T R E N D SI M M U N O L O G Y TO D AY

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ucosal tissues of variousorgans show similarpowerful innate andadaptive defense mecha-

nisms against environmental pathogens,resulting in the concept of mucosa-associatedlymphoid tissue (MALT). However, thereare also organ-specific variations, and strik-ing differences in the mechanism of antigenhandling was one of the major themes ofthis workshop. Investigating organ-specificvariations in MALT has improved ourknowledge of mucosal tolerance induction.Several mucous membranes can be used forthe induction of tolerance, and a series ofpresentations dealt with models and clinicaltrials of oral tolerance induction.

Physiology of antigen handling bymucous membranesAntigen uptake in the intestine is mediatedprimarily through specialized epithelial M cells, which deliver luminal antigens toorganized lymphoid tissue in Peyer’s patchesfor processing1. Distinct regional differencesoccur throughout the intestine in the num-bers of M cells and the magnitude of Peyer’spatch uptake of antigens, as well as in thepopulations of dendritic and T cells. Peyer’spatch lymphocytes may induce the trans-formation of enterocytes into M cells2, andM cells, in turn, may serve as a portal ofentry for the invasion of specific patho-gens1. Antigen handling in the gut may alsobe mediated by alternative transcellular orparacellular routes, including epithelialdendritic cells (P.W. Bland, Bristol). Crypto-patches in murine intestinal mucosa3 mayplay a role in antigen processing but areprobably more important as lymphoid tis-sues where lymphohematopoietic progeni-tors for T and/or B cells are generated.

The method of antigen handling on theocular surface is unclear. M cells have notbeen detected in conjunctival epithelium,

the functional role of conjunctival lymphoidfollicles is unknown and antigens do notappear to undergo retrograde transport tothe lacrimal gland (D.A. Sullivan, Boston,MA). It may be that antigens are sampled inthe stratified conjunctival epithelium viadendritic cell processes (J.P. Kraehenbuhl,Epalinges), and that lymphatic sinusoidsadjacent to conjunctival lymphoid aggre-gates are involved in the local immune re-sponse (R. Pabst, Hannover). It also seemsthat the ocular IgA response to infectious ortoxic substances may require antigenicclearance through the nasolacrimal duct,stimulation of pharyngeal or gut-associatedlymphoid tissue and consequent IgA lym-phoblast migration to the lacrimal gland(Sullivan).

Mucosal epithelial cells may also modu-late antigen processing by gut-associatedprofessional antigen-presenting cells (APCs)– for example, by secretion of cytokines(Bland) or, in the eye, by release of hormonessuch as androgens (Sullivan).

Antigen handling at intraocular as wellas mucosal sites may be further modulatedby programmed cell death. Fas ligand ex-pression in the cornea, for instance, servesto protect this tissue against antigenic chal-lenge and subsequent immigration of in-flammatory cells (e.g. from the conjunctivaand anterior chamber) by inducing theapoptosis of activated Fas1 immune cells4

(T.A. Ferguson, St Louis, MO).

Cellular and surface defensemechanisms of mucosal tissueThe type of proteins involved in ocular de-fense varies among species. These differ-ences are mainly reflected in distinct ocularinnate rather than adaptive defense mecha-nisms. Secretory (S) antibodies produced by

local plasma cells in the lacrimal glands aretransported into secretions by the polymericIg (poly-Ig) receptor. The major immuno-globulin isotype is S-IgM in lower verte-brates and S-IgA in mammals. This evolu-tionary switch probably occurred to facilitatethe diffusion of a smaller immunoglobulinacross the epithelial basal lamina to interactwith the epithelial poly-Ig receptor.

New data have emerged concerning thecontrol of the epithelial poly-Ig receptorgene (P. Brandtzaeg, Oslo). An E-box in thepromoter is involved in constitutive ex-pression whereas upregulation may occurvia interferon g (IFN-g)-responsive elements.Concentration of S-IgA on the human ocu-lar surface is highly dependent on the flowrate of tears. During sleep, lacrimal glandfluid secretion ceases and the concentrationof S-IgA can increase to as high as 10 mg ml21 (Ref. 5). This is associated witha rapid influx of polymorphonuclear cells.The combination of polymorphonuclearneutrophil action via its IgA Fc receptor andthe high local S-IgA concentration providesprotection against residual infectiousmicroorganisms during the night.

In the event that exclusion of foreignantigens at the mucosal surface fails, entryto the deeper layers of the tissue is met witha powerful array of macrophages and T cells.Lamina propria in the gut as well as con-junctival tissue (H.S. Dua, Nottingham)contain T cells that home to these sites afterinteraction with adhesion molecules thatmay express tissue specificity. Analysis ofthe function of lamina propria T cells hasshown proliferation of these cells via theCD2 but not the CD3 pathway (A. Stallmach,Homburg). Study of the fine specificity ofgluten-specific T cells in coeliac disease hasled to the identification of a natural immuno-stimulatory pepsin-derived fragment ofgluten (F. Koning, Leiden). Truncation of thispeptide resulted in the production of pep-tide antigens that can block activation of the

Mucosal immunology and the eyeManfred Zierhut, Charles O. Elson, John V. Forrester, Aize Kijlstra,

Jean-Pierre Kraehenbuhl and David A. Sullivan

A recent meeting* compared the

immunoregulatory mechanisms of

the ocular mucosa with those of the

mucosal membranes of other

tissues.M

*The workshop ‘Mucosal Immunology and Ocular Disease’ was held at Ettal,Germany, on 23–24 October 1997.

T-cell clone by the original peptide, offeringexciting prospects for the treatment ofcoeliac disease.

Effector mechanisms of the mucosal im-mune response are partly regulated by localcytokine production, and both corneal andconjunctival cells have been shown to pro-duce many cytokines and chemokines,including interleukin 1 (IL-1), IL-6, IL-8,monocyte chemoattractant protein (MCP),GRO-a and a wide range of growth factors(A. Kijlstra, Amsterdam). Cytokines on theocular surface play a role in the continuousphysiological renewal and wound healingof corneal and conjunctival epithelium, aswell as the control of angiogenesis, apop-tosis and leukocyte recruitment. Since IL-1is one of the main triggers of these func-tions, the finding of a high constitutive ex-pression of the IL-1 receptor antagonist inboth human and rat corneas is not com-pletely surprising6. Other anti-inflammatorycytokines produced by resident cells of thecornea include IL-10 (Ref. 7). IL-10 may playa role in controlling keratitis induced byherpes simplex virus8.

Tolerance induction via the mucousmembranesTolerance can be induced via mucous mem-branes. There appears to be a clear dosedependency: low-dose tolerance involvesregulatory cells (‘active suppression’), whilehigh-dose tolerance involves anergy (N.Staines, London). With oral tolerance, theinduction of regulatory cells appeared to belost with very large doses, but the deliveryof high doses through other mucous mem-branes such as the conjunctiva did not resultin a decrease in induction of regulatory cells(J.G. Woodward, Lexington, KY). Usingantigen coupled to cholera toxin B subunit,there was no decrease in induction of regu-latory cells at high antigen doses eventhrough the oral mucous membrane (J.Y.Niederkorn, Dallas, TX). Active regulationmay be mediated by cytokines, especiallyIL-10 and transforming growth factor b

(TGF-b), and in this respect resembledmany of the other tolerizing processes dis-cussed at the meeting (R. Caspi, Bethesda,MD; Ferguson). The possible role of regu-latory T cells was discussed in some depth,

with CD41 T cells emerging as the mostlikely candidate – the role of CD81 T cells or gd T cells as regulatory cells in mucosaltolerance remains unclear.

The cellular mechanisms required for theinduction of mucosal tolerance were ad-dressed by several speakers. Agreementcentred on the fact that antigen uptake andprocessing probably occurred at the site ofentry (i.e. on the mucous membrane), butthereafter the site of antigen presentation tothe putative regulatory cells differed be-tween models. For oral tolerance in mice, an intact spleen appears to be necessary(Caspi; Staines), as it is for anterior chamber-associated immune deviation (Ferguson).However, splenectomy has no effect onnasal or conjunctival tolerance (D.C. Wraith,Bristol; Woodward). Presentation of antigenin these tissues appears to occur in thedraining lymph nodes9 (Woodward; Wraith;A. Dick, Aberdeen) and seems to be medi-ated by antigen-specific apoptosis-inducingmechanisms occurring on re-exposure tothe tolerizing antigen in adjuvant (Wraith;Dick). Little is known about the transport ofantigen from the mucous membrane to thelymphoid organs. Wraith described somerecent data using labeled antigens thatshowed tritiated peptide delivered throughnasal passages was rapidly cleared from thelungs and reached a single peak in thebloodstream, whereas it appeared to have abiphasic peak in the lymphatics.

Although antigen specificity is a majorcharacteristic of mucosal tolerance, by-stander suppression (i.e. where tolerance toone peptide can confer protection to thewhole molecule or even to unrelated mol-ecules) was also inducible in some cases.Intramolecular bystander suppression wasmore readily induced than intermolecularbystander suppression. For instance, a sub-dominant peptide of the collagen type IImolecule was effective in the induction oftolerance to the entire collagen molecule(Staines) but a crude extract of retinal anti-gens was protective for some antigens (e.g. S antigen) but not others [e.g. inter-photoreceptor retinoid-binding protein(IRBP)10] (Dick).

A major issue was whether mucosal tolerance was effective in the presence ofactive inflammation. Although Niederkorn

found it was in the context of corneal transplantation11, others (Wraith; Caspi;Dick) agreed that it was not possible toachieve good immunosuppression, at leastexperimentally, when there was active inflammation. Initial studies using a combi-nation therapy of mucosal tolerance withspecific antigen and immunosuppressivedrugs in experimental autoimmune uveitis(EAU)12 and in the treatment of multiplesclerosis appeared to be encouraging (A. Slavin, Boston, MA).

The factors that determine whether toler-ance or immunity results from a mucosalantigen exposure remain unclear. The mu-cosal adjuvant cholera toxin can switch a response from tolerance to immunity. Themechanism appears to involve the upregu-lation of costimulatory molecules like B7-2 onAPCs, as well as stimulation of costimulatorycytokines (Elson, Birmingham, AL). The latter include IL-10; thus, this cytokine can beinvolved in either tolerance or immunity.

Vaccines are therapeutic alternatives totolerance induction. Essential to the devel-opment of recombinant vaccines are consid-erations of effective presentation of vaccineantigen in vivo. Recombinant multiple T-cellepitope vaccines against rat experimentalautoimmune encephalomyelitis, experi-mental autoimmune neuritis and EAU werediscussed by Schluesener (Tübingen),demonstrating that use of peptides that ‘direct’ T-cell epitopes to specialized lymph-oid compartments increased the potency ofvaccines. Combinatorial peptide phage-dis-play and RNA libraries are used to defineligands inducing effective transport acrossmucosal barriers.

Concluding remarksThe study of mucosal tissues demonstratesorgan-specific variance. While M cells seemto be unique for Peyer’s patches, the anti-gen uptake at the conjunctiva depends ondendritic cells and lymphatic sinusoids ad-jacent to conjunctival lymphoid aggregates.Investigations of the mechanisms that par-ticularly involve the use of adjuvants forgenerating oral, nasal or conjunctival tolerance may help to institute improvedstudy protocols for the induction of toler-ance in autoimmune disorders.

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he neonatal period is markedby a high susceptibility to infections. Pathogens consid-ered to be essential targets for

early immune intervention in industrialized(P. Wright, Nashville, TN) and developing(K. Mulholland, Geneva) countries are summarized in Table 1.

Innate immunity in early life Innate immunity represents the first barrierto infections and plays a pivotal role in theinduction of adaptive immunity. Decreasedbone marrow reserve, reduced adherence/chemotaxis and lower enzymatic activityand signalling of neonatal neutrophils con-tribute to the risk of rapid systemic spreadfrom any localized infection (R.B. Johnston,New Haven, CT). Further limitations occurat the levels of monocytes/macrophages,the complement system and natural killerand lymphocyte-activated killer cell cyto-toxicity, which remain below adult levels

even after induction with exogenous inter-leukin 12 (IL-12)/IL-15 (Ref. 1). Cord blood(CB) dendritic cells (DCs) seem to be less ef-fective than adult DCs in supporting prolif-eration of T cells in response to antigenicstimulation. This could be attributed to reduced expression of HLA molecules, co-stimulatory/adhesion molecules and acti-vation markers. Increasing the mitogen concentration or number of DCs in culture,or adding interferon g (IFN-g), allowed partial restoration of neonatal DC function2

(R.E. Petty, Vancouver).The major components of innate im-

munity thus appear weakened in newborninfants. The main limitation of reportedstudies was identified as the use of CBrather than peripheral blood cells fromhealthy newborns, which hampers the

distinction between developmental effectsand consequences of perinatal stress. Essen-tially, unanswered questions concern thetime required for maturation to adult-likefunctions and the potential physiologicalrole of the limitation of innate immunity inthe maternofetal and early-life context.

Neonatal T-cell responsesThe patterns of certain viral infections inearly life strongly suggest that impaired T-cell responses limit viral clearance in vivo.When assessed in vitro, the majority ofhuman CB CD41/CD81 T cells present theCD45RA1 ‘naive’ phenotype, whereas boththe CD45RA1 and the ‘memory/mature’CD45R01 phenotypes are represented inequal numbers in adult peripheral bloodmononuclear cells (APBMCs). Naive T cellswere shown to require much longer periodsof T-cell receptor (TCR) triggering com-pared with mature effector cells3 (A. Lanzavecchia, Basel). The CD40 ligand

Manfred Zierhut is at the Dept of Ophthalmol-ogy, University of Tübingen, 72076 Tübingen,Germany; Charles Elson is at the Division ofGastroenterology, University of Alabama, Birmingham, AL 35294-0007, USA; John Forrester is at the Dept of Ophthalmology, Uni-versity of Aberdeen, Aberdeen, UK AB9 2ZD;Aize Kijlstra is at The Netherlands OphthalmicResearch Institute, University of Amsterdam,1100 AC Amsterdam, The Netherlands; Jean-Pierre Kraehenbuhl is at the Swiss Institute forExperimental Cancer Research, University ofLausanne, 066 Epalinges, Switzerland; DavidSullivan is at the Dept of Ophthalmology, Schepens Eye Research Institute, Boston, MA02114-2500, USA.

T R E N D SI M M U N O L O G Y TO D AY

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Immunity in early lifeJiri Kovarik and Claire-Anne Siegrist

A recent meeting* focused on the

functional development of the

immune system in the neonate,

with the aim of identifying specific

immunization approaches for

protection in early life.T

*The International Symposium on Immunity in Early Life was held at Annecy,France, on 17–19 November 1997.