CITOQUINAS Y AUTOINMUNIDAD

© 2001 Macmillan Magazines Ltd

Autoimmune diseases, which affect approximately 5%of the population and disproportionately affectwomen, comprise a heterogeneous group of poorlyunderstood disorders1,2. In general, these illnesses arecharacterized by the presence of abnormal lymphocyteactivation, although we now know that non-lymphoidcells, especially antigen-presenting cells such asmacrophages and dendritic cells (DCs), also haveimportant contributions to disease pathogenesis. It iswell established that cytokines have essential roles inimmune cell development, immunoregulation andimmune effector functions. Accordingly, the role ofcytokines has been extensively studied in the pathogene-sis of autoimmune disease3. The surprise has been thatalteration of cytokine levels and responsiveness does notnecessarily have the expected effect — cytokines thoughtto promote inflammatory and immune responsivenesshave unexpected, but essential, immunosuppressive,actions. The idea that pro-inflammatory cytokines pro-mote autoimmune disease, whereas regulatory cytokinessuppress it, is too simple to explain the mechanismsunderlying autoimmunity. Emerging data indicate that,as the Dave Mason song goes,“there ain’t no good guys,there ain’t no bad guys”, but rather a combination ofcytokines exert variable effects at different times duringthe evolution of autoimmune disease. The complex roleof cytokines in autoimmunity, especially their unantici-pated immunosuppressive functions, is the focus of thethis review.We have focused on interleukin-2 (IL-2), but

will also draw on lessons learned from interferons(IFNs) and tumour-necrosis factor (TNF) to illustratethese points.

A simple, but flawed, view of cytokinesIt is recognized that T-lymphocyte function contributesto the pathogenesis of many autoimmune diseases. Theaccepted model is that naive T cells become activated byantigen and produce IL-2, which, in turn, causes clonalexpansion and induces the production of other pro-inflammatory cytokines, such as TNF. Administrationof IL-2 to humans results in expansion of lymphoidcells, but its clinical application is limited by its toxicity.IL-2 administration is also associated with a variety ofautoimmune disorders, including immune thyroiditis,rheumatoid arthritis and other arthropathies4.

CD4+ T cells differentiate into at least two subsetsof helper cells, T helper 1 (T

H1) and T

H2 cells5 (FIG. 1).

The former produce IFN-γ (also known as type 2 IFN)and lymphotoxin-α. This differentiation is regulatedby IL-12, which activates the transcription factorSTAT4 (signal transducer and activator of transcrip-tion 4) in lymphocytes6. T

H1 responses have been

implicated in many autoimmune disorders, as shownby elevated levels of IFN-γ in tissues, by the ameliora-tion of disease with anti-cytokine treatment, and fromstudies of IFN-γ-, IL-12- and Stat4-deficient mice.In addition, a promoter polymorphism associatedwith overexpression of an IFN-inducible gene (Ifi202)

CYTOKINES AND AUTOIMMUNITYJohn J. O’Shea*, Averil Ma‡ and Peter Lipsky*

Cytokines have crucial functions in the development, differentiation and regulation of immunecells. As a result, dysregulation of cytokine production or action is thought to have a central rolein the development of autoimmunity and autoimmune disease. Some cytokines, such asinterleukin-2, tumour-necrosis factor and interferons — ostensibly, the ‘bad guys’ in terms ofdisease pathogenesis — are well known for the promotion of immune and inflammatoryresponses. However, these cytokines also have crucial immunosuppressive functions and so,paradoxically, can also be ‘good guys’. The balance between the pro-inflammatory andimmunosuppressive functions of these well-known cytokines and the implications for thepathogenesis of autoimmune disease is the focus of this review.

NATURE REVIEWS | IMMUNOLOGY VOLUME 2 | JANUARY 2002 | 37

*Lymphocyte Cell BiologySection, Arthritis andRheumatism Branch andAutoimmunity Branch,National Institute of Arthritisand Musculoskeletal andSkin Diseases,National Institutes of Health,Building 10 Room 9N252,10 Center Drive,MSC-1820, Bethesda,Maryland 20892, USA.‡Departments of Medicineand Ben May Institute forCancer Research,University of Chicago,Chicago, Illinois 60637, USA.Correspondence to J.J.O.e-mail: [email protected] DOI: 10.1038/nri702

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CROHN’S DISEASE

One of the two predominantforms of inflammatory boweldisease that afflicts humanpatients. The pathophysiology is unknown, but is presumedto be autoimmune in nature.

TOLERANCE

Denotes lymphocyte non-responsiveness to antigen, butimplies an active process, notsimply a passive lack of response.

PERIPHERAL TOLERANCE

This form of tolerance refers tothe lack of responsiveness ofmature lymphocytes.

38 | JANUARY 2002 | VOLUME 2 www.nature.com/reviews/immunol

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responses might contribute to disease. Additionally, ourconclusions regarding the role of cytokines in autoim-munity largely arise from animal models of disease, andthe extent to which these models accurately reflectmechanisms in human disease varies. More intriguing,however, is the fact that the cytokines themselves havecomplex actions. It is well known that cytokines arepleiotropic, redundant to some degree, and induce theproduction of other cytokines. More pertinent for thisreview, however, is that they have both immunostimu-latory and immunosuppressive actions; cytokines suchas IL-2, IFN-γ and TNF really are double-edged swords,both enhancing and limiting autoimmune disease. Thisis not to say that these are the only cytokines that havesuch complex actions. The actions of other cytokinesare also not simple. However, because the positiveeffects of IL-2, IFN-γ and TNF on immune responsesare so well appreciated, we have focused on just thesethree cytokines.

Autoimmunity associated with IL-2 deficiencyThe pathology in IL-2 and IL-2 receptor (IL-2R)knockout mice necessitated a major revision in ourthinking about the function of IL-2 (REFS 11,12 and FIG. 2).Because of its well-known role in lymphoid prolifera-tion, it was anticipated that the main phenotype associ-ated with impaired IL-2 signalling would primarily be aform of immunodeficiency. Instead, the mice exhibitlymphoid hyperplasia and produce high levels of multi-ple autoantibodies, including anti-DNA antibodies.About half of the mice die of autoimmune haemolyticanaemia before 2 months of age, and the survivorsdevelop inflammatory bowel disease. Similar to CROHN’S

DISEASE, the cytokines present in the colitis associatedwith IL-2 deficiency are typical of a T

H1 response, and

disease is attenuated by interfering with IL-12 and IFN-γ.Importantly, the pathology is also corrected by theaddition of exogenous IL-2 and adoptive transfer oflymphocytes capable of producing IL-2- or IL-2-treatedcells from IL-2-deficient mice12. Although the disorderis improved when IL-2-deficient mice are kept in germ-free conditions, colitis does not occur in more pro-foundly immunodeficient, recombination-activatinggene (Rag) knockout animals, which indicates that theinflammatory bowel disease is not solely the result ofimpaired host defence or an opportunistic infection.Instead, the autoimmune disease in Il2–/– mice indicatesthat, despite the positive effects of this cytokine on lym-phocyte proliferation in vitro, the non-redundant roleof IL-2 in vivo is to constrain lymphoid growth andmaintain PERIPHERAL TOLERANCE. What, then, is the mecha-nism(s) through which IL-2 maintains TOLERANCE, andwhat is the relevance to human disease?

IL-2 and activation-induced cell death. One mechanismthat has been pursued is the ability of IL-2 to promoteprogrammed cell death13. Apoptosis of lymphocytes hasemerged as a principal means of controlling immuneresponse, but there are two biochemically distinct path-ways that contribute to this process. The first type ofapoptosis, referred to as passive cell death, results from

has been shown to be associated with mouse lupus,and a polymorphism in the IL-12p40 gene has beenreported as a disease-associated allele in type I diabetesin humans7,8.

Conversely, TH2 cells, which produce IL-4, promote

allergic responses and constrain cell-mediated immu-nity. Accordingly, T

H2 cytokines can inhibit autoim-

mune disease. Two important immunosuppressivecytokines, IL-10 and transforming growth factor-β(TGF-β) are produced by both subsets of T

Hcells,

although T cells that are the main producers of thesecytokines have also been denoted as regulatory T cells;interfering with these cytokines results in severe inflam-mation and autoimmune disease. IL-10 activates Stat3,and absence of Stat3 in myeloid cells also leads toautoimmune disease9. Therefore, it would seem reason-able to speculate that the balance between the ‘bad guys’(IL-2, IFN-γ and TNF) and the ‘good guys’ (IL-4, IL-10and TGF-β) would determine the propensity for thedevelopment of autoimmune disease. If only this simplemodel were true — unfortunately, this scenario is toosimple to reflect reality. Indeed, if it were true, then onemight expect that the rising incidence of T

H2-driven

allergic diseases in developed countries would be associ-ated with a concomitant reduction in T

H1-diseases, and

this is evidently not the case10.There are several reasons underlying this complexity.

First, not all autoimmune diseases are strictly TH1 dis-

eases; ulcerative colitis, and to an extent, systemic lupuserythematosus, are two examples in which T

H2

IL-12

IL-4IL-5IL-6IL-10IL-13

B cell

Macrophage

LTIFN-γ

Cell-mediatedimmunity

Allergicresponses

Anti-helminthresponses

IL-12STAT4

STAT6 IL-4

TH

TH1

TH2

Eosinophil

IgE

Figure 1 | Cytokines and helper T-cell differentiation. In the standard model, helper T cellsare polarized to one of two subsets, TH1 and TH2, by the cytokines interleukin (IL)-12 and IL-4,respectively. TH1 cells promote cell-mediated immunity and host defence against intracellularorganisms, but also contribute to the pathogenesis of autoimmune diseases such asrheumatoid arthritis, Crohn’s disease and multiple sclerosis. TH2 cytokines, especially IL-4,antagonize TH1 differentiation and attenuate cell-mediated immunity but promote allergic andanti-helminth responses. Although this model has been extremely useful and explains a greatdeal, the role of cytokines in autoimmune disease is more complicated than is indicated by thissimple model. IFN-γ, interferon-γ; IgE, immunoglobulin E; LT, lymphotoxin; STAT, signaltransducer and activator of transcription.

© 2001 Macmillan Magazines LtdNATURE REVIEWS | IMMUNOLOGY VOLUME 2 | JANUARY 2002 | 39

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These mice have profound LYMPHADENOPATHY, autoanti-body production and nephritis. No defects in thymicdeletion were found in Fas-deficient mice; instead, thisdisorder is an example of impaired peripheral toler-ance. Fas also has an important role in the removal ofautoreactive B cells. The human disease autoimmunelymphoproliferative disease (ALPS) is caused bymutations of Fas14. Mutations of Fas and other com-ponents of this pathway have been sought, but havenot been found in lupus or other common humanautoimmune diseases.

As mentioned above, the relevance of defects in theFas pathway to the autoimmunity associated with IL-2deficiency is that IL-2 promotes Fas-dependent AICDby upregulating Fas, FasL and FADD transcription15.This is dependent on Stat5 (REF. 15). Conversely, FLIP(FLICE/caspase-8 inhibitory protein) interferes withFas-mediated apoptosis, and IL-2 downregulates FLIPexpression. Transfer of cells that constitutively expressFLIP results in autoimmunity16.

IL-2R is a multimeric complex that comprises IL-2Rα (CD25), IL-2Rβ and a shared receptor cytokinesubunit, termed the common γ-chain (γ

c). Other

cytokines that use γcinclude: IL-4, IL-7, IL-9, IL-15 and

IL-21. IL-15, in fact, binds to two receptor subunits alsobound by IL-2, IL-2Rβ and γ

c; IL-2 and IL-15 use sepa-

rate α-chains. One striking finding is that although allthe γ

ccytokines induce lymphocyte proliferation, the

promotion of AICD seems to be a unique action ofIL-2, at least at physiological levels of cytokine. The lackof effect of IL-15 is particularly difficult to understand,given that no discernible differences in IL-2 and IL-15signalling have been unearthed. Non-mutually exclu-sive factors that might account for the distinct activitiesof IL-15 and IL-2 include in vivo receptor and ligandbioavailability or the location of signalling events.These possibilities are appealing given the distinct tis-sue expression of the ligands. Alternatively, the ligandsmight differ in their relative strength of signalling.

Despite the clear importance of Fas and AICD, is thisthe whole explanation for the autoimmunity associatedwith IL-2 deficiency? Evidently not, as the phenotype ofFas- and IL-2-deficient mice are quite distinct even instrain-matched mice. For instance, neither lpr mice norpatients with ALPS develop colitis. These differencesindicate that Fas deficiency is not the sole explanationfor the autoimmunity that occurs in the absence of IL-2.Furthermore, despite in vitro results, there are still nocompelling in vivo data that establish IL-2-dependentAICD as the primary mechanism underlying the role ofIL-2 in maintaining peripheral tolerance.

Regulatory T cells — is there an IL-2 connection? A sec-ond possible mechanism underlying the role of IL-2 inmaintaining peripheral tolerance relates to the genera-tion of regulatory cells17,18. The controversial notion that‘suppressor’ T cells are important in physiology andpathology has undergone a recent renaissance. Studiesinitiated more than 20 years ago on the generation oforgan-specific autoimmunity by neonatal thymectomyhave lead to the conclusion that regulatory T cells reside

the lack of crucial pro-survival signals. Absence ofcytokine signalling results in increased mitochondrialpermeability and cytochrome c release, which, alongwith apoptotic protease-activating factor 1 (APAF1),activates caspase-9 and downstream effector caspases.Proteins in the BCL-2 family interfere with this mode ofprogrammed cell death, and cytokines such as IL-7(and IL-2) upregulate anti-apoptotic members of theBCL-2 family.

The second type of apoptosis, which is probably moreimportant in terms of the pathogenesis of autoimmunity,is activation-induced cell death (AICD). This is mediatedpredominantly by the death-domain-containing receptorFas (CD95) and, importantly, IL-2 has essential functionsin promoting Fas-dependent AICD (FIG. 2). Occupancyof Fas results in the recruitment of proteins containingdeath domains and death effector domains; for exam-ple, Fas-associated death domain (FADD), whichrecruit and activate caspase-8, which then activatesdownstream caspases, inducing programmed celldeath12. The importance of Fas/Fas ligand (FasL;CD178) was established by the demonstration of theirdeficiency in autoimmune lpr and gld mice, respectively.

LYMPHADENOPATHY

Enlargement of lymph nodes.

TH1 Granzyme

Perforin

Apoptosis

CytotoxicityPerforinGranzymes TH2 cells

Apoptosis

ImmunoregulatoryT cells

Cytokines

ProliferationCyclinsCell-cycle inhibitorsFOSMYC

Anti-apoptoticBCL-2BCL-XL

FasFasLFLIP

IFN-γ

TNF

Perforin

IL-4

Immunostimulatory Immunosuppressive

IL-10

TGF-β

TH1

TH2

IL-2

CD25

CD152

CD4Treg

Figure 2 | Summary of the immunostimulatory and immunosuppressive effects of IL-2.Although interleukin-2 (IL-2) is well known for its ability to promote proliferation, to inhibit apoptosisand to induce cytokines, the main, non-redundant in vivo function of IL-2 is to limit lymphoidexpansion and promote peripheral tolerance. IL-2 is also important for the differentiation of T-helper(TH)2 cells. The importance of CD4+CD25+ regulatory T cells is increasingly recognized. These cellsare absent in Il2–/– mice, but the exact role in the development and/or function of these cells is notknown. FasL, Fas ligand; FLIP, FLICE/caspase-8 inhibitory protein; IFN-γ, interferon-γ; TGF-β;transforming growth factor-β; TNF, tumour-necrosis factor; Treg, T-regulatory cell.

© 2001 Macmillan Magazines Ltd40 | JANUARY 2002 | VOLUME 2 www.nature.com/reviews/immunol

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fatal lymphoproliferative disease that is more aggressivethan either IL-2 or Fas deficiency21. Another mechanismby which CD4+CD25+ regulatory T cells mediate sup-pression is by the production of immunosuppressivecytokines such as IL-4, TGF-β and IL-10. In some, butnot all, systems, the effect of the CD4+CD25+ cells isreversed by anti-cytokine interventions, but the roles ofthese cytokines in suppression remain controversial anddepend on the experimental systems used. IL-2 has beenreported to be required for differentiation of CD4+ cellsinto IL-4-producing T

H2 cells11,22. Additionally, it should

be noted that IL-2 induces TNF and IFN-γ, which, aswill be discussed, also have immunosuppressive actionsin addition to the well-known pro-inflammatory effects.

So, there are several mechanisms by which IL-2 sig-nals might negatively regulate activated T cells andimmune responses. But why do IL-2-deficient micespecifically develop colitis? Recently, missense mutationsof the NOD2 gene have been found to be associatedwith Crohn’s disease and BLAU SYNDROME23–25. But is therean IL-2/NOD2 connection? There are no data thataddress this issue presently, although both moleculescan regulate apoptosis. Still, exactly how IL-2 deficiencypredisposes to colitis remains uncertain. Nonetheless, itis clear that IL-2 has an essential immunoregulatory roleand its absence is accompanied by autoimmune mani-festations (see BOX 1 for a discussion of the role of IL-2and human autoimmune disease).

Immunosuppression by TNFOne of the most exciting breakthroughs for physicianscaring for patients with autoimmune diseases is thedevelopment of agents that interfere with TNF, eithermonoclonal antibodies or soluble receptors. Inhibitionof TNF has proven to be remarkably effective in treatingrheumatoid arthritis and Crohn’s disease, and this strat-egy has a strong scientific basis given the abundance ofliterature substantiating the pro-inflammatory role ofTNF. TNF activates macrophages, endothelial cells, syn-oviocytes and other cells, and induces the production ofother pro-inflammatory cytokines (for example, IL-1and IL-6) and chemokines (FIG. 3). Its overproduction

within the CD4+CD25+ T-cell population that exits thethymus within the first few days of life. Transfer ofCD4+ cells that are depleted of CD25+ cells intoimmunodeficient recipients leads to a spectrum ofautoimmune diseases17, the phenotype of whichdepend on background genes. However, transfer ofCD4+CD25+ cells within 10 days of the transfer ofCD4+CD25– T cells prevents disease. CD4+CD25+ lym-phocytes are present in the thymus, but it is not knownwhether the regulatory properties of this subset areacquired within the thymus or in the periphery,although there are data that indicate the former is thecase17,18. Importantly, these cells do not fit neatly intoeither the T

H1 or T

H2 category.

Although the protective effect of this subpopulationof lymphocytes has been well documented, the mecha-nism by which these cells attenuate autoimmunity isunclear, and an unequivocal connection with IL-2action has not been made. IL-2 has important functionsin controlling the expression of its high-affinity receptor,but is CD25 a marker indicating that the cells haveundergone activation or does IL-2 actually have animportant role in the physiology of these cells? Severallines of evidence support the latter possibility. The sup-pression that is mediated by CD4+CD25+ cells is associ-ated with inhibition of IL-2 production. Conversely, itcan be overcome by exogenous IL-2. Finally, and per-haps of most relevance, CD4+CD25+ cells are absent inIL-2-deficient mice. Transfer of normal T cells into IL-2or IL-2Rβ knockout mice results in removal of theabnormally activated T cells present in these mice19,20;in the former case, transfer of CD4+CD25+ cells sup-presses lymphoproliferation, despite the fact that theseregulatory cells produce little IL-2.

An interesting connection is that the CD4+CD25+

subset is the only population in the normal mouse thatconstitutively expresses CTLA-4 (cytotoxic T-lympho-cyte-associated protein 4; CD152) and IL-2 is known toupregulate CTLA-4 expression20. CTLA-4 binds B7.1(CD80) and B7.2 (CD86) on antigen-presenting cellsand, in contrast to CD28, has a vital negative regulatoryrole in T-cell function. CTLA-4-deficient mice develop a

BLAU SYNDROME

A rare, autosomal-dominantdisorder characterized bygranulomatous arthritis,uveitis, skin rash and cranial neuropathy.

Box 1 | IL-2 and human autoimmune disease

Although there is strong evidence supporting the role of interleukin (IL)-2 in maintaining lymphoid homeostasis andperipheral tolerance, there is a paucity of evidence indicating that IL-2 deficiency is the primary mechanism underlyingcommon autoimmune diseases. Mutation in the common γ-chain (γ

c) and Janus kinase 3 (JAK3; the tyrosine kinase that

binds γc) typically results in profound immunodeficiency, probably owing to the impairment in IL-7 signalling6.

Nonetheless, some patients with mutations in the IL-2 receptor and JAK3 have features of autoimmune disease41,42.In a recently described JAK3-deficient patient with severe combined immunodeficiency (SCID), this was found to beassociated with impaired expression of Fas ligand in the lymphocytes produced41–44. Lymphocytes from these patients,and from the respective knockout mice, have activation markers, but have defective in vitro proliferation — a phenotypethat is shared with STAT5-deficient cells44. This apparent paradox of poor in vitro proliferation associated with evidenceof in vivo activation is also found in cells from patients with systemic lupus erythematosus and rheumatoid arthritis45.These cells typically fail to produce normal IL-2 levels in vitro, but there is no evidence that this is owing to an intrinsicabnormality that is a causal defect in these disorders. The IL2 gene has also been linked to type I diabetes and resideswithin the Idd3 locus identified in non-obese diabetic mice. Disease susceptibility and resistance are linked with distinctpatterns of IL-2 glycosylation. Although some data indicate that this modification might alter the activity of IL-2, thishas not been firmly established46,47.


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clear examples of the immunosuppressive functions ofTNF. The issue, however, is not simply of academic inter-est. Despite the impressive positive effects of blockade ofTNF in rheumatoid arthritis, and even the salutatoryeffects of TNF antagonism in experimental autoimmuneencephalomyelitis (the mouse model for multiple sclero-sis), anti-TNF agents worsened disease in patients withmultiple sclerosis and precipitated demyelination in oth-ers26.Additionally, enhanced production of anti-nuclear,anti-DNA and anti-cardiolipin antibodies is associatedwith anti-TNF therapy, although systemic lupus erythe-matosus is still very uncommon. Therefore, understand-ing the counterintuitive consequences of inhibition ofTNF is of very practical concern.

There are several examples in which TNF amelio-rates animal models of lupus, but more unexpected wasthat TNF can also reduce the severity of prototypicmodels of T

H1 diseases, including diabetes, experimen-

tal autoimmune encephalomyelitis and arthritis. TNF

has been documented in rheumatoid arthritis, multiplesclerosis and in many animal models of autoimmunedisease26,27. Additionally, transgenic overexpression ofTNF results in arthritis, demyelinating disease, inflam-matory bowel disease and diabetes26,27. The arthritisassociated with transgenic expression of TNF, whichresults in synovial proliferation and erosive arthritis, isparticularly impressive as it occurs even in the absenceof lymphocytes.

However, about one-third of patients with rheuma-toid arthritis treated with anti-TNF agents do notrespond. Interestingly, collagen-induced arthritis can alsooccur in TNF-deficient mice, albeit with somewhatreduced severity28. Intriguingly, however, this arthropathyis associated with splenomegaly, lymphadenopathy andincreased memory CD4+ T cells. There is also an increasein activated lymph node B cells in TNF-deficient mice,indicating immunosuppressive functions of TNF.In fact, there are a number of model systems that provide

IL-6

Acute-phaseresponse

IL-1

Fever

Activation of APCsMHCAdhesion molecules

ShockiNOSActivation of neutrophils

Activation of endothelial cells Apoptosis

Inhibition of DCsDefectiveco-stimulation

Inhibition ofT-cell signalling

Cytokine inductionCytokine induction

Macrophage

Endothelial cells

O2•–

Immunostimulatory Immunosuppressive

Leukotriene

IL-6IL-10

TGF-β

Dendriticcell

LymphopaeniaSuppressionof proliferation

TH2

TNF

Figure 3 | Summary of immunosuppressive effects of TNF, the prototypic pro-inflammatory cytokine. The pro-inflammatoryactions of tumour-necrosis factor (TNF) on haematopoietic and non-haematopoietic cells are well recognized; however, it is also clearthat TNF can attenuate immune responses by inhibiting T-cell receptor signalling, promoting lymphoid T-cell apoptosis, inhibitingdendritic cell (DC) co-stimulation and inducing other cytokines that can inhibit cell-mediated immunity. APCs, antigen-presenting cells;IL, interleukin; iNOS, inducible nitric oxide synthase; MHC, major histocompatibility complex, O2

.–, superoxide; TGF-β, transforminggrowth factor-β.


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activates nuclear factor-κB (NF-κB), a key anti-apoptotictranscription factor. TNF has also been shown to inhibitT-cell-receptor signalling and even a short course of anti-TNF therapy can reverse this28 (FIG. 3). TNF also inducesthe expression IL-6, a pro-inflammatory cytokine thatalso promotes T

H2 differentiation through IL-4 induc-

tion. Additionally, IL-6 attenuates TH1 differentiation,

independent of IL-4 but dependent on suppressor ofcytokine signalling 1 (SOCS1; see below).

Finally, TNF also influences the function of antigen-presenting cells (APCs), but, again, its effects are compli-cated28. In some circumstances, TNF can activate APCs,augment antigen-presentation capability and upregulatethe expression of co-stimulatory molecules. However, itcan also inhibit the function of mature DCs, and mightinduce their apoptosis and impair antigen presentation.

So, although it is clear that TNF has profound pro-inflammatory effects, there is also no question that itcan have equally impressive immunosuppressiveeffects in vivo and in vitro; precisely which effect willpredominate is regulated by the timing and the durationof the exposure.

Complex effects of IFNs on autoimmunityBoth type 1 (IFN-α and IFN-β) and type 2 (IFN-γ)IFNs are used to treat infectious diseases33,34, but canprecipitate autoimmune disease (4–19% of patientstreated with these cytokines)4. Complications associ-ated with IFN treatment include autoimmune thy-roiditis, the development of antinuclear antibodies orlupus, rheumatoid arthritis and other arthropathies.Paradoxically, IFNs, especially type 1 IFNs are also usedto treat autoimmune disease; in fact, IFN-β is a corner-stone of therapy in multiple sclerosis35. In humans, type 1IFNs can induce IFN-γ production, promote T

H1 differ-

entiation and activate STAT4, although this is not thecase in the mouse (TABLE 1)36. Type 1 IFNs also inducesIL-15, which promotes the differentiation and develop-ment of natural killer and memory T cells. Additionally,type 1 IFNs induce production of chemokines in APCsand enhance DC maturation. Notably, type 1 IFNs aremade at substantial levels by some DC subsets37.Moreover, type 1 IFNs enhance DC maturation and canincrease antigen presentation. Why then, would such acytokine be useful in ameliorating a putative T

H1 dis-

ease? The predominant mechanism underlying the ther-apeutic activity of IFN-β in multiple sclerosis is notknown, but it is notable that type 1 IFNs inhibit IL-12

expression as a transgene in pancreatic islet cells or bysystemic administration can either precipitate diabetesor be protective. Specifically, TNF expression under thecontrol of the rat insulin promoter (RIP) along withexpression of CD80 in C57BL/6 mice caused dia-betes29,30. In non-obese diabetic mice (also abbreviatedto NOD, but this has nothing to do with NOD2, theCrohn’s susceptibility gene!), TNF expression in neona-tal islets alone resulted in acceleration of spontaneousdiabetes. Early systemic administration of TNF in thesemice also exacerbated disease. By contrast, transgenicexpression of TNF or systemic administration of TNF inadult NOD mice protected against diabetes, illustratingthe immunosuppressive actions of TNF and arguing fora role in promoting tolerance. The basis for these differ-ential effects of TNF has been attributed to factors suchas the effect of background genes. However, more recentdetailed studies indicate that both the timing and theduration of TNF expression are important in determin-ing pathogenic versus protective roles of TNF. Usingtransgenic mice in which pancreatic TNF expression iscontrolled by a tetracycline-responsive promoter, itwas found that only if TNF is expressed for more than21 days did CD8+ lymphocytes enter islets and destroythem26. Importantly, this was the case in both neonataland adult animals. A second model used TNF expres-sion controlled in the same manner in a model ofinsulitis in which lymphocytic choriomenigitis viralglycoprotein was expressed as an autoantigen under thecontrol of RIP31. Expression of TNF early in the diseaseenhanced inflammation, whereas late expression of theTNF transgene abrogated diabetes with a markeddecrease in autoreactive CD8+ lymphocytes.

A number of possible mechanisms for theimmunosuppressive effects of TNF have emerged andlymphocytes have been implicated as targets of thisimmunosuppressive action32. Administration of TNFcan result in lymphopaenia and TNF can clearly induceapoptosis – similar to Fas, the TNF receptor 1 (TNFR1)is a death-domain-containing receptor. Ligand bindingcauses recruitment of the death-domain-containingadaptor protein TRADD, which recruits FADD, leadingto caspase activation. Several groups have shown a roleof TNF in lymphocyte apoptosis, especially for CD8+

cells. However, TNF, TNFRI and TNFR2 knockout micedo not show spontaneous lymphadenopathy as found ingld or lpr mice28. Perhaps this is because, similar to IL-2,TNF has both pro- and anti-apoptotic actions; TNF

Table 1 | Examples of the complex and even counterintuitive effects of interferons

Cytokine IFN-α/β IFN-γ

Immunostimulatory Antiviral; increased antigen presentation; Activation of macrophages; increasedTH1 differentiation*; induction of IL-15, which antigen presentation and expression promotes NK and memory T-cell development of MHC class II; promotion of TH1and differentiation; increased DC maturation. differentiation.

Immunosuppressive Increased Fas; increased IL-10; decreased IL-12; Upregulation of SOCS1; anti-proliferativeinhibition of IL-12-signalling; anti-proliferative effects on myeloid and lymphoid cells.effects; differentiation of TR cells.

*IFN-α/β (Type I IFN) induces TH1 differentiation of human cells, but not mouse cells. DC, dendritic cell; IFN-γ, interferon-γ; IL, interleukin;MHC, major histocompatibility complex, NK, natural killer; SOCS1, suppressor of cytokine signalling 1; TCR, T-cell receptor; TH1, T helper 1;TR cells, T-regulatory cells.


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exogenous IFN-γ can serve as a protective factor inmodels of autoimmune disease. Possible mechanismsby which IFN-γ can attenuate autoimmunity will there-fore be briefly considered. A principal immunosup-pressive action of IFN-γ and other cytokines relates totheir ability to induce members of the SOCS family.These proteins contain Src homology 2 (SH2) domainsand function as classic feedback inhibitors6. They bindcytokine receptors and Janus-activated kinases (JAKs)to interfere with cytokine signal transduction, andmight also promote degradation of activated signallingmolecules by a ubiquitin/proteasome-dependent path-way. SOCS1 is of particular importance in attenuatingthe effects of IFN-γ. Mice in which the Socs1 gene hasbeen knocked out have extensive IFN-γ-dependentpathology and show excessive IFN-γ responses6.SOCS1 seems to be a key modulator of IFN-γ action

production and enhance IL-10 secretion. Additionally,recombinant IFN can inhibit IL-12 signalling. Theinhibitory effect is STAT1 dependent; in fact, in theabsence of STAT1, IFN-α induces IFN-γ production.Furthermore, IFN-α with IL-10 can promote the differ-entiation of CD25+CD4+ regulatory T cells38. Clearly,more work needs to be done to explain these compleximmunoregulatory effects of type 1 IFNs.

IFN-γ, the cytokine that defines TH1 differentiation,

has been extensively studied in both mouse and man.IFN-γ activates APCs and promotes T

H1 differentiation

by upregulating the transcription factor T-bet (TABLE 1)39.Again, despite extensive evidence incriminating it as amediator of disease, there is also evidence for protectiveroles; this seemingly conflicting information has beenreviewed in REFS 27,40. Suffice to say, that despite the roleof IFN-γ as the canonical T

H1 cytokine, endogenous and

IL-4 IL-10 TGF-βIL-2 TNF

IL-10 TGF-β

a Traditional view

b Revised view

Pro-inflammatory Anti-inflammatory

IL-4 IL-10 TGF-β

IL-10IL-4TGF-β

TNFIFN-γ

Pro-inflammatory Anti-inflammatory

Pro-inflammatory

Normal Autoimmunity

Normal Autoimmunity

Anti-inflammatory Pro-inflammatory Anti-inflammatory

TNFIFN-γIL-2

IL-2

IFN-γ

IL-2TNF IFN-γ

IFN-α/β IFN-α/β

?

IL-2 TNF IFN-γ IL-4

IFN-γ

TNF

Figure 4 | Cytokines in autoimmunity — good guys and bad guys switch sides! a | The traditional view of cytokines isdepicted.The cytokines that promote inflammation and immune responses, including interleukin (IL)-2, tumour-necrosis factor (TNF)and interferon-γ (IFN-γ), are shown wearing red shirts. The good guys, IL-10, transforming growth factor-β (TGF-β) and IL-4, inhibitcell-mediated immune responses and are depicted wearing blue shirts. Normally, there is an equilibrium between these cytokines(left), but in autoimmune disease (right), the bad guys overpower the good guys. b | A more realistic view is that these cytokines arenot especially loyal to their team; they switch sides, meaning that they do not have simple pro- or anti-inflammatory actions; this isshown by cytokines such as IL-2 and IFN-α/β wearing both red and blue shirts. The cartoon is meant to emphasize that manycytokines have complex actions. The same cytokine can promote immune and inflammatory responses in some circumstances andinhibit responses in other settings.


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sobering lesson that it is so easy to generate autoim-mune disease. Given the large numbers of apparentlyessential negative regulators, one might conclude thatautoimmunity does not routinely occur only because ofthe constant operation of a variety of inhibitory mecha-nisms. Indeed, the immune response seems to be in aconstant balancing act with itself.

With the many layers of inhibitory mechanisms usedby the vertebrate immune system, it comes as no surprisethat cytokines, even those thought to promote immuneresponses, have essential inhibitory functions.Apparently, they push on both ends of the balance in thiseffort to maintain effective host defence mechanismswhile preventing autoimmunity. As we rush to the clinicwith new therapies in hand, we need to keep in mindthat blocking cytokines are used to prevent autoimmunedisease in most experimental systems, and are not usedto treat established disease. Often paradoxical effectsoccur when cytokines are administered during thecourse of disease; the exact timing and duration of expo-sure to these cytokines can evidently have variable andunexpected effects. Apparently, cytokines can switchsides in the balancing act in surprising ways (FIG. 4). Nowthat cytokine and anti-cytokine therapies are increasinglyused, we need to learn in detail the complex action ofthese cytokines, including both their pro-inflammatoryand immunosuppressive properties. Prior to DaveMason, another Bard also put it well: “There is nothingeither good or bad, but thinking makes it so”(Hamlet Actii, Sc. 2). Clearly, there is much left to be done to dissectthe complex and highly interactive effects of cytokinessuch as IL-2, TNF and the IFNs — old players that onewould have thought we understood very well.

that permits the protective effects of this cytokine to occurwithout the risk of associated pathological responses.Once again, invoking the notion of timing and durationof cytokine exposure in the pathogenesis of autoimmunedisease, one can imagine scenarios in which low-levelexposure to IFN-γ or other cytokines might attenuatesubsequent responsiveness to these cytokines. Therefore,the net effect would be inhibitory. Additionally, it hasbeen noted that the variability of the effects of IFN-γdepend on the model used. In models in which completeFreund’s adjuvant (CFA) is used to induce disease, it hasbeen proposed that IFN-γ can have protective effects byinhibiting CFA-induced proliferation of myeloid cells.40

ConclusionsAlthough thymic selection and the deletion of autoreac-tive clones (CENTRAL TOLERANCE) has been put forth as aprincipal regulator of autoimmunity, we now know thatcentral tolerance is incomplete and that some degree ofautoreactivity is normal. Conversely, there are manyrecent examples in which loss of any one of an array ofinhibitory molecules results in autoimmune disease. Formany of these inhibitors, their function in the peripheryis of greater significance than their thymic function,which emphasizes the importance of peripheral toler-ance. Experiments with knockout mice indicate thatmany of these mechanisms are remarkably non-redun-dant because, despite the apparently vast number ofinhibitory pathways, the absence of even one of theseregulators results in inflammatory and autoimmunedisease. Although it is conceivable that species differ-ences or the use of inbred mice might overestimate thepropensity toward autoimmunity, it is nevertheless a

CENTRAL TOLERANCE

This form of tolerance refers tothe lack of self-responsivenessfound as lymphoid cellsdevelop, and is associated withthe deletion of autoreactiveclones. For T cells, this occurs in the thymus.

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AcknowledgementsWe thank B. Diamond, M. Lenardo, P. Plotz and W. Strober forreading this manuscript and providing helpful suggestions.

Online links

DATABASESThe following terms in this article are linked online to:LocusLink: http://www.ncbi.nlm.nih.gov/LocusLink/APAF1 | caspase-9 | CD28 | CD80 | CD86 | CD95 | CD152 |CD178 | FADD | FLIP | γc | Ifi202 | IFN-α | IFN-β | IFN-γ | IL-1 | IL-2 |IL-2R | IL-4 | IL-6 | IL-7 | IL-9 | IL-10 | IL-12 | Il-12p40 | IL-15 | IL-21 |lymphotoxin-α | NOD2 | Rag | Socs1 | SOCS1 | STAT1 | Stat3 |STAT4 | Stat5 | T-bet | TGF-β | TNFR1 | TNFR2 | TRADDOMIM: http://www.ncbi.nlm.nih.gov/Omim/autoimmune diseases | autoimmune lymphoproliferative disease |inflammatory bowel disease | multiple sclerosis | rheumatoidarthritis | SCID | systemic lupus erythematosus | type I diabetes |ulcerative colitis

FURTHER INFORMATIONEncyclopedia of Life Sciences: http:www.els.netautoimmune disease: pathogenesis | cytokines | cytokines asmediators of disease | immunoregulation | interferonsAccess to this interactive links box is free online.