HLA-DR3 restricted environmental epitopes from the bacterium Clostridium tetani have T cell cross-reactivity to the SLE-related autoantigen SmD.

HLA-DR3 (DR3) is one of the dominant HLA-DR alleles associated with systemic lupus erythematosus (SLE) susceptibility. Our previous studies showed multiple intramolecular DR3 restricted T cell epitopes in the Smith D (SmD) protein, from which we generated a non-homologous, bacterial epitope mimics library. From this library we identified ABC247-261 Mimic as one new DR3 restricted bacterial T cell epitope from the ABC transporter ATP-binding protein in Clostridium tetani. It activated and induced autoreactive SmD66-80-specific T cells and induced autoantibodies to lupus-related autoantigens in vivo. Compared to healthy donors, SLE patients have a greater percentage of cross-reactive T cells to ABC247-261 Mimic and SmD66-80. In addition, we analyzed the ability of single DR3 restricted Tetanus toxoid (TT) T cell epitopes to induce autoimmune T cells. We found that the immunodominant TT epitope TT826-845 stimulated SmD66-80 reactive T cells but failed to induce persistent anti-SmD autoantibodies compared to the ABC247-261 Mimic. Thus, exposure to the ABC247-261 Mimic epitope may contribute to autoimmunity in susceptible DR3 individuals.

Tfh cells with NLRP3 inflammasome activation are essential for high-affinity antibody generation, germinal centre formation and autoimmunity.

OBJECTIVE: NLRP3 inflammasome regulates T cell responses. This study examined the roles of NLRP3 inflammasome activation in the regulation of T follicular helper (Tfh) cells during humoral response to T dependent antigens and in systemic lupus erythematosus (SLE). METHODS: NLRP3 inflammasome activation of Tfh cells was studied in B6, MRL/lpr and NZM2328 mice and in SLE patients and healthy controls using a fluorescence-labelled caspase-1 inhibitor probe. MCC950, a selective inhibitor of NLRP3, was used to investigate the relation between NLRP3 inflammasome activation and germinal centre (GC) reaction, Ab responses to immunisation, and autoantibody production. RESULTS: NLRP3 inflammasome activation in Tfh cells after immunisation was identified in B6 mice. MCC950 inhibited humoral responses to sheep red blood cell and NP-CGG with reduction of the GC reaction. B6 mice with lymphoid cell-specific deletion of NLRP3 or Casp1 mounted suboptimal humoral responses with impaired GC formation and defective affinity maturation. In MRL/lpr and NZM2328 mice, inhibition of NLRP3 activation suppressed NLRP3 activated Tfh cell expansion as well as attenuated lupus-like phenotypes. Tfh cells with activated NLRP3 inflammasome exhibited increased expression of molecules for Tfh cell function and differentiation, and had greater ability to activate B cells. In SLE patients, disease activity was positively correlated with an increase in the activated NLRP3+ Tfh population and this population was markedly reduced in response to therapy. CONCLUSIONS: The activation of NLRP3 inflammasome in Tfh cells is an integral part of responses to immunisation. The activated NLRP3+ Tfh population is essential for optimal humoral responses, GC formation and autoimmunity.

Autoimmune experimental orchitis and chronic glomerulonephritis with end stage renal disease are controlled by Cgnz1 for susceptibility to end organ damage.

Cgnz1 on chromosome 1 mapped into a 1.34 Mb region of chromosome 1 in NZM2328 confers the progression of immune complex (IC)-mediated glomerulonephritis (GN) from acute GN (aGN) to chronic GN (cGN) with severe proteinuria and end stage renal disease in female mice. This genetic locus mediates podocyte susceptibility to IC-mediated damage. Taking advantage of the published observation that Cgnz1 is derived from NZW and that NZW is susceptible to orchitis, epididymitis and vasitis while C57L/J is resistant to these diseases, the possibility that this genetic region also confers germ cells susceptible to damage with aspermatogenesis and sterility in an active experimental autoimmune orchitis (EAO) model was investigated. Male mice from multiple intrachromosome (chromosome 1) recombinant strains were subjected to immunization with a sperm homogenate in CFA with concomitant administration of Bordetella pertussis toxin. There was concordance of the progression from aGN to cGN, severe proteinuria and end stage renal disease with susceptibility of EAO in NZM2328 and its congenic strains with various chromosome 1 genetic intervals introgressed from C57L/J to NZM2328. Both resistant and susceptible strains made comparable anti-testis and anti-sperm Abs. Thus the genetic interval that determines susceptibility to EAO is identical to that of Cgnz1 and mapped to the 1.34 Mb region in chromosone 1. This region likely confers germ cells in the male gonad susceptible to damage by immunologically mediated inflammation. This region has been tentatively renamed Cgnz1/Eaoz1. These observations further emphasize the importance of end organ susceptibility to damage in the pathogenesis of both systemic and organ specific autoimmune diseases.

Pathogenesis of lupus nephritis: RIP3 dependent necroptosis and NLRP3 inflammasome activation.

RIP3 activation leads to activation of necroptosis and the NLRP3 inflammasome pathways. The activation of RIP3 in lupus nephritis (LN) has not been investigated. In this study, RIP3 and necroptosis pathway activations were demonstrated in podocytes in renal biopsies from patients with class IV LN and in the diseased kidneys from lupus-prone NZM2328 and MRL/lpr mice. RIP3 activation was accompanied with the activation of MLKL, the effector molecule of the necroptosis pathway, and activation of caspase-1, the effector of the NLRP3 inflammasome pathway. Podocyte activation of RIP3 was detected readily with the development of LN in NZM2328 mice, suggesting this activation may play a significant role in the pathogenesis of LN. GSK872, a RIP3 specific inhibitor, inhibited the development of LN in MRL/lpr mice with down-regulation of RIP3 activation in podocytes, decreased the splenic sizes and weights and anti-dsDNA antibody titers. IgG from pooled sera of diseased NZM2328 mice succumbing to LN induced both the necroptosis pathway and NLRP3 inflammasome activation in a podocyte cell line and this activation was specifically blocked by GSK872. These results indicate that the necroptosis pathway and the RIP3 dependent NLRP3 inflammasome pathway are activated in podocytes during LN. Inhibition of RIP3 kinase may be a novel therapeutic approach to treat LN and systemic lupus erythematosus (SLE).

Innate lymphoid cell disturbance with increase in ILC1 in systemic lupus erythematosus.

The innate lymphoid cell (ILC) is a group of effector cells with diverse important cellular functions in both health and disease states. In comparison with healthy controls, there were increases in circulating ILC in SLE patients. The proportion of ILC1 significantly increased with significant decreases of ILC2 in SLE patients and ILC3 in SLE patients with moderate to severe activity. IL-12, IL-18, IL-25, IL-33, IL-23, IL-1ß and IFN-γ were significantly increased in SLE patients. Moreover, IL-12, IL-18 and IL-1ß but not IFN-γ correlated significantly with SLEDAI. Successful treatments rapidly reduced them and with certain normalization of the ILC subsets. In addition to increases in ILC1 numbers, ~ 80% of the ILC1 in SLE patients were positive for synthesis of IFN-γ. Plasma from SLE patients were shown to be potent in inducing ILC1. Thus, increased circulating ILC1 might contribute to the pathogenesis of SLE through mounting type 1 immune response.

Nature of T cell epitopes in lupus antigens and HLA-DR determines autoantibody initiation and diversification.

OBJECTIVES: The generation of systemic lupus erythematosus (SLE)-related autoantibodies have been shown to be T cell dependent and antigen driven with HLA-DR restriction. In this study, the initiating antigen(s) and the mechanism of autoantibody diversification were investigated. METHODS: T cell epitopes (T-epitopes) of SmD1 (SmD) were mapped by T-T hybridomas generated from DR3+AE0 mice immunised with SmD and with SmD overlapping peptides. TCRs from the reactive hybridomas were sequenced. The core epitopes were determined. Bacterial mimics were identified by bioinformatics. Sera from DR3+AE0 mice immunised with SmD peptides and their mimics were analysed for their reactivity by ELISA and immunohistochemistry. Samples of blood donors were analysed for HLA-DR and autoantibody specificities. RESULTS: Multiple HLA-DR3 restricted T-epitopes within SmD were identified. Many T-T hybridomas reacted with more than one epitope. Some of them were cross-reactive with other snRNP peptides and with proteins in the Ro60/La/Ro52 complex. The reactive hybridomas used unique TCRs. Multiple T-epitope mimics were identified in commensal and environmental bacteria. Certain bacterial mimics shared both T and B cell epitopes with the related SmD peptide. Bacterial mimics induced autoantibodies to lupus-related antigens and to different tissues. HLA-DR3+ blood donors made significantly more SLE-related autoantibodies. CONCLUSIONS: The unique antigenic structures of the lupus-related autoantigens provide the basis for being targeted and for T and B cell epitope spreading and autoantibody diversification with unique patterns. SLE-related autoantibodies are likely generated from responses to commensal and/or environmental microbes due to incomplete negative selection for autoreactive T cells. The production of SLE-related antibodies is inevitable in normal individuals. The findings in this investigation have significant implications in autoimmunity in general.

The ratio of circulating follicular T helper cell to follicular T regulatory cell is correlated with disease activity in systemic lupus erythematosus.

Follicular T regulatory (Tfr) cells inhibit follicular T helper (Tfh) cells mediated B cell responses. Tfh cells are involved in the pathogenesis of systemic lupus erythematosus (SLE). However, the role of Tfr cells in SLE remains unclear. The frequency of circulating Tfr and Tfh cells were examined in SLE patients and healthy controls. The frequency of circulating Tfr cell decreased and Tfh/Tfr ratio increased in SLE patients. Serum anti-dsDNA antibody level positively correlated with frequency of Tfh cells and Tfh/Tfr ratios but negatively correlated with the frequency of Tfr cells. Moreover, the frequency of Tfr and Tfh/Tfr ratio but not that of Tfh was correlated with diseases activity. In addition, increase in Tfr cell numbers and decrease in the Tfh/Tfr ratios were observed with successful treatments. Thus, Tfr cells should be considered as a biomarker for SLE and their role in the pathogenesis of SLE warrants further investigation.

Podocyte Activation of NLRP3 Inflammasomes Contributes to the Development of Proteinuria in Lupus Nephritis.

OBJECTIVE: Development of proteinuria in lupus nephritis (LN) is associated with podocyte dysfunction. The NLRP3 inflammasome has been implicated in the pathogenesis of LN. The purpose of this study was to investigate whether NLRP3 inflammasome activation is involved in the development of podocyte injury in LN. METHODS: A fluorescence-labeled caspase 1 inhibitor probe was used to detect the activation of NLRP3 inflammasomes in podocytes derived from lupus-prone NZM2328 mice and from renal biopsy tissues obtained from patients with LN. MCC950, a selective inhibitor of NLRP3, was used to treat NZM2328 mice. Proteinuria, podocyte ultrastructure, and renal pathology were evaluated. In vitro, sera from diseased NZM2328 mice were used to stimulate a podocyte cell line, and the cells were analyzed by flow cytometry. RESULTS: NLRP3 inflammasomes were activated in podocytes from lupus-prone mice and from patients with LN. Inhibition of NLRP3 with MCC950 ameliorated proteinuria, renal histologic lesions, and podocyte foot process effacement in lupus-prone mice. In vitro, sera from diseased NZM2328 mice activated NLRP3 inflammasomes in the podocyte cell line through the production of reactive oxygen species. CONCLUSION: NLRP3 inflammasomes were activated in podocytes from lupus-prone mice and from LN patients. Activation of NLRP3 is involved in the pathogenesis of podocyte injuries and the development of proteinuria in LN.

Anti-dsDNA Antibodies are one of the many autoantibodies in systemic lupus erythematosus.

Anti-dsDNA antibodies are the most studied antibodies of the lupus-related autoantibodies. The dogma is that these are the most important autoantibodies in systemic lupus erythematosus. In this review, evidence is presented to show that these antibodies (as measured by modern clinical laboratories) are not the most important autoantibodies in the diagnosis of systemic lupus erythematosus, and are of limited value in clinical correlation and in predicting disease flares. In addition, they are not likely to be the initiating autoantibodies in lupus nephritis. Thus, several pervasively held beliefs on anti-dsDNA antibodies are not valid. We suggest that anti-dsDNA antibodies should be considered as just one of the many autoantibodies associated with systemic lupus erythematosus.

Myeloid-derived suppressor cells contribute to bone erosion in collagen-induced arthritis by differentiating to osteoclasts.

Bone erosion is a sign of severe rheumatoid arthritis and osteoclasts play a major role in the bone resorption. Recently, myeloid-derived suppressor cells (MDSC) has been reported to be increased in collagen-induced arthritis (CIA). The number of circulating MDSCs is shown to correlate with rheumatoid arthritis. These findings suggest that MDSCs are precursor cells involved in bone erosion. In this study, MDSCs isolated from mice with CIA stimulated with M-CSF and RANKL in vitro expressed osteoclast markers and acquired osteoclast bone resorption function. MDSCs sorted from CIA mice were transferred into the tibia of normal DBA/1J mice and bones were subjected to histological and Micro CT analyses. The transferred CIA-MDSCs were shown to differentiate into TRAP(+) osteoclasts that were capable of bone resorption in vivo. MDSCs isolated from normal mice had more potent suppressor activity and much less capability to differentiate to osteoclast. Additional experiments showed that NF-κB inhibitor Bay 11-7082 or IκB inhibitor peptide blocked the differentiation of MDSCs to osteoclast and bone resorption. IL-1Ra also blocked this differentiation. In contrast, the addition of IL-1α further enhanced osteoclast differentiation and bone resorption. These results suggest that MDSCs are a source of osteoclast precursors and inflammatory cytokines such as IL-1, contributing significantly to erosive changes seen in rheumatoid arthritis and related disorders.

Myeloid-derived suppressor cells are proinflammatory and regulate collagen-induced arthritis through manipulating Th17 cell differentiation.

Myeloid-derived suppressor cells (MDSC) and Th17 cells were found to expand in collagen-induced arthritis (CIA) significantly. Two subsets of MDSC, polymorphonuclear (PMN) and mononuclear (MO), were detected and their ratios varied during the development of CIA. The depletion of MDSC in vivo resulted in suppression of T-cell proliferation and decreased IL-17A and IL-1ß production. The adoptive transfer of MDSC restored the severity of arthritis and Th17 cell differentiation. The depletion of MDSCs on day 35 resulted in arthritis amelioration without reaching a significant difference. Furthermore, MDSCs from CIA mice had higher production of IL-1ß and promoted Th17 cell differentiation. The expansion of MDSCs in the peripheral blood of rheumatoid arthritis (RA) patients was in correlation with increased Th17 cells and disease activity DAS28. These results support the hypothesis that MDSC may play a significant proinflammatory role in the pathogenesis of CIA and RA by inducing Th17 development in an IL-1ß-dependent manner.

Lupus nephritis: glycogen synthase kinase 3ß promotion of renal damage through activation of the NLRP3 inflammasome in lupus-prone mice.

OBJECTIVE: Glycogen synthase kinase 3ß (GSK-3ß) has been demonstrated to be involved in immune and inflammatory responses via multiple signaling pathways, leading to the production of proinflammatory cytokines. The purpose of this study was to investigate the role of GSK-3ß in the pathogenesis of lupus nephritis in 2 mouse models. METHODS: Thiadiazolidinone 8 (TDZD-8), a selective inhibitor of GSK-3ß, was administered intraperitoneally to 12-week-old MRL/lpr mice for 8 weeks or to 22-week-old (NZB × NZW)F1 mice for 12 weeks. The expression of GSK-3ß and NLRP3 inflammasome components was analyzed. Proteinuria, biochemical parameters, proinflammatory cytokines, anti-double-stranded DNA (anti-dsDNA) antibody levels, and renal pathology were examined. In vitro, the effect of GSK-3ß-directed small interfering RNA (siRNA) on NLRP3 inflammasome activation was evaluated in bone marrow-derived macrophages (BMMs) from the mice and in the J774A.1 macrophage cell line. RESULTS: The incidence of severe proteinuria and renal inflammation was significantly attenuated in both models, with a significant reduction in anti-dsDNA antibody production, immune complex deposition in the kidney, and circulating proinflammatory cytokine levels. TDZD-8 inhibited the activation of GSK-3ß and caspase 1, with a concomitant decrease in interleukin-1ß (IL-1ß) synthesis. In vitro, GSK-3ß siRNA transfection of mouse BMMs and the J774A.1 cell line with GSK-3ß siRNA inhibited the expression of GSK-3ß, the activation of caspase 1, and the production of IL-1ß. CONCLUSION: These results show that GSK-3ß promotes lupus nephritis at least partly by activating the NLRP3/IL-1ß pathway. The linking of GSK-3ß to the NLRP3/IL-1ß pathway is a novel observation in our study. Our results suggest that the GSK-3ß/NLRP3/IL-1ß pathway may be a potential therapeutic target for lupus in humans.

Genetics of systemic lupus erythematosus: immune responses and end organ resistance to damage.

Systemic lupus erythematosus (SLE) is a prototypic systemic autoimmune disorder. Considerable progress has been made to delineate the genetic control of this complex disorder. In this review, selected aspects of human and mouse genetics related to SLE are reviewed with emphasis on genes that contribute to both innate and adaptive immunity and to genes that contribute directly to susceptibility to end organ damage. It is concluded that the interactions among these two major pathways will provide further insight into the pathogenesis of SLE. An interactive model of the two major pathways is proposed without emphasis on the importance of breaking tolerance to autoantigens.

Interferon alpha on NZM2328.Lc1R27: enhancing autoimmunity and immune complex-mediated glomerulonephritis without end stage renal failure.

Interferon alpha (IFNα) may play a significant role in systemic lupus erythematosus (SLE) pathogenesis. Recent literature suggests that IFNα does not correlate with disease activities and blockade of IFNα is not effective in treating SLE. This study aims to delineate further the role of IFNα in SLE. 12-week old NZM2328 and its congenic NZM2328.Lc1R27 (R27) female mice were challenged with adenovirus-IFNα (adeno-IFNα) or adenovirus-LacZ (adeno-LacZ). Only adeno-IFNα treated NZM2328 developed severe proteinuria and died of chronic glomerulonephritis (GN) and end stage renal disease. Adeno-IFNα treated R27 did develop immune complex-mediated GN but had normal renal function. Adeno-LacZ treated NZM2328 showed enlarged glomeruli and increased cellularity without immune complex deposition. Adeno-LacZ treated R27 did not show serological and histological abnormalities. Adeno-IFNα induced anti-dsDNA and anti-kidney autoantibodies in NZM2328 and R27. These results suggest that end organ damage is host-dependent and less related to autoimmunity and may have significant implications in SLE pathogenesis.

Cgnz1 allele confers kidney resistance to damage preventing progression of immune complex-mediated acute lupus glomerulonephritis.

Cgnz1 and Agnz1 on the distal region of mouse chromosome 1 are associated with chronic glomerulonephritis (cGN) and acute GN (aGN). NZM2328.Lc1R27 (R27) was generated by introgressing a C57L/J region where Cgnz1 is located to NZM2328. R27 female mice developed aGN mediated by immune complex (IC) deposition and complement activation without progression to cGN with severe proteinuria. End stage renal disease (ESRD) was not seen in R27 mice as old as 15 mo. Thus, aGN and cGN are under separate genetic control, and IC-mediated proliferative GN need not progress to cGN and ESRD. NZM2328 and R27 female mice have comparable immune and inflammatory parameters. In contrast to NZM2328, R27 mice were resistant to sheep anti-mouse GBM serum-induced nephritis, supporting the hypothesis that aGN is mediated by autoimmunity and resistance to the development of cGN is mediated by end organ resistance to damage. Thus, autoimmunity should be considered distinct from end organ damage. The Cgnz1 region has been mapped to a 1.34 MB region with 45 genes. Nine candidate genes were identified. Clinical relevance of these observations is supported by case studies. Clinical implications and the significance to human lupus and other diseases are presented.

P2X7 blockade attenuates murine lupus nephritis by inhibiting activation of the NLRP3/ASC/caspase 1 pathway.

OBJECTIVE: The NLRP3 inflammasome plays key roles in inflammation and autoimmunity, and purinergic receptor P2X7 has been proposed to be upstream of NLRP3 activation. The aim of the present study, using murine models, was to investigate whether the P2X7 /NLRP3 inflammasome pathway contributes to the pathogenesis of lupus nephritis (LN). METHODS: MRL/lpr mice were treated with the selective P2X7 antagonist brilliant blue G (BBG) for 8 weeks. Following treatment, the severity of renal lesions, production of anti-double-stranded DNA (anti-dsDNA) antibodies, rate of survival, activation of the NLRP3/ASC/caspase 1 inflammasome pathway, and ratio of Th17 cells to Treg cells were evaluated. P2X7 -targeted small interfering RNA (siRNA) was also used for in vivo intervention. Similar evaluations were carried out in NZM2328 mice, a model of LN in which the disease was accelerated by administration of adenovirus-expressing interferon-α (AdIFNα). RESULTS: Significant up-regulation of P2X7 /NLRP3 inflammasome signaling molecules was detected in the kidneys of MLR/lpr mice as compared with normal control mice. Blockade of P2X7 activation by BBG suppressed NLRP3/ASC/caspase 1 assembly and the subsequent release of interleukin-1ß (IL-1ß), resulting in a significant reduction in the severity of nephritis and circulating anti-dsDNA antibodies. The lifespan of the treated mice was significantly prolonged. BBG treatment reduced the serum levels of IL-1ß and IL-17 and the Th17:Treg cell ratio. Similar results were obtained by specific siRNA silencing of P2X7 in vivo. The effectiveness of BBG treatment in modulating LN was confirmed in NZM2328 mice with AdIFNα-accelerated disease. CONCLUSION: Activation of the P2X7 signaling pathway accelerates murine LN by activating the NLRP3/ASC/caspase 1 inflammasome, resulting in increased IL-1ß production and enhanced Th17 cell polarization. Thus, targeting of the P2X7 /NLRP3 pathway should be considered as a novel therapeutic strategy in patients with lupus.

Autoimmunity, end organ damage, and the origin of autoantibodies and autoreactive T cells in systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is a prototype of systemic autoimmunity affecting many systems. Both antibodies and autoreactive T cells play significant roles in its pathogenesis. Experimental data and clinical observations indicate that autoimmunity and end organ damage are under separate genetic controls and that there are significant interactions between these two pathways. Experimental evidence has been obtained to support the hypothesis that autoantibodies and autoreactive T effector cells may be initiated by environmental factors through molecular mimicry and the inherent polyreactive nature of antigen receptors. A unified hypothesis has been postulated for the pathogenesis of SLE that has practical implications.

IL-2 controls trafficking receptor gene expression and Th2 response for skin and lung inflammation.

Both Il2(-/-) mice and Scurfy (Sf) mutant mice that are deficient in FoxP3, develop multi-organ inflammation but only the latter display severe skin and lung inflammation. In contrast, Sf.Il2(-/-) double mutant mice do not display skin inflammation and markedly reduced lung inflammation. In this review, we summarize our recent findings based on microarray, q-PCR and functional studies of 10 Sf double mutant mice. These studies revealed novel pro-inflammatory functions of IL-2 in regulating inflammation in an organ-specific manner. IL-2 exerts its "organ-specific" pro-inflammatory function by regulating the migration and retention of CD4(+) T-cells (both Th1 and Th2) specifically to the skin and lung. In addition, IL-2 is also required for regulating the Th2 cytokine response during T-cell activation. Further studies on these IL-2-regulated genes will help in identifying novel targets for intervention in inflammatory diseases of skin and lung.

A novel function of IL-2: chemokine/chemoattractant/retention receptor genes induction in Th subsets for skin and lung inflammation.

The Foxp3(+)CD4(+) regulatory T-cell (Treg)-deficient Scurfy (Sf) mice rapidly develop severe inflammation in the skin and lungs with expanded Th subsets bearing increased expression of various chemokine/chemoattractant/retention receptor genes (CRG). Nine different double mutants were generated to elucidate their roles in the skin and lung inflammation. The expanded Th2 response and the increased expression of several CRG for the skin and lung inflammation were inhibited in Sf.Il2(-/-) mice as previously described using microarray analysis. Herein in a reciprocal approach, we demonstrated that Sf.Il4(-/-) and Sf.Stat6(-/-) mice, despite lacking Th2 cytokines IL-4, IL-5, and IL-13, as well as the IL-4/STAT6-dependent CRG expression, the inflammation in the skin and lungs remained. The effect of the other Th1 cytokine IFN-γ was studied in Sf.Ifng(-/-) mice in which the multi-organ inflammation (MOI) was delayed but fully developed afterward with enhanced CRG expression except for the IFN-γ-dependent Cxcr3 in CD4(+) T-cells. Similarly, a transient delay of MOI was observed for Sf.Itgae(-/-) mice but their Th subsets and the critical CRG expansion remained. Ltb4r1(-/-), Alox5(-/-), Cx3cr1(gfp/gfp), or Il10(-/-) mutant genes also failed to effectively block inflammation in the skin and lungs in Sf mice. Our study has identified a novel function of IL-2 as a powerful Th1 cytokine that induces a panel of CRG in Th subsets required for skin and lung inflammation in Sf mice. The CRG panel induced by IL-2 but not by IL-4 or IFN-γ explains the apparent "organ-specific" display of the skin and lung inflammation in Sf mice.