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.

Interactions among glomerulus infiltrating macrophages and intrinsic cells via cytokines in chronic lupus glomerulonephritis.

Inflammation plays a key role in the pathogenesis of lupus nephritis (LN) and inflammatory cytokines within the glomeruli are critical in this process. However, little information is available for the identities of the cell types that are primarily responsible for the production and function of the various cytokines. We have devised a novel method to visualize cytokine signals in the kidney by confocal microscopy and found that cytokine production within the glomerulus is cell type-specific and under translational control. In the lupus-prone NZM2328 mice with chronic glomerulonephritis, IL-6, IL-1ß, and TNF-α in the glomerulus were produced predominantly by mesangial cells, podocytes, and glomerulus-infiltrating blood-derived macrophages, respectively. Microarray and RNASeq analyses showed that these cells expressed the receptors for these cytokines. Together the 3 cell types form a cytokine circuit in amplifying cytokine responses in LN. The intrinsic cells and infiltrating macrophages also produced other cytokines including M-CSF, SCF, and IL-34 that constituted within the enclosed glomerular space the soluble effector milieu which may mediate cellular damage and proliferation, and cytokine transcriptional and translation regulation. IL-10 and IL-1ß were translationally regulated in the glomeruli in the intact kidney in a cell type-specific manner. The production of these 2 cytokines by infiltrating macrophages was undetectable in a visualization system for in situ protein accumulation despite high mRNA expression levels. However, these macrophages in isolated glomeruli which are released from Bowman's capsules produced large amounts of IL-10 and IL-1ß. These data reveal the complexity of cytokine regulation, production, and function in the glomerulus and provide a model in which cytokine blocking may be beneficial in LN treatment.

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.

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).

IL233, an IL-2-IL-33 hybrid cytokine induces prolonged remission of mouse lupus nephritis by targeting Treg cells as a single therapeutic agent.

Lupus glomerulonephritis (GN) is an autoimmune disease characterized by immune complex-deposition, complement activation and glomerular inflammation. In lupus-prone NZM2328 mice, the occurrence of lupus GN was accompanied by a decrease in Treg cells and an increase in proinflammatory cytokine-producing T cells. Because IL-33 in addition to IL-2 has been shown to be important for Treg cell proliferation and ST2 (IL-33 receptor) positive Treg cells are more potent in suppressor activity, a hybrid cytokine with active domains of IL-2 and IL-33 was generated to target the ST2+ Treg cells as a therapeutic agent to treat lupus GN. Three mouse models were used: spontaneous and Ad-IFNα- accelerated lupus GN in NZM2328 and the lymphoproliferative autoimmune GN in MRL/lpr mice. Daily injections of IL233 for 5 days prevented Ad-IFNα-induced lupus GN and induced remission of spontaneous lupus GN. The remission was permanent in that no relapses were detected. The remission was accompanied by persistent elevation of Treg cells in the renal lymph nodes. IL233 is more potent than IL-2 and IL-33 either singly or in combination in the treatment of lupus GN. The results of this study support the thesis that IL233 should be considered as a novel agent for treating lupus GN.

Dependence of Glomerulonephritis Induction on Novel Intraglomerular Alternatively Activated Bone Marrow-Derived Macrophages and Mac-1 and PD-L1 in Lupus-Prone NZM2328 Mice.

Glomerular damage mediated by glomerulus-infiltrating myeloid-derived cells is a key pathogenic event in lupus nephritis (LN), but the process is poorly understood. Confocal microscopy of kidney sections and flow cytometry analysis of glomerular cells from magnetic bead-purified glomeruli have identified glomerulus-infiltrating leukocyte populations in NZM2328 (NZM) lupus-prone mice with spontaneous chronic glomerulonephritis (GN) and anti-glomerular basement membrane-induced nephritis. The occurrence of a major glomerulus-infiltrating CD11b+F4/80-I-A- macrophage population exhibiting the markers programmed death ligand-1 (PD-L1), Mac-2, and macrophage mannose receptor (CD206) and producing Klf4, Il10, Retnla, Tnf, and Il6 mRNA, which are known to be expressed by alternatively activated (M2b) macrophages, correlated with proteinuria status. In NZM mice with spontaneous LN, glomerular macrophage infiltration is predominant. CD11b+F4/80-I-A- intraglomerular macrophages and polymorphonuclear neutrophils (PMN) are important in inducing GN, as anti-CD11b and -ICAM-1 mAb inhibited both proteinuria and macrophage and PMN infiltration. The predominant and high expression of PD-L1 by CD11b+F4/80-I-A- glomerular macrophages in kidneys of mice with GN and the inhibition of proteinuria by anti-PD-L1 mAb supported the pathogenic role of these macrophages but not the PD-L1- PMN in GN development and in inducing podocyte damage. In NZM mice with spontaneous chronic GN and severe proteinuria, few glomerulus-infiltrating PMN were found, leaving macrophages and, to a less extent, dendritic cells as the major infiltrating leukocytes. Taken together, these data support the important pathogenic effect of CD11b+F4/80-I-A- M2b-like glomerulus-infiltrating macrophages in LN and reinforce macrophages as a promising target for GN treatment.

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.

A Central Role for HLA-DR3 in Anti-Smith Antibody Responses and Glomerulonephritis in a Transgenic Mouse Model of Spontaneous Lupus.

MHC, especially HLA-DR3 and HLA-DR2, is one of the most important genetic susceptibility regions for systemic lupus erythematosus. Human studies to understand the role of specific HLA alleles in disease pathogenesis have been hampered by the presence of strong linkage disequilibrium in this region. To overcome this, we produced transgenic mice expressing HLA-DR3 (DRß1*0301) and devoid of endogenous class II (both I-A and I-E genes, AE(0)) on a lupus-prone NZM2328 background (NZM2328.DR3(+)AE(0)). Both NZM2328 and NZM2328.DR3(+)AE(0) mice developed anti-dsDNA and glomerulonephritis, but anti-dsDNA titers were higher in the latter. Although kidney histological scores were similar in NZM2328 and NZM2328.DR3(+)AE(0) mice (7.2 ± 4.3 and 8.6 ± 5.7, respectively, p = 0.48), the onset of severe proteinuria occurred earlier in NZM2328.DR3(+)AE(0) mice compared with NZM2328 mice (median, 5 and 9 mo respectively, p < 0.001). Periarterial lymphoid aggregates, classic wire loop lesions, and occasional crescents were seen only in kidneys from NZM2328.DR3(+)AE(0) mice. Interestingly, NZM2328.DR3(+)AE(0) mice, but not NZM2328 mice, spontaneously developed anti-Smith (Sm) Abs. The anti-Sm Abs were seen in NZM2328.DR3(+)AE(0) mice that were completely devoid of endogenous class II (AE(-/) (-)) but not in mice homozygous (AE(+/+)) or heterozygous (AE(+/-)) for endogenous MHC class II. It appears that only HLA-DR3 molecules can preferentially select SmD-reactive CD4(+) T cells for generation of the spontaneous anti-Sm immune response. Thus, our mouse model unravels a critical role for HLA-DR3 in generating an autoimmune response to SmD and lupus nephritis in the NZM2328 background.

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.

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.

T cell epitope mimicry between Sjögren's syndrome Antigen A (SSA)/Ro60 and oral, gut, skin and vaginal bacteria.

This study was undertaken to test the hypothesis that Sjogren's syndrome Antigen A (SSA)/Ro60-reactive T cells are activated by peptides originating from oral and gut bacteria. T cell hybridomas generated from HLA-DR3 transgenic mice recognized 3 regions on Ro60, with core epitopes mapped to amino acids 228-238, 246-256 and 371-381. BLAST analysis identified several mimicry peptides, originating from human oral, intestinal, skin and vaginal bacteria, as well as environmental bacteria. Amongst these, a peptide from the von Willebrand factor type A domain protein (vWFA) from the oral microbe Capnocytophaga ochracea was the most potent activator. Further, Ro60-reactive T cells were activated by recombinant vWFA protein and whole Escherichia coli expressing this protein. These results demonstrate that peptides derived from normal human microbiota can activate Ro60-reactive T cells. Thus, immune responses to commensal microbiota and opportunistic pathogens should be explored as potential triggers for initiating autoimmunity in SLE and Sjögren's syndrome.

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.

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.

IL-2-controlled expression of multiple T cell trafficking genes and Th2 cytokines in the regulatory T cell-deficient scurfy mice: implication to multiorgan inflammation and control of skin and lung inflammation.

Scurfy (Sf) mice bear a mutation in the Foxp3 transcription factor, lack regulatory T cells (Treg), develop multiorgan inflammation, and die prematurely. The major target organs affected are skin, lungs, and liver. "Sf mice lacking the Il2 gene (Sf.Il2­/­), despite being devoid of Treg, did not develop skin and lung inflammation, but the inflammation in liver remained [corrected]. Genome-wide microarray analysis revealed hundreds of genes that were differentially regulated among Sf, Sf.Il2(-/-), and B6 CD4(+) T cells, but the most significant changes were those encoding receptors for trafficking/chemotaxis/retention and cytokines. Our study suggests that IL-2 controls the skin and lung inflammation in Sf mice in an apparent "organ-specific" manner through two novel mechanisms: by regulating the expression of genes encoding a variety of receptors for T cell trafficking/chemotaxis/retention and by regulating Th2 cell expansion and cytokine production. Thus, IL-2 is potentially a master regulator for multiorgan inflammation and an underlying etiological factor for various diseases associated with skin and lung inflammation.

Molecular mechanisms governing the progression of nephritis in lupus prone mice and human lupus patients.

Introduction: Pathologic inflammation is a major driver of kidney damage in lupus nephritis (LN), but the immune mechanisms of disease progression and risk factors for end organ damage are poorly understood. Methods: To characterize molecular profiles through the development of LN, we carried out gene expression analysis of microdissected kidneys from lupus-prone NZM2328 mice. We examined male mice and the congenic NZM2328.R27 strain as a means to define mechanisms associated with resistance to chronic nephritis. Gene expression profiles in lupus mice were compared with those in human LN. Results: NZM2328 mice exhibited progress from acute to transitional and then to chronic glomerulonephritis (GN). Each stage manifested a unique molecular profile. Neither male mice nor R27 mice progressed past the acute GN stage, with the former exhibiting minimal immune infiltration and the latter enrichment of immunoregulatory gene signatures in conjunction with robust kidney tubule cell profiles indicative of resistance to cellular damage. The gene expression profiles of human LN were similar to those noted in the NZM2328 mouse suggesting comparable stages of LN progression. Conclusions: Overall, this work provides a comprehensive examination of the immune processes involved in progression of murine LN and thus contributes to our understanding of the risk factors for end-stage renal disease. In addition, this work presents a foundation for improved classification of LN and illustrates the applicability of murine models to identify the stages of human disease.

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.