Developing connections amongst key cytokines and dysregulated germinal centers in autoimmunity

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Developing connections amongst key cytokines and dysregulated germinal centers in autoimmunity Rebecca A Sweet, Sau K Lee and Carola G Vinuesa Systemic autoimmunity owing to overactivity of Tfh and dysregulated germinal centers has been described in mice and humans. Cytokines such as IL-21, IFN-g, IL-6 and IL-17 are elevated in the plasma of mouse models of lupus, arthritis, and multiple sclerosis, and in subsets of patients with autoimmune disease. Monoclonal antibodies targeting these cytokines are entering clinical trials. While these cytokines exert pleiotropic effects on immune cells and organs, it is becoming clear that each and all of them can profoundly regulate Tfh numbers and/ or function and induce or maintain the aberrant germinal center reactions that lead to pathogenic autoantibody formation. Here we review recent discoveries into the roles of IL-21, IFN-g, IL-6, and IL-17 in germinal center responses and antibody-driven autoimmunity. These new insights used in conjunction with biomarkers of an overactive Tfh pathway may help stratify patients to rationalize the use of emerging monoclonal anticytokine antibody therapies. Address Department of Pathogens and Immunity, John Curtin School of Medical Research, The Australian National University, Building 131, Garran Road, Canberra, ACT, Australia Corresponding author: Vinuesa, Carola G ([email protected])

Current Opinion in Immunology 2012, 24:658–664 This review comes from a themed issue on Autoimmunity Edited by Yanick J Crow and Edward Wakeland For a complete overview see the Issue and the Editorial Available online 1st November 2012

B cells and their antibody products are critical to the disease manifestations of lupus as demonstrated in humans and mice [2]. Initial activation of autoreactive B cells can be B cell autonomous if a TLR7 or TLR9 signal serves as co-stimulation [3] or can be triggered by aberrant activation and/or survival of self-reactive T cells [4]. Rituximab depletes mature B cells, but it is not universally effective, and patients experience side effects including greater susceptibility to infection [5]. Belimumab, the only new FDA-approved lupus treatment introduced in the clinic in over 50 years, is an anti-BAFF monoclonal antibody that reduces B cell survival. However, it has only modest effects on clinical manifestations [6]. The greatly needed development of more targeted and effective therapies hinges on improving our understanding of disease pathogenesis. There is accumulating evidence that dysregulation of germinal centers (GC) is a prominent, albeit not exclusive, pathway to lupus development [7]. GCs are specialized microenvironments that form within follicles of secondary lymphoid tissue that promote antigen-dependent affinity maturation of B cells. Within GCs, proliferating antigen-specific B cells acquire random somatic mutations in the Ig V region genes. Follicular helper T (Tfh) cells located within GCs drive positive selection of B cells that have acquired high affinity receptors [8]. Recently described T follicular regulatory (Tfr) cells limit GC B cell reactions and limit selection of non-antigenspecific and self-reactive B cells [9].

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Introduction The incidence of autoimmune diseases has been increasing over the past few decades. In the prototypic systemic autoimmune disease, Systemic Lupus Erythematosus (SLE), activation of self-reactive T and B cells triggers an inflammatory cascade that results in immune complex deposition often resulting in kidney disease that can be life-threatening [1]. Other symptoms include skin rashes, hematologic abnormalities, arthritis, and cardiovascular and neuropsychiatric complications. Broadly immunosuppressive treatments are widely available but these often fail to control the most serious disease manifestations and cause serious side effects, highlighting the need for more targeted therapies. One of the main stumbling blocks is the marked clinical heterogeneity of SLE, likely to be underpinned by more than one pathway to disease. Current Opinion in Immunology 2012, 24:658–664

Dysregulation of GCs is often caused by aberrant accumulation, overactivity and/or aberrant function of Tfh cells [7,10]. In several mouse models of lupus, SAP deficiency, which eliminates GC Tfh cells, reduces or even abolishes disease [11–13]. Furthermore, subsets of patients with lupus and other autoimmune disorders have recently been shown to bear biomarkers of an overactive Tfh/GC pathway [14,15]. In MRLlpr mice, in which B cells grow and mutate at extrafollicular sites, extrafollicular helper T (Tefh) cells, that share many attributes of Tfh cells, sustain autoantibody responses [16]. It is becoming apparent that the cytokine milieu profoundly influences Tfh and GC numbers and function. Increased expression of IFN-g, IL-17, IL-21, and IL-6 have all been shown to be associated with systemic autoimmunity [17] and have been shown to directly influence Tfh and germinal center responses (Figure 1a–f). Here we review recently uncovered links between the dysregulation www.sciencedirect.com

Developing connections amongst key cytokines and dysregulated germinal centers Sweet, Lee and Vinuesa 659

Figure 1

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(b) B cell priming & Ig switching IL-21

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Mechanisms by which IL-6, IFN-g, IL-21 and IL-6 may contribute to dysregulation of Tfh and GC responses, and autoimmunity. (a) IFN-g promotes proliferation of early activated T cells and similar to IL-21 and IL-6, can induce Bcl-6 expression. (b) IL-21 produced by Tfh cells acts at the time of T cell priming to promote B cell activation and differentiation along both the extrafollicular and GC pathway. IFN-g promotes Ig switching to the most pathogenic isotype, IgG2a in mice. (c) Excessive IL-6, IL-21 or IFN-g cause accumulation of Tfh cells that sustain numerous and enlarged GCs. IL-6 and IFN-g can both cause Bcl-6 overexpression in GC Tfh cells and/or their precursors. This may cause retention and possibly proliferation of Tfh cells. (d) IL-6 can repress the function of Tregs and thus it is possible that it also repress Tfr cells that normally limit Tfh cell and GC B cell accumulation. Tfr cells may act via repression of IFN-g in GCs. (e) IL-17 aberrantly produced by Tfh cells in autoimmune-prone mice can promote GC B cell retention in GC, which may consequently undergo more rounds of somatic mutation and become self-reactive. IL-21 acts directly on GC B cells to maximize Bcl-6 expression and thus promote their growth and survival. (f) Gut Th17 cells formed in the presence of certain strains of commensal bacteria (i.e. SFB) can promote the formation of ectopic follicles and GCs at distant sites such as the brain and joints in mouse models of multiple sclerosis and rheumatoid arthritis respectively.

of these four cytokines and GC-driven autoimmunity. A number of other cytokines and growth factors such as type 1 interferons, IL-10, IL-27 and BAFF are also important players in systemic autoimmunity, but their roles in Tfh and GC biology are either indirect or remain to be fully elucidated; they will not be discussed here owing to the focused nature of this review.

IL-21 boosts GC B cells and can induce Tfh formation Tfh cells, critical helpers of B cells and drivers of autoimmune disease via GCs [13] are major producers of IL21 [18]. Tfh and NKTfh cells produce IL-21 to enhance B cell differentiation toward both extrafollicular and GC pathways [19–22] (Figure 1b). Within GCs, IL-21 signals directly in GC B cells to maximize Bcl-6 expression and sustain the GC [23,24] (Figure 1e). IL-21 signaling, although not obligatory for Tfh differentiation, boosts www.sciencedirect.com

Tfh formation to some but not all protein antigens [23– 27] (Figure 1a). Given the prominent roles of IL-21 in Tfh and GC biology, it is not surprising that IL-21 plays a major role in autoimmunity. IL-21 production is increased in lupus mouse models [28,29] and SLE patients [30] and has been found in the brains of MS patients [31]. IL-21 blockade ameliorates disease in mouse models of lupus: IL-21 receptor deficiency or neutralization of IL-21 in BXSBYaa or MRL/lpr mice caused a reduction in IgG and kidney disease [28,32,33,34]. Furthermore, IL-21 receptor deficiency in BXSB-Yaa mice rescued the diseaseassociated increase in Tfh cells [28]. Similarly, deficiency in IL-21 receptor in MRL/lpr mice also led to a reduction in numbers of Tfh and Tefh, GC B cells, plasma cells, and plasmablasts [34]. IL-21 blockade also reduced symptoms in collagen-induced arthritis [35], a disease also recently shown to be driven by GCs and Tfh cells [36]. Current Opinion in Immunology 2012, 24:658–664

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By contrast, IL-21 deficiency had no effect on the Tfhdriven lupus of sanroque mice [13].

IL-17 promotes the retention of GC B cells and formation of ectopic GCs Of increasing interest in autoimmunity is the role of Th17 cells and their key cytokine IL-17 in its association with the GC. Neither human tonsilar Tfh cells nor murine Tfh cells induced by immunization with protein antigens produce IL-17 [37] and this may be key to preventing pathologic GC responses: Evidence is accumulating that aberrant production of IL-17 may drive pathogenic antibody responses through its ability to induce ectopic GC development and alter GC kinetics (Figure 1e and f). IL17 receptor deficiency reduced the percentage of dysregulated GCs in the spleens of BXD2 lupus prone mice [38]. By contrast, when mice were infected with an adenovirus expressing IL-17, GCs were increased. The authors attributed this effect to decreased chemotactic migration of B cells that were retained within GCs (Figure 1e). Increased retention time probably increased accumulation of harmful muatations as IL-17 delivery also increased anti-nuclear antibodies in BXD2 mice. Additionally, IL-17 producing T cells were found within the GC and B cells were found to express IL-17R [38]. Furthermore, in the autoantibody driven K/BxN model of arthritis, splenic GCs were greatly reduced following neutralization of IL-17, and GC B cell expansion was impaired if they lacked the IL-17 receptor [39]. Ectopic GCs can be found in the brain in MS [40], in the joints in RA [41], and in the kidney in SLE [42]. In work by Kuchroo and colleagues, transfer of in vitro skewed CNS-specific Th17 cells, but not Th1, Th2, or Th9 cells, induced ectopic B cell follicles in the CNS upon induction of EAE [43] (Figure 1f). Interestingly, CNS infiltrating T cells originally skewed to Th17 had surface markers similar to Tfh as they expressed Bcl6 and high levels of GL-7 and ICOS. Thus, aberrant production of IL-17 by Tfh cells may lead to dysregulated GC biology and autoantibody production.

IFN-g drives pathogenic accumulation of Tfh cells in lupus IFN-g is intimately connected to autoimmunity [44]. Tfh cells have the capacity to produce IFN-g in the context of infections [18,45,46]. However, human tonsil and mouse Tfh cells produce less IFN-g than their nonTfh effector counterparts [37,46,47]; Bcl-6, the transcription factor that drives Tfh formation, has been shown to repress IFN-g expression [37,48,49]. This suggests a physiological need to limit IFN-g production in GCs. Although IFN-g has long been associated with lupus, it was thought to act predominantly via stimulation of myeloid cells to release BAFF [50] and through the induction of Ig switching to IgG2a in mice, an isotype Current Opinion in Immunology 2012, 24:658–664

that can potently activate downstream inflammatory cascades and is thus more pathogenic [51]. More recently, excessive IFN-g signaling in T cells owing to impaired posttranscriptional repression by Roquinsan has been shown to be the cause for the pathogenic accumulation of Tfh cells and lupus in sanroque mice [52]. Deficiency in IFN-gR or IFN-g prevented aberrant Tfh and GC formation, splenomegaly, antinuclear antibody formation and nephritis. By contrast, deficiency in ICOS, T-bet and IL-21 failed to rescue the autoimmune manifestations. Furthermore, excessive IFN-gR signaling in T cells was sufficient to drive Tfh and autoantibody formation and IFN-g blockade for 3 weeks was sufficient to reduce Tfh cells and decrease autoantibodies to wild-type levels. Thus, IFN-g overproduction drives and sustains the aberrant Tfh response and lupus. Increased IFN-gR signaling promoted T cell proliferation from the naı¨ve stages and caused overexpression of Bcl-6 in Tfh cells and their precursors (Figure 1a–c). In vitro, IFN-g can induce upregulation of Bcl-6 in anti-CD3-activated T cells within 24 hours [52,53], and this is thought to act via a STAT-1 binding site in Bcl6 [53]. This work highlights the importance of repressing IFN-g signaling in Tfh cells and their precursors to prevent autoimmunity.

IL-6 induces Tfh formation and accumulation In addition to its role in promoting Th17 differentiation [54], IL-6 has pleiotropic effects on immune cells and increased IL-6 has been associated with SLE, RA, and MS in humans and mice [55,56]. While the role of IL-6 in Tfh formation has been known for several years [8], only recently, an important role for this cytokine in Tfh expansion and maintenance has been described (Figure 1a and c). IL-6 produced by radioresistant cells, is responsible for the increase in Tfh cells seen in chronic LCMV infection, and reported to be critical to control the virus [57]. IL-6 also appears to contribute to the aberrant Tfh accumulation seen in HIV infection at least partly though direct signaling to Tfh cells [58]. FDCs have been previously shown to produce IL-6 and interestingly, this IL-6 production was enhanced after IFN-g activation [59]. Thus, it is tempting to speculate that the autoimmunity associated with excessive IL-6 may be partly owing to accumulation of Tfh cells and its downstream effects on GC selection and autoantibody formation. Given that IL-6 has been shown to limit Treg suppression [60], it is possible that the IL-6-mediated increase in Tfh cells is partly an indirect effect of IL-6 on Tfr cells (Figure 1d).

Causes of cytokine dysregulation Many factors can contribute to abnormal production of one or several GC-promoting cytokines. Single nucleotide polymorphisms (SNPs) in IL21 [61,62], IL21R [63], IFNG [64] and IL6 [65] have also been associated with autoimmune disease. The SNP in IFNG increases binding affinity for NFkB [66] and the SNP in IL6 increases www.sciencedirect.com

Developing connections amongst key cytokines and dysregulated germinal centers Sweet, Lee and Vinuesa 661

IL-6 production [67]. Specific patterns of expression of IFN-g receptors 1 and 2 are also associated with disease [68]. Chronic viral, bacterial or fungal infections can lead to elevated levels of IFN-g, IL-6, or IL-17, and thus also trigger and sustain pathogenic accumulation of Tfh cells. In this context, repression of Ifng mRNA described above may be an important mechanism to prevent excessive IFN-g signaling [52]. In the cases of lupus and multiple sclerosis, enhanced TLR signaling is often directly or indirectly upstream of dysregulated cytokine production [3,69]. Autoantigenic targets in SLE center around nucleic acids. TLRs that sense self and foreign nucleic acids contribute to the activation of B cells that produce anti-nucleic acid autoantibodies [3], and of DCs and myeloid cells that produce Tfh-inducing cytokines such as IL-6 [70] and prime T cells to produce IFN-g [71]. In particular, TLR7 signaling is highly pathogenic in lupus and can synergize with IL-21 during B cell responses [72]. Interestingly, pristane, which induces experimental lupus via TLR7, enhanced TLR7 induced IL-6 [73]. Additionally, TLR7 can trigger IFN-g secretion [74]. TLR2, which has been implicated in sensing hyaluronan in lesions of MS patients [75], can induce IL-17 during bacterial infection [76] and IL-6 from microglia, the major immunological cell type in the CNS [77]. Composition of commensal flora is likely to also affect the balance of cytokine-producing T cells and thus trigger dysregulation of GCs. In the K/BxN model of arthritis, germ-free mice were protected from arthritis [39]. Strikingly, single colonization with segmented filamentous bacteria drove IL-17-producing cells in the gut, and increased antibody secretion, resulting in arthritis (Figure 1f). Similarly, in MOG-induced EAE, germ free mice were protected, and this protection was lost after single colonization with SFB and induction of IL-17 producing T cells in the CNS [78] (Figure 1f). It appears that the species of commensal flora can determine the outcome: In MOG-induced EAE, treatment of mice with Lactobacillus reduced IL-17 and induced regulatory T cells that suppressed disease via an IL-10 dependent mechanism [79]. Tregs are also known to control the cytokine milieu. For example, ablation of FoxP3 leads to increased IFN-g that rapidly triggers autoimmune diabetes [80]. This mechanism may also apply to Tfr cells, the specialized regulatory cells that are located within the GC and are thus poised to control dysregulated Tfh [9] (Figure 1d). IL-10 is produced abundantly by Tfr cells and is known to indirectly suppress IFN-g [81]. Additionally, Qa-1 restricted CD8+ regulatory cells, which specifically suppress Tfh cells [82], do not suppress Ab responses efficiently when they originate from B6-Yaa lupus prone mice [83], which have increased GC and Tfh. The mechanism of suppression of these CD8+ regulatory cells www.sciencedirect.com

is dependent on IL-15 rather than IL-10. It is possible that IL-15 competes with IL-21 in use of the common gamma chain for receptor signaling.

Conclusions Although GC-driven autoimmunity is complex and probably results from the intersection of many dysregulated processes, cytokine imbalance is a common theme, particularly centered around IL-21, IL-17, IL-6 and IFN-g. Given the elevated levels of these cytokines in autoimmune disease [17] and their role in promoting GCs that give rise to autoantibodies, it is likely that monoclonal antibodies that target these molecules, currently entering clinical trials, will be effective in selected groups of patients. Stratification of patients according to plasma levels of individual cytokines, together with signs of Tfh overactivity in the blood or ectopic GC formation in the synovium, may offer the possibility of rationalizing the use of anti-cytokine mAb therapy in autoimmunity. Also, monitoring the frequency of effector GC Tfh cells using improved biomarkers in the blood promises to provide novel readouts for the assessment of treatment efficacy.

Acknowledgements R.A.S. is a Sir Keith Murdoch Fellow of the American Australian Association. C.G.V. is supported by an Elizabeth Blackburn NHMRC Fellowship. The Vinuesa Lab is supported by NHMRC program and project grants.

References and recommended reading Papers of particular interest, published within the period of review, have been highlighted as:  of special interest  of outstanding interest 1.

Kotzin BL: Systemic lupus erythematosus. Cell 1996, 85:303-306.

2.

Townsend MJ, Monroe JG, Chan AC: B-cell targeted therapies in human autoimmune diseases: an updated perspective. Immunol Rev 2010, 237:264-283.

3.

Shlomchik MJ: Activating systemic autoimmunity: B’s, T’s, and tolls. Curr Opin Immunol 2009, 21:626-633.

4.

Ohashi PS: T-cell signalling and autoimmunity: molecular mechanisms of disease. Nat Rev Immunol 2002, 2:427-438.

5.

Ramos-Casals M, Sanz I, Bosch X, Stone JH, Khamashta MA: Bcell-depleting therapy in systemic lupus erythematosus. Am J Med 2012, 125:327-336.

6.

Manzi S, Sanchez-Guerrero J, Merrill JT, Furie R, Gladman D, Navarra SV, Ginzler EM, D’Cruz DP, Doria A, Cooper S et al.: Effects of belimumab, a B lymphocyte stimulator-specific inhibitor, on disease activity across multiple organ domains in patients with systemic lupus erythematosus: combined results from two phase III trials. Ann Rheum Dis 2012, 71:1833-1838.

7.

Vinuesa CG, Sanz I, Cook MC: Dysregulation of germinal centres in autoimmune disease. Nat Rev Immunol 2009, 9:845-857.

8.

Crotty S: Follicular helper CD4 T cells (TFH). Annu Rev Immunol 2011, 29:621-663.

9.

Linterman MA, Liston A, Vinuesa CG: T-follicular helper cell differentiation and the co-option of this pathway by nonhelper cells. Immunol Rev 2012, 247:143-159. Current Opinion in Immunology 2012, 24:658–664

662 Autoimmunity

10. Craft JE: Follicular helper T cells in immunity and systemic autoimmunity. Nat Rev Rheumatol 2012, 8:337-347. 11. Hron JD, Caplan L, Gerth AJ, Schwartzberg PL, Peng SL: SH2D1A regulates T-dependent humoral autoimmunity. J Exp Med 2004, 200:261-266. 12. Komori H, Furukawa H, Mori S, Ito MR, Terada M, Zhang MC, Ishii N, Sakuma N, Nose M, Ono M: A signal adaptor SLAMassociated protein regulates spontaneous autoimmunity and Fas-dependent lymphoproliferation in MRL-Faslpr lupus mice. J Immunol 2006, 176:395-400. 13. Linterman MA, Rigby RJ, Wong RK, Yu D, Brink R, Cannons JL, Schwartzberg PL, Cook MC, Walters GD, Vinuesa CG: Follicular helper T cells are required for systemic autoimmunity. J Exp Med 2009, 206:561-576. 14. Morita R, Schmitt N, Bentebibel SE, Ranganathan R, Bourdery L, Zurawski G, Foucat E, Dullaers M, Oh S, Sabzghabaei N et al.: Human blood CXCR5(+)CD4(+) T cells are counterparts of T follicular cells and contain specific subsets that differentially support antibody secretion. Immunity 2011, 34:108-121. 15. Simpson N, Gatenby PA, Wilson A, Malik S, Fulcher DA, Tangye SG, Manku H, Vyse TJ, Roncador G, Huttley GA et al.: Expansion of circulating T cells resembling follicular helper T cells is a fixed phenotype that identifies a subset of severe systemic lupus erythematosus. Arthritis Rheum 2010, 62:234-244. 16. Odegard JM, Marks BR, DiPlacido LD, Poholek AC, Kono DH, Dong C, Flavell RA, Craft J: ICOS-dependent extrafollicular helper T cells elicit IgG production via IL-21 in systemic autoimmunity. J Exp Med 2008, 205:2873-2886. 17. Moudgil KD, Choubey D: Cytokines in autoimmunity: role in induction, regulation, and treatment. J Interferon Cytokine Res 2011, 31:695-703. 18. Lu¨thje K, Kallies A, Shimohakamada Y, Belz GT, Light A,  Tarlinton DM, Nutt SL: The development and fate of follicular helper T cells defined by an IL-21 reporter mouse. Nat Immunol 2012, 13:491-498. This paper showed definitively the plasticity of Tfh cells including their ability to secrete IFN-g. 19. Lee SK, Rigby RJ, Zotos D, Tsai LM, Kawamoto S, Marshall JL, Ramiscal RR, Chan TD, Gatto D, Brink R et al.: B cell priming for extrafollicular antibody responses requires Bcl-6 expression by T cells. J Exp Med 2011, 208:1377-1388. 20. Chang PP, Barral P, Fitch J, Pratama A, Ma CS, Kallies A, Hogan JJ, Cerundolo V, Tangye SG, Bittman R et al.: Identification of Bcl-6-dependent follicular helper NKT cells that provide cognate help for B cell responses. Nat Immunol 2012, 13:35-43. 21. King IL, Fortier A, Tighe M, Dibble J, Watts GF, Veerapen N, Haberman AM, Besra GS, Mohrs M, Brenner MB et al.: Invariant natural killer T cells direct B cell responses to cognate lipid antigen in an IL-21-dependent manner. Nat Immunol 2012, 13:44-50. 22. Ozaki K, Spolski R, Ettinger R, Kim HP, Wang G, Qi CF, Hwu P, Shaffer DJ, Akilesh S, Roopenian DC et al.: Regulation of B cell differentiation and plasma cell generation by IL-21, a novel inducer of Blimp-1 and Bcl-6. J Immunol 2004, 173:5361-5371. 23. Linterman MA, Beaton L, Yu D, Ramiscal RR, Srivastava M, Hogan JJ, Verma NK, Smyth MJ, Rigby RJ, Vinuesa CG: IL-21 acts directly on B cells to regulate Bcl-6 expression and germinal center responses. J Exp Med 2010, 207:353-363.

26. Vogelzang A, McGuire HM, Yu D, Sprent J, Mackay CR, King C: A fundamental role for interleukin-21 in the generation of T follicular helper cells. Immunity 2008, 29:127-137. 27. Eto D, Lao C, DiToro D, Barnett B, Escobar TC, Kageyama R, Yusuf I, Crotty S: IL-21 and IL-6 are critical for different aspects of B cell immunity and redundantly induce optimal follicular helper CD4 T cell (Tfh) differentiation. PLoS ONE 2011, 6:e17739. 28. Bubier JA, Sproule TJ, Foreman O, Spolski R, Shaffer DJ, Morse HC 3rd, Leonard WJ, Roopenian DC: A critical role for IL-21 receptor signaling in the pathogenesis of systemic lupus erythematosus in BXSB-Yaa mice. Proc Natl Acad Sci USA 2009, 106:1518-1523. 29. Vinuesa CG, Cook MC, Angelucci C, Athanasopoulos V, Rui L, Hill KM, Yu D, Domaschenz H, Whittle B, Lambe T et al.: A RINGtype ubiquitin ligase family member required to repress follicular helper T cells and autoimmunity. Nature 2005, 435:452-458. 30. Dolff S, Abdulahad WH, Westra J, Doornbos-van der Meer B, Limburg PC, Kallenberg CG, Bijl M: Increase in IL-21 producing T-cells in patients with systemic lupus erythematosus. Arthritis Res Ther 2011, 13:R157. 31. Tzartos JS, Craner MJ, Friese MA, Jakobsen KB, Newcombe J, Esiri MM, Fugger L: IL-21 and IL-21 receptor expression in lymphocytes and neurons in multiple sclerosis brain. Am J Pathol 2011, 178:794-802. 32. Bubier JA, Bennett SM, Sproule TJ, Lyons BL, Olland S, Young DA, Roopenian DC: Treatment of BXSB-Yaa mice with IL-21R-Fc fusion protein minimally attenuates systemic lupus erythematosus. Ann NY Acad Sci 2007, 1110:590-601. 33. Herber D, Brown TP, Liang S, Young DA, Collins M, DunussiJoannopoulos K: IL-21 has a pathogenic role in a lupus-prone mouse model and its blockade with IL-21R.Fc reduces disease progression. J Immunol 2007, 178:3822-3830. 34. Rankin AL, Guay H, Herber D, Bertino SA, Duzanski TA, Carrier Y,  Keegan S, Senices M, Stedman N, Ryan M et al.: IL-21 receptor is required for the systemic accumulation of activated B and T lymphocytes in MRL/MpJ-Fas(lpr/lpr)/J mice. J Immunol 2012, 188:1656-1667. This work provides a comprehensive characterization of disease amelioration in the absence of IL-21 in MRL/lpr mice. 35. Young DA, Hegen M, Ma HL, Whitters MJ, Albert LM, Lowe L, Senices M, Wu PW, Sibley B, Leathurby Y et al.: Blockade of the interleukin-21/interleukin-21 receptor pathway ameliorates disease in animal models of rheumatoid arthritis. Arthritis Rheum 2007, 56:1152-1163. 36. Hu YL, Metz DP, Chung J, Siu G, Zhang M: B7RP-1 blockade ameliorates autoimmunity through regulation of follicular helper T cells. J Immunol 2009, 182:1421-1428. 37. Yu D, Rao S, Tsai LM, Lee SK, He Y, Sutcliffe EL, Srivastava M, Linterman M, Zheng L, Simpson N et al.: The transcriptional repressor Bcl-6 directs T follicular helper cell lineage commitment. Immunity 2009, 31:457-468. 38. Hsu HC, Yang P, Wang J, Wu Q, Myers R, Chen J, Yi J, Guentert T,  Tousson A, Stanus AL et al.: Interleukin 17-producing T helper cells and interleukin 17 orchestrate autoreactive germinal center development in autoimmune BXD2 mice. Nat Immunol 2008, 9:166-175. This study brought to the forefront the importance of repressing IL-17 in germinal centers to prevent lupus.

24. Zotos D, Coquet JM, Zhang Y, Light A, D’Costa K, Kallies A, Corcoran LM, Godfrey DI, Toellner KM, Smyth MJ et al.: IL-21 regulates germinal center B cell differentiation and proliferation through a B cell-intrinsic mechanism. J Exp Med 2010, 207:365-378.

39. Wu HJ, Ivanov II, Darce J, Hattori K, Shima T, Umesaki Y,  Littman DR, Benoist C, Mathis D: Gut-residing segmented filamentous bacteria drive autoimmune arthritis via T helper 17 cells. Immunity 2010, 32:815-827. This paper was the first to show that single colonization by SFB could induce autoantibody-driven autoimmunity at a distal site via the induction of pathogenic Th17 cells.

25. Nurieva RI, Chung Y, Hwang D, Yang XO, Kang HS, Ma L, Wang YH, Watowich SS, Jetten AM, Tian Q et al.: Generation of T follicular helper cells is mediated by interleukin-21 but independent of T helper 1, 2, or 17 cell lineages. Immunity 2008, 29:138-149.

40. Magliozzi R, Howell O, Vora A, Serafini B, Nicholas R, Puopolo M, Reynolds R, Aloisi F: Meningeal B-cell follicles in secondary progressive multiple sclerosis associate with early onset of disease and severe cortical pathology. Brain 2007, 130:1089-1104.

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Developing connections amongst key cytokines and dysregulated germinal centers Sweet, Lee and Vinuesa 663

41. Humby F, Bombardieri M, Manzo A, Kelly S, Blades MC, Kirkham B, Spencer J, Pitzalis C: Ectopic lymphoid structures support ongoing production of class-switched autoantibodies in rheumatoid synovium. PLoS Med 2009, 6:e1. 42. Chang A, Henderson SG, Brandt D, Liu N, Guttikonda R, Hsieh C, Kaverina N, Utset TO, Meehan SM, Quigg RJ et al.: In situ B cellmediated immune responses and tubulointerstitial inflammation in human lupus nephritis. J Immunol 2011, 186:1849-1860. 43. Peters A, Pitcher LA, Sullivan JM, Mitsdoerffer M, Acton SE,  Franz B, Wucherpfennig K, Turley S, Carroll MC, Sobel RA et al.: Th17 cells induce ectopic lymphoid follicles in central nervous system tissue inflammation. Immunity 2011, 35:986-996. This study was the first to show that ectopic follicles in the CNS could be induced by Th17 cells with characteristics of Tfh cells. 44. Billiau A: Interferon-gamma in autoimmunity. Cytokine Growth Factor Rev 1996, 7:25-34. 45. Reinhardt RL, Liang HE, Locksley RM: Cytokine-secreting follicular T cells shape the antibody repertoire. Nat Immunol 2009, 10:385-393. 46. Yusuf I, Kageyama R, Monticelli L, Johnston RJ, Ditoro D, Hansen K, Barnett B, Crotty S: Germinal center T follicular helper cell IL-4 production is dependent on signaling lymphocytic activation molecule receptor (CD150). J Immunol 2010, 185:190-202. 47. Ma CS, Suryani S, Avery DT, Chan A, Nanan R, Santner-Nanan B, Deenick EK, Tangye SG: Early commitment of naive human CD4(+) T cells to the T follicular helper (T(FH)) cell lineage is induced by IL-12. Immunol Cell Biol 2009, 87:590-600. 48. Nurieva RI, Chung Y, Martinez GJ, Yang XO, Tanaka S, Matskevitch TD, Wang YH, Dong C: Bcl6 mediates the development of T follicular helper cells. Science 2009, 325:1001-1005. 49. Oestreich KJ, Huang AC, Weinmann AS: The lineage-defining factors T-bet and Bcl-6 collaborate to regulate Th1 gene expression patterns. J Exp Med 2011, 208:1001-1013. 50. Harigai M, Kawamoto M, Hara M, Kubota T, Kamatani N, Miyasaka N: Excessive production of IFN-gamma in patients with systemic lupus erythematosus and its contribution to induction of B lymphocyte stimulator/B cell-activating factor/ TNF ligand superfamily-13B. J Immunol 2008, 181:2211-2219. 51. Baudino L, Azeredo da Silveira S, Nakata M, Izui S: Molecular and cellular basis for pathogenicity of autoantibodies: lessons from murine monoclonal autoantibodies. Springer Semin Immunopathol 2006, 28:175-184. 52. Lee SK, Silva DG, Martin JL, Pratama A, Hu X, Chang PP. Walters  G, Vinuesa CG: Interferon-gamma excess leads to pathogenic accumulation of follicular helper T cells and germinal centers. Immunity 2012, in press. This paper revealed the importance of limiting IFN-g production to prevent Tfh cell accumulation and autoimmunity. 53. Zhou G, Ono SJ: Induction of BCL-6 gene expression by interferon-gamma and identification of an IRE in exon I. Exp Mol Pathol 2005, 78:25-35. 54. Kimura A, Kishimoto T: IL-6: regulator of Treg/Th17 balance. Eur J Immunol 2010, 40:1830-1835. 55. Ohl K, Tenbrock K: Inflammatory cytokines in systemic lupus erythematosus. J Biomed Biotechnol 2011, 2011:432595. 56. Ishihara K, Hirano T: IL-6 in autoimmune disease and chronic inflammatory proliferative disease. Cytokine Growth Factor Rev 2002, 13:357-368. 57. Harker JA, Lewis GM, Mack L, Zuniga EI: Late interleukin-6 escalates T follicular helper cell responses and controls a chronic viral infection. Science 2011, 334:825-829. 58. Petrovas C, Yamamoto T, Gerner MY, Boswell KL, Wloka K, Smith EC, Ambrozak DR, Sandler NG, Timmer KJ, Sun X et al.: CD4 T follicular helper cell dynamics during SIV infection. J Clin Invest 2012, 122(9):3281-3294 http://dx.doi.org/10.1172/ JCI63039 Epub 2012 Aug 27. www.sciencedirect.com

59. Clark EA, Grabstein KH, Shu GL: Cultured human follicular dendritic cells. Growth characteristics and interactions with B lymphocytes. J Immunol 1992, 148:3327-3335. 60. Pasare C, Medzhitov R: Toll pathway-dependent blockade of CD4+CD25+ T cell-mediated suppression by dendritic cells. Science 2003, 299:1033-1036. 61. Ding L, Wang S, Chen GM, Leng RX, Pan HF, Ye DQ: A single nucleotide polymorphism of IL-21 gene is associated with systemic lupus erythematosus in a Chinese population. Inflammation 2012. [Epub ahead of print]. 62. Hughes T, Kim-Howard X, Kelly JA, Kaufman KM, Langefeld CD, Ziegler J, Sanchez E, Kimberly RP, Edberg JC, RamseyGoldman R et al.: Fine-mapping and transethnic genotyping establish IL2/IL21 genetic association with lupus and localize this genetic effect to IL21. Arthritis Rheum 2011, 63:1689-1697. 63. Webb R, Merrill JT, Kelly JA, Sestak A, Kaufman KM, Langefeld CD, Ziegler J, Kimberly RP, Edberg JC, RamseyGoldman R et al.: A polymorphism within IL21R confers risk for systemic lupus erythematosus. Arthritis Rheum 2009, 60:24022407. 64. Kim K, Park SY, Kim T, Kang YM, Shim SC, Suh CH, Park YB, Kim CS, Kang C, Bae SC: Replicated association of a regulatory polymorphism in the interferon gamma gene with lupus susceptibility. Ann Rheum Dis 2011, 70:1878-1879. 65. Lee YH, Lee HS, Choi SJ, Ji JD, Song GG: The association between interleukin-6 polymorphisms and systemic lupus erythematosus: a meta-analysis. Lupus 2012, 21:60-67. 66. Pravica V, Perrey C, Stevens A, Lee JH, Hutchinson IV: A single nucleotide polymorphism in the first intron of the human IFNgamma gene: absolute correlation with a polymorphic CA microsatellite marker of high IFN-gamma production. Hum Immunol 2000, 61:863-866. 67. Fishman D, Faulds G, Jeffery R, Mohamed-Ali V, Yudkin JS, Humphries S, Woo P: The effect of novel polymorphisms in the interleukin-6 (IL-6) gene on IL-6 transcription and plasma IL-6 levels, and an association with systemic-onset juvenile chronic arthritis. J Clin Invest 1998, 102:1369-1376. 68. Nakashima H, Inoue H, Akahoshi M, Tanaka Y, Yamaoka K, Ogami E, Nagano S, Arinobu Y, Niiro H, Otsuka T et al.: The combination of polymorphisms within interferon-gamma receptor 1 and receptor 2 associated with the risk of systemic lupus erythematosus. FEBS Lett 1999, 453:187-190. 69. Gambuzza M, Licata N, Palella E, Celi D, Foti Cuzzola V, Italiano D, Marino S, Bramanti P: Targeting Toll-like receptors: emerging therapeutics for multiple sclerosis management. J Neuroimmunol 2011, 239:1-12. 70. Yasuda K, Richez C, Maciaszek JW, Agrawal N, Akira S, MarshakRothstein A, Rifkin IR: Murine dendritic cell type I IFN production induced by human IgG-RNA immune complexes is IFN regulatory factor (IRF)5 and IRF7 dependent and is required for IL-6 production. J Immunol 2007, 178:6876-6885. 71. Teichmann LL, Ols ML, Kashgarian M, Reizis B, Kaplan DH, Shlomchik MJ: Dendritic cells in lupus are not required for activation of T and B cells but promote their expansion, resulting in tissue damage. Immunity 2010, 33:967-978. 72. Bessa J, Kopf M, Bachmann MF: Cutting edge: IL-21 and TLR signaling regulate germinal center responses in a B cellintrinsic manner. J Immunol 2010, 184:4615-4619. 73. Lee PY, Kumagai Y, Li Y, Takeuchi O, Yoshida H, Weinstein J, Kellner ES, Nacionales D, Barker T, Kelly-Scumpia K et al.: TLR7dependent and FcgammaR-independent production of type I interferon in experimental mouse lupus. J Exp Med 2008, 205:2995-3006. 74. Lombardi V, Van Overtvelt L, Horiot S, Moingeon P: Human dendritic cells stimulated via TLR7 and/or TLR8 induce the sequential production of Il-10, IFN-gamma, and IL-17A by naive CD4+ T cells. J Immunol 2009, 182:3372-3379. 75. Sloane JA, Batt C, Ma Y, Harris ZM, Trapp B, Vartanian T: Hyaluronan blocks oligodendrocyte progenitor maturation Current Opinion in Immunology 2012, 24:658–664

664 Autoimmunity

and remyelination through TLR2. Proc Natl Acad Sci USA 2010, 107:11555-11560. 76. Teixeira-Coelho M, Cruz A, Carmona J, Sousa C, RamosPereira D, Saraiva AL, Veldhoen M, Pedrosa J, Castro AG, Saraiva M: TLR2 deficiency by compromising p19 (IL-23) expression limits Th 17 cell responses to Mycobacterium tuberculosis. Int Immunol 2011, 23:89-96. 77. Lin HY, Tang CH, Chen JH, Chuang JY, Huang SM, Tan TW, Lai CH, Lu DY: Peptidoglycan induces interleukin-6 expression through the TLR2 receptor, JNK, c-Jun, and AP-1 pathways in microglia. J Cell Physiol 2011, 226:1573-1582. 78. Lee YK, Menezes JS, Umesaki Y, Mazmanian SK: Proinflammatory T-cell responses to gut microbiota promote experimental autoimmune encephalomyelitis. Proc Natl Acad Sci USA 2011, 108(Suppl 1):4615-4622. 79. Lavasani S, Dzhambazov B, Nouri M, Fak F, Buske S, Molin G, Thorlacius H, Alenfall J, Jeppsson B, Westrom B: A novel probiotic mixture exerts a therapeutic effect on experimental

Current Opinion in Immunology 2012, 24:658–664

autoimmune encephalomyelitis mediated by IL-10 producing regulatory T cells. PLoS ONE 2010, 5:e9009. 80. Feuerer M, Shen Y, Littman DR, Benoist C, Mathis D: How punctual ablation of regulatory T cells unleashes an autoimmune lesion within the pancreatic islets. Immunity 2009, 31:654-664. 81. D’Andrea A, Aste-Amezaga M, Valiante NM, Ma X, Kubin M, Trinchieri G: Interleukin 10 (IL-10) inhibits human lymphocyte interferon gamma-production by suppressing natural killer cell stimulatory factor/IL-12 synthesis in accessory cells. J Exp Med 1993, 178:1041-1048. 82. Kim HJ, Verbinnen B, Tang X, Lu L, Cantor H: Inhibition of follicular T-helper cells by CD8(+) regulatory T cells is essential for self tolerance. Nature 2010, 467:328-332. 83. Kim HJ, Wang X, Radfar S, Sproule TJ, Roopenian DC, Cantor H: CD8+ T regulatory cells express the Ly49 Class I MHC receptor and are defective in autoimmune prone B6-Yaa mice. Proc Natl Acad Sci USA 2011, 108:2010-2015.

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