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Can type-1 responses against intracellular pathogens be T helper 2 cytokine dependent?
Microbes and Infection 10 (2008) 953e959 www.elsevier.com/locate/micinf
Original article
Can type-1 responses against intracellular pathogens be T helper 2 cytokine dependent? James Alexander*, Emma McFarlane Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, The John Arbuthnott Building, 27 Taylor St, Glasgow G4 ONR, UK Available online 13 August 2008
Abstract While control of intracellular pathogens, such as the protozoan Leishmania, is dependent on the generation of type-1 immune responses, the role of T helper 2 cytokines in the disease process is more controversial. Traditionally these cytokines were perceived as counter-regulating type1 responses and promoting disease exacerbation. Nevertheless a substantial body of evidence now exists suggesting that the development of effective type-1 immunity can involve the significant involvement of the Th2 cytokines IL-4 and IL-13. This article reviews, using Leishmania species in particular, the circumstances under which these cytokines can promote protective type-1 immunity. Ó 2008 Elsevier Masson SAS. All rights reserved. Keywords: Th1; Th2; IL-4; IL-13; Leishmania
1. Introduction The seminal work of Mosmann, Coffman and colleagues [1] describing two distinct CD4þ T helper cell populations in 1986 ultimately gave rise to the Th1/Th2 paradigm of susceptibility/resistance to intracellular infection. This has provided an excellent foundation from which immunologists can further dissect the immune response and design appropriate therapeutic and intervention strategies. However, not only has the more recent characterisation of CD4þ T cell regulatory populations as well as a further effector CD4þ T helper population (Th17) [2] added to the complexity of hoste pathogen interactions but there is also a substantial body of evidence derived from both infectious and inflammatory disease models indicating that the archetypal Th2 cytokines IL-4 and IL-13 need not necessarily counter-regulate but could actually drive, facilitate or promote a Th1 response (Table 1). Consequently, IL-4/IL-13 has been shown to be associated
* Corresponding author. Tel.: þ44 141 548 3925; fax: þ44 141 552 3562. E-mail address: [email protected] (J. Alexander). 1286-4579/$ - see front matter Ó 2008 Elsevier Masson SAS. All rights reserved. doi:10.1016/j.micinf.2008.07.038
with driving protective type-1 responses against, Toxoplasma gondii [3], Listeria monocytogenes [4], Cryptosporidium parvum [5], Leishmania major [6] and Leishmania donovani [7e11] as well as aggravating Th1 responses in experimental autoimmune uveoretinitis and a murine model of colitis [12,13]. IL-4/IL-13 may mediate their effects by inducing macrophage and dendritic cell IL-12 production [6,14], enhancing IFN-g production [15] or synergising with IFN-g for enhanced anti-microbial activity [16,17]. Leishmania spp. have long been considered ideal models to study chronic intracellular infections as protective immunity is globally recognised as being type-1 dependent. However, the causative agents of both Old World and New World cutaneous leishmaniasis as well as visceral leishmaniasis diverged in evolutionary terms 40e80 million years ago (reviewed in [18]). Therefore it is hardly surprising that the separate species have developed different strategies for evading and interacting with the immune response. Consequently they provide excellent tools to dissect different pathways leading to, or preventing, a protective type-1 response. In this review, we explore, using different species of Leishmania, the spectrum of Th2 cytokine activities operating in the disease process.
J. Alexander, E. McFarlane / Microbes and Infection 10 (2008) 953e959 Mencacci et al., 1998 [5] McDonald et al., 2004 [6] Biedermann et al., 2001 [7] Alexander et al., 2000 [8] Stager et al., 2003 [9] Stager et al., 2003 [10] Murray et al., 2006 [11] McFarlane et al., unpublished
Flesch et al., 1997 [4]
Suzuki et al., 1996 [3]
Mice succumbed to infection and brain parasite burden increased Number of viable bacteria in spleen and liver reduced Mice succumb to infection Susceptibility to infection increased Footpad lesion size reduced Parasite burdens increased Parasite burdens increased Parasite burdens reduced Retarded granuloma formation Parasite burdens increased and retarded granuloma formation IL-13/ IL-13/
IL-4/ IL-4Ra/
BALB/c BALB/c BALB/c BALB/c BALB/c BALB/c BALB/c BALB/c Secondary infection Primary infection rIL-4 treatment Chemotherapy Primary infection Vaccine Primary infection Primary infection þ þ Candida albicans Cryptosporidium parvum Leishmania major Leishmania donovani Leishmania donovani Leishmania donovani Leishmania donovani Leishmania donovani
IL-4 IL-4 IL-4 IL-4 IL-4Ra IL-4 IL-13 IL-13
rIL-13 treatment þ IL-13 Listeria monocytogenes
C57BL/6
IL-4/ IL-4/
IFNg production from spleen cells reduced Frequency of IL-12 producing cells increased Defective IFNg and IL-12 production IL-12 and IFNg downregulated Enhanced IL-12 production by APCs IL-12 and IFNg downregulated IFNg downregulated CD8þT cells producing IFNg IFNg downregulated IFNg downregulated Primary infection IL-4 Toxoplasma gondii
129/J IL-4
Disease Immunology Mouse strain Context Cytokine Microbial species
Table 1 Studies where IL-4 and IL-13 have been shown to promote Th1 responses and protection against microbial infection
/
Reference
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2. Leishmania major: is non-healing disease IL-4/IL-13 dependent or independent or can these cytokines be protective? The classic studies by Locksley, Coffman and colleagues on Leishmania major which demonstrated that an IL-4 driven type-2 response and associated cytokines counter-regulated type-1 responses and consequently promoted non-healing infection have in large part given rise to the Th1/Th2 paradigm of resistance/susceptibility to intracellular parasitism [19]. Further studies in susceptible BALB/c mice demonstrated that a single T-cell epitope derived from the parasite LACK antigen (Leishmania homologue of receptors for activated C kinase) induced rapid IL-4 production by Vb4Va8CD4þ T cells which apparently rendered T cells unresponsive to IL-12 and correlated with lesion development [20]. Thus a particularly strong case for the predominant role of IL-4 and a Th2 response in non-healing cutaneous disease was established. Nevertheless, other studies on resistant C57BL/6 mice noted that this strain also produced IL-4 early during infection and yet developed an unimpaired Th1 biased response [21]. Subsequently studies utilizing IL-4/ mice seriously questioned the paramount role of IL-4 in the non-healing response of BALB/c mice to L. major as a number of apparently contradictory reports determining the significance of IL-4 were published [22e24]. This posed the question as to whether other cytokines could be regulating Th1 development. As IL-4 and IL-13 share a common signaling pathway through the IL-4 receptor a (IL-4Ra) chain and as IL-4Ra deficient mice were found to be more resistant than IL-4/ mice in one study this suggested a compensatory role for IL-13 in the absence of IL-4 [25]. However, in a further study IL-13 was also found to be able to act independently of IL-4 and it was surmised that the effects of IL-4 and IL-13 were additive [26]. Nevertheless, in another study IL-4Ra/ mice were found to be more susceptible than IL-4/ animals indicating a protective role for IL-13 in preventing disease dissemination in the absence of IL-4 signaling [27]. While question marks remain over whether IL-4 and IL-13 play a significant role in L. major disease progression in BALB/c mice IL-10 unquestionably does promote parasite persistence in resistant mice and multiplication in susceptible mice [28]. IL-10 could be generated via either Fcg-mediated uptake of antibody coated amastigotes [29], or produced by Th2 cells, or CD4þ CD25þ T regulatory cells [30] or indeed a recently described CD4þCD25FoxP3 Th1 population [31]. In the absence of a definitive role for IL-4/IL-13 in nonhealing disease it has been proposed that defective T cell responses as a consequence of intrinsic defects in antigen presenting cell function or alternatively in T cell development could be the ultimate underlying problem determining the exquisite susceptibility of BALB/c mice to infection with L. major. While several studies suggest that non-healing disease is a consequence of the failure to produce or respond to IL-12 [32,33] this could be irrespective of the presence of a Th2 response and although IL-4 can downregulate IL-12 production and expression of IL-12Rb2 on CD4þ T cells
J. Alexander, E. McFarlane / Microbes and Infection 10 (2008) 953e959
[20,33,34], this could also be by IL-4 independent mechanisms [34]. In this context it is significant that IL-12b2 expression remains impaired in IL-4Ra/ BALB/c mice compared with resistant C57BL/6 mice infected with L. major [24]. Defects in antigen presenting cell function reported for BALB/c mice include a comparative deficiency in dendritic cell IL-1 production [35] compared with L. major resistant strains. IL-1 has been shown to upregulate dendritic cell IL-12 production as well as MHC Class II and co-stimulatory molecule expression [36] as well to enhance IFN-g mediated inhibition of Th2 cell proliferation [37]. However, as transgenic IL-12Rb2 BALB/c mice exhibit a non-healing phenotype despite intact IL-12 signaling the importance of an APC defect remains questionable [38]. Irrespective of any potential defect in APC function BALB/c CD4þ T cells have been demonstrated as having an intrinsic IL-4 independent Th2 bias [39]. A commitment to Th2 development could be a consequence of a defective co-polarisation of the TCR and IFN-g receptor complex of naive CD4þ T cells at the immunological APC/T cell precursor synapse in BALB/c mice compared with C57BL/6 mice [40]. Nevertheless, immune regulatory mechanisms as well as intrinsic T cell defects can serve to impair Th1 responses in BALB/c mice as IL-4 signaling via STAT-6 also serves to block co-localisation of this complex. Recent studies may serve to help reconcile the observations above. CD4þ T cell specific IL-4Ra/ BALB/c mice unlike global IL-4Ra/ animals, which develop disseminating disease, are as resistant as C57BL/6 mice to infection with L. major [41]. Consequently it has been concluded that IL-4/IL13 responsive non-CD4þ T cells induce protection against L. major in the absence of CD4þ T cells responding to IL-4 [42]. Under such circumstances it can be hypothesised that the reported intrinsic BALB/c T cell defect is not enough in the absence of CD4þ T cell responding to IL-4 to prevent the generation of a Th1 response. Conversely C57BL/6 mice, like BALB/c mice, also produce IL-4 early in infection but in the absence of the intrinsic T cell defect that operates in BALB/c mice C57BL/6 mice would proceed to resolve infection as a Th1 response then develops. Overall this would suggest a potential significant role for IL-4Ra responsive non-CD4þ T cells in generating a Th1 biased response in L. major resistant strains. Studies utilizing macrophage specific BALB/c IL-4Ra deficient mice have demonstrated that IL-4/IL-13 operates through this population to enhance parasite growth via alternative macrophage activation [42] and consequently these are unlikely to be the population driving a Th1 response in CD4þIL-4Ra/ mice. However, IL-4 treatment of BALB/c mice pre T cell priming has previously been demonstrated to instruct dendritic cells to produce IL-12 and facilitate a protective Th1 response against L. major [7]. This would indicate that dendritic cells are the IL-4/IL-13 responsive cells facilitating protection in the absence of IL-4Ra responsive CD4þ T cells in BALB/c mice. A potential contribution of IL4/IL-13 responsive non-CD4þcells to protective immunity may explain why Leishmania vaccines incorporating IFN-gpromoting adjuvants are relatively ineffective compared with those that also utilize ALUM [43,44].
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3. Leishmania donovani: is healing IL-4/IL13 dependent or independent? Infection with the intracellular protozoan parasite Leishmania donovani causes a potentially fatal visceral infection if left untreated, and has resulted in up to 100,000 deaths in recent epidemics in Sudan and India [45]. Protective immunity against this parasite, as with species causing cutaneous leishmaniasis, is dependent on an IL-12 driven type-1 response and IFN-g production [46,47], which results in the induction of parasite killing by macrophages primarily via the production of reactive nitrogen and oxygen intermediates [48]. Studies, not only in mice [49], but also humans [50e 52], suggested that cure was independent of the differential production of Th1 and Th2 cytokines and both IFN-g and IL-4 producing T cells have been isolated from asymptomatic and cured patients. Indeed in the mouse model it has been shown that the induction of enhanced protection is related to increasing the frequency of cytokine-producing cells rather than altering the IFN-g /IL-4 balance [53]. In addition, while a recent study has shown that the control of L. donovani in the susceptible BALB/c mouse is IL-12 dependent, this cytokine was found to promote the expansion of Th2 as well as Th1 responses [46]. Both human [50,53] and murine studies [54] indicate that IL-10 is the major immunosuppressive cytokine in visceral leishmaniasis. Furthermore it has been demonstrated that the source of this IL-10 is not traditional Th2 cells [53] but more likely a regulatory T cell population [55]. Thus it would probably be the general consensus of opinion that the Th2 response does not contribute adversely to the immunological control of visceral leishmaniasis. Indeed studies utilising gene deficient mice have identified a protective role for IL-4 and IL-4Ra signalling (Fig. 1) during primary L. donovani infections [9,56]. Furthermore the increased susceptibility of IL-4Ra/ mice compared with IL-4/ BALB/c mice as measured by hepatic parasite burdens [9] also suggested a protective role for IL-13 as both IL-4 and IL-13 activities are dependent on the expression of IL-4Ra. Control of parasite growth in the liver was associated with the ability to produce sterile granulomas [9], a mechanism driven by T cell derived IFNg [57]. Enhanced susceptibility of IL-4/ and IL-4Ra/ mice was associated with downregulated type-1 responses and markedly retarded granuloma maturation [8,9]. While decreased IFN-g and retarded granuloma maturation have also been observed in IL-13/ mice this was not associated with a reduced parasite burden [11]. The recent availability of tissue specific IL-4Ra/ mice will facilitate the dissection of the IL-4 and IL-13 effects during visceral leishmaniasis and so far we have examined the course of L. donovani in macrophage/neutrophil specific IL-4Ra/ and CD4þ Tcell specific IL-4Ra/ BALB/c mice. The protective activity of IL-4/IL-13 during visceral leishmaniasis is not via these cell populations (McFarlane, Carter, Kaye, Brombacher and Alexander, unpublished). In addition to playing a significant protective role during primary infection with L. donovani effective sodium
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Fig. 1. Liver parasite burdens (A) in wild-type and IL-4Ra/ mice at day 40 post-infection with (1e2)107 L. donovani amastigotes. Hepatic granuloma maturation in L. donovani infected wild-type (B) and IL-4Ra/ (C) mice at day 40 post-infection. Wild-type mice (B) show well developed mature granulomas (arrows) surrounding a core of infected Kupffer cells whilst IL-4Ra/ mice (C) display abrogated granuloma development characterised by the presence of immature granulomas and amastigotes within the cytoplasm of infected Kupffer cells (arrows). Magnification 400.
stibogluconate (SSG) chemotherapy is dependent on Th2 cytokines [8]. Previous studies have demonstrated that successful treatment of visceral leishmaniasis with SSG requires the presence of both CD4þ and CD8þ T cells [58] accompanied by the type-1 cytokines IL-12 and IFN-g [57]. We have observed that SSG chemotherapy of L. donovani in IL-4/ [8] or IL-13/ (unpublished) BALB/c mice compared with wild-type mice resulted in downregulated IFN-g production and increased IL-10 production. This would suggest a requirement for either of these cytokines in instructing a type-1 response. Significantly, successful immunotherapeutic intervention in the BALB/c mouse using a hybrid cell vaccine which was mediated by a strong CD8þ CTL response was associated with downregulation of IL-10 but significantly enhanced IL-4 and IL-13 expression as well as IFN-g production [59] providing further strong circumstantial evidence for these Th2 cytokines facilitating the generation of type-1 immunity. Similarly the protection afforded by a kinetoplast membrane protein protein-11 DNA vaccine which protected hamsters against visceral leishmaniasis correlated with decreased IL-10 levels but with increased IL-4 production and an elevated mixed Th1/Th2
response [60]. However, vaccination studies utilizing HASPB-1 have provide the most concrete evidence for a Th2 cytokine requirement to instruct a protective type-1 response mediated by CD8þ T cells against visceral leishmaniasis [10] as this vaccine generates a mixed Th1/Th2 response and is totally ineffective in IL-4 or IL-4Ra/ mice. The primary source of IL-4 has been defined as a CD11bþCD11clo phagocyte [10]. 4. Leishmania mexicana: is non-healing disease IL-4/IL13 dependent? New world cutaneous leishmaniasis resulting from infection with parasites of the mexicana complex is under different genetic and immunoregulatory controls to those controlling L. major infection and cysteine peptidases would appear to be the major virulence factors with evidence that they can both promote a Th2 response as well as inhibit a protective Th1 response [61]. Also unlike L. major the majority of mouse strains are susceptible to this parasite. As with the other Leishmania species protective immunity is the result of a STAT4 dependent type-1 immune response although this apparently
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can be generated independently of IL-12 [62]. Early studies on Leishmania mexicana complex parasites that examined IL-4 mRNA transcript expression in the draining lymph nodes of resistant and susceptible mice indicated that IL-4 played no role in the non-healing response of mice to this parasite [63]. Nevertheless, follow up experiments using IL-4/ mice to confirm this observation demonstrated that IL-4 was after all the predominant cytokine responsible for non-healing disease [56,64]. However, these apparently contradictory results are now easy to reconcile as it is now well established that IL-4 transcript expression is not a reliable indicator of IL-4 production [65]. In addition studies using IL-13/ mice also indicate that IL-13 may play a role in preventing disease resolution by inhibiting IL-12Rb2 expression [66]. The paramount role of IL-4 in promoting disease progression is, however, mouse strain and site of infection dependent. Thus the growth of both L. mexicana and L. amazonensis is primarily IL4 dependent in rump infections in BALB/c and C57BL/6 mice [18]. By contrast the growth of these parasites in the footpad of C57BL/6 mice, though not BALB/c mice, is IL-4 independent [18,67] and the contribution of IL-10 to this activity may also depend on whether L. amazonensis or L. mexicana is studied [68e70]. A recent elegant study associates L. mexicana with not only the ability to inhibit Th1 differentiation but also to impaired lymph node expansion [71]. Overall there is no evidence whatsoever that Th2 cytokines can be beneficial during infection with New World cutaneous leishmaniasis and often they play the classic counter-protective role. 5. Conclusions IL-4 and IL-13 are pleiotropic cytokines and numerous cell types express their receptors. Consequently it is not surprising that their activity can vary significantly depending on the microbe initiating infection, the tissue site or cells involved, and the chronicity of infection. The three Leishmania species/ complexes discussed above are all controlled by a type-1 response. Nevertheless they not only differ significantly in how they can evade, or manipulate the immune response to induce non-healing disease but also the immunological pathways leading to protective type-1 responses would appear to be under different controls for each species. Consequently IL-4, IL-13 and IL-10 all promote infection with L. mexicana complex parasites and there is no evidence whatsoever for a protective role for the Th2 cytokines. Conversely, strong evidence has now been accumulating suggesting that Th2 cytokines not only do not contribute to the progression of visceral leishmaniasis but that they also actually promote the induction of type-1 immunity. Indeed IL-4 used judiciously can also be used to drive a protective type-1 response against L. major. The recent generation of spatial (tissue specific) and temporal (switch on/ switch off) IL-4Ra/ mice has provided new powerful tools by which the specific role of IL-4/IL-13 responding cells can be dissected from global effects. This will ultimately facilitate the design of more appropriate and effective anti-microbial therapeutic and prophylactic strategies.
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Acknowledgements J.A. is in receipt of funding from the Wellcome Trust and Royal Society. E.McF. is an E.P.S.R.C. sponsored Research Fellow. References [1] T.R. Mosmann, H. Cherwinski, M.W. Bond, M.A. Giedlin, R.L. Coffman, Two types of murine helper T cell clone. 1. Definition according to profiles of lymphokine activities and secreted proteins, J. Immunol. 136 (1986) 2348e2357. [2] E. Bettelli, M. Oukka, V.K. Kuchroo, T(H)-17 cells in the circle of immunity and autoimmunity, Nat. Immunol. 8 (2007) 345e350. [3] Y. Suzuki, Q. Yang, S. Yang, N. Nguyen, S. Lim, O. Liesenfeld, T. Kojima, J.S. Remington, IL-4 is protective against development of toxoplasmic encephalitis, J. Immunol. 157 (1996) 2564e2569. [4] I.E. Flesch, A. Wandersee, S.H. Kaufmann, Effects of IL-13 on murine listeriosis, Int. Immunol. 9 (1997) 467e474. [5] A. Mencacci, G. Del Sero, E. Cenci, C.F. d’Ostiani, A. Bacci, C. Montagnoli, M. Kopf, L. Romani, Endogenous interleukin 4 is required for development of protective CD4þ T helper type 1 responses to Candida albicans, J. Exp. Med. 187 (1998) 307e317. [6] S.A. McDonald, J.E. O’Grady, M. Bajaj-Elliott, C.A. Notley, J. Alexander, F. Brombacher, V. McDonald, Protection against the early acute phase of cryptosporidium parvum infection conferred by Interleukin-4-induced expression of T Helper 1 cytokines, J. Infect. Dis. 190 (2004) 1019e1025. [7] T. Biedermann, S. Zimmermann, H. Himmelrich, A. Gumy, O. Egeter, A.K. Sakrauski, I. Seegmu¨ller, H. Voigt, P. Launois, A.D. Levine, H. Wagner, K. Heeg, J.A. Louis, M. Rocken, IL-4 instructs TH1 responses and resistance to Leishmania major in susceptible BALB/c mice, Nat. Immunol. 2 (2001) 1054e1060. [8] J. Alexander, K.C. Carter, N. Al-Fasi, A. Satoskar, F. Brombacher, Endogenous IL-4 is necessary for effective drug therapy against visceral leishmaniasis, Eur. J. Immunol. 30 (2000) 2935e2943. [9] S. Stager, J. Alexander, K.C. Carter, F. Brombacher, P.M. Kaye, Both interleukin-4 (IL-4) and IL-4 receptor alpha signaling contribute to the development of hepatic granulomas with optimal antileishmanial activity, Infect. Immun. 71 (2003) 4804e4807. [10] S. Stager, J. Alexander, A.C. Kirby, M. Botto, N.V. Rooijen, D.F. Smith, F. Brombacher, P.M. Kaye, Natural antibodies and complement are endogenous adjuvants for vaccine-induced CD8þ T-cell responses, Nat. Med. 9 (2003) 1287e1292. [11] H.M. Murray, C.W. Tsai, J. Liu, X. Ma, Visceral Leishmania donovani infection in interleukin-13/ mice, Infect. Immun. 74 (2006) 2487e 2490. [12] S. Ramanathan, Y. de Kosak, A. Saoudi, O. Goureau, P.H. Van der Meide, P. Druet, B. Bellon, Recombinant IL-4 aggravates experimental autoimmune uveoretinitis in rats, J. Immunol. 157 (1996) 2209e2215. [13] M.M. Fort, R. Lesley, N.J. Davidson, S. Menon, F. Brombacher, M.W. Leach, D.M. Rennick, IL-4 exacerbates disease in a Th1 cell transfer model of colitis, J. Immunol. 166 (2001) 2793e2800. [14] H. Hochrein, M. O’Keefe, T. Luft, S. Vandenabeele, R.J. Grumont, E. Maraskovsky, K. Shortman, Interleukin (IL-4) is a major regulatory cytokine governing bioactive IL-12 production by mouse and human dendritic cells, J. Exp. Med. 192 (2000) 823e833. [15] A. Noble, D.M. Kemeny, Interleukin-4 enhances interferon-g synthesis but inhibits development of interferon-g-producing cells, Immunology 85 (1995) 357e363. [16] I.-S. Lean, S.A. McDonald, M. Bajaj-Elliott, R.C.G. Pollok, M.J.G. Farthing, V. McDonald, Interleukin-4 and transforming growth factor-beta have opposing regulatory effects on gamma interferonmediated inhibition of Cryptosporidium parvum reproduction, Infect. Immun. 71 (2003) 4580e4585. [17] C. Bogdan, S. Stenger, M. Rollinghoff, W. Solbach, Cytokine interactions in experimental cutaneous leishmaniasis. Interleukin 4
958
[18]
[19]
[20]
[21]
[22]
[23] [24]
[25]
[26]
[27]
[28]
[29] [30]
[31]
[32]
[33]
[34]
[35]
[36]
J. Alexander, E. McFarlane / Microbes and Infection 10 (2008) 953e959 synergizes with interferon-gamma to activate murine macrophages for killing of Leishmania major amastigotes, Eur. J. Immunol. 21 (1993) 327e333. D. McMahon-Pratt, J. Alexander, Does the Leishmania major paradigm of pathogenesis and protection hold for New World cutaneous leishmaniases or the visceral disease? Immunol. Rev. 201 (2004) 206e224. F.P. Heinzel, M.D. Sadick, B.J. Holaday, R.L. Coffman, R.M. Locksley, Reciprocal expression of interferon gamma or interleukin 4 during the resolution or progression of murine leishmaniasis. Evidence for expansion of distinct helper T cell subsets, J. Exp. Med. 169 (1989) 59e72. P. Launois, T. Ohteki, K. Swihart, H.R. MacDonald, J.A. Louis, In susceptible mice, Leishmania major induce very rapid interleukin-4 production by CD4þ T cells which are NK1.1, Eur. J. Immunol. 25 (1995) 3298e3007. P. Scott, A. Eaton, W.C. Gause, X. di Zhou, B. Hondowicz, Early IL-4 production does not predict susceptibility to Leishmania major, Exp. Parasitol. 84 (1996) 178e187. M. Kopf, F. Brombacher, G. Kohler, G. Kienzle, K.H. Widmann, K. Lefrang, B. Humborg, B. Lederman, W. Solbach, IL-4 deficient mice resist infection with Leishmania major, J. Exp. Med. 184 (1996) 1127e1136. N. Noben-Trauth, P. Kropf, I. Muller, Susceptibility to Leishmania major infection in interleukin-4-deficient mice, Science 271 (1996) 987e990. M. Mohrs, C. Holscher, F. Brombacher, Interleukin-4 receptor alpha deficient BALB/c mice show an unimpaired T helper 2 polarization in response to Leishmania major infection, Infect. Immun. 68 (2000) 1773e 1780. N. Noben-Trauth, W.E. Paul, D.L. Sacks, IL-4- and IL-4 receptor deficient BALB/c mice reveal differences in susceptibility to Leishmania major parasite substrains, J. Immunol. 162 (1999) 6132e6140. D.J. Matthews, C.L. Emson, G.J. McKenzie, H.E. Jolin, J.M. Blackwell, A.N. McKenzie, IL-13 is a susceptibility factor for Leishmania major infection, J. Immunol. 164 (2000) 1458e1462. M. Mohrs, B. Ledermann, G. Kohler, A. Dorfmuller, A. Gessner, F. Brombacher, Differences between IL-4- and IL-4 receptor alphadeficient mice in chronic leishmaniasis reveal a protective role for IL-13 receptor signaling, J. Immunol. 162 (1999) 7302e7308. N. Noben-Trauth, R. Lira, H. Nagase, W.E. Paul, D.L. Sacks, The relative contribution of IL-4 receptor signaling and IL-10 to susceptibility to Leishmania major, J. Immunol. 170 (2003) 5152e5158. M.M. Kane, D.M. Mosser, The role of IL-10 in promoting disease progression in leishmaniasis, J. Immunol. 166 (2001) 1141e1147. D. Sacks, C. Anderson, Re-examination of the immunosuppressive mechanisms mediating non-cure of Leishmania infection in mice, Immunol. Rev. 201 (2004) 225e238. C.F. Anderson, M. Oukka, V.J. Kuchroo, D.L. Sacks, CD4þCD25Foxp3-Th1 cells are the source of IL-10-mediated immune suppression in chronic cutaneous leishmaniasis, J. Exp. Med. 204 (2007) 285e297. S.L. Reiner, S. Zheng, Z.E. Wang, L. Stowring, R.M. Locksley, Leishmania promastigotes evade interleukin 12 (IL-12) induction by macrophages and stimulate a broad range of cytokines from CD4þ T cells during initiation of infection, J. Exp. Med. 179 (1994) 447e456. H. Himmelrich, C. Parra-Lopez, F. Tacchini-Cottier, J.A. Louis, P. Launois, The IL-4 rapidly produced in BALB/c mice after infection with Leishmania major down-regulates IL-12 receptor beta 2-chain expression on CD4þ T cells resulting in a state of unresponsiveness to IL-12, J. Immunol. 161 (1998) 6156e6163. D. Jones, M.M. Elloso, L. Showe, D. Williams, G. Trinchieri, P. Scott, Differential regulation of the interleukin-12 receptor during the innate immune response to Leishmania major, Infect. Immun. 66 (1998) 3818e3824. E. Von Stebut, J.M. Ehrchen, Y. Belkaid, S.L. Kostka, K. Molle, J. Knop, C. Sunderkotter, M.C. Udey, Interleukin 1alpha promotes Th1 differentiation and inhibits disease progression in Leishmania major susceptible BALB/c mice, J. Exp. Med. 198 (2003) 191e199. U. Eriksson, M.O. Kurrer, I. Sonderegger, G. Iezzi, A. Tafuri, L. Hunziker, S. Suzuki, K. Bachmaier, R.M. Bingisser, J.M. Penninger, M. Kopf, Activation of dendritic cells through the interleukin 1 receptor 1
[37]
[38]
[39]
[40]
[41]
[42]
[43]
[44]
[45]
[46]
[47]
[48]
[49]
[50]
[51] [52]
[53]
[54]
is critical for the induction of autoimmune myocarditis, J. Exp. Med. 197 (2003) 323e331. T.B. Oriss, S.A. McCarthy, B.F. Morel, M.A. Campana, P.A. Morel, Cross-regulation between T helper cell (Th)1 and Th2: inhibition of Th2 proliferation by IFN-g involves interference with IL-1, J. Immunol. 158 (1997) 3666e3672. R. Nishikomori, S. Gurunathan, K. Nishikomori, W. Strober, BALB/c mice bearing a transgenic IL-12 receptor beta 2 gene exhibit a nonhealing phenotype to Leishmania major infection despite intact IL-12 signaling, J. Immunol. 166 (2001) 6776e6783. M. Bix, Z.E. Wang, B. Thiel, N.J. Schork, R.M. Locksley, Genetic regulation of commitment to interleukin 4 production by a CD4þ T cellintrinsic mechanism, J. Exp. Med. 188 (1998) 2289e2299. R.A. Maldonado, D.J. Irvine, R. Schreiber, L.H. Glimcher, A role for the immunological synapse in lineage commitment of CD4 lymphocytes, Nature 431 (2004) 527e532. M. Radwanska, A.J. Cutler, J.C. Hoving, S. Magez, C. Holscher, A. Bohms, B. Arendse, R. Kirsch, T. Hunig, J. Alexander, P.M. Kaye, F. Brombacher, Deletion of IL-4Ralpha on CD4 T cells renders BALB/c mice resistant to Leishmania major infection, PLoS Pathog. 3 (5) (2007) e68. C. Holscher, B. Arendse, A. Schwegmann, E. Myburgh, F. Brombacher, Impairment of alternative macrophage activation delays cutaneous leishmaniasis in non-healing BALB/c mice, J. Immunol. 176 (2006) 1115e1121. M.M. Gicheru, J.O. Olobo, C.O. Anjili, F. Modabber, P. Scott, Vervet monkeys vaccinated with killed Leishmania major parasites and interleukin-12 develop a type 1 immune response but are not protected against challenge infection, Infect. Immun. 69 (2001) 245e251. R.T. Kenny, D.L. Sacks, J.P. Sypek, L. Vilela, A.A. Gam, K. EvansDavis, Protective immunity using recombinant human IL-12 and alum as adjuvants in a primate model of cutaneous leishmaniasis, J. Immunol. 163 (1999) 4481e4488. P.C. Melby, G.B. Ogden, H.A. Flores, W. Zhao, C. Geldmacher, N.M. Biediger, S.K. Ahuja, J. Uranga, M. Melendez, Identification of vaccine candidates for experimental visceral leishmaniasis by immunization with sequential fractions of a cDNA expression library, Infect. Immun. 68 (2000) 5595e5602. C.R. Engwerda, M.L. Murphy, S.L. Cotterell, S.C. Smelt, P.M. Kaye, Neutralization of IL-12 demonstrates the existence of discrete organspecific phases in the control of Leishmania donovani, Eur. J. Immunol. 28 (1998) 669e680. H.W. Murray, J. Hariprashad, R.L. Coffman, Behaviour of visceral Leishmania donovani in an experimentally induced T helper associated response model, J. Exp. Med. 185 (1997) 867e874. H.W. Murray, C.F. Nathan, Macrophage microbicidal mechanisms in vivo: reactive nitrogen versus oxygen intermediates in the killing of intracellular visceral Leishmania donovani, J. Exp. Med. 189 (1999) 741e746. P.M. Kaye, A.J. Curry, J.M. Blackwell, Differential production of Th1and Th2-derived cytokines does not determine the genetically controlled or vaccine-induced rate of cure in murine visceral leishmaniasis, J. Immunol. 146 (1991) 2763e2770. M. Kemp, J.A. Kurtzhals, K. Bendtzen, L.K. Poulsen, M.B. Hansen, D.K. Koech, A. Kharazmi, T.G. Theander, Leishmania donovani reactive Th1- and Th2-like T-cell clones from individuals who have recovered from visceral leishmaniasis, Infect. Immun. 61 (1993) 1069e1073. K. Kemp, Cytokine-producing T cell subsets in human leishmaniasis, Arch. Immunol. Ther. Exp. 48 (2000) 173e177. C.L. Karp, S.H. el-Safi, T.A. Wynn, M.M. Satti, A.M. Kordofani, F.A. Hashim, M. Hag-Ali, F.A. Neva, T.B. Nutman, D.L. Sacks, In vivo cytokine profiles in patients with kala-azar. Marked elevation of both interleukin-10 and interferon-gamma, J. Clin. Invest 91 (1993) 1644e1648. M.L. Murphy, S.E.J. Cotterell, P.M.A. Gorak, C.R. Engwerda, P.M. Kaye, Blockade of CTLA4 enhances host resistance to the intracellular pathogen, Leishmania donovani, J. Immunol. 166 (1998) 4153e4160. M.L. Murphy, U. Wille, E.N. Villegas, C.A. Hunter, J.P. Farrell, IL-10 mediates susceptibility to Leishmania donovani infection, Eur. J. Immunol. 31 (2001) 848e856.
J. Alexander, E. McFarlane / Microbes and Infection 10 (2008) 953e959 [55] M. Bodas, N. Jain, A. Awasthi, S. Martin, R. Kumar, P. Loka, D. Dandekar, D. Mitra, B. Saha, Inhibition of IL-2 induced IL-10 production as a principle of phase-specific immunotherapy, J. Immunol. 177 (2006) 4636e4643. [56] A. Satoskar, H. Bluethmann, J. Alexander, Disruption of the murine interleukin-4 gene inhibits disease progression during Leishmania mexicana infection but does not increase control of Leishmania donovani infection, Infect. Immun. 63 (1995) 4894e4899. [57] H.W. Murray, C. Montelibano, R. Peterson, J.P. Sypek, Interleukin-12 regulates the response to chemotherapy in experimental visceral leishmaniasis, J. Infect. Dis. 182 (2000) 1497e1502. [58] H.W. Murray, M.J. Oca, A.M. Granger, R.D. Schreiber, Requirement for T cells and effect of lymphokines in successful chemotherapy for an intracellular infection, J. Clin. Invest 83 (1989) 1253e1257. [59] R. Basu, S. Bhaumik, A.K. Halder, K. Naskar, T. De, S.K. Dana, P. Walden, S. Roy, Hybrid cell vaccination resolves Leishmania donovani infection by eliciting a strong CD8þ cytotoxic T-lymphocyte response with concomitant suppression of interleukin-10 (IL-10) but not IL-4 or IL-13, Infect. Immun. 75 (2007) 5956e5966. [60] R. Basu, S. Bhaumik, J.M. Basu, K. Naskar, T. De, S. Roy, Kinetoplastid membrane protein-11 DNA vaccination induces complete protection against both pentavalent antimonial-sensitive and -resistant strains of Leishmania donovani that correlates with inducible nitric oxide synthase activity and IL-4 generation: evidence for mixed Th1- and Th2-like responses in visceral leishmaniasis, J. Immunol. 174 (2005) 160e171. [61] J.C. Mottram, G.H. Coombs, J. Alexander, Cysteine peptidases as virulence factors of Leishmania, Curr. Opin. Microbiol. 7 (2004) 375e381. [62] L.U. Buxbaum, J.E. Uzonna, M.H. Goldschmidt, P. Scott, Control of New World cutaneous leishmaniasis is IL-12 independent but STAT4 dependent, Eur. J. Immun. 32 (2002) 3206e3615.
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[63] A. Satoskar, J. Alexander, Sex determined susceptibility and differential IFN-g and TNF-a in mRNA expression in DBA/2 mice infected with Leishmania mexicana, Immunology 84 (1995) 1e4. [64] A. Satoskar, F. Brombacher, W.J. Dai, I. McInnes, F.Y. Liew, J. Alexander, W. Walker, SCID mice reconstituted with IL-4-deficient lymphocytes, but not immunocompetent lymphocytes, are resistant to cutaneous leishmaniasis, J. Immunol. 159 (1997) 5005e5013. [65] K. Mohrs, A.E. Wakil, N. Killeen, R.M. Locksley, M. Mohrs, A two-step process for cytokine production revealed by IL-4 dual-reporter mice, Immunity 23 (2005) 419e429. [66] J. Alexander, F. Brombacher, H.A. McGachy, A.N. McKenzie, W. Walker, K.C. Carter, An essential role for IL-13 in maintaining a nonhealing response following Leishmania mexicana infection, Eur. J. Immunol. 32 (2002) 2923e2933. [67] D.E. Jones, L.U. Buxbaum, P. Scott, IL-4-independent inhibition of IL12 responsiveness during Leishmania amazonensis infection, J. Immunol. 165 (2000) 364e372. [68] D.E. Jones, M.R. Ackermann, U. Wille, C.A. Hunter, P. Scott, Early enhanced Th1 response after Leishmania amazonensis infection of C57BL/6 interleukin-10-deficient mice does not lead to resolution of infection, Infect. Immun. 70 (2002) 2151e2058. [69] U.M. Padigel, J. Alexander, J.P. Farrell, The role of interleukin-10 in susceptibility of BALB/c mice to infection with Leishmania mexicana and Leishmania amazonensis, J. Immunol. 171 (2003) 3705e3710. [70] L.U. Buxbaum, P. Scott, Interleukin 10 and Fc gamma receptor-deficient mice resolve Leishmania mexicana lesions, Infect. Immun. 73 (2005) 101e108. [71] A.C. Hsu, P. Scott, Leishmania mexicana infection induces impaired lymph node expansion and Th1 cell differentiation despite normal T cell proliferation, J. Immunol. 179 (2007) 8200e8207.
Original article
Can type-1 responses against intracellular pathogens be T helper 2 cytokine dependent? James Alexander*, Emma McFarlane Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, The John Arbuthnott Building, 27 Taylor St, Glasgow G4 ONR, UK Available online 13 August 2008
Abstract While control of intracellular pathogens, such as the protozoan Leishmania, is dependent on the generation of type-1 immune responses, the role of T helper 2 cytokines in the disease process is more controversial. Traditionally these cytokines were perceived as counter-regulating type1 responses and promoting disease exacerbation. Nevertheless a substantial body of evidence now exists suggesting that the development of effective type-1 immunity can involve the significant involvement of the Th2 cytokines IL-4 and IL-13. This article reviews, using Leishmania species in particular, the circumstances under which these cytokines can promote protective type-1 immunity. Ó 2008 Elsevier Masson SAS. All rights reserved. Keywords: Th1; Th2; IL-4; IL-13; Leishmania
1. Introduction The seminal work of Mosmann, Coffman and colleagues [1] describing two distinct CD4þ T helper cell populations in 1986 ultimately gave rise to the Th1/Th2 paradigm of susceptibility/resistance to intracellular infection. This has provided an excellent foundation from which immunologists can further dissect the immune response and design appropriate therapeutic and intervention strategies. However, not only has the more recent characterisation of CD4þ T cell regulatory populations as well as a further effector CD4þ T helper population (Th17) [2] added to the complexity of hoste pathogen interactions but there is also a substantial body of evidence derived from both infectious and inflammatory disease models indicating that the archetypal Th2 cytokines IL-4 and IL-13 need not necessarily counter-regulate but could actually drive, facilitate or promote a Th1 response (Table 1). Consequently, IL-4/IL-13 has been shown to be associated
* Corresponding author. Tel.: þ44 141 548 3925; fax: þ44 141 552 3562. E-mail address: [email protected] (J. Alexander). 1286-4579/$ - see front matter Ó 2008 Elsevier Masson SAS. All rights reserved. doi:10.1016/j.micinf.2008.07.038
with driving protective type-1 responses against, Toxoplasma gondii [3], Listeria monocytogenes [4], Cryptosporidium parvum [5], Leishmania major [6] and Leishmania donovani [7e11] as well as aggravating Th1 responses in experimental autoimmune uveoretinitis and a murine model of colitis [12,13]. IL-4/IL-13 may mediate their effects by inducing macrophage and dendritic cell IL-12 production [6,14], enhancing IFN-g production [15] or synergising with IFN-g for enhanced anti-microbial activity [16,17]. Leishmania spp. have long been considered ideal models to study chronic intracellular infections as protective immunity is globally recognised as being type-1 dependent. However, the causative agents of both Old World and New World cutaneous leishmaniasis as well as visceral leishmaniasis diverged in evolutionary terms 40e80 million years ago (reviewed in [18]). Therefore it is hardly surprising that the separate species have developed different strategies for evading and interacting with the immune response. Consequently they provide excellent tools to dissect different pathways leading to, or preventing, a protective type-1 response. In this review, we explore, using different species of Leishmania, the spectrum of Th2 cytokine activities operating in the disease process.
J. Alexander, E. McFarlane / Microbes and Infection 10 (2008) 953e959 Mencacci et al., 1998 [5] McDonald et al., 2004 [6] Biedermann et al., 2001 [7] Alexander et al., 2000 [8] Stager et al., 2003 [9] Stager et al., 2003 [10] Murray et al., 2006 [11] McFarlane et al., unpublished
Flesch et al., 1997 [4]
Suzuki et al., 1996 [3]
Mice succumbed to infection and brain parasite burden increased Number of viable bacteria in spleen and liver reduced Mice succumb to infection Susceptibility to infection increased Footpad lesion size reduced Parasite burdens increased Parasite burdens increased Parasite burdens reduced Retarded granuloma formation Parasite burdens increased and retarded granuloma formation IL-13/ IL-13/
IL-4/ IL-4Ra/
BALB/c BALB/c BALB/c BALB/c BALB/c BALB/c BALB/c BALB/c Secondary infection Primary infection rIL-4 treatment Chemotherapy Primary infection Vaccine Primary infection Primary infection þ þ Candida albicans Cryptosporidium parvum Leishmania major Leishmania donovani Leishmania donovani Leishmania donovani Leishmania donovani Leishmania donovani
IL-4 IL-4 IL-4 IL-4 IL-4Ra IL-4 IL-13 IL-13
rIL-13 treatment þ IL-13 Listeria monocytogenes
C57BL/6
IL-4/ IL-4/
IFNg production from spleen cells reduced Frequency of IL-12 producing cells increased Defective IFNg and IL-12 production IL-12 and IFNg downregulated Enhanced IL-12 production by APCs IL-12 and IFNg downregulated IFNg downregulated CD8þT cells producing IFNg IFNg downregulated IFNg downregulated Primary infection IL-4 Toxoplasma gondii
129/J IL-4
Disease Immunology Mouse strain Context Cytokine Microbial species
Table 1 Studies where IL-4 and IL-13 have been shown to promote Th1 responses and protection against microbial infection
/
Reference
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2. Leishmania major: is non-healing disease IL-4/IL-13 dependent or independent or can these cytokines be protective? The classic studies by Locksley, Coffman and colleagues on Leishmania major which demonstrated that an IL-4 driven type-2 response and associated cytokines counter-regulated type-1 responses and consequently promoted non-healing infection have in large part given rise to the Th1/Th2 paradigm of resistance/susceptibility to intracellular parasitism [19]. Further studies in susceptible BALB/c mice demonstrated that a single T-cell epitope derived from the parasite LACK antigen (Leishmania homologue of receptors for activated C kinase) induced rapid IL-4 production by Vb4Va8CD4þ T cells which apparently rendered T cells unresponsive to IL-12 and correlated with lesion development [20]. Thus a particularly strong case for the predominant role of IL-4 and a Th2 response in non-healing cutaneous disease was established. Nevertheless, other studies on resistant C57BL/6 mice noted that this strain also produced IL-4 early during infection and yet developed an unimpaired Th1 biased response [21]. Subsequently studies utilizing IL-4/ mice seriously questioned the paramount role of IL-4 in the non-healing response of BALB/c mice to L. major as a number of apparently contradictory reports determining the significance of IL-4 were published [22e24]. This posed the question as to whether other cytokines could be regulating Th1 development. As IL-4 and IL-13 share a common signaling pathway through the IL-4 receptor a (IL-4Ra) chain and as IL-4Ra deficient mice were found to be more resistant than IL-4/ mice in one study this suggested a compensatory role for IL-13 in the absence of IL-4 [25]. However, in a further study IL-13 was also found to be able to act independently of IL-4 and it was surmised that the effects of IL-4 and IL-13 were additive [26]. Nevertheless, in another study IL-4Ra/ mice were found to be more susceptible than IL-4/ animals indicating a protective role for IL-13 in preventing disease dissemination in the absence of IL-4 signaling [27]. While question marks remain over whether IL-4 and IL-13 play a significant role in L. major disease progression in BALB/c mice IL-10 unquestionably does promote parasite persistence in resistant mice and multiplication in susceptible mice [28]. IL-10 could be generated via either Fcg-mediated uptake of antibody coated amastigotes [29], or produced by Th2 cells, or CD4þ CD25þ T regulatory cells [30] or indeed a recently described CD4þCD25FoxP3 Th1 population [31]. In the absence of a definitive role for IL-4/IL-13 in nonhealing disease it has been proposed that defective T cell responses as a consequence of intrinsic defects in antigen presenting cell function or alternatively in T cell development could be the ultimate underlying problem determining the exquisite susceptibility of BALB/c mice to infection with L. major. While several studies suggest that non-healing disease is a consequence of the failure to produce or respond to IL-12 [32,33] this could be irrespective of the presence of a Th2 response and although IL-4 can downregulate IL-12 production and expression of IL-12Rb2 on CD4þ T cells
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[20,33,34], this could also be by IL-4 independent mechanisms [34]. In this context it is significant that IL-12b2 expression remains impaired in IL-4Ra/ BALB/c mice compared with resistant C57BL/6 mice infected with L. major [24]. Defects in antigen presenting cell function reported for BALB/c mice include a comparative deficiency in dendritic cell IL-1 production [35] compared with L. major resistant strains. IL-1 has been shown to upregulate dendritic cell IL-12 production as well as MHC Class II and co-stimulatory molecule expression [36] as well to enhance IFN-g mediated inhibition of Th2 cell proliferation [37]. However, as transgenic IL-12Rb2 BALB/c mice exhibit a non-healing phenotype despite intact IL-12 signaling the importance of an APC defect remains questionable [38]. Irrespective of any potential defect in APC function BALB/c CD4þ T cells have been demonstrated as having an intrinsic IL-4 independent Th2 bias [39]. A commitment to Th2 development could be a consequence of a defective co-polarisation of the TCR and IFN-g receptor complex of naive CD4þ T cells at the immunological APC/T cell precursor synapse in BALB/c mice compared with C57BL/6 mice [40]. Nevertheless, immune regulatory mechanisms as well as intrinsic T cell defects can serve to impair Th1 responses in BALB/c mice as IL-4 signaling via STAT-6 also serves to block co-localisation of this complex. Recent studies may serve to help reconcile the observations above. CD4þ T cell specific IL-4Ra/ BALB/c mice unlike global IL-4Ra/ animals, which develop disseminating disease, are as resistant as C57BL/6 mice to infection with L. major [41]. Consequently it has been concluded that IL-4/IL13 responsive non-CD4þ T cells induce protection against L. major in the absence of CD4þ T cells responding to IL-4 [42]. Under such circumstances it can be hypothesised that the reported intrinsic BALB/c T cell defect is not enough in the absence of CD4þ T cell responding to IL-4 to prevent the generation of a Th1 response. Conversely C57BL/6 mice, like BALB/c mice, also produce IL-4 early in infection but in the absence of the intrinsic T cell defect that operates in BALB/c mice C57BL/6 mice would proceed to resolve infection as a Th1 response then develops. Overall this would suggest a potential significant role for IL-4Ra responsive non-CD4þ T cells in generating a Th1 biased response in L. major resistant strains. Studies utilizing macrophage specific BALB/c IL-4Ra deficient mice have demonstrated that IL-4/IL-13 operates through this population to enhance parasite growth via alternative macrophage activation [42] and consequently these are unlikely to be the population driving a Th1 response in CD4þIL-4Ra/ mice. However, IL-4 treatment of BALB/c mice pre T cell priming has previously been demonstrated to instruct dendritic cells to produce IL-12 and facilitate a protective Th1 response against L. major [7]. This would indicate that dendritic cells are the IL-4/IL-13 responsive cells facilitating protection in the absence of IL-4Ra responsive CD4þ T cells in BALB/c mice. A potential contribution of IL4/IL-13 responsive non-CD4þcells to protective immunity may explain why Leishmania vaccines incorporating IFN-gpromoting adjuvants are relatively ineffective compared with those that also utilize ALUM [43,44].
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3. Leishmania donovani: is healing IL-4/IL13 dependent or independent? Infection with the intracellular protozoan parasite Leishmania donovani causes a potentially fatal visceral infection if left untreated, and has resulted in up to 100,000 deaths in recent epidemics in Sudan and India [45]. Protective immunity against this parasite, as with species causing cutaneous leishmaniasis, is dependent on an IL-12 driven type-1 response and IFN-g production [46,47], which results in the induction of parasite killing by macrophages primarily via the production of reactive nitrogen and oxygen intermediates [48]. Studies, not only in mice [49], but also humans [50e 52], suggested that cure was independent of the differential production of Th1 and Th2 cytokines and both IFN-g and IL-4 producing T cells have been isolated from asymptomatic and cured patients. Indeed in the mouse model it has been shown that the induction of enhanced protection is related to increasing the frequency of cytokine-producing cells rather than altering the IFN-g /IL-4 balance [53]. In addition, while a recent study has shown that the control of L. donovani in the susceptible BALB/c mouse is IL-12 dependent, this cytokine was found to promote the expansion of Th2 as well as Th1 responses [46]. Both human [50,53] and murine studies [54] indicate that IL-10 is the major immunosuppressive cytokine in visceral leishmaniasis. Furthermore it has been demonstrated that the source of this IL-10 is not traditional Th2 cells [53] but more likely a regulatory T cell population [55]. Thus it would probably be the general consensus of opinion that the Th2 response does not contribute adversely to the immunological control of visceral leishmaniasis. Indeed studies utilising gene deficient mice have identified a protective role for IL-4 and IL-4Ra signalling (Fig. 1) during primary L. donovani infections [9,56]. Furthermore the increased susceptibility of IL-4Ra/ mice compared with IL-4/ BALB/c mice as measured by hepatic parasite burdens [9] also suggested a protective role for IL-13 as both IL-4 and IL-13 activities are dependent on the expression of IL-4Ra. Control of parasite growth in the liver was associated with the ability to produce sterile granulomas [9], a mechanism driven by T cell derived IFNg [57]. Enhanced susceptibility of IL-4/ and IL-4Ra/ mice was associated with downregulated type-1 responses and markedly retarded granuloma maturation [8,9]. While decreased IFN-g and retarded granuloma maturation have also been observed in IL-13/ mice this was not associated with a reduced parasite burden [11]. The recent availability of tissue specific IL-4Ra/ mice will facilitate the dissection of the IL-4 and IL-13 effects during visceral leishmaniasis and so far we have examined the course of L. donovani in macrophage/neutrophil specific IL-4Ra/ and CD4þ Tcell specific IL-4Ra/ BALB/c mice. The protective activity of IL-4/IL-13 during visceral leishmaniasis is not via these cell populations (McFarlane, Carter, Kaye, Brombacher and Alexander, unpublished). In addition to playing a significant protective role during primary infection with L. donovani effective sodium
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Fig. 1. Liver parasite burdens (A) in wild-type and IL-4Ra/ mice at day 40 post-infection with (1e2)107 L. donovani amastigotes. Hepatic granuloma maturation in L. donovani infected wild-type (B) and IL-4Ra/ (C) mice at day 40 post-infection. Wild-type mice (B) show well developed mature granulomas (arrows) surrounding a core of infected Kupffer cells whilst IL-4Ra/ mice (C) display abrogated granuloma development characterised by the presence of immature granulomas and amastigotes within the cytoplasm of infected Kupffer cells (arrows). Magnification 400.
stibogluconate (SSG) chemotherapy is dependent on Th2 cytokines [8]. Previous studies have demonstrated that successful treatment of visceral leishmaniasis with SSG requires the presence of both CD4þ and CD8þ T cells [58] accompanied by the type-1 cytokines IL-12 and IFN-g [57]. We have observed that SSG chemotherapy of L. donovani in IL-4/ [8] or IL-13/ (unpublished) BALB/c mice compared with wild-type mice resulted in downregulated IFN-g production and increased IL-10 production. This would suggest a requirement for either of these cytokines in instructing a type-1 response. Significantly, successful immunotherapeutic intervention in the BALB/c mouse using a hybrid cell vaccine which was mediated by a strong CD8þ CTL response was associated with downregulation of IL-10 but significantly enhanced IL-4 and IL-13 expression as well as IFN-g production [59] providing further strong circumstantial evidence for these Th2 cytokines facilitating the generation of type-1 immunity. Similarly the protection afforded by a kinetoplast membrane protein protein-11 DNA vaccine which protected hamsters against visceral leishmaniasis correlated with decreased IL-10 levels but with increased IL-4 production and an elevated mixed Th1/Th2
response [60]. However, vaccination studies utilizing HASPB-1 have provide the most concrete evidence for a Th2 cytokine requirement to instruct a protective type-1 response mediated by CD8þ T cells against visceral leishmaniasis [10] as this vaccine generates a mixed Th1/Th2 response and is totally ineffective in IL-4 or IL-4Ra/ mice. The primary source of IL-4 has been defined as a CD11bþCD11clo phagocyte [10]. 4. Leishmania mexicana: is non-healing disease IL-4/IL13 dependent? New world cutaneous leishmaniasis resulting from infection with parasites of the mexicana complex is under different genetic and immunoregulatory controls to those controlling L. major infection and cysteine peptidases would appear to be the major virulence factors with evidence that they can both promote a Th2 response as well as inhibit a protective Th1 response [61]. Also unlike L. major the majority of mouse strains are susceptible to this parasite. As with the other Leishmania species protective immunity is the result of a STAT4 dependent type-1 immune response although this apparently
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can be generated independently of IL-12 [62]. Early studies on Leishmania mexicana complex parasites that examined IL-4 mRNA transcript expression in the draining lymph nodes of resistant and susceptible mice indicated that IL-4 played no role in the non-healing response of mice to this parasite [63]. Nevertheless, follow up experiments using IL-4/ mice to confirm this observation demonstrated that IL-4 was after all the predominant cytokine responsible for non-healing disease [56,64]. However, these apparently contradictory results are now easy to reconcile as it is now well established that IL-4 transcript expression is not a reliable indicator of IL-4 production [65]. In addition studies using IL-13/ mice also indicate that IL-13 may play a role in preventing disease resolution by inhibiting IL-12Rb2 expression [66]. The paramount role of IL-4 in promoting disease progression is, however, mouse strain and site of infection dependent. Thus the growth of both L. mexicana and L. amazonensis is primarily IL4 dependent in rump infections in BALB/c and C57BL/6 mice [18]. By contrast the growth of these parasites in the footpad of C57BL/6 mice, though not BALB/c mice, is IL-4 independent [18,67] and the contribution of IL-10 to this activity may also depend on whether L. amazonensis or L. mexicana is studied [68e70]. A recent elegant study associates L. mexicana with not only the ability to inhibit Th1 differentiation but also to impaired lymph node expansion [71]. Overall there is no evidence whatsoever that Th2 cytokines can be beneficial during infection with New World cutaneous leishmaniasis and often they play the classic counter-protective role. 5. Conclusions IL-4 and IL-13 are pleiotropic cytokines and numerous cell types express their receptors. Consequently it is not surprising that their activity can vary significantly depending on the microbe initiating infection, the tissue site or cells involved, and the chronicity of infection. The three Leishmania species/ complexes discussed above are all controlled by a type-1 response. Nevertheless they not only differ significantly in how they can evade, or manipulate the immune response to induce non-healing disease but also the immunological pathways leading to protective type-1 responses would appear to be under different controls for each species. Consequently IL-4, IL-13 and IL-10 all promote infection with L. mexicana complex parasites and there is no evidence whatsoever for a protective role for the Th2 cytokines. Conversely, strong evidence has now been accumulating suggesting that Th2 cytokines not only do not contribute to the progression of visceral leishmaniasis but that they also actually promote the induction of type-1 immunity. Indeed IL-4 used judiciously can also be used to drive a protective type-1 response against L. major. The recent generation of spatial (tissue specific) and temporal (switch on/ switch off) IL-4Ra/ mice has provided new powerful tools by which the specific role of IL-4/IL-13 responding cells can be dissected from global effects. This will ultimately facilitate the design of more appropriate and effective anti-microbial therapeutic and prophylactic strategies.
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Acknowledgements J.A. is in receipt of funding from the Wellcome Trust and Royal Society. E.McF. is an E.P.S.R.C. sponsored Research Fellow. References [1] T.R. Mosmann, H. Cherwinski, M.W. Bond, M.A. Giedlin, R.L. Coffman, Two types of murine helper T cell clone. 1. Definition according to profiles of lymphokine activities and secreted proteins, J. Immunol. 136 (1986) 2348e2357. [2] E. Bettelli, M. Oukka, V.K. Kuchroo, T(H)-17 cells in the circle of immunity and autoimmunity, Nat. Immunol. 8 (2007) 345e350. [3] Y. Suzuki, Q. Yang, S. Yang, N. Nguyen, S. Lim, O. Liesenfeld, T. Kojima, J.S. Remington, IL-4 is protective against development of toxoplasmic encephalitis, J. Immunol. 157 (1996) 2564e2569. [4] I.E. Flesch, A. Wandersee, S.H. Kaufmann, Effects of IL-13 on murine listeriosis, Int. Immunol. 9 (1997) 467e474. [5] A. Mencacci, G. Del Sero, E. Cenci, C.F. d’Ostiani, A. Bacci, C. Montagnoli, M. Kopf, L. Romani, Endogenous interleukin 4 is required for development of protective CD4þ T helper type 1 responses to Candida albicans, J. Exp. Med. 187 (1998) 307e317. [6] S.A. McDonald, J.E. O’Grady, M. Bajaj-Elliott, C.A. Notley, J. Alexander, F. Brombacher, V. McDonald, Protection against the early acute phase of cryptosporidium parvum infection conferred by Interleukin-4-induced expression of T Helper 1 cytokines, J. Infect. Dis. 190 (2004) 1019e1025. [7] T. Biedermann, S. Zimmermann, H. Himmelrich, A. Gumy, O. Egeter, A.K. Sakrauski, I. Seegmu¨ller, H. Voigt, P. Launois, A.D. Levine, H. Wagner, K. Heeg, J.A. Louis, M. Rocken, IL-4 instructs TH1 responses and resistance to Leishmania major in susceptible BALB/c mice, Nat. Immunol. 2 (2001) 1054e1060. [8] J. Alexander, K.C. Carter, N. Al-Fasi, A. Satoskar, F. Brombacher, Endogenous IL-4 is necessary for effective drug therapy against visceral leishmaniasis, Eur. J. Immunol. 30 (2000) 2935e2943. [9] S. Stager, J. Alexander, K.C. Carter, F. Brombacher, P.M. Kaye, Both interleukin-4 (IL-4) and IL-4 receptor alpha signaling contribute to the development of hepatic granulomas with optimal antileishmanial activity, Infect. Immun. 71 (2003) 4804e4807. [10] S. Stager, J. Alexander, A.C. Kirby, M. Botto, N.V. Rooijen, D.F. Smith, F. Brombacher, P.M. Kaye, Natural antibodies and complement are endogenous adjuvants for vaccine-induced CD8þ T-cell responses, Nat. Med. 9 (2003) 1287e1292. [11] H.M. Murray, C.W. Tsai, J. Liu, X. Ma, Visceral Leishmania donovani infection in interleukin-13/ mice, Infect. Immun. 74 (2006) 2487e 2490. [12] S. Ramanathan, Y. de Kosak, A. Saoudi, O. Goureau, P.H. Van der Meide, P. Druet, B. Bellon, Recombinant IL-4 aggravates experimental autoimmune uveoretinitis in rats, J. Immunol. 157 (1996) 2209e2215. [13] M.M. Fort, R. Lesley, N.J. Davidson, S. Menon, F. Brombacher, M.W. Leach, D.M. Rennick, IL-4 exacerbates disease in a Th1 cell transfer model of colitis, J. Immunol. 166 (2001) 2793e2800. [14] H. Hochrein, M. O’Keefe, T. Luft, S. Vandenabeele, R.J. Grumont, E. Maraskovsky, K. Shortman, Interleukin (IL-4) is a major regulatory cytokine governing bioactive IL-12 production by mouse and human dendritic cells, J. Exp. Med. 192 (2000) 823e833. [15] A. Noble, D.M. Kemeny, Interleukin-4 enhances interferon-g synthesis but inhibits development of interferon-g-producing cells, Immunology 85 (1995) 357e363. [16] I.-S. Lean, S.A. McDonald, M. Bajaj-Elliott, R.C.G. Pollok, M.J.G. Farthing, V. McDonald, Interleukin-4 and transforming growth factor-beta have opposing regulatory effects on gamma interferonmediated inhibition of Cryptosporidium parvum reproduction, Infect. Immun. 71 (2003) 4580e4585. [17] C. Bogdan, S. Stenger, M. Rollinghoff, W. Solbach, Cytokine interactions in experimental cutaneous leishmaniasis. Interleukin 4
958
[18]
[19]
[20]
[21]
[22]
[23] [24]
[25]
[26]
[27]
[28]
[29] [30]
[31]
[32]
[33]
[34]
[35]
[36]
J. Alexander, E. McFarlane / Microbes and Infection 10 (2008) 953e959 synergizes with interferon-gamma to activate murine macrophages for killing of Leishmania major amastigotes, Eur. J. Immunol. 21 (1993) 327e333. D. McMahon-Pratt, J. Alexander, Does the Leishmania major paradigm of pathogenesis and protection hold for New World cutaneous leishmaniases or the visceral disease? Immunol. Rev. 201 (2004) 206e224. F.P. Heinzel, M.D. Sadick, B.J. Holaday, R.L. Coffman, R.M. Locksley, Reciprocal expression of interferon gamma or interleukin 4 during the resolution or progression of murine leishmaniasis. Evidence for expansion of distinct helper T cell subsets, J. Exp. Med. 169 (1989) 59e72. P. Launois, T. Ohteki, K. Swihart, H.R. MacDonald, J.A. Louis, In susceptible mice, Leishmania major induce very rapid interleukin-4 production by CD4þ T cells which are NK1.1, Eur. J. Immunol. 25 (1995) 3298e3007. P. Scott, A. Eaton, W.C. Gause, X. di Zhou, B. Hondowicz, Early IL-4 production does not predict susceptibility to Leishmania major, Exp. Parasitol. 84 (1996) 178e187. M. Kopf, F. Brombacher, G. Kohler, G. Kienzle, K.H. Widmann, K. Lefrang, B. Humborg, B. Lederman, W. Solbach, IL-4 deficient mice resist infection with Leishmania major, J. Exp. Med. 184 (1996) 1127e1136. N. Noben-Trauth, P. Kropf, I. Muller, Susceptibility to Leishmania major infection in interleukin-4-deficient mice, Science 271 (1996) 987e990. M. Mohrs, C. Holscher, F. Brombacher, Interleukin-4 receptor alpha deficient BALB/c mice show an unimpaired T helper 2 polarization in response to Leishmania major infection, Infect. Immun. 68 (2000) 1773e 1780. N. Noben-Trauth, W.E. Paul, D.L. Sacks, IL-4- and IL-4 receptor deficient BALB/c mice reveal differences in susceptibility to Leishmania major parasite substrains, J. Immunol. 162 (1999) 6132e6140. D.J. Matthews, C.L. Emson, G.J. McKenzie, H.E. Jolin, J.M. Blackwell, A.N. McKenzie, IL-13 is a susceptibility factor for Leishmania major infection, J. Immunol. 164 (2000) 1458e1462. M. Mohrs, B. Ledermann, G. Kohler, A. Dorfmuller, A. Gessner, F. Brombacher, Differences between IL-4- and IL-4 receptor alphadeficient mice in chronic leishmaniasis reveal a protective role for IL-13 receptor signaling, J. Immunol. 162 (1999) 7302e7308. N. Noben-Trauth, R. Lira, H. Nagase, W.E. Paul, D.L. Sacks, The relative contribution of IL-4 receptor signaling and IL-10 to susceptibility to Leishmania major, J. Immunol. 170 (2003) 5152e5158. M.M. Kane, D.M. Mosser, The role of IL-10 in promoting disease progression in leishmaniasis, J. Immunol. 166 (2001) 1141e1147. D. Sacks, C. Anderson, Re-examination of the immunosuppressive mechanisms mediating non-cure of Leishmania infection in mice, Immunol. Rev. 201 (2004) 225e238. C.F. Anderson, M. Oukka, V.J. Kuchroo, D.L. Sacks, CD4þCD25Foxp3-Th1 cells are the source of IL-10-mediated immune suppression in chronic cutaneous leishmaniasis, J. Exp. Med. 204 (2007) 285e297. S.L. Reiner, S. Zheng, Z.E. Wang, L. Stowring, R.M. Locksley, Leishmania promastigotes evade interleukin 12 (IL-12) induction by macrophages and stimulate a broad range of cytokines from CD4þ T cells during initiation of infection, J. Exp. Med. 179 (1994) 447e456. H. Himmelrich, C. Parra-Lopez, F. Tacchini-Cottier, J.A. Louis, P. Launois, The IL-4 rapidly produced in BALB/c mice after infection with Leishmania major down-regulates IL-12 receptor beta 2-chain expression on CD4þ T cells resulting in a state of unresponsiveness to IL-12, J. Immunol. 161 (1998) 6156e6163. D. Jones, M.M. Elloso, L. Showe, D. Williams, G. Trinchieri, P. Scott, Differential regulation of the interleukin-12 receptor during the innate immune response to Leishmania major, Infect. Immun. 66 (1998) 3818e3824. E. Von Stebut, J.M. Ehrchen, Y. Belkaid, S.L. Kostka, K. Molle, J. Knop, C. Sunderkotter, M.C. Udey, Interleukin 1alpha promotes Th1 differentiation and inhibits disease progression in Leishmania major susceptible BALB/c mice, J. Exp. Med. 198 (2003) 191e199. U. Eriksson, M.O. Kurrer, I. Sonderegger, G. Iezzi, A. Tafuri, L. Hunziker, S. Suzuki, K. Bachmaier, R.M. Bingisser, J.M. Penninger, M. Kopf, Activation of dendritic cells through the interleukin 1 receptor 1
[37]
[38]
[39]
[40]
[41]
[42]
[43]
[44]
[45]
[46]
[47]
[48]
[49]
[50]
[51] [52]
[53]
[54]
is critical for the induction of autoimmune myocarditis, J. Exp. Med. 197 (2003) 323e331. T.B. Oriss, S.A. McCarthy, B.F. Morel, M.A. Campana, P.A. Morel, Cross-regulation between T helper cell (Th)1 and Th2: inhibition of Th2 proliferation by IFN-g involves interference with IL-1, J. Immunol. 158 (1997) 3666e3672. R. Nishikomori, S. Gurunathan, K. Nishikomori, W. Strober, BALB/c mice bearing a transgenic IL-12 receptor beta 2 gene exhibit a nonhealing phenotype to Leishmania major infection despite intact IL-12 signaling, J. Immunol. 166 (2001) 6776e6783. M. Bix, Z.E. Wang, B. Thiel, N.J. Schork, R.M. Locksley, Genetic regulation of commitment to interleukin 4 production by a CD4þ T cellintrinsic mechanism, J. Exp. Med. 188 (1998) 2289e2299. R.A. Maldonado, D.J. Irvine, R. Schreiber, L.H. Glimcher, A role for the immunological synapse in lineage commitment of CD4 lymphocytes, Nature 431 (2004) 527e532. M. Radwanska, A.J. Cutler, J.C. Hoving, S. Magez, C. Holscher, A. Bohms, B. Arendse, R. Kirsch, T. Hunig, J. Alexander, P.M. Kaye, F. Brombacher, Deletion of IL-4Ralpha on CD4 T cells renders BALB/c mice resistant to Leishmania major infection, PLoS Pathog. 3 (5) (2007) e68. C. Holscher, B. Arendse, A. Schwegmann, E. Myburgh, F. Brombacher, Impairment of alternative macrophage activation delays cutaneous leishmaniasis in non-healing BALB/c mice, J. Immunol. 176 (2006) 1115e1121. M.M. Gicheru, J.O. Olobo, C.O. Anjili, F. Modabber, P. Scott, Vervet monkeys vaccinated with killed Leishmania major parasites and interleukin-12 develop a type 1 immune response but are not protected against challenge infection, Infect. Immun. 69 (2001) 245e251. R.T. Kenny, D.L. Sacks, J.P. Sypek, L. Vilela, A.A. Gam, K. EvansDavis, Protective immunity using recombinant human IL-12 and alum as adjuvants in a primate model of cutaneous leishmaniasis, J. Immunol. 163 (1999) 4481e4488. P.C. Melby, G.B. Ogden, H.A. Flores, W. Zhao, C. Geldmacher, N.M. Biediger, S.K. Ahuja, J. Uranga, M. Melendez, Identification of vaccine candidates for experimental visceral leishmaniasis by immunization with sequential fractions of a cDNA expression library, Infect. Immun. 68 (2000) 5595e5602. C.R. Engwerda, M.L. Murphy, S.L. Cotterell, S.C. Smelt, P.M. Kaye, Neutralization of IL-12 demonstrates the existence of discrete organspecific phases in the control of Leishmania donovani, Eur. J. Immunol. 28 (1998) 669e680. H.W. Murray, J. Hariprashad, R.L. Coffman, Behaviour of visceral Leishmania donovani in an experimentally induced T helper associated response model, J. Exp. Med. 185 (1997) 867e874. H.W. Murray, C.F. Nathan, Macrophage microbicidal mechanisms in vivo: reactive nitrogen versus oxygen intermediates in the killing of intracellular visceral Leishmania donovani, J. Exp. Med. 189 (1999) 741e746. P.M. Kaye, A.J. Curry, J.M. Blackwell, Differential production of Th1and Th2-derived cytokines does not determine the genetically controlled or vaccine-induced rate of cure in murine visceral leishmaniasis, J. Immunol. 146 (1991) 2763e2770. M. Kemp, J.A. Kurtzhals, K. Bendtzen, L.K. Poulsen, M.B. Hansen, D.K. Koech, A. Kharazmi, T.G. Theander, Leishmania donovani reactive Th1- and Th2-like T-cell clones from individuals who have recovered from visceral leishmaniasis, Infect. Immun. 61 (1993) 1069e1073. K. Kemp, Cytokine-producing T cell subsets in human leishmaniasis, Arch. Immunol. Ther. Exp. 48 (2000) 173e177. C.L. Karp, S.H. el-Safi, T.A. Wynn, M.M. Satti, A.M. Kordofani, F.A. Hashim, M. Hag-Ali, F.A. Neva, T.B. Nutman, D.L. Sacks, In vivo cytokine profiles in patients with kala-azar. Marked elevation of both interleukin-10 and interferon-gamma, J. Clin. Invest 91 (1993) 1644e1648. M.L. Murphy, S.E.J. Cotterell, P.M.A. Gorak, C.R. Engwerda, P.M. Kaye, Blockade of CTLA4 enhances host resistance to the intracellular pathogen, Leishmania donovani, J. Immunol. 166 (1998) 4153e4160. M.L. Murphy, U. Wille, E.N. Villegas, C.A. Hunter, J.P. Farrell, IL-10 mediates susceptibility to Leishmania donovani infection, Eur. J. Immunol. 31 (2001) 848e856.
J. Alexander, E. McFarlane / Microbes and Infection 10 (2008) 953e959 [55] M. Bodas, N. Jain, A. Awasthi, S. Martin, R. Kumar, P. Loka, D. Dandekar, D. Mitra, B. Saha, Inhibition of IL-2 induced IL-10 production as a principle of phase-specific immunotherapy, J. Immunol. 177 (2006) 4636e4643. [56] A. Satoskar, H. Bluethmann, J. Alexander, Disruption of the murine interleukin-4 gene inhibits disease progression during Leishmania mexicana infection but does not increase control of Leishmania donovani infection, Infect. Immun. 63 (1995) 4894e4899. [57] H.W. Murray, C. Montelibano, R. Peterson, J.P. Sypek, Interleukin-12 regulates the response to chemotherapy in experimental visceral leishmaniasis, J. Infect. Dis. 182 (2000) 1497e1502. [58] H.W. Murray, M.J. Oca, A.M. Granger, R.D. Schreiber, Requirement for T cells and effect of lymphokines in successful chemotherapy for an intracellular infection, J. Clin. Invest 83 (1989) 1253e1257. [59] R. Basu, S. Bhaumik, A.K. Halder, K. Naskar, T. De, S.K. Dana, P. Walden, S. Roy, Hybrid cell vaccination resolves Leishmania donovani infection by eliciting a strong CD8þ cytotoxic T-lymphocyte response with concomitant suppression of interleukin-10 (IL-10) but not IL-4 or IL-13, Infect. Immun. 75 (2007) 5956e5966. [60] R. Basu, S. Bhaumik, J.M. Basu, K. Naskar, T. De, S. Roy, Kinetoplastid membrane protein-11 DNA vaccination induces complete protection against both pentavalent antimonial-sensitive and -resistant strains of Leishmania donovani that correlates with inducible nitric oxide synthase activity and IL-4 generation: evidence for mixed Th1- and Th2-like responses in visceral leishmaniasis, J. Immunol. 174 (2005) 160e171. [61] J.C. Mottram, G.H. Coombs, J. Alexander, Cysteine peptidases as virulence factors of Leishmania, Curr. Opin. Microbiol. 7 (2004) 375e381. [62] L.U. Buxbaum, J.E. Uzonna, M.H. Goldschmidt, P. Scott, Control of New World cutaneous leishmaniasis is IL-12 independent but STAT4 dependent, Eur. J. Immun. 32 (2002) 3206e3615.
959
[63] A. Satoskar, J. Alexander, Sex determined susceptibility and differential IFN-g and TNF-a in mRNA expression in DBA/2 mice infected with Leishmania mexicana, Immunology 84 (1995) 1e4. [64] A. Satoskar, F. Brombacher, W.J. Dai, I. McInnes, F.Y. Liew, J. Alexander, W. Walker, SCID mice reconstituted with IL-4-deficient lymphocytes, but not immunocompetent lymphocytes, are resistant to cutaneous leishmaniasis, J. Immunol. 159 (1997) 5005e5013. [65] K. Mohrs, A.E. Wakil, N. Killeen, R.M. Locksley, M. Mohrs, A two-step process for cytokine production revealed by IL-4 dual-reporter mice, Immunity 23 (2005) 419e429. [66] J. Alexander, F. Brombacher, H.A. McGachy, A.N. McKenzie, W. Walker, K.C. Carter, An essential role for IL-13 in maintaining a nonhealing response following Leishmania mexicana infection, Eur. J. Immunol. 32 (2002) 2923e2933. [67] D.E. Jones, L.U. Buxbaum, P. Scott, IL-4-independent inhibition of IL12 responsiveness during Leishmania amazonensis infection, J. Immunol. 165 (2000) 364e372. [68] D.E. Jones, M.R. Ackermann, U. Wille, C.A. Hunter, P. Scott, Early enhanced Th1 response after Leishmania amazonensis infection of C57BL/6 interleukin-10-deficient mice does not lead to resolution of infection, Infect. Immun. 70 (2002) 2151e2058. [69] U.M. Padigel, J. Alexander, J.P. Farrell, The role of interleukin-10 in susceptibility of BALB/c mice to infection with Leishmania mexicana and Leishmania amazonensis, J. Immunol. 171 (2003) 3705e3710. [70] L.U. Buxbaum, P. Scott, Interleukin 10 and Fc gamma receptor-deficient mice resolve Leishmania mexicana lesions, Infect. Immun. 73 (2005) 101e108. [71] A.C. Hsu, P. Scott, Leishmania mexicana infection induces impaired lymph node expansion and Th1 cell differentiation despite normal T cell proliferation, J. Immunol. 179 (2007) 8200e8207.