Regulatory T cells friend or foe in immunity to infection?

REVIEWS REGULATORY T CELLS: FRIEND OR FOE IN IMMUNITY TO INFECTION? Kingston H. G. Mills Abstract | Homeostasis in the immune system depends on a balance between the responses that control infection and tumour growth and the reciprocal responses that prevent inflammation and autoimmune diseases. It is now recognized that regulatory T cells have a crucial role in suppressing immune responses to self-antigens and in preventing autoimmune diseases. Evidence is also emerging that regulatory T cells control immune responses to bacteria, viruses, parasites and fungi. This article explores the possibility that regulatory T cells can be both beneficial to the host, through limiting the immunopathology associated with anti-pathogen immune responses, and beneficial to the pathogen, through subversion of the protective immune responses of the host. CHRONIC INFECTIONS

Infections that persist for a long time, often indefinitely, and might not be cleared following the development of anti-pathogen immune responses. These include infection with HIV, hepatitis C virus and many parasites.

Immune Regulation Research Group, Department of Biochemistry, Trinity College, Dublin 2, Ireland. e-mail: [email protected] doi:10.1038/nri1485

Protection against infection is fundamental to the survival of all animals and is mediated by the immune system, which has evolved both innate and adaptive mechanisms to deal with invading microorganisms. The effector mechanisms used by the host to control infection include production of pro-inflammatory cytokines and chemokines, recruitment of inflammatory cells to the site of infection and activation of cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells, which lyse infected host cells (FIG. 1). Although these responses help to eliminate or slow the spread of the pathogen, if they are not tightly controlled, they can result in severe inflammation and collateral tissue damage1. A further potential for damage arises because the cells and molecules of the immune system that respond to pathogen-derived antigens can also respond to self-antigens, and if this reactivity is uncontrolled, it can result in autoimmune disease2. Inflammation and the immune response to pathogens are regulated by various host suppressor mechanisms, including the production of antiinflammatory cytokines by cells of the innate immune system in response to conserved pathogen-derived products3,4. However, recent evidence indicates that the adaptive immune system might also help to control infection-induced immunopathology through the generation of antigen-specific regulatory T cells (FIG. 1).

NATURE REVIEWS | IMMUNOLOGY

Therefore, regulatory T cells might have a host-protective role in immunity to infection. It is also possible that pathogens can exploit regulatory T cells to subvert the protective immune responses of the host. Although the infections caused by many pathogens are self-limiting in immunocompetent hosts, other pathogens can persist and cause CHRONIC INFECTIONS. In infections such as those caused by HIV, hepatitis C virus (HCV) and many parasites, the pathogen persists because the appropriate immune response required for pathogen elimination either fails to develop or is suppressed. Furthermore, there is evidence that the incidence of atopic diseases (such as allergy and asthma) and autoimmune diseases is lower in individuals infected with helminth parasites or exposed to microbial products as children 5,6. It seems that many, and possibly all, pathogens that cause PERSISTENT INFECTIONS or chronic infections have evolved strategies to subvert the immune responses of the host. These strategies include evasion of humoral and cellular immunity by ANTIGENIC VARIATION, interference with antigen processing or presentation, and subversion of phagocytosis and killing by cells of the innate immune system7. However, of most relevance to this discussion is a common immune-subversion strategy used by many pathogens that involves increasing the production of anti-inflammatory or immunosuppressive

VOLUME 4 | NOVEMBER 2004 | 8 4 1 ©2004 Nature Publishing Group

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PERSISTENT INFECTIONS

Non-lethal infections (such as infection with Bordetella pertussis) that are not cleared immediately (lasting for weeks or months rather than days) and are usually associated with the delayed development or suppression of anti-pathogen immune responses. In persistent viral infections, virus production occurs in a cell that is not killed by the virus (non-lytic); this includes chronic, latent and transforming infections.

responses, which normally function to control or terminate the protective effector immune responses of the host. This can be achieved in the following ways: through the production of molecules with homology to human cytokines, such as viral interleukin-10 (IL-10); through the direct induction of host immunosuppressive cytokines, such as IL-10 and transforming growth factor-β (TGF-β), which are produced by innate immune cells in response to pathogen-derived molecules3,8; or indirectly through the generation of regulatory T cells.

Our understanding of the role of regulatory T cells in immune homeostasis is far from complete, and there are several important unanswered questions. How do regulatory mechanisms control the development of autoimmunity but allow the same type of immune response to mediate protection against infection? What is the advantage to the host of inducing microorganism-activated regulatory T cells that suppress the immune responses that facilitate pathogen elimination? However, our knowledge has increased in the past few years, and several studies, mainly in mouse models of

• TH1-type pro-inflammatory response • Inflammation • Tissue damage • (Autoimmunity)

• TH2-type response • Inflammation • Tissue damage • (Allergic reactions)

ANTIGENIC VARIATION

Changes in the composition, structure or amino-acid sequence of antigenic components of pathogens recognized by T or B cells, which allow the microorganism to escape recognition by the adaptive immune response.

Pathogen invasion

TLR

Macrophage

Dendritic cell PRR

Pathogen

CD80/CD86

MHC class II

CD28 IgA

IL-1β, TNF and chemokines

TCR

Naive T cell

IgG

Neutrophil recruitment

B cell IFN-γ IgE IL-4, IL-5 and IL-6

TH2 cell

TH1 cell

CD8+ T cell

Eosinophil recruitment Epithelial cell

Cell lysis Regulatory T cell

Figure 1 | Immunity to infection. Innate immune effector cells, including macrophages, dendritic cells (DCs), neutrophils and natural killer (NK) cells (not shown), together with various protein components of the complement system, provide the first line of defence against invading microorganisms. Binding of conserved pathogen-derived molecules to pattern-recognition receptors (PRRs), such as Toll-like receptors (TLRs), on the cell surface of macrophages and DCs activates the production of pro-inflammatory cytokines and chemokines, which help to attract other effector cells to the site of infection. Pathogenactivated DCs present pathogen-derived antigens to T cells and promote the differentiation of naive T cells to various subtypes of effector CD4+ and CD8+ T cell. CD4+ T helper 1 (TH1) cells secrete interferon-γ (IFN-γ), which activates the anti-microbial activity of macrophages and helps B-cell production of IgG2a antibodies, whereas TH2 cells provide help for B-cell production of IgG1, IgA and IgE. CD8+ T cells lyse host cells infected with viruses, intracellular bacteria or parasites. Many of these responses can cause host tissue damage — for example, excessive inflammation from uncontrolled pro-inflammatory cytokine and chemokine production by innate immune cells and TH1 cells, eosinophilia and allergic reactions from uncontrolled TH2-cell responses, and killing of host cells by CD8+ cytotoxic T lymphocytes (CTLs) and NK cells. In normal individuals, regulatory T cells (both natural regulatory T cells circulating in the periphery and those induced by infection) help to control these effector functions and the associated damage to host tissues. IL, interleukin; TCR, T-cell receptor; TNF, tumour-necrosis factor.

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| NOVEMBER 2004 | VOLUME 4

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ATHYMIC

Mice that lack a thymus and are therefore deficient in T cells. NUDE

A mutation in mice that causes both hairlessness and defective formation of the thymus, which results in a lack of mature T cells.

infection with bacteria, viruses, parasites or fungi, have shown that regulatory T cells specific for pathogenderived antigens are induced during infection. Furthermore, studies involving depletion or transfer of CD4+CD25+ regulatory T cells (TReg cells) have provided evidence that natural TReg cells can influence the immune response to pathogens and the outcome of infectious disease. This article reviews the recent evidence for pathogen-specific regulatory T cells and their role in infection, focusing on the protective role of these cells in immunity to pathogens, as a means of limiting infection-induced immunopathology, as well as the exploitation of regulatory T cells by pathogens, as an immune-subversion mechanism to prolong pathogen survival in the host. The biology of regulatory T cells

Natural regulatory T cell

Natural and inducible regulatory T cells. It is now firmly established that there are both natural (or constitutive) and inducible (or adaptive) populations of regulatory T cells (FIG. 2), which probably have complementary and overlapping functions in the control of immune responses. However, the lineage relationship, if any, between these subtypes remains to be defined. The failure to identify definitive cell-surface markers for either population has compromised advances in the field and has led to some confusion about the precise nature of

CD4+ CD25+ FOXP3+ TReg cell Antigen (foreign or self)

Naive CD8+ CD25– T cell

CD8+ regulatory T cell

Antigen (foreign or self)

Naive CD4+ CD25– T cell

TH3 cell

Inducible regulatory T cells

Thymus

TR1 cell

Figure 2 | Natural and inducible regulatory T cells. Natural regulatory T cells express the cell-surface marker CD25 and the transcriptional repressor FOXP3 (forkhead box P3). These cells mature and migrate from the thymus and constitute 5–10% of peripheral T cells in normal mice. Other populations of antigen-specific regulatory T cells can be induced from naive CD4+CD25– or CD8+CD25– T cells in the periphery under the influence of semi-mature dendritic cells, interleukin-10 (IL-10), transforming growth factor-β (TGF-β) and possibly interferon-α (IFN-α). The inducible populations of regulatory T cells include distinct subtypes of CD4+ T cell: T regulatory 1 (TR1) cells, which secrete high levels of IL-10, no IL-4 and no or low levels of IFN-γ; and T helper 3 (TH3) cells, which secrete high levels of TGF-β. Although CD8+ T cells are normally associated with cytotoxic T-lymphocyte function and IFN-γ production, these cells or a subtype of these cells can secrete IL-10 and have been called CD8+ regulatory T cells.


the cells being studied in different laboratories. It seems that natural self-antigen-reactive CD4+CD25+ TReg cells develop in the thymus and then enter peripheral tissues, where they suppress the activation of other self-reactive T cells9,10. By contrast, IL-10- or TGF-β-secreting regulatory T cells, which are known as T regulatory 1 (TR1) or T helper 3 (TH3) cells respectively, are generated from naive T cells in the periphery after encounter with antigen presented by dendritic cells (DCs) that have an activation status distinct from those DCs that promote the differentiation of TH1 or TH2 cells. In addition to these well-defined populations of CD4+ regulatory T cells, there is also evidence for an immunosuppressive function of CD8+ regulatory T cells that secrete either IL-10 or TGF-β11,12. Furthermore, antigen-activated CD8+ γδ T cells can prevent insulin-dependent diabetes in mice13, and IL-10- and TGF-β-producing regulatory γδ T cells can suppress the anti-tumour activity of CTLs and NK cells14. In addition, natural killer T (NKT) cells, which co-express NK-cell and T-cell markers, can secrete regulatory cytokines, including IL-10 (REF. 15). Therefore, NKT and γδ T cells can also be categorized as regulatory T cells. In this review, ‘TReg cells’ denotes only CD4+CD25+ regulatory T cells and ‘T R1 cells’ denotes T regulatory 1 cells. When referring to other types of regulatory T cell or regulatory T cells in general, no abbreviations are used. Natural CD4+CD25+ TReg cells were first defined in 1995 by Sakaguchi and colleagues16, who showed that the transfer into ATHYMIC NUDE mice of lymphoid-cell populations from which CD4+ T cells expressing the α-chain of the IL-2 receptor (IL-2Rα; also known as CD25) had been removed caused spontaneous development of various T-cell-mediated autoimmune diseases. Furthermore, reconstitution with CD4+CD25+ T cells prevented the development of autoimmunity. This discovery, together with the work of Powrie and colleagues on a CD45RBlow population of T cells17, challenged traditional theories about clonal deletion being the only mechanism of self-tolerance and provided convincing evidence that self-antigen-reactive T cells that cause autoimmune diseases can be controlled through active suppression by natural TReg cells. CD4+CD25+ TReg cells, which constitute 5–10% of peripheral T cells in mice, are continuously produced in the thymus as a functionally mature T-cell population that includes cells with immunosuppressive activity. However, CD25 is not a definitive marker of natural regulatory T cells; CD25 is an activation marker for T cells and is therefore also expressed by effector TH1 and TH2 cells, and suppressive function has also been documented for CD25– T cells. These observations led to attempts to find alternative markers for regulatory T cells. Putative markers for regulatory T cells include cell-surface expression of CD38, CD62L, CD103 or glucocorticoid-induced tumournecrosis factor (TNF) receptor (GITR), or low levels of cell-surface CD45RB expression or intracellular expression of the transcriptional repressor FOXP3 (forkhead box P3)17–19. FOXP3 seems to be the most promising marker of natural regulatory T cells, and recent studies have shown that transfection of CD4+CD25– T cells


REVIEWS

ANERGIC

A state of unresponsiveness by T or B cells to antigens. After stimulation, anergic T cells cannot produce interleukin-2 or proliferate, even in the presence of co-stimulatory signals.

with Foxp3 confers them with intracellular regulatory activity20. T-cell receptor (TCR) engagement seems to be necessary for optimal suppressive activity, and it has been assumed that circulating CD4+CD25+ TReg cells are activated by the recognition of self-antigens in vivo9. However, evidence that natural regulatory T cells are antigen specific is still limited. A unique cytokine-production profile, rather than the expression of cell-surface markers, has been used to define at least two populations of inducible regulatory T cells. Although it had been recognized for some time that T cells with suppressive or ANERGIC activity could be generated in vivo in certain situations — for example, in oral tolerance 21,22 or during infection with certain pathogens, such as rabies virus23, Brugia malayi 24 and Mycobacterium tuberculosis 25 — it was not until the mid-1990s that these cells were given a unified nomenclature. Weiner and colleagues showed that the induction of oral tolerance and the prevention of TH1-cell-mediated autoimmune diseases by feeding self-antigens were associated with the generation of TGF-β-secreting T cells in the gut26. These T cells, which were distinct from TH2 cells in that they produced large amounts of TGF-β and varying amounts of IL-4 and IL-10, were named TH3 cells. In 1997, Groux et al. showed that repeated in vitro stimulation of T cells isolated from ovalbumin-specific TCR-transgenic mice with their cognate antigen in the presence of IL-10 resulted in the expansion of a population of regulatory T cells that produced large amounts of IL-10 and could suppress TH1-cell responses and TH1-cell-mediated autoimmune diseases27; they called these cells TR1 cells. More recently, it has been shown that antigen-specific TR1 cells can be generated in vivo during certain infections and that IL-10 might be a differentiation factor rather than a growth factor for these cells28. Because TH2 cells secrete the immunosuppressive or antiinflammatory cytokines IL-10 and IL-4, these cells might also have regulatory function, as well as effector function, but they are distinguished from T H3 and TR1 cells by the production of large amounts of IL-4 and smaller amounts of IL-10, as well as a lack of TGF-β production. Targets of suppressor activity. Immunity to intracellular pathogens is mediated by CD4+ TH1 cells and CD8+ CTLs, whereas immunity to extracellular pathogens is mediated by antibodies and TH2 cells. Innate immune responses also have a protective role early in infection and instruct the adaptive immune response (FIG. 1). Each of these effector mechanisms can be suppressed by natural and inducible regulatory T cells. It has been shown that T R1 cells and CD4+CD25+ TReg cells can suppress the proliferation of and cytokine production by naive CD4 +CD25– T cells or antigen-specific TH1 or TH2 cells in vitro28–33. There is more-limited evidence that regulatory T cells can suppress pathogen-specific T cells in vivo; existing evidence includes TR1-cell-mediated suppression of interferon-γ (IFN-γ) production by TH1 cells in response to Bordetella pertussis 28 and CD4+CD25+

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TReg-cell-mediated suppression of CD4+CD25– T cells responding to Leishmania major 29. More recently, it has been shown that CD25+ T cells can suppress the activation of CD8+ T cells in vitro 34, as well as secondary CD8+ T-cell responses to Listeria monocytogenes 35 and herpes simplex virus (HSV)36 in vivo. Finally, there is evidence that regulatory T cells can suppress the recruitment and activation of innate immune cells induced by Helicobacter hepaticus that leads to inflammatory pathology in the colon37. Therefore, the targets of suppressor activity by regulatory T cells are immune responses that confer protection against infection with microorganisms and also responses that can cause collateral damage to host tissue during infection. Mechanisms of suppression. The mechanism of the suppressive function of natural and inducible regulatory T cells is still debated, but in different model systems, suppressive activity has been shown to be mediated either through secretion of immunosuppressive cytokines or through cell–cell contact (FIG. 3). Many studies have shown that the suppression mediated by TR1 or TH3 cells can be reversed using antibodies specific for IL-10 or TGF-β. IL-10 inhibits the production of TNF and IL-12 by DCs and macrophages, whereas TGF-β inhibits TH1-cell responses through its effects on expression of the transcription factor T-bet and the IL-12R38–40. It has been reported that the production of TGF-β by regulatory T cells induces IL-10 production by TH1 cells through SMAD4-induced activation of the IL-10 promoter41. This indicates that there might be interdependent, as well as distinct, roles for IL-10 and TGF-β in the immunosuppressive function of inducible regulatory T cells. Cytokine-mediated suppression might also operate at the level of the antigen-presenting cell, because IL-10 and regulatory T cells can inhibit the expression of MHC class II and co-stimulatory molecules by DCs40,42. The suppressive mechanisms of CD4+CD25+ TReg cells are not clear, but there is evidence that cell–cell contact is required and that expression of the inhibitory costimulatory molecule CTLA4 (CTL antigen 4) might be involved43. However there is also conflicting evidence concerning roles for IL-10 and secreted or cell-surface TGF-β29,43,44. Finally, it has also been proposed that regulatory T cells might inhibit pathogenic effector T-cell responses by competing for shared resources in the normal immune system45. So, although the mechanisms of suppression by TR1 and TH3 cells seem to be mediated mainly by cytokines, CD4+CD25+ TReg cells might use many and as-yet-unidentified mechanisms to mediate suppression. Pathogen-specific regulatory T cells

Studies involving cell depletion and transfer, as well as cytokine-knockout or -inhibition experiments, have provided considerable indirect evidence of a role for inducible (TABLE 1) and natural (TABLE 2) regulatory T cells during infection. However, there are still only a small number of reports showing that regulatory T cells are specific for pathogen-derived antigens.


REVIEWS

Natural regulatory T cells

Inducible regulatory T cells

TH3 cell

TReg cell (CD4+ CD25+FOXP3+)

TR1 cell

CD8+ regulatory T cell

?

IL-10 and/or TGF-β

TCR

↓ MHC and co-stimulatory molecules ↓ APC function ↓ Inflammatory cytokines

MHC class II

Dendritic cell

CTLA4

CD80/ CD86 CD28

Cell–cell contact

CD25– cell

↓ Proliferation

TH1 cell

TH2 cell

↓ Proliferation ↓ IFN-γ

↓ Proliferation ↓ IL-4

CD8+ cell ↓ CTL activity ↓ IFN-γ

Figure 3 | Targets of regulatory T cells and mechanisms of suppression. CD4+CD25+FOXP3+ (forkhead box P3) natural regulatory T cells (TReg cells) inhibit the proliferation of CD25– T cells. The mechanism of suppression seems to be multifactorial and includes cell–cell contact. CD4+CD25+ TReg cells express cytotoxic T-lymphocyte antigen 4 (CTLA4), which interacts with CD80 and/or CD86 on the surface of antigen-presenting cells (APCs) such as dendritic cells (DCs), and this interaction delivers a negative signal for T-cell activation. There is also some evidence that secreted or cell-surface transforming growth factor-β (TGF-β) or secreted interleukin-10 (IL-10) might have a role in suppression mediated by natural regulatory T cells. Natural killer T (NKT) cells (not shown) and inducible populations of regulatory T cells, which include T regulatory 1 (TR1) cells, T helper 3 (TH3) cells and CD8+ regulatory T cells, secrete IL-10 and/or TGF-β. These immunosuppressive cytokines inhibit the proliferation of and cytokine production by effector T cells, including TH1 cells, TH2 cells and CD8+ cytotoxic T lymphocytes (CTLs), either directly or through their inhibitory influence on the maturation and activation of DCs or other APCs. IFN-γ, interferon-γ; TCR, T-cell receptor.

TR1 cells and TH3 cells. Although many studies have shown that pathogens, in particular those that cause chronic infections or are associated with immunosuppression, induce production of the regulatory cytokines IL-10 and TGF-β, the cellular source of these cytokines has not always been defined. In some cases, it has been shown that innate immune cells, usually macrophages or more rarely DCs, are the source, whereas in other studies, it has been shown that these cytokines are produced by T cells8. However, the distinction between IL-10 or TGF-β production by TH2 cells versus regulatory T cells has not always been made. The definitive demonstration of antigen-specific regulatory T cells depends on the generation of antigen-specific T-cell clones or on careful ex vivo intracellular cytokine staining of antigen-stimulated T cells, showing high levels of IL-10 production, no IL-4 production and low (human) or no (mouse) IFN-γ production. The first definitive observations of inducible antigenspecific TR1-cell clones generated during infection were made using mice infected with B. pertussis 28 and


humans infected with HCV46 or the nematode parasite Onchocerca volvulus 47,48. The study involving B. pertussis 28 showed direct evidence of suppression of TH1 cells by TR1-cell clones specific for bacterial antigens, and the human studies46–48 showed indirect evidence of a role for TR1 cells through increased IFN-γ production in the presence of IL-10-specific antibodies. The studies with B. pertussis28 and of virus-specific CD8+ regulatory T cells in chronic HCV infection49 indicate that antigen-specific regulatory T cells are recruited to the site of infection in mucosal tissues. More recently, antigen-specific TR1 cells have been described in several other chronic infections, including infection with Epstein–Barr virus (EBV)50, M. tuberculosis 25,51,52, HIV53 and murine leukaemia virus, which is a mouse model of AIDS54. It has also been shown that IL-10-producing regulatory T cells can be induced in vitro by DCs stimulated with phosphatidylserine isolated from Schistosoma mansoni 55. Although the problems of cultivating and cloning antigen-specific regulatory T cells in vitro have hampered advances in this area, it is tempting to speculate


REVIEWS that regulatory T cells are induced during infection with most, if not all, pathogens, in particular those that cause persistent or chronic infections. Natural regulatory T cells. Most studies of CD4+CD25+ TReg cells in infection have shown a role for these cells in controlling anti-pathogen immunity, but few studies have shown that they are specific for pathogen-derived antigens (TABLE 2). CD4+CD25+ TReg cells specific for pathogen-derived antigens have been shown to accumulate at the site of infection in the dermis soon after infection with L. major and to suppress IFN-γ production and the ability of effector T cells to eliminate the parasite from the host29. CD4+CD45RBlow regulatory T cells from mice infected with H. hepaticus prevent the development of intestinal inflammation induced by the transfer of CD4+ T cells from IL-10-deficient mice to recombinationactivating gene (RAG)-deficient mice56. The observations that CD4+CD45RBlow regulatory T cells from H. hepaticusinfected wild-type mice inhibit IFN-γ production by

T cells from IL-10-deficient mice and produce IL-10 after exposure to H. hepaticus-derived antigens in vitro indicate that these regulatory T cells, rather than being endogenous, might be a memory population resulting from previous exposure to bacterial antigens. Protective role of regulatory T cells in infection

There is convincing evidence of a protective role for regulatory T cells against autoimmune diseases, allograft rejection and allergy, in these situations, they suppress potentially pathogenic immune responses mediated by effector TH1 cells, TH2 cells or CTLs2,16,27,57–59. As these effector T-cell responses also have important roles in protection against pathogens, it might seem counterintuitive that regulatory T cells could have a protective role in infection. However, in many infectious diseases, immune responses to the pathogen can result in collateral damage to host tissues, and immunoregulatory mechanisms, including the induction of regulatory T cells, are essential to control this immunopathology.

Table 1 | Pathogen-induced regulatory T cells and their role in infection Pathogen

Cell type

Antigen T-cell specific?* clones

Cytokine secreted

Responses suppressed

Manipulation of regulatory cells

Effect on immune response, immunopathology and pathogen load

References

Friend virus

Mouse TR1 cell

ND

ND

IL-10

CD8+ T-cell IFN-γ production

Depletion with GITR-specific antibody

Increases IFN-γ-secreting CD8+ T cells and reduces viral load

84

Murine leukaemia virus

Mouse TR1 CD4+CD25+ cell

ND

ND

IL-10

ND

CD25+ T-cell depletion

Prevents spleen pathology and disease progression but has no effect on viral load

54

HCV

Human TR1cell

Yes

Yes

IL-10

PBMC IFN-γ production

ND

ND

46,60

HCV

Human CD8+ T cell

Yes

ND

IL-10

Antigen-specific PBMC proliferation

ND

ND

49

EBV

Human TR1 cell

Yes

IL-10

T-cell proliferation ND and IFN-γ production to recall antigen

ND

50

HIV

Human CD8+ T cell

Yes

ND

TGF-β

Vaccinia-virusspecific CD8+ T-cell IFN-γ production

ND

11

Bordetella pertussis

Mouse TR1 CD4+CD25+ cell

Yes

Yes

IL-10 +/– TGF-β

Antigen-specific Cell transfer TH1-cell proliferation and IFN-γ production in vitro and in vivo

Suppresses TH1-cell response and increases bacterial load

28

Bordetella pertussis

Mouse TR1 cell

Yes

Yes

IL-10

TH1-cell IFN-γ production

Defective TR1 cells in TLR4-deficient mice

Increases TH1-cell response, lung inflammation and bacterial load

67

Helicobacter hepaticus

Mouse CD45RBlow T cell

Yes

ND

IL-10

Antigen-specific IL-10-deficient CD4+ T-cell IFN-γ production

CD45RBlow-cell transfer

Prevents colitis in IL-10deficient mice

56

Yes

ND

IL-10

Allogeneic CD4+ T-cell proliferation

ND

ND

25,51

Yes Human TH3 and/or TR1 cell

Yes

IL-10 +/– TGF-β

PBMC proliferation

ND

ND

47,48

Viruses

ND

Bacteria

Mycobacterium Human tuberculosis TR1 cell Parasites Onchocerca volvulus

*Demonstration that regulatory cells respond to the pathogen or pathogen-derived antigens in vitro. EBV, Epstein–Barr virus; GITR, glucocorticoid-induced tumour-necrosis factor receptor-related protein; HCV, hepatitis C virus; IFN-γ, interferon-γ; IL-10, interleukin-10; ND, not determined; PBMC, peripheral-blood mononuclear cell; TGF-β, transforming growth factor-β; TH, T helper; TLR4, Toll-like receptor 4; TR1, T regulatory 1.

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REVIEWS

MIXED CRYOGLOBULINAEMIA

Cryoglobulins are antibodies that precipitate at cold temperatures and dissolve on warming. Mixed cryoglobulinaemia is a B-cell proliferative disorder that is characterized by polyclonal B-cell activation and autoantibody production. Patients with mixed cryoglobulinaemia have circulating cryoproteins and inflammation of small blood vessels, with inflammatory changes prominent in the skin (vasculitis), and might have renal and neurological involvement.

Viruses. IL-10-producing CD4+ and CD8+ regulatory T cells have been shown in HCV infection, and there is indirect evidence that both CD4+ and CD8+ T-cell populations can inhibit HCV-specific T cells in chronically infected individuals46,49,60. However, it has been suggested that HCV-specific CTLs that home to the liver produce IL-10 and help to reduce liver inflammation49. Furthermore, HCV-infected patients with reduced numbers of CD4+CD25+ T cells often develop an autoimmune syndrome, known as MIXED CRYOGLOBULINAEMIA, which is characterized by B-cell proliferation and autoantibody production 61. So, although regulatory T cells can prevent viral clearance, they also prevent immunopathology and the development of autoimmunity.

More-direct evidence of a role for regulatory T cells in preventing immunopathology has come from studies using mouse models of viral infection. Infection of mice with Theiler’s virus induces a DEMYELINATING disease mediated by CD4+ T cells, and the transfer of virus-specific CD8+ regulatory T cells has been shown to prevent inflammation and the pathogenic effects of the CD4+ T cells12. In footpad infection of mice with HSV, removal of CD25+ T cells increases the virus-specific CD8+ T-cell response and improves viral clearance36. However, TH1-cell responses and the severity of T-cell-mediated lesions in the cornea of HSV-infected mice were increased if mice were depleted of CD25+ T cells before infection62. CD4+CD25+ TReg cells therefore seem to reduce the severity of immune-mediated inflammatory

Table 2 | Natural CD4+CD25+ TReg cells and their role in infection Pathogen

Species

Antigen specific?*

Cytokine secreted

Responses suppressed‡

Manipulation of regulatory T cells

Effect on immune response, immunopathology and pathogen load

References

Herpes simplex virus

Mouse

ND

IL-10

Antigen-specific CD4+ T-cell IFN-γ production

In vivo depletion

Increases TH1-cell response, CD4+ T-cell infiltration and stromal keratitis

62

HIV

Human

ND

ND

Antigen-specific CD4+ and CD8+ T-cell proliferation and cytokine production

In vitro depletion

Increases HIV-specific CD8+ T-cell IFN-γ production

81,82

Helicobacter hepaticus

Mouse

ND

ND

Innate immune responses in vivo

Transfer

Prevents H. hepaticus-induced intestinal inflammation (reversed by IL-10- or TGF-β-specific antibody) but has no effect on bacterial colonization

37

Helicobacter pylori

Mouse

ND

ND

ND

In vivo depletion

Increases CD4+ T-cell IFN-γ production and gastritis but decreases bacterial load

68

Helicobacter pylori

Human

ND

ND

Antigen-specific CD25low T-cell proliferation and IFN-γ production

In vitro depletion

Increases antigen-specific T-cell proliferation

30

Schistosoma mansoni

Mouse

ND

IL-10

Naive T-cell proliferation

Transfer

Reduces liver damage and increases survival

77

Leishmania major

Mouse

Yes

IL-10

CD25– T-cell proliferation and IFN-γ production in vitro and in vivo

Transfer to IL-10deficient or wildtype mice

Increases non-healing skin lesions and parasite load

29

Leishmania major

Mouse

ND

ND

ND

Depletion from splenocytes before transfer to SCID mice

Increases antigen-specific lymph node IFN-γ and IL-4, severity of colon lesions and parasite load

78

Plasmodium yoelii

Mouse

ND

ND

ND

In vivo depletion

Increases splenocyte proliferation and decreases parasite load

91

Plasmodium berghei

Mouse

ND

ND

ND

In vivo depletion

Decreases parasite load

92

Pneumocystis carinii

Mouse

ND

ND

ND

Transfer

Increases pathogen load

69

Candida albicans

Mouse

ND

IL-10, IL-4 and TGF-β

ND

Transfer

Decreases IFN-γ-secreting cells in vivo and decreases fungal load

70

Viruses

Bacteria

Parasites

Fungi

*Demonstration that regulatory cells respond to the pathogen or pathogen-derived antigens in vitro. None of these studies showed antigen-specific CD4+CD25+ regulatory T (TReg) cells. ‡In vitro unless otherwise stated. IFN-γ, interferon-γ; IL, interleukin; ND, not determined; SCID, severe combined immunodeficient; TGF-β, transforming growth factor-β; TH1, T helper 1.



REVIEWS

DEMYELINATING

Causing damage to the myelin sheath surrounding nerves in the brain and spinal cord, which affects the function of the nerves involved. Demyelination occurs in multiple sclerosis, a chronic disease of the nervous system affecting young and middleaged adults, and in experimental autoimmune encephalomyelitis, which is a mouse model of multiple sclerosis. SECONDARY INFECTION

An infection in a host already infected with another pathogen, often caused by opportunistic pathogens in immunodeficient or immunosuppressed hosts. TOLL-LIKE RECEPTOR

(TLR). A member of a family of receptors that recognize pathogen-associated molecular patterns. TLRs recognize conserved molecular patterns that are common to large groups of microorganisms and/or viruses.

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lesions by preventing the induction of pathogenic CD4+ T cells and by limiting the migration of these cells to inflammatory sites. Therefore, in chronic viral infections, regulatory T cells might be beneficial to the host by maintaining a balance between efficient effectors and memory responses, but with a low level of inflammation that causes minimal damage to the host. Bacteria. Indirect evidence of a protective role for IL-10producing regulatory T cells in host defence against bacteria-induced immune-mediated pathology has come from studies showing that disease severity is increased in IL-10-deficient mice. IL-10-deficient mice succumb more readily to primary and SECONDARY INFECTION with L. monocytogenes than control mice; the IL-10-deficient mice have an increased number of cells in the inflammatory infiltrate, increased production of pro-inflammatory cytokines in the brain and increased severity of brain lesions63. Peritonitis and mortality from infection with Escherichia coli is increased in IL-10-deficient mice, despite accelerated clearance of the bacteria compared with wild-type animals64. Colonization of the gastric mucosa by Helicobacter pylori is reduced in IL-10deficient mice, but the severity of chronic active gastritis is significantly greater than in wild-type mice65. Similarly, IL-10-deficient mice infected with H. hepaticus develop severe inflammation that is associated with IL-12 production and TH1-cell responses66. The IL-10 that helps to limit inflammation during bacterial infection might, in part, be derived from innate immune cells. However, it has been shown that the induction of IL-10 production by macrophages and DCs in response to certain pathogen-derived molecules facilitates the induction of TR1 cells, thereby amplifying the effect of IL-10 produced by innate immune cells 28. Indeed, studies with mice that lack functional TOLL-LIKE RECEPTOR 4 (TLR4-defective mice) have indicated that IL-10 produced by both innate immune cells and TR1 cells might help to limit inflammatory pathology in the lungs induced by B. pertussis infection. B. pertussis stimulates IL-10 production by DCs and macrophages and generates TR1 cells in the respiratory tract of infected mice. However, TLR4-defective mice have reduced IL-10 production by DCs and macrophages and do not generate TR1 cells when infected with B. pertussis67. These mice have significantly greater cellular infiltrates, lung damage and bacterial loads than wild-type mice, which has led to the hypothesis that the induction of TR1 cells helps to limit inflammatory pathology and thereby improves pathogen elimination by preventing damage to the ciliated epithelial cells required for removal of the bacteria from the lungs. Direct evidence of a role for CD4+CD25+ TReg cells in the prevention of intestinal inflammation has come from the demonstration that infection with H. hepaticus induces a population of CD4+CD45RBlow regulatory T cells that inhibit the development of colitis in IL-10-deficient mice56. Removal of CD25+ T cells from the lymph-node cells used to reconstitute athymic mice before infection with H. pylori reduced the bacterial load but increased the severity of gastritis 68.


Furthermore, transfer of CD4+CD25+ TReg cells from normal mice to Rag2 –/– mice prevented intestinal inflammation induced by infection with H. hepaticus, through IL-10- and TGF-β-dependent mechanisms37. The regulatory T cells did not affect bacterial colonization in the gut; instead, the protective effect of the regulatory T cells seemed to be mediated by suppressing T-cell-dependent and innate inflammatory responses, including the recruitment of neutrophils and macrophages and the activation of NK cells in the intestine37. Fungi. Pneumocystis carinii causes pneumonia in immunocompromised individuals. In a mouse model, transfer of CD4+CD25– T cells to Rag2–/– mice infected with P. carinii reduced the pathogen load, but these mice developed severe lung inflammation and a fatal wasting disease. Co-transfer of CD4+CD25+ T cells prevented lung inflammation and the development of disease induced by CD4+CD25– T cells but increased the pathogen load69. A similar situation has been reported for infection with Candida albicans. TH1 cells mediate protection against C. albicans, and in the absence of CD4+CD25+ T cells, which are not induced in CD86- or CD28-deficient mice or in situations in which IL-10mediated signalling is deficient, the fungal growth is reduced, but inflammatory pathology is increased70. Furthermore, transfer of IL-10- and TGF-β-secreting CD4+CD25+ T cells decreased inflammation in CD86deficient mice. In a separate study, an absence of TLR2 was associated with impaired IL-10 production, fewer CD4+CD25+ TReg cells and more inflammatory infiltrate, but lower pathogen load in C. albicans-infected mice71. So, although regulatory T cells might compromise fungal clearance, they can also be beneficial to the host by limiting infection-induced pathology. Parasites. In human malaria, polymorphisms in the TNF promoter have been associated with disease severity; among children with severe malaria, those with the TNF-308A allele had lower plasma levels of IL-10 than of TNF72. Furthermore, higher ratios of IL-10 to TNF in children with mild malaria compared with those who have severe malaria indicate a role for IL-10 in controlling the excessive inflammatory activities of TNF73. Although these studies do not provide direct evidence that IL-10 is produced by parasite-specific regulatory T cells, it has been suggested that regulatory T cells might contribute to the anti-inflammatory cytokine pool that controls TNF-mediated inflammation in malaria1. These observations are complemented by studies in mice showing that infection with Plasmodium chabaudi chabaudi is more severe in IL-10-deficient mice than in wild-type mice and that this is associated with increased inflammation, including increased production of TNF, IFN-γ and IL-12 (REF. 74). Treatment of these infected, IL-10-deficient mice with TNF-specific antibodies increases survival74. Furthermore, treatment of infected mice with a neutralizing antibody specific for TGF-β exacerbated infection with Plasmodium berghei and P. chabaudi chabaudi, and treatment with recombinant


REVIEWS

SEVERE COMBINED IMMUNODEFICIENT

(SCID). Mice with this immunesystem defect do not have B or T cells and therefore can accept tumour cells from another species without rejection.

TGF-β slowed the rate of parasite growth and increased survival75. CD4+CD25+ T cells and CD8+ T cells from malaria-infected mice secreted high levels of TGF-β in response to parasite-derived antigens in vitro76, which indicates that antigen-specific CD8+ regulatory T cells might help to control malaria-induced inflammation. IL-10-producing CD4+CD25+ T cells are induced in mice during infection with S. mansoni, and these regulatory T cells (as well as the production of IL-10 by innate immune cells) help to protect the host from the hepatocyte damage induced by S. mansoni eggs and to prevent death from the infection through immune-mediated pathology77. Similarly, depletion of CD4+CD25+ T cells increased the parasite load, severity of colon lesions and colitis in L. major-infected SEVERE COMBINED IMMUNODEFICIENT (SCID) mice adoptively transferred with splenocytes78. Although infection of IL-10-deficient mice with L. major is associated with increased parasite-specific immune responses and pathogen clearance from the host, this sterilizing cure results in a loss of immune memory and therefore of resistance to re-infection by the same parasite29. Therefore, regulatory T cells seem to control the immune response sufficiently to contain but not eradicate the infection, thereby suppressing potentially pathogenic T-cell effector responses but allowing the maintenance of T-cell memory. Pathogen immune evasion

Although regulatory T cells are beneficial to the host by preventing immunopathology and enabling the development of immune memory, they can also be beneficial to the pathogen, enabling it to establish a chronic infection. Many pathogens have evolved strategies that facilitate their persistence, largely through their ability to evade or subvert the host immune response. One strategy is to induce a state of immunosuppression, either through direct interference with host immune effector mechanisms or through the production of immunosuppressive cytokines. Many viruses produce antagonists of pro-inflammatory cytokines or their receptors, or molecules that are homologous to host IL-10 or TGF-β, or they stimulate the production of anti-inflammatory cytokines by host macrophages or other innate immune cells3,8. It has recently been recognized that parasite-induced immunosuppression can also be extended to the induction of T cells with suppressor activity, including natural and inducible regulatory T cells. Viruses. Most patients infected with HCV remain persistently infected despite the induction of HCV-specific antibodies and T-cell responses. Many chronically infected patients remain disease free for decades; others go on to develop cirrhosis of the liver and, in certain cases, hepatocarcinoma. It has recently been shown that patients with chronic HCV infection have circulating HCV-specific CD4+ TR1 cells46 and CD8+ regulatory T cells49. These regulatory T cells seem to be capable of inhibiting anti-viral immunity, because the addition of a neutralizing IL-10-specific antibody significantly increased HCV-specific IFN-γ production by T cells


in vitro60. Furthermore, there is a higher frequency of CD4+CD25+ TReg cells in patients with chronic infections than in those that have cleared the infection79. These CD4+CD25+ TReg cells could suppress HCV-specific CTL responses, indicating that natural TReg cells could also contribute to chronic infection by suppressing protective immune responses. This is consistent with findings in HSV-infected mice, in which CD4+CD25+ TReg cells suppress virus-specific CD8+ T-cell responses and delay viral clearance36. Retroviruses, such as HIV, usually persist for the lifetime of the infected host and escape immunity by antigenic variation. The immunodeficiency syndrome in the later stages of AIDS is the direct result of a reduction in the number of CD4+ T cells. However, even before the numbers of CD4+ T cells start to decline, immune responses to HIV and unrelated pathogens are suppressed. One explanation for this is the switch from TH1- to TH2-cell-dominated responses that has been observed to occur during disease progression80. Alternatively, activation of regulatory T cells that inhibit TH1-cell and CTL responses in vivo might explain the immunosuppression that occurs during retroviral infection before depletion of CD4+ T cells. Individuals with progressive or active HIV replication have a high frequency of IL-10-producing CD4+ T cells; these cells include those that produce IL-10 constitutively and those that only produce it after stimulation with the HIV protein Gag (group-specific antigen)53. Furthermore, CD4+CD25+ T cells from HIV-infected individuals suppress the proliferation of and cytokine production by CD8+ and CD4+ T cells in response to HIV antigens81, and depletion of CD4+CD25+ TReg cells from peripheral-blood mononuclear cells (PBMCs) increases the frequency of CD8+ and CD4+ T cells that secrete IFN-γ in response to HIV81,82 and cytomegalovirus82 antigens. In addition, HIV antigens induce TGF-β-secreting CD8+ regulatory T cells that inhibit IFN-γ secretion by CD8+ T cells specific for vaccinia virus. So, regulatory T cells specific for HIV antigens might contribute to general immunosuppression during retroviral infection11, and this conclusion is supported by studies using animal models of retroviral infection. Infection of cats with feline immunodeficiency virus is associated with the activation of CD4+CD25+CTLA4+ TReg cells that inhibit the proliferation of and IL-2 production by CD4+CD25– T cells from normal cats31. Furthermore, ablation of IL-10-secreting regulatory T cells in mice prevented the progression of mouse AIDS (an immunodeficiency syndrome induced by murine leukaemia virus)54. Persistent infection of mice with the Friend retrovirus is associated with a decreased ability to develop anti-tumour immune responses83. IL-10-producing CD4+ T cells from mice persistently infected with Friend virus suppress IFN-γ production by CD8+ T cells84. Similarly, in humans infected with EBV, TR1 cells are induced that are specific for LMP1 (latent membrane protein 1) of EBV, and these cells inhibit TH1-cell responses to other EBV proteins, which might facilitate viral persistence and promote the induction of EBV-associated tumours50.


REVIEWS These findings indicate that, in some cases, virus-specific regulatory T cells not only prevent pathogen elimination but also can promote a generalized state of immune suppression in vivo such that the host is more susceptible to secondary infections with other pathogens or has reduced resistance to tumours. Bacteria. A proportion of individuals who are infected with M. tuberculosis do not have positive skin-test responses to mycobacterial purified protein derivative (PPD), and this absence of delayed-type hypersensitivity responses to mycobacterial antigens is associated with a poorer clinical outcome. T cells from patients with positive PPD skin tests proliferated and secreted IFN-γ and IL-10 in response to PPD, whereas T cells from non-responding patients produced IL-10 but not IFN-γ 25,51. Furthermore, IL-10-specific antibodies increased PPD-specific IFN-γ production by T cells from non-responding patients25. This indicates that TR1 cells that suppress TH1-cell responses to PPD through IL-10 production mediate T-cell suppression in patients with tuberculosis. In addition, recent studies using mice transgenic for the gene encoding IL-10 show that reactivation of chronic M. tuberculosis infection and suppression of protective TH1-cell responses is strongly influenced by the expression of IL-10 during the latent phase of infection85. Furthermore, cell-mediated immunity to Mycobacterium bovis bacillus CalmetteGuérin (BCG) is increased in IL-10-deficient mice and these mice eliminate the bacteria faster than wildtype mice86. Collectively, these findings indicate that IL-10-producing cells, probably TR1 cells as well as innate immune cells, contribute to the chronic state of mycobacterial infections. Similarly, during infection with Yersinia enterocolitica, the V antigen of this pathogen stimulates IL-10 production by macrophages, which suppresses production of the host-protective cytokine TNF, and IL-10-deficient mice are highly resistant to infection with Y. enterocolitica 87. Although Y. enterocolitica-specific regulatory T cells have not yet been documented, it is probable that the bacteriatriggered IL-10 production by macrophages facilitates suppression of protective immunity either directly or indirectly through the induction of regulatory T cells. This conclusion is supported by studies with B. pertussis, in which two bacterial virulence factors — filamentous haemagglutinin (FHA) and adenylate cyclase toxin (CyaA), which stimulate IL-10 production by macrophages and DCs — direct the induction of TR1 cells in vivo 28,88. Suppression of local TH1-cell responses early in infection with B. pertussis 89 seems to result from the induction of regulatory T cells, which can be detected in the lung during acute infection28, even before the appearance of TH1 cells (P. McGuirk and K.H.G.M., unpublished observations). Furthermore, co-transfer of B. pertussis-specific TR1-cell clones with TH1 cells from convalescent mice to naive mice before infection with B. pertussis suppressed TH1-cell responses and exacerbated infection28. Therefore, it seems that the persistence of infection with certain bacteria might be associated with the induction of IL-10-producing CD4+ TR1 cells 850


and that this might be a strategy used by the bacteria to evade host immune responses. Indirect evidence of a suppressive role for CD4+ regulatory T cells in the development of memory CD8+ T cells has also been provided by studies using mice infected with L. monocytogenes, in which removal of CD4+ T cells increases the generation of CD8+ T-cell responses and increases protection against infection induced by immunization with killed L. monocytogenes 90. Parasites. Infection with malaria parasites is persistent and is associated with suppressed immune responses both to the parasite and to unrelated antigens. In a mouse model of malaria, depletion of CD4+CD25+ TReg cells protected mice against lethal infection with Plasmodium yoelii 91 and reduced the parasite load in naive and immunized mice infected with P. berghei 92. CD25+ T-cell depletion also reversed the defect in the proliferative response of splenocytes from P. yoelii-infected mice to parasitized erythrocytes91. Furthermore, treatment of P. yoelii-infected mice with antibodies specific for TGF-β and IL-10 reduced parasitaemia and increased survival76. Similarly, infection of IL-10-deficient mice with L. major results in more rapid clearance of the infection93. Although susceptibility to L. major infection in BALB/c mice has been associated with IL-4 production and TH2-cell polarization, recent evidence indicates that IL-10 production by regulatory T cells might have an important role in persistence of L. major infection in these mice94. IL-10-deficient mice clear L. major infection more rapidly than wild-type mice. Furthermore, CD4+CD25+ TReg cells specific for L. major antigens have been shown to accumulate rapidly at the site of infection in the dermis. These regulatory T cells suppress the ability of effector T cells to eliminate the parasite from the host29. Therefore, pathogens have evolved strategies for persistence by the subversion of host-protective immune responses through activation of anti-inflammatory cytokine production by innate immune cells and through activation of natural and inducible regulatory T cells. Pathogen-activated DCs induce regulatory T cells

During infection, the differentiation of naive T cells to distinct effector CD4+ T-cell subtypes is controlled by DCs and regulatory cytokines produced by innate immune cells (FIG. 4). Following the binding of conserved pathogen-derived molecules to pattern-recognition receptors, such as TLRs, on the surface of immature DCs at the site of infection, the DCs mature and migrate to the lymph nodes, where they present antigen to naive T cells. The differentiation of naive T cells to TH1 cells is promoted by the production of IL-12, IL-23 and IL-27, whereas differentiation to TH2 cells is promoted by IL-4 and IL-6. Although there is some evidence that immature DCs can selectively activate regulatory T cells95, it seems that T cells induced by immature DCs are anergic rather than regulatory and that the DCs that direct the induction of regulatory T cells have an intermediate phenotype, including increased expression of MHC class II


REVIEWS molecules and CD86 but low levels of expression of CD40 and intercellular adhesion molecule 1 (ICAM1)32,88,96. This is supported by studies showing that DCs lacking surface expression of CD40 can suppress a primed immune response and induce IL-10secreting CD4+ regulatory T cells96. Furthermore, the interaction between ICAM1 and leukocyte functionassociated antigen 1 (LFA1) is thought to promote the induction of T H1 cells independently of IL-12. So, DCs in which CD40 and ICAM1 expression is suppressed but CD80 and CD86 expression is increased might promote the induction of TR1 cells but block the differentiation of TH1 cells 8. The cytokine environment that promotes the differentiation of regulatory T cells is also distinct from that which drives TH1- and TH2-cell differentiation.

TH1-cell-promoting pathogen molecules (such as the TLR ligands LPS, CpG-containing DNA and viral RNA)

Pathogen molecules that inhibit IL-12 production and increase IL-10 production have been shown to promote the induction of TR1 cells in vitro and in vivo8. FHA from B. pertussis induces IL-10 production and inhibits IL-12 production by DCs and macrophages, and FHA-stimulated DCs promote the clonal expansion of IL-10-secreting T cells from naive T cells in vitro28. Furthermore, incubation of FHA-stimulated DCs with an IL-10-specific antibody prevents the induction of TR1 cells, which indicates that IL-10 is a differentiation factor for TR1 cells. Cholera toxin and B. pertussis CyaA also inhibit IL-12 production and CD40 expression by DCs and synergize with TLR ligands in activating IL-10 production by DCs and macrophages32,88. DCs activated by these pathogen-derived molecules induce TR1 and TH2 cells.

TH2-cell-promoting pathogen molecules (helminth products, yeast hyphae, cholera toxin, LT and TLR2 ligands)

TR1/TH3-cell-promoting pathogen molecules (FHA, CyaA, cholera toxin, NS4, and S. mansoni phosphatidylserine)

Innate immune cell

TLR Macrophage

CD11b–CD18 Semi-mature DC (CD40–CD80+)

Mature DC (CD40+CD80+CD86+)

Mature DC (CD86+OX40L+) PRR

IL-12 and IL-27

CD40

MHC class II

CD40L

TCR

IL-10 and TGF-β

CD28 Regulatory T cell

TH1 cell

IL-1β and TNF

IFN-γ

• TH1-type responses • Pro-inflammatory responses • Immunity to intracellular pathogens • (Autoimmune diseases)

IL-4 and IL-6

CD80/ CD86

IL-10 and/or TGF-β • Immune regulation • Anti-inflammatory responses

OX40L OX40

TH2 cell

IL-4, IL-5, IL-6, IL-10 and IL-13 • TH2-type responses • Immunity to extracellular pathogens • (Allergy)

Figure 4 | Role of pathogen-derived molecules in promoting the induction of regulatory T cells versus TH1 and TH2 cells. Pathogens produce a range of conserved molecules that interact with pattern-recognition receptors (PRRs), such as Toll-like receptors (TLRs), on the surface of innate immune cells, including macrophages and dendritic cells (DCs). Most TLR ligands, including lipopolysaccharide (LPS), CpG-containing DNA and viral RNA, activate DC maturation (that is, upregulate cell-surface expression of CD40, CD80 and CD86) and production of interleukin-12 (IL-12) and IL-27, leading to T helper 1 (TH1)-cell induction. Distinct families of pathogen-derived molecules — including filamentous haemagglutinin (FHA) and adenylate cyclase toxin (CyaA) from Bordetella pertussis, cholera toxin, hepatitis C virus non-structural protein 4 (NS4) and Schistosoma mansoni-specific phosphatidylserine — interact with PRRs, including CD11b–CD18, ganglioside GM1 or TLR2 on the surface of DCs. This stimulates IL-10 production and inhibits IL-12 production by macrophages and DCs and activate DCs to a semi-mature or intermediate phenotype, which promotes the induction of T regulatory 1 (TR1) and/or TH3 cells. Finally, yeast hyphae, cholera toxin, CyaA, Escherichia coli heat-labile enterotoxin (LT) and products of helminth parasites, which stimulate the production of IL-4 and/or IL-6 by DCs or other innate immune cells, promote the induction of TH2 cells. IL-10 and transforming growth factor-β (TGF-β) produced by regulatory T cells and by innate immune cells inhibit the activation of TH1 cells, which mediate immunity to intracellular pathogens, and the activation of TH2 cells, which mediate immunity to extracellular pathogens. However, these immunosuppressive cytokines also prevent innate inflammatory responses, autoimmunity and allergy, which are mediated by pathogenic TH1 and TH2 cells, respectively. CD40L, CD40 ligand; IFN-γ, interferon-γ; OX40L, OX40 ligand; TCR, T-cell receptor; TNF, tumour-necrosis factor.



REVIEWS

Endothelial cell

a

Pathogen persistence Pathogen

TH1/2 TCR MHC class II Naive T cell

CD28 CD80/CD86 Semi-mature DC Effector T cells IL-10 or TGF-β Regulatory T cell

Thymus

Natural regulatory T cell

Natural and inducible regulatory T cells Mature DC

b

Pathogen clearance with immunopathology

IL-4 or IL-12

Pathogen clearance with limited immunopathology and memory development

c

IL-4 or IL-12

IL-10 or TGF-β

Figure 5 | Protective immunity versus immunopathology depends on a balance between regulatory and effector T cells. I propose a model in which certain pathogens in different individuals can induce three different responses. a | Pathogens can stimulate potent pathogen-specific regulatory T-cell responses, through the selective induction of interleukin-10 (IL-10) or transforming growth factor-β (TGF-β) production by innate immune cells, which together with natural CD4+CD25+ regulatory T (TReg) cells can inhibit the generation and function of effector T cells and prevent clearance of the microorganism. This immuneevasion strategy is used by many pathogens that cause chronic infections. b | Pathogens can induce T helper 1 (TH1)-cell-biased or TH2-cell-biased immune responses in certain individuals, by activating innate IL-12 or IL-4 production, respectively. In cases in which the number of regulatory T cells is limiting, as a result of either a defect in natural CD4+CD25+ TReg cells or the limited induction of inducible regulatory T cells — T regulatory 1 (TR1) cells and/or TH3 cells — by the pathogen, these effector cells can mediate clearance of the microorganism. However, the absence of control by an appropriate complement of regulatory T cells allows these effector T cells to cause pathological damage to host tissue. c | Pathogens can induce a balanced number of regulatory and effector T cells, possibly by stimulating both IL-10/TGF-β and IL-4/IL-12 production by innate immune cells, thereby allowing effector T-cell-mediated clearance of the microorganism, together with control of the inflammatory response by regulatory T cells, which limits damage to host tissue. DC, dendritic cell; TCR, T-cell receptor.

852



REVIEWS

ALLOREACTIVE

Responding to antigens that are distinct between members of the same species, such as MHC molecules or blood-group antigens.

Lactobacillus paracasei inhibits the proliferation of and the TH1 and TH2 cytokine production by ALLOREACTIVE T cells, but it increases IL-10 and TGF-β production, indicating that these bacteria probably promote the induction of regulatory T cells97. C. albicans hyphae induce IL-4 and IL-10 production by DCs, and hyphaepulsed DCs can induce the clonal expansion of CD4+CD25+ T cells in vivo. Furthermore, an antibody specific for the IL-10R prevented the clonal expansion of CD4+CD25+ T cells in C. albicans-infected mice70. It seems, however, that the source of innate IL-10 that promotes the induction of TR1 cells is not confined to DCs. Non-structural protein 4 from HCV stimulates IL-10 production by monocytes (but not DCs), which in turn activates DCs to induce TR1 cells at the expense of TH1 cells60. EBV infection is associated with the induction of TR1-type cells specific for LMP1. EBV produces a viral homologue of mammalian IL-10, which is expressed, together with LMP1, during the lytic cycle. Because LMP1-specific IL-10-secreting T cells are induced in EBV-infected individuals, viral IL-10 might help to promote the differentiation of these TR1 cells in vivo50. Therefore, the production of immunoregulatory cytokines — IL-10 and TGF-β, and possibly IFN-α — by innate immune cells in response to certain pathogenderived products, together with the suppression of IL-12 production and the selective activation of co-stimulatory molecule expression by DCs, might have an important influence on the induction of regulatory T cells during infection. Finally, it has also been proposed that regulatory T cells might respond directly to physiological or pathogenic ligand interaction with TLR4 (REF. 98) or CD46 (REF. 99) expressed on the surface of T cells. Conclusions and therapeutic prospects

The study of regulatory T cells in the context of infection has shown that these cells form an essential component of the protective armoury of the host immune system through their ability to limit immunopathology and allow the development of immunological memory. However, regulatory T cells can also be detrimental to the host because they can be exploited by pathogens to facilitate pathogen persistence by suppressing anti-pathogen protective immune responses. This review has provided evidence from many studies that regulatory T cells can be both beneficial and detrimental to the host in response to the same pathogen. An explanation for these apparently contradictory findings might lie in the balance between regulatory and effector cells in different individuals, disease settings and experimental systems. The different outcomes of infection — persistence, resolution with excessive collateral damage, or resolution with limited immunopathology and development of immune memory — might be influenced by the ratio of regulatory T cells to effector T cells (FIG. 5). This hypothesis is supported by a recent report showing that re-infection of mice with L. major at a secondary site increased the number of regulatory T cells, resulting in disease reactivation at the primary site of infection; the equilibrium between effector and regulatory T cells controlled the efficiency of recall immune responses and disease reactivation100.


These studies have also helped to increase our understanding of the role of regulatory T cells in immune homeostasis and how they could be manipulated for the treatment of human diseases. In a normal healthy individual, the immune system must be capable of preventing the development of autoimmune diseases by suppressing immune responses to self-antigens. It must also be able to mount immune responses that control infections with a range of pathogenic organisms. Immune homeostasis is achieved through a careful balance between effector and suppressor responses, possibly through an appropriate frequency of TH1 cells, TH2 cells and CTLs versus natural and induced regulatory T cells (FIG. 5). The development of autoimmunity and allergy might, in part, arise from a deficit in regulatory T cells, whereas the development of cancer and chronic infections might be associated with an excess of these cells. Therefore, manipulation of this balance has opened up new approaches to therapy for a range of human diseases. The clonal expansion of regulatory T cells using strategies traditionally associated with the induction of tolerance has had some success in reducing symptoms of autoimmune disease in animal models26,101. However, this approach requires further development before it can be routinely applied to humans. Studies of animal tumour models have shown that altering the ratio of regulatory T cells to TH1 cells and CTLs can affect tumour survival; removal of CD4+CD25+ T cells increases anti-tumour immunity102, and therapy with pathogen-derived molecules that promote TH1-cell and CTL responses versus TR1-cell responses results in reduced versus increased tumour survival, respectively (A. Jarnicki, J. Lysaght, S. Todryk and K.H.G.M., unpublished observations). In mouse models of infectious disease, there is some evidence that removal of CD4+CD25+ regulatory T cells can help to resolve infection36,91. However, the application of this approach to humans will not be straightforward or without risk, and there are many unanswered questions. Will it be possible to deplete CD4+CD25+ TReg cells in vivo using monoclonal antibodies? Will the transient removal of CD4+CD25+ TReg cells be sufficient for resolution of infection and, if not, will the longer-term removal of these cells increase the risk of developing autoimmunity? Inhibition of pathogen-induced TR1 or TH3 cells or the cytokines they secrete is an alternative approach for the treatment of chronic infections. Studies with PBMCs from patients infected with HCV60 or with M. tuberculosis 25 have shown that antigen-specific IFN-γ secretion can be increased in vitro by the addition of IL-10-specific antibodies. Targeting IL-10 and TGF-β has the advantage of inhibiting the innate cytokines that induce regulatory T cells, as well as the products of these cells that mediate suppression. However, this is also not without risk because antiinflammatory cytokines can favour T cells that mediate pathogen clearance, but if uncontrolled, the same T cells can contribute to immunopathology. Therefore, the key to success with immunotherapeutic approaches will be to elicit the correct balance of effector/pathogenic and regulatory T cells.


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23.

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Artavanis-Tsakonas, K., Tongren, J. E. & Riley, E. M. The war between the malaria parasite and the immune system: immunity, immunoregulation and immunopathology. Clin. Exp. Immunol. 133, 145–152 (2003). von Herrath, M. G. & Harrison, L. C. Antigen-induced regulatory T cells in autoimmunity. Nature Rev. Immunol. 3, 223–232 (2003). Redpath, S., Ghazal, P. & Gascoigne, N. R. Hijacking and exploitation of IL-10 by intracellular pathogens. Trends Microbiol. 9, 86–92 (2001). Reed, S. G. TGF-β in infections and infectious diseases. Microbes Infect. 1, 1313–1325 (1999). Yazdanbakhsh, M., Kremsner, P. G. & van Ree, R. Allergy, parasites, and the hygiene hypothesis. Science 296, 490–494 (2002). Braun-Fahrlander, C. et al. Environmental exposure to endotoxin and its relation to asthma in school-age children. N. Engl. J. Med. 347, 869–877 (2002). Mills, K. H. G. & Boyd, A. in Topley and Wilsons’s Microbiology and Microbial Infections 10th edn (eds Kaufmann, S. H. & Steward, M.) (Edward Arnold Publishers Ltd, London, in press). McGuirk, P. & Mills, K. H. G. Pathogen-specific regulatory T cells provoke a shift in the TH1/TH2 paradigm in immunity to infectious diseases. Trends Immunol. 23, 450–455 (2002). Cozzo, C., Larkin, J. & Caton, A. J. Self-peptides drive the peripheral expansion of CD4+CD25+ regulatory T cells. J. Immunol. 171, 5678–5682 (2003). Bluestone, J. A. & Abbas, A. K. Natural versus adaptive regulatory T cells. Nature Rev. Immunol. 3, 253–257 (2003). Garba, M. L., Pilcher, C. D., Bingham, A. L., Eron, J. & Frelinger, J. A. HIV antigens can induce TGF-β1-producing immunoregulatory CD8+ T cells. J. Immunol. 168, 2247–2254 (2002). Haynes, L. M., Vanderlugt, C. L., Dal Canto, M. C., Melvold, R. W. & Miller, S. D. CD8+ T cells from Theiler’s virus-resistant BALB/cByJ mice downregulate pathogenic virus-specific CD4+ T cells. J. Neuroimmunol. 106, 43–52 (2000). Harrison, L. C., Dempsey-Collier, M., Kramer, D. R. & Takahashi, K. Aerosol insulin induces regulatory CD8 γδ T cells that prevent murine insulin-dependent diabetes. J. Exp. Med. 184, 2167–2174 (1996). Seo, N., Tokura, Y., Takigawa, M. & Egawa, K. Depletion of IL-10- and TGF-β-producing regulatory γδ T cells by administering a daunomycin-conjugated specific monoclonal antibody in early tumor lesions augments the activity of CTLs and NK cells. J. Immunol. 163, 242–249 (1999). Sonoda, K. H. et al. NK T cell-derived IL-10 is essential for the differentiation of antigen-specific T regulatory cells in systemic tolerance. J. Immunol. 166, 42–50 (2001). Sakaguchi, S., Sakaguchi, N., Asano, M., Itoh, M. & Toda, M. Immunologic self-tolerance maintained by activated T cells expressing IL-2 receptor α-chains (CD25). Breakdown of a single mechanism of self-tolerance causes various autoimmune diseases. J. Immunol. 155, 1151–1164 (1995). The first description of CD25 as a marker for TReg cells and the demonstration that these cells can suppress immune responses that mediate autoimmune diseases. Powrie, F., Carlino, J., Leach, M. W., Mauze, S. & Coffman, R. L. A critical role for transforming growth factor-β but not interleukin 4 in the suppression of T helper type 1mediated colitis by CD45RBlowCD4+ T cells. J. Exp. Med. 183, 2669–2674 (1996). Shimizu, J., Yamazaki, S., Takahashi, T., Ishida, Y. & Sakaguchi, S. Stimulation of CD25+CD4+ regulatory T cells through GITR breaks immunological self-tolerance. Nature Immunol. 3, 135–142 (2002). Fontenot, J. D., Gavin, M. A. & Rudensky, A. Y. Foxp3 programs the development and function of CD4+CD25+ regulatory T cells. Nature Immunol. 4, 330–336 (2003). Hori, S., Nomura, T. & Sakaguchi, S. Control of regulatory T cell development by the transcription factor Foxp3. Science 299, 1057–1061 (2003). Richman, L. K., Chiller, J. M., Brown, W. R., Hanson, D. G. & Vaz, N. M. Enterically induced immunologic tolerance. I. Induction of suppressor T lymphocytes by intragastric administration of soluble proteins. J. Immunol. 121, 2429–2434 (1978). Miller, A., Lider, O., Roberts, A. B., Sporn, M. B. & Weiner, H. L. Suppressor T cells generated by oral tolerization to myelin basic protein suppress both in vitro and in vivo immune responses by the release of transforming growth factor-β after antigen-specific triggering. Proc. Natl Acad. Sci. USA 89, 421–425 (1992). Hirai, K. et al. Suppression of cell-mediated immunity by street rabies virus infection. Microbiol. Immunol. 36, 1277–1290 (1992).

24. Mahanty, S. et al. High levels of spontaneous and parasite antigen-driven interleukin-10 production are associated with antigen-specific hyporesponsiveness in human lymphatic filariasis. J. Infect. Dis. 173, 769–773 (1996). 25. Boussiotis, V. A. et al. IL-10-producing T cells suppress immune responses in anergic tuberculosis patients. J. Clin. Invest. 105, 1317–1325 (2000). 26. Chen, Y., Kuchroo, V. K., Inobe, J., Hafler, D. A. & Weiner, H. L. Regulatory T cell clones induced by oral tolerance: suppression of autoimmune encephalomyelitis. Science 265, 1237–1240 (1994). 27. Groux, H. et al. A CD4+ T-cell subset inhibits antigen-specific T-cell responses and prevents colitis. Nature 389, 737–742 (1997). The first description of TR1-cell clones and their ability to prevent inflammatory disease. 28. McGuirk, P., McCann, C. & Mills, K. H. G. Pathogen-specific T regulatory 1 cells induced in the respiratory tract by a bacterial molecule that stimulates interleukin 10 production by dendritic cells: a novel strategy for evasion of protective T helper type 1 responses by Bordetella pertussis. J. Exp. Med. 195, 221–231 (2002). The first demonstration of TR1 cells specific for a bacterial antigen, their cloning from a mucosal site of infection and a role for IL-10 produced by DCs in their induction. 29. Belkaid, Y., Piccirillo, C. A., Mendez, S., Shevach, E. M. & Sacks, D. L. CD4+CD25+ regulatory T cells control Leishmania major persistence and immunity. Nature 420, 502–507 (2002). The first description of a role for CD4+CD25+ TReg cells in protective immunity to infection and in the development of immunological memory to a parasite. 30. Lundgren, A., Suri-Payer, E., Enarsson, K., Svennerholm, A. M. & Lundin, B. S. Helicobacter pylorispecific CD4+CD25high regulatory T cells suppress memory T-cell responses to H. pylori in infected individuals. Infect. Immun. 71, 1755–1762 (2003). 31. Vahlenkamp, T. W., Tompkins, M. B. & Tompkins, W. A. Feline immunodeficiency virus infection phenotypically and functionally activates immunosuppressive CD4+CD25+ T regulatory cells. J. Immunol. 172, 4752–4761 (2004). 32. Lavelle, E. C. et al. Cholera toxin promotes the induction of regulatory T cells specific for bystander antigens by modulating dendritic cell activation. J. Immunol. 171, 2384–2392 (2003). 33. Cottrez, F., Hurst, S. D., Coffman, R. L. & Groux, H. T regulatory cells 1 inhibit a TH2-specific response in vivo. J. Immunol. 165, 4848–4853 (2000). 34. Piccirillo, C. A. & Shevach, E. M. Control of CD8+ T cell activation by CD4+CD25+ immunoregulatory cells. J. Immunol. 167, 1137–1140 (2001). 35. Kursar, M. et al. Regulatory CD4+CD25+ T cells restrict memory CD8+ T cell responses. J. Exp. Med. 196, 1585–1592 (2002). 36. Suvas, S., Kumaraguru, U., Pack, C. D., Lee, S. & Rouse, B. T. CD4+CD25+ T cells regulate virus-specific primary and memory CD8+ T cell responses. J. Exp. Med. 198, 889–901 (2003). 37. Maloy, K. J. et al. CD4+CD25+ TR cells suppress innate immune pathology through cytokine-dependent mechanisms. J. Exp. Med. 197, 111–119 (2003). This paper describes the role of CD45RBhi TReg cells in the prevention of bacteria-induced inflammation in the intestine. 38. Kitani, A., Chua, K., Nakamura, K. & Strober, W. Activated self-MHC-reactive T cells have the cytokine phenotype of TH3/T regulatory cell 1 T cells. J. Immunol. 165, 691–702 (2000). 39. Gorelik, L., Constant, S. & Flavell, R. A. Mechanism of transforming growth factor β-induced inhibition of T helper type 1 differentiation. J. Exp. Med. 195, 1499–1505 (2002). 40. Moore, K. W., de Waal Malefyt, R., Coffman, R. L. & O’Garra, A. Interleukin-10 and the interleukin-10 receptor. Annu. Rev. Immunol. 19, 683–765 (2001). 41. Kitani, A. et al. Transforming growth factor (TGF)-β1producing regulatory T cells induce Smad-mediated interleukin 10 secretion that facilitates coordinated immunoregulatory activity and amelioration of TGF-β1mediated fibrosis. J. Exp. Med. 198, 1179–1188 (2003). 42. Grundstrom, S., Cederbom, L., Sundstedt, A., Scheipers, P. & Ivars, F. Superantigen-induced regulatory T cells display different suppressive functions in the presence or absence of natural CD4+CD25+ regulatory T cells in vivo. J. Immunol. 170, 5008–5017 (2003). 43. Read, S., Malmstrom, V. & Powrie, F. Cytotoxic T lymphocyteassociated antigen 4 plays an essential role in the function of CD25+CD4+ regulatory cells that control intestinal inflammation. J. Exp. Med. 192, 295–302 (2000).


44. Nakamura, K., Kitani, A. & Strober, W. Cell contactdependent immunosuppression by CD4+CD25+ regulatory T cells is mediated by cell surface-bound transforming growth factor-β. J. Exp. Med. 194, 629–644 (2001). 45. Barthlott, T., Kassiotis, G. & Stockinger, B. T cell regulation as a side effect of homeostasis and competition. J. Exp. Med. 197, 451–460 (2003). 46. MacDonald, A. J. et al. CD4 T helper type 1 and regulatory T cells induced against the same epitopes on the core protein in hepatitis C virus-infected persons. J. Infect. Dis. 185, 720–727 (2002). 47. Doetze, A. et al. Antigen-specific cellular hyporesponsiveness in a chronic human helminth infection is mediated by TH3/TR1type cytokines IL-10 and transforming growth factor-β but not by a TH1 to TH2 shift. Int. Immunol. 12, 623–630 (2000). The first demonstration of parasite-specific TR1 and TH3 cells at the clonal level in humans. 48. Satoguina, J. et al. Antigen-specific T regulatory-1 cells are associated with immunosuppression in a chronic helminth infection (onchocerciasis). Microbes Infect. 4, 1291–1300 (2002). 49. Accapezzato, D. et al. Hepatic expansion of a virus-specific regulatory CD8+ T cell population in chronic hepatitis C virus infection. J. Clin. Invest. 113, 963–972 (2004). 50. Marshall, N. A., Vickers, M. A. & Barker, R. N. Regulatory T cells secreting IL-10 dominate the immune response to EBV latent membrane protein 1. J. Immunol. 170, 6183–6189 (2003). 51. Delgado, J. C. et al. Antigen-specific and persistent tuberculin anergy in a cohort of pulmonary tuberculosis patients from rural Cambodia. Proc. Natl Acad. Sci. USA 99, 7576–7581 (2002). 52. Gerosa, F. et al. CD4+ T cell clones producing both interferon-γ and interleukin-10 predominate in bronchoalveolar lavages of active pulmonary tuberculosis patients. Clin. Immunol. 92, 224–234 (1999). 53. Ostrowski, M. A. et al. Quantitative and qualitative assessment of human immunodeficiency virus type 1 (HIV-1)-specific CD4+ T cell immunity to gag in HIV-1infected individuals with differential disease progression: reciprocal interferon-γ and interleukin-10 responses. J. Infect. Dis. 184, 1268–1278 (2001). 54. Beilharz, M. W. et al. Timed ablation of regulatory CD4+ T cells can prevent murine AIDS progression. J. Immunol. 172, 4917–4925 (2004). 55. van der Kleij, D. et al. A novel host–parasite lipid cross-talk. Schistosomal lyso-phosphatidylserine activates Toll-like receptor 2 and affects immune polarization. J. Biol. Chem. 277, 48122–48129 (2002). 56. Kullberg, M. C. et al. Bacteria-triggered CD4+ T regulatory cells suppress Helicobacter hepaticus-induced colitis. J. Exp. Med. 196, 505–515 (2002). 57. Graca, L., Le Moine, A., Cobbold, S. P. & Waldmann, H. Dominant transplantation tolerance. Curr. Opin. Immunol. 15, 499–506 (2003). 58. Chen, C., Lee, W. H., Yun, P., Snow, P. & Liu, C. P. Induction of autoantigen-specific TH2 and TR1 regulatory T cells and modulation of autoimmune diabetes. J. Immunol. 171, 733–744 (2003). 59. Akdis, M. et al. Immune responses in healthy and allergic individuals are characterized by a fine balance between allergen-specific T regulatory 1 and T helper 2 cells. J. Exp. Med. 199, 1567–1575 (2004). 60. Brady, M. T., MacDonald, A. J., Rowan, A. G. & Mills, K. H. Hepatitis C virus non-structural protein 4 suppresses TH1 responses by stimulating IL-10 production from monocytes. Eur. J. Immunol. 33, 3448–3457 (2003). 61. Boyer, O. et al. CD4+CD25+ regulatory T-cell deficiency in patients with hepatitis C-mixed cryoglobulinemia vasculitis. Blood 103, 3428–3430 (2004). 62. Suvas, S., Azkur, A. K., Kim, B. S., Kumaraguru, U. & Rouse, B. T. CD4+CD25+ regulatory T cells control the severity of viral immunoinflammatory lesions. J. Immunol. 172, 4123–4132 (2004). This study shows that CD25+ TReg cells suppress CD4+ T-cell responses in vivo and limit inflammation during corneal infection with HSV. 63. Deckert, M. et al. Endogenous interleukin-10 is required for prevention of a hyperinflammatory intracerebral immune response in Listeria monocytogenes meningoencephalitis. Infect. Immun. 69, 4561–4571 (2001). 64. Sewnath, M. E. et al. IL-10-deficient mice demonstrate multiple organ failure and increased mortality during Escherichia coli peritonitis despite an accelerated bacterial clearance. J. Immunol. 166, 6323–6331 (2001). 65. Chen, W., Shu, D. & Chadwick, V. S. Helicobacter pylori infection: mechanism of colonization and functional dyspepsia. Reduced colonization of gastric mucosa by Helicobacter pylori in mice deficient in interleukin-10. J. Gastroenterol. Hepatol. 16, 377–383 (2001).


REVIEWS 66. Kullberg, M. C. et al. Helicobacter hepaticus-induced colitis in interleukin-10-deficient mice: cytokine requirements for the induction and maintenance of intestinal inflammation. Infect. Immun. 69, 4232–4241 (2001). 67. Higgins, S. C. et al. Toll-like receptor 4-mediated innate IL-10 activates antigen-specific regulatory T cells and confers resistance to Bordetella pertussis by inhibiting inflammatory pathology. J. Immunol. 171, 3119–3127 (2003). This paper describes a role for TLR4 in the induction of IL-10 production by innate immune cells and regulatory T cells and its role in preventing bacteriainduced lung inflammation. 68. Raghavan, S., Fredriksson, M., Svennerholm, A. M., Holmgren, J. & Suri-Payer, E. Absence of CD4+CD25+ regulatory T cells is associated with a loss of regulation leading to increased pathology in Helicobacter pyloriinfected mice. Clin. Exp. Immunol. 132, 393–400 (2003). 69. Hori, S., Carvalho, T. L. & Demengeot, J. CD25+CD4+ regulatory T cells suppress CD4+ T cell-mediated pulmonary hyperinflammation driven by Pneumocystis carinii in immunodeficient mice. Eur. J. Immunol. 32, 1282–1291 (2002). This paper shows that regulatory T cells can increase the pathogen load but protect the host by limiting lung inflammation and the wasting syndrome induced by P. carinii. 70. Montagnoli, C. et al. B7/CD28-dependent CD4+CD25+ regulatory T cells are essential components of the memoryprotective immunity to Candida albicans. J. Immunol. 169, 6298–6308 (2002). 71. Netea, M. G. et al. Toll-like receptor 2 suppresses immunity against Candida albicans through induction of IL-10 and regulatory T cells. J. Immunol. 172, 3712–3718 (2004). 72. May, J., Lell, B., Luty, A. J., Meyer, C. G. & Kremsner, P. G. Plasma interleukin-10: tumor necrosis factor (TNF)-α ratio is associated with TNF promoter variants and predicts malarial complications. J. Infect. Dis. 182, 1570–1573 (2000). 73. Othoro, C. et al. A low interleukin-10 tumor necrosis factor-α ratio is associated with malaria anemia in children residing in a holoendemic malaria region in western Kenya. J. Infect. Dis. 179, 279–282 (1999). 74. Li, C., Corraliza, I. & Langhorne, J. A defect in interleukin-10 leads to enhanced malarial disease in Plasmodium chabaudi chabaudi infection in mice. Infect. Immun. 67, 4435–4442 (1999). 75. Omer, F. M. & Riley, E. M. Transforming growth factor-β production is inversely correlated with severity of murine malaria infection. J. Exp. Med. 188, 39–48 (1998). 76. Omer, F. M., de Souza, J. B. & Riley, E. M. Differential induction of TGF-β regulates proinflammatory cytokine production and determines the outcome of lethal and nonlethal Plasmodium yoelii infections. J. Immunol. 171, 5430–5436 (2003). 77. Hesse, M. et al. The pathogenesis of schistosomiasis is controlled by cooperating IL-10-producing innate effector and regulatory T cells. J. Immunol. 172, 3157–3166 (2004). 78. Xu, D. et al. CD4+CD25+ regulatory T cells suppress differentiation and functions of TH1 and TH2 cells, Leishmania major infection and colitis in mice. J. Immunol. 170, 394–399 (2003). 79. Sugimoto, K. et al. Suppression of HCV-specific T cells without differential hierarchy demonstrated ex vivo in persistent HCV infection. Hepatology 38, 1437–1448 (2003).

80. Clerici, M. & Shearer, G. M. The TH1–TH2 hypothesis of HIV infection: new insights. Immunol. Today 15, 575–581 (1994). 81. Kinter, A. L. et al. CD25+CD4+ regulatory T cells from the peripheral blood of asymptomatic HIV-infected individuals regulate CD4+ and CD8+ HIV-specific T cell immune responses in vitro and are associated with favorable clinical markers of disease status. J. Exp. Med. 200, 331–343 (2004). 82. Aandahl, E. M., Michaelsson, J., Moretto, W. J., Hecht, F. M. & Nixon, D. F. Human CD4+CD25+ regulatory T cells control T-cell responses to human immunodeficiency virus and cytomegalovirus antigens. J. Virol. 78, 2454–2459 (2004). 83. Iwashiro, M. et al. Immunosuppression by CD4+ regulatory T cells induced by chronic retroviral infection. Proc. Natl Acad. Sci. USA 98, 9226–9230 (2001). 84. Dittmer, U. et al. Functional impairment of CD8+ T cells by regulatory T cells during persistent retroviral infection. Immunity 20, 293–303 (2004). This study describes a role for IL-10-secreting CD4+ regulatory T cells in suppressing the protective effect of CD8+ CTLs in a viral infection. 85. Turner, J. et al. In vivo IL-10 production reactivates chronic pulmonary tuberculosis in C57BL/6 mice. J. Immunol. 169, 6343–6351 (2002). 86. Jacobs, M., Brown, N., Allie, N., Gulert, R. & Ryffel, B. Increased resistance to mycobacterial infection in the absence of interleukin-10. Immunology 100, 494–501 (2000). 87. Sing, A., Roggenkamp, A., Geiger, A. M. & Heesemann, J. Yersinia enterocolitica evasion of the host innate immune response by V antigen-induced IL-10 production of macrophages is abrogated in IL-10-deficient mice. J. Immunol. 168, 1315–1321 (2002). 88. Ross, P. J., Lavelle, E. C., Mills, K. H. G. & Boyd, A. P. Adenylate cyclase toxin from Bordetella pertussis synergises with lipopolysaccaride to promote innate IL-10 production and enhance the induction of TH2 and regulatory T cells. Infect. Immun. 72, 1568–1579 (2003). 89. McGuirk, P., Mahon, B. P., Griffin, F. & Mills, K. H. G. Compartmentalization of T cell responses following respiratory infection with Bordetella pertussis: hyporesponsiveness of lung T cells is associated with modulated expression of the co-stimulatory molecule CD28. Eur. J. Immunol. 28, 153–163 (1998). 90. Kursar, M., Kohler, A., Kaufmann, S. H. & Mittrucker, H. W. Depletion of CD4+ T cells during immunization with nonviable Listeria monocytogenes causes enhanced CD8+ T cell-mediated protection against listeriosis. J. Immunol. 172, 3167–3172 (2004). 91. Hisaeda, H. et al. Escape of malaria parasites from host immunity requires CD4+ CD25+ regulatory T cells. Nature Med. 10, 29–30 (2004). This study shows that depletion of natural regulatory T cells allows the host immune response to clear a parasite infection. 92. Long, T. T., Nakazawa, S., Onizuka, S., Huaman, M. C. & Kanbara, H. Influence of CD4+CD25+ T cells on Plasmodium berghei NK65 infection in BALB/c mice. Int. J. Parasitol. 33, 175–183 (2003). 93. Noben-Trauth, N., Lira, R., Nagase, H., Paul, W. E. & Sacks, D. L. The relative contribution of IL-4 receptor signaling and IL-10 to susceptibility to Leishmania major. J. Immunol. 170, 5152–5158 (2003).


94. Sacks, D. & Noben-Trauth, N. The immunology of susceptibility and resistance to Leishmania major in mice. Nature Rev. Immunol. 2, 845–858 (2002). 95. Jonuleit, H., Schmitt, E., Schuler, G., Knop, J. & Enk, A. H. Induction of interleukin 10-producing, nonproliferating CD4+ T cells with regulatory properties by repetitive stimulation with allogeneic immature human dendritic cells. J. Exp. Med. 192, 1213–1222 (2000). 96. Martin, E., O’Sullivan, B., Low, P. & Thomas, R. Antigenspecific suppression of a primed immune response by dendritic cells mediated by regulatory T cells secreting interleukin-10. Immunity 18, 155–167 (2003). 97. von der Weid, T., Bulliard, C. & Schiffrin, E. J. Induction by a lactic acid bacterium of a population of CD4+ T cells with low proliferative capacity that produce transforming growth factor-β and interleukin-10. Clin. Diagn. Lab. Immunol. 8, 695–701 (2001). 98. Caramalho, I. et al. Regulatory T cells selectively express Toll-like receptors and are activated by lipopolysaccharide. J. Exp. Med. 197, 403–411 (2003). 99. Kemper, C. et al. Activation of human CD4+ cells with CD3 and CD46 induces a T-regulatory cell 1 phenotype. Nature 421, 388–392 (2003). 100. Mendez, S., Reckling, S. K., Piccirillo, C. A., Sacks, D. & Belkaid, Y. Role for CD4+CD25+ regulatory T cells in reactivation of persistent leishmaniasis and control of concomitant immunity. J. Exp. Med. 200, 201–210 (2004). 101. Bynoe, M. S., Evans, J. T., Viret, C. & Janeway, C. A. Jr. Epicutaneous immunization with autoantigenic peptides induces T suppressor cells that prevent experimental allergic encephalomyelitis. Immunity 19, 317–328 (2003). 102. Golgher, D., Jones, E., Powrie, F., Elliott, T. & Gallimore, A. Depletion of CD25+ regulatory cells uncovers immune responses to shared murine tumor rejection antigens. Eur. J. Immunol. 32, 3267–3275 (2002).

Acknowledgements I acknowledge support from Science Foundation Ireland, The Irish Health Research Board and Enterprise Ireland, and I am grateful to P. McGuirk and E. Lavelle for helpful discussions.

Competing interests statement The author declares competing financial interests: see Web version for details.

Online links DATABASES The following terms in this article are linked online to: Entrez Gene: http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=gene CD4 | CD25 | CD28 | CD38 | CD40 | CD45 | CD46 | CD62L | CD86 | CD103 | CTLA4 | FOXP3 | GITR | ICAM1 | IFN-α | IFN-γ | IL-4 | IL-6 | IL-10 | IL-10R | IL-12 | IL-12R | IL-23 | IL-27 | LFA1 | RAG | SMAD4 | T-bet | TGF-β | TLR2 | TLR4 | TNF Infectious Disease Information: http://www.cdc.gov/ncidod/diseases/index.htm Bordetella pertussis | Brugia malayi | Candida albicans | EBV | Escherichia coli | HCV | HIV | Leishmania major | Listeria monocytogenes | malaria | Mycobacterium tuberculosis | Onchocerca volvulus | Pneumocystis carinii | rabies virus | Schistosoma mansoni | Yersinia enterocolitica Access to this interactive links box is free online.
