: ‘Taken together,these observations indicate that the lamina propria(inside or outside ILFs) might be a site where T-cell-independent responses are generated’ [11]. This exemplifieshow excellent science can be erroneously communicated,thereby confusing the issue. The suggestion that classswitch to IgE occurs in the nasal mucosa of allergicpatients [37] could likewise be explained by the presenceof ILFs [38]. There is always a risk that already switchedB cells from germinal centers contaminate tissue suspen-sions when attempts are made to sample the LP [39].Notably, a recent study based on similar molecularmethods, but also including well-controlled microdissec-tion, clearly demonstrated that no IgA class switch was


detectable in the murine nasal or intestinal mucosaoutside of MALT structures [40].

Potential sources of switched mucosal B cells beyond

GALT

Although GALT is the largest and best defined part ofMALT, an additional major inductive site for mucosalB-cell responses is NALT, which was originally defined inrodents and spelled out as ‘nasal-associated lymphoidtissue’ [41]. Grammatically, this is a misnomer, and someauthors have therefore changed it to ‘nose-associatedlymphoid tissue’. We recommend, instead, ‘nasopharynx-associated lymphoid tissue’, which in humans wouldcomprise the unpaired nasopharyngeal tonsil (also called‘adenoids’ – note, in pleural), the paired palatine tonsilsand other smaller lymphoid structures of Waldeyer’spharyngeal ring [42].

Rodents lack tonsils, and their NALT structures occuron both sides of the nasopharyngeal duct dorsal to thecartilaginous soft palate [41]. Application of nasal vaccinein mice elicited a regionalized protective IgA response[43]. It has furthermore been shown that murine NALTcan generate an IgA-skewed enrichment of high-affinitymemory–effector B cells, and additionally give rise to amajor germinal center population of IgG-producing cells[44], which is quite similar to the response seen in humantonsils [30,45]. In contrast to tonsils, however, the anlagenof which appears at the same fetal age as that of PPs [46],the organogenesis of murine NALT begins after birth [47],as for murine ILFs [29].

In mice, proliferating T cells rapidly obtain gut-homingproperties during antigen priming in MLNs [48]. Thesame apparently applies to B cells because these nodesare enriched sources of precursors for intestinal IgA-producing PCs [19,23]. It is therefore most likely thatmucosa-draining LNs generally share immune-inductiveproperties with regional MALT and receive similarantigens via afferent lymph and carried by DCs, such asCLNs in relation to NALT (Figure 1). Numerous DCsoccur at epithelial surfaces (Figure 3), where they cansample luminal antigens by penetrating tight junctionswith their processes [49]. Importantly, the human nasalmucosa is extremely rich in various APC phenotypes, bothwithin and beneath the epithelium [50], and a subepi-thelial band of similar cells is seen below the intestinalsurface epithelium and the FAE of PPs [51].

The peritoneal cavity is yet another source of B cells inmice, reportedly providing 40–50% of the intestinal IgAC

PCs [52]. The precursors are self-renewing sIgMC B1(CD5C) cells generating polyreactive (‘natural’) SIgAantibodies directed against commensal bacteria as a resultof a T-cell-independent switch to IgA [53], which is claimedto take place in the LP [11]. However, another studyshowed that murine sIgMCIgAK B1 cells were alreadyclass-switched to IgA at the DNA level within theperitoneal cavity [54]. Equally importantly, a recentstudy provided evidence that both total and bacteria-specific intestinal IgA in the mouse is produced mainly byGALT-derived B2 cells [55]. Finally, no evidence exists tosuggest that B1 cells are significantly involved inintestinal IgA production in humans [10,39,56], despite



substantial levels of polyreactive SIgA antibodies recog-nizing both self- and microbial antigens [57].

The immunological potency of effector sites is

determined by topical antigen exposure

Excluding the LP from the MALT terminology does notmean that we neglect its role in shaping mucosal immuneresponses. Antigen-driven proliferation of intestinal IgAcells, apparently outside of GALT structures, has beenobserved in experimental animals [10,58,59], and scat-tered sIgAC memory–effector cells with a proliferativepotential occur in the human LP [39]. Interestingly, rapidwidespread dissemination of fed antigen, followed byactivation of cognate T cells, has been described in T-cell-receptor transgenic mice [60]. This observation, togetherwith previous results of adoptive B-cell transfer insyngeneic rats [61], supports the notion that localantigen-driven T-cell activation provides important sec-ondary signals for retention, proliferation and terminaldifferentiation of B cells at secretory effector sites. Inaddition to paracellular or transcellular uptake of anti-gens through surface epithelia, DCsmight actively sampleluminal antigens [49], and specialized M-cell-like epi-thelial cells identified in the villi of mouse gut couldcontribute to this sampling [62].

In keeping with such activity at mucosal effector sites,the density of IgAC PCs in a particular secretory tissuegenerally reflects the level of topical antigen exposure,being seven times higher in the human colon (Figure 2c),with its enormous microbial load, than in parotid andlactating mammary glands [63]. Also notably, the lacrimalglands, which are connected by many short ducts to theexcessively aeroantigen-exposed conjunctiva, show adensity of IgAC PCs approaching that of the colon [63].

Conclusions

Mucosal immunologists should standardize their termi-nology to set an example for the rest of the immunologicalbrotherhood. The drawing of a correct map of immunecompartments requires knowledge of the actual ‘landscape’.It is also important to know the definition of lymphoidtissue, which appropriately applies only to inductiveMALT structures. The mucosal effector sites, representedby the LP and surface epithelium, exert distinct functionsand cannot, by established classical definition, be calledlymphoid tissue; they should hence not be referred to as‘MALT’. Also notably, different cues are operating inmucosal inductive and effector compartments, both fororganogenesis and immune-cell extravasation, prolifer-ation and differentiation. Such knowledge is mainlyderived from genetically manipulated mice, and scientistsshould be aware of the fact that fundamental speciesdifferences exist within the mucosal immune system.

Acknowledgements

Studies in the laboratory of P.B. are supported by the University of Oslo,the Research Council of Norway, and the Norwegian Cancer Society. Theresearch of R.P. is funded by the German Research Foundation (DFG). Wethank Hege Eliassen and Erik K. Hagen for excellent assistance with themanuscript preparation.


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