AM14 rheumatoid factor (RF) B cells in the MRL/lpr mice are activated by dual BCR and TLR7/9 ligation and differentiate into plasmablasts via an extrafollicular (EF) route. It was not known whether this mechanism of activation of RF B cells applied to other lupus-prone mouse models. We investigated the mechanisms by which RF B cells break tolerance in the NZM2410-derived B6.Sle1.Sle2.Sle3 (TC) strain in comparison with C57BL/6 (B6) controls, each expressing the AM14 H chain transgene in the presence or absence of the IgG2a(a) autoantigen. The TC, but not B6, genetic background promotes the differentiation of RF B cells into Ab-forming cells (AFCs) in the presence of the autoantigen. Activated RF B cells preferentially differentiated into plasmablasts in EF zones. Contrary to the MRL/lpr strain, TC RF B cells were also located within germinal centers, but only the formation of EF foci was positively correlated with the production of RF AFCs. Immunization of young TC.AM14 H chain transgenic mice with IgG2a(a) anti-chromatin immune complexes (ICs) activated RF B cells in a BCR- and TLR9-dependent manner. However, these IC immunizations did not result in the production of RF AFCs. These results show that RF B cells break tolerance with the same general mechanisms in the TC and the MRL/lpr lupus-prone genetic backgrounds, namely the dual activation of the BCR and TLR9 pathways. There are also distinct differences, such as the presence of RF B cells in GCs and the requirement of chronic IgG2a(a) anti-chromatin ICs for full differentiation of RF AFCs.
A subset of patients with severe hemophilia B, the X-linked bleeding disorder resulting from absence of coagulation factor IX (FIX), develop pathogenic antibody responses during replacement therapy. These inhibitors block standard therapy and are often associated with anaphylactic reactions to FIX. Established clinical immune tolerance induction protocols often fail for FIX inhibitors. In a murine model of this immune complication, retrovirally transduced primary B cells expressing FIX antigen fused with immunoglobulin-G heavy chain prevented antibody formation to FIX and was also highly effective in desensitizing animals with preexisting response. In contrast, transplant of B cells that received the identical expression cassette via nucleofection of plasmid vector substantially heightened antibody formation against FIX, a response that could be blocked by toll-like receptor 9 (TLR9) inhibition. While innate responses to TLR4 activation or to retrovirus were minimal in B cells, plasmid DNA activated TLR9, resulting in CpG-dependent NF-?B activation/IL-6 expression and adaptor protein 3 dependent, CpG-independent induction of IFN-I. Neither response was seen in TLR9-deficient B cells. Therefore, TLR9 signaling in B cells, in particular in response to plasmid vector, is highly immunogenic and has to be avoided in design of tolerance protocols.
Induced mouse models of systemic lupus erythematosus (SLE) have been developed to complement the spontaneous models. This chapter describes the methods used in the pristane-induced model and the chronic graft-versus-host disease (cGVHD) model, both of which have been extensively used. We will also outline the specific mechanisms of systemic autoimmunity that can be best characterized using each of these models.
The breakdown in tolerance of autoreactive B cells in the lupus-prone NZM2410-derived B6.Sle1.Sle2.Sle3 (TC) mice results in the secretion of autoantibodies. TC dendritic cells (DCs) enhance B cell proliferation and antibody secretion in a cytokine-dependent manner. However, the specific cytokine milieu by which TC DCs activate B cells was not known. In this study, we compared TC and C57BL/6 (B6) control for the distribution of DC subsets and for their production of cytokines affecting B cell responses. We show that TC DCs enhanced B cell proliferation through the production of IL-6 and IFN-?, while antibody secretion was only dependent on IL-6. Pre-disease TC mice showed an expanded PDCA1(+) cells prior to disease onset that was localized to the marginal zone and further expanded with age. The presence of PDCA1(+) cells in the marginal zone correlated with a Type I Interferon (IFN) signature in marginal zone B cells, and this response was higher in TC than B6 mice. In vivo administration of anti-chromatin immune complexes upregulated IL-6 and IFN-? production by splenic DCs from TC but not B6 mice. The production of BAFF and APRIL was decreased upon TC DC stimulation both in vitro and in vivo, indicating that these B cell survival factors do not play a role in B cell modulation by TC DCs. Finally, TC B cells were defective at downregulating IL-6 expression in response to anti-inflammatory apoptotic cell exposure. Overall, these results show that the TC autoimmune genetic background induces the production of B cell-modulating inflammatory cytokines by DCs, which are regulated by the microenvironment as well as the interplay between DC.
Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by the production of autoantibodies. This review summarizes first the results obtained in the mouse that have revealed how B cell tolerance is breached in SLE. We then review the B cell subsets, in addition to the autoAb producing cells, which contribute to SLE pathogenesis, focusing on marginal zone B cells, B-1 cells and regulatory B cells. Finally, we review the interactions between B cells and other immune cells that have been implicated in SLE, such as dendritic cells, macrophages, neutrophils and T cells.
INTRODUCTION: An NZB-derived genetic locus (Sle2c2) that suppresses autoantibody production in a mouse model of induced systemic lupus erythematosus contains a polymorphism in the gene encoding the G-CSF receptor. This study was designed to test the hypothesis that the Sle2c2 suppression is associated with an impaired G-CSF receptor function that can be overcome by exogenous G-CSF. METHODS: Leukocytes from B6.Sle2c2 and B6 congenic mice, which carry a different allele of the G-CSF receptor, were compared for their responses to G-CSF. Autoantibody production was induced with the chronic graft-versus-host-disease (cGVHD) model by adoptive transfer of B6.bm12 splenocytes. Different treatment regimens varying the amount and frequency of G-CSF (Neulasta®) or carrier control were tested on cGVHD outcomes. Autoantibody production, immune cell activation, and reactive oxygen species (ROS) production were compared between the two strains with the various treatments. In addition, the effect of G-CSF treatment was examined on the production autoantibodies in the B6.Sle1.Sle2.Sle3 (B6.TC) spontaneous model of lupus. RESULTS: B6.Sle2c2 and B6 leukocytes responded differently to G-CSF. G-CSF binding by B6.Sle2c2 leukocytes was reduced as compared to B6, which was associated with a reduced expansion in response to in vivo G-CSF treatment. G-CSF in vivo treatment also failed to mobilize bone-marrow B6.Sle2c2 neutrophils as it did for B6 neutrophils. In contrast, the expression of G-CSF responsive genes indicated a higher G-CSF receptor signaling in B6.Sle2c2 cells. G-CSF treatment restored the ability of B6.Sle2c2 mice to produce autoantibodies in a dose-dependent manner upon cGVHD induction, which correlated with restored CD4+ T cells activation, as well as dendritic cell and granulocyte expansion. Steady-state ROS production was higher in B6.Sle2c2 than in B6 mice. cGVHD induction resulted in a larger increase in ROS production in B6 than in B6.Sle2c2 mice, and this difference was eliminated with G-CSF treatment. Finally, a low dose G-CSF treatment accelerated the production of anti-dsDNA IgG in young B6.TC mice. CONCLUSION: The different in vivo and in vitro responses of B6.Sle2c2 leukocytes are consistent with the mutation in the G-CSFR having functional consequences. The elimination of Sle2c2 suppression of autoantibody production by exogenous G-CSF indicates that Sle2c2 corresponds to a loss of function of G-CSF receptor. This result was corroborated by the increased anti-dsDNA IgG production in G-CSF-treated B6.TC mice, which also carry the Sle2c2 locus. Overall, these results suggest that the G-CSF pathway regulates the production of autoantibodies in murine models of lupus.
Hematopoietic stem cells (HSCs) are protected in a metabolically dormant state within the bone marrow stem cell niche. Inflammation has been shown to disrupt HSC dormancy and cause multiple functional changes. Here, we investigated whether HSC functions were altered in systemic lupus erythematosus (SLE)-prone mice and whether this contributed to clinical manifestations of SLE. We found that HSCs were significantly expanded in lupus mice. The increase in HSC cellularity was caused by both genetic lupus risk factors and inflammatory cytokines in lupus mice. In addition, the inflammatory conditions of lupus led to HSC mobilization and lineage-biased hematopoiesis. Strikingly, these functionally altered HSCs possessed robust self-renewal capacity and exhibited repopulating advantages over wild-type HSCs. A single-nucleotide polymorphism in the cdkn2c gene encoding p18(INK4c) within a SLE susceptibility locus was found to account for reduced p18(INK4c) expression and the increase in HSC self-renewal capacity in lupus mice. Lupus HSCs with enhanced self-renewal capacity and resistance to stress may compete out transplanted healthy HSCs, thereby leading to relapses after HSC transplantation.
Sle1a.1 is part of the Sle1 susceptibility locus, which has the strongest association with lupus nephritis in the NZM2410 mouse model. In this study, we show that Sle1a.1 results in the production of activated and autoreactive CD4(+) T cells. Additionally, Sle1a.1 expression reduces the peripheral regulatory T cell pool, as well as induces a defective response of CD4(+) T cells to the retinoic acid expansion of TGF-?-induced regulatory T cells. At the molecular level, Sle1a.1 corresponds to an increased expression of a novel splice isoform of Pbx1, Pbx1-d. Pbx1-d overexpression is sufficient to induce an activated/inflammatory phenotype in Jurkat T cells and to decrease their apoptotic response to retinoic acid. PBX1-d is expressed more frequently in the CD4(+) T cells from lupus patients than from healthy controls, and its presence correlates with an increased central memory T cell population. These findings indicate that Pbx1 is a novel lupus susceptibility gene that regulates T cell activation and tolerance.
Tissue and organ differentiation is tightly controlled to ensure proper development and function of the growing embryo as well as cells such as lymphocytes that differentiate throughout the adult stage. Therefore it is vital that the genes and the protein they encode that are involved in these processes function accurately. Hence, any mutation or error that occurs along the way can result in extensive damage, which is expressed in various ways in the embryo and can result in immune pathogenesis, including immunodeficiency and autoimmune diseases, when lymphocyte development is altered. A number of studies have been carried out to look at the genes regulating transcription in tissue differentiation, including the transcription factors Pbx1. This gene is of particular interest to us as we have identified that it is associated with systemic lupus erythematosus susceptibility (Cuda et al., in press). This perspective summarizes the known roles of Pbx1 in tissue differentiation as well as our recent findings associating genetic variations in Pbx1 to lupus susceptibility, and we will speculate on how this gene controls the maintenance of immune tolerance in T cells.
CD25(+) FOXP3(+) CD4(+) regulatory T cells (Tregs) are induced by transforming growth factor ? (TGF?) and further expanded by retinoic acid (RA). We have previously shown that this process was defective in T cells from lupus-prone mice expressing the novel isoform of the Pbx1 gene, Pbx1-d. This study tested the hypothesis that CD4(+) T cells from systemic lupus erythematosus (SLE) patients exhibited similar defects in Treg induction in response to TGF? and RA, and that PBX1-d expression is associated with this defect.
Sle2c1 is an NZM2410- and NZB-derived lupus susceptibility locus that induces an expansion of the B1a cell compartment. B1a cells have a repertoire enriched for autoreactivity, and an expansion of this B cell subset occurs in several mouse models of lupus. A combination of genetic mapping and candidate gene analysis presents Cdkn2c, a gene encoding for cyclin-dependent kinase inhibitor p18(INK4c) (p18), as the top candidate gene for inducing the Slec2c1-associated expansion of B1a cells. A novel single nucleotide polymorphism in the NZB allele of the Cdkn2c promoter is associated with a significantly reduced Cdkn2c expression in the splenic B cells and peritoneal cavity B1a cells from Sle2c1-carrying mice, which leads to a defective G1 cell cycle arrest in splenic B cells and increased proliferation of peritoneal cavity B1a cells. As the cell cycle is differentially regulated in B1a and B2 cells, these results suggest that Cdkn2c plays a critical role in B1a cell self-renewal and that its impaired expression leads to an accumulation of these cells with high autoreactive potential.
Studies of human systemic lupus erythematosus patients and of murine congenic mouse strains associate genes in a DNA segment on chromosome 1 with a genetic predisposition for this disease. The systematic analysis of lupus-prone congenic mouse strains suggests a role for two isoforms of the Ly108 receptor in the pathogenesis of the disease. In this study, we demonstrate that Ly108 is involved in the pathogenesis of lupus-related autoimmunity in mice. More importantly, we identified a third protein isoform, Ly108-H1, which is absent in two lupus-prone congenic animals. Introduction of an Ly108-H1-expressing transgene markedly diminishes T cell-dependent autoimmunity in congenic B6.Sle1b mice. Thus, an immune response-suppressing isoform of Ly108 can regulate the pathogenesis of lupus.
Sle2 is a lupus susceptibility locus that has been linked to glomerulonephritis in the NZM2410 mouse. By itself, Sle2 does not induce any autoimmune pathology but results in the accumulation of B-1a cells. This study was designed to assess the contribution of Sle2 to the pathogenesis of autoimmunity.
The development of lupus pathogenesis results from the integration of susceptibility and resistance genes. We have used a chronic graft-versus-host disease (cGVHD) model to characterize a suppressive locus at the telomeric end of the NZM2410-derived Sle2 susceptibility locus, which we named Sle2c2. cGVHD is induced normally in Sle2c2-expressing mice, but it is not sustained. The analysis of mixed bone marrow chimeras revealed that cGVHD resistance was eliminated by non-B non-T hematopoietic cells expressing the B6 allele, suggesting that resistance is mediated by this same cell type. Furthermore, Sle2c2 expression was associated with an increased number and activation of the CD11b(+) GR-1(+) subset of granulocytes before and in the early stage of cGVHD induction. We have mapped the Sle2c2 critical interval to a 6-Mb region that contains the Cfs3r gene, which encodes for the G-CSFR, and its NZM2410 allele carries a nonsynonymous mutation. The G-CSFR-G-CSF pathway has been previously implicated in the regulation of GVHD, and our functional data on Sle2c2 suppression suggest a novel regulation of T cell-induced systemic autoimmunity through myeloid-derived suppressor cells. The validation of Csf3r as the causative gene for Sle2c2 and the further characterization of the Sle2c2 MDSCs promise to unveil new mechanisms by which lupus pathogenesis is regulated.
Systemic lupus erythematosus (SLE) is a multifactorial autoimmune disorder. The study of diverse mouse models of lupus has provided clues to the etiology of SLE. Spontaneous mouse models of lupus have led to identification of numerous susceptibility loci from which several candidate genes have emerged. Meanwhile, induced models of lupus have provided insight into the role of environmental factors in lupus pathogenesis as well as provided a better understanding of cellular mechanisms involved in the onset and progression of disease. The SLE-like phenotypes present in these models have also served to screen numerous potential SLE therapies. Due to the complex nature of SLE, it is necessary to understand the effect specific targeted therapies have on immune homeostasis. Furthermore, knowledge gained from mouse models will provide novel therapy targets for the treatment of SLE.
Systemic lupus erythematosus (SLE) is a chronic autoimmune disease with a multifaceted range of symptoms affecting almost every organ system. The prototypical pathology of SLE involves the production of antinuclear antibodies and the deposition of immune complexes in basement membranes throughout the body where they induce inflammatory responses. The genetic and environmental etiologies of this process are being intensively sought, and recently, T( H )17 cells have been implicated in the pathogenesis of SLE. T( H )17 cells are CD4+ memory T cells that behave as both helper and effector cell populations functioning through their signature IL-17 cytokines. Their differentiation is distinct to either the T( H )1 or T( H )2 cell lineage, but strongly influences development of adaptive responses, including autoimmunity. This paper details the biological functions and regulation of T( H )17 cells, followed by an update of their expanding role in SLE.
Marginal zone B cells have been implicated in the production of autoantibodies in murine models of lupus. It has been suggested that they contribute to lupus immunopathogenesis through their enhanced effector functions and their repertoire that is biased toward autoreactive specificities. In the B6.NZM2410.Sle.Sle2.Sle3 (B6.TC) model of lupus, the majority of marginal zone B cells are located outside the marginal zone and inside the follicles. Genetic alterations of this strain have shown a correlation between autoimmune pathogenesis and the presence of intrafollicular marginal zone B cells. This study was designed first to strengthen our original observations and to determine how the marginal zone B cells from the lupus-prone mice respond to stimulations and interact with T cells.
Reactive oxygen species are used by the immune system to eliminate infections; however, they may also serve as signaling intermediates to coordinate the efforts of the innate and adaptive immune systems. In this study, we show that by eliminating macrophage and T cell superoxide production through the NADPH oxidase (NOX), T cell polarization was altered. After stimulation with immobilized anti-CD3 and anti-CD28 or priming recall, T cells from NOX-deficient mice exhibited a skewed Th17 phenotype, whereas NOX-intact cells produced cytokines indicative of a Th1 response. These findings were corroborated in vivo by studying two different autoimmune diseases mediated by Th17 or Th1 pathogenic T cell responses. NOX-deficient NOD mice were Th17 prone with a concomitant susceptibility to experimental allergic encephalomyelitis and significant protection against type 1 diabetes. These data validate the role of superoxide in shaping Th responses and as a signaling intermediate to modulate Th17 and Th1 T cell responses.
The past two years have brought great progress in the genetics of systemic lupus erythematosus (SLE) heralded by the publication of genome-wide association studies in humans and the identification of susceptibility genes in mouse models of spontaneous lupus. This influx of new information has revealed an ever-increasing interdependence between the mouse and human systems for unraveling the genetic basis of lupus susceptibility. SLE is a complex disease in which defects in several functional pathways have been identified. Genetic variants in a number of genes in these pathways have now been directly associated with lupus in both species. These discoveries have lead to a better understanding of the mechanisms of disease, and offer potential novel target for therapeutic intervention. As a large number of susceptibility genes are identified, lupus genetics will focus on mechanistic and molecular studies, in which mouse models will continue to serve a pre-eminent role.
The Sle1c subinterval on distal murine chromosome 1 confers loss of tolerance to chromatin. Cr2, which encodes complement receptors 1 and 2 (CR1/CR2; CD35/CD21), is a strong candidate gene for lupus susceptibility within this interval based on structural and functional alterations in its protein products. CR1-related protein/gene Y (Crry) lies 10 kb from Cr2 and encodes a ubiquitously expressed complement regulatory protein that could also play a role in the pathogenesis of systemic lupus erythematosus. Crry derived from B6.Sle1c congenic mice migrated at a higher m.w. by SDS-PAGE compared with B6 Crry, as a result of differential glycosylation. A single-nucleotide polymorphism in the first short consensus repeat of Sle1c Crry introduced a novel N-linked glycosylation site likely responsible for this structural alteration. Five additional single-nucleotide polymorphisms in the signal peptide and short consensus repeat 1 of Sle1c Crry were identified. However, the cellular expression of B6 and B6.Sle1c Crry and their ability to regulate the classical pathway of complement were not significantly different. Although soluble Sle1c Crry regulated the alternative pathway of complement more efficiently than B6 Crry, as a membrane protein, it regulated the alternative pathway equivalently to B6 Crry. These data fail to provide evidence for a functional effect of the structural alterations in Sle1c Crry and suggest that the role of Cr2 in the Sle1c autoimmune phenotypes can be isolated in recombinant congenic mice containing both genes.
We report a case of infection with coagulase-positive Staphylococcus pseudintermedius related to the implantation of a cardioverter-defribrillator device. This species is usually isolated from infected animals, and contact with a dog was the probable source of infection in this patient. This isolate produced a leukotoxin effective against human polymorphonuclear leukocytes.
Great progress has been made in the field of lupus genetics in the past few years, notably with the publication of genome-wide association studies in humans and the identification of susceptibility genes (including Fcgr2b, Ly108, Kallikrein genes and Coronin-1A) in mouse models of spontaneous lupus. This influx of new information has revealed an ever-increasing interdependence between the mouse and human systems for unraveling the genetic basis of lupus susceptibility. Studies in the 1980s and 1990s established that mice prone to spontaneous lupus constitute excellent models of the genetic architecture of human systemic lupus erythematosus (SLE). This notion has been greatly strengthened by the convergence of the functional pathways that are defective in both human and murine lupus. Within these pathways, variants in a number of genes have now been shown to be directly associated with lupus in both species. Consequently, mouse models will continue to serve a pre-eminent role in lupus genetics research, with an increased emphasis on mechanistic and molecular studies of human susceptibility alleles.
Paratuberculosis (Johnes disease) imposes a significant problem to the world dairy and beef industries and today is considered a potential zoonosis. The disease is caused by Mycobacterium avium subsp. paratuberculosis and is characterized by progressive weight loss and profuse diarrhoea. Susceptibility to infection is suspected to have a genetic component, and moderated values for heritability of infection have been reported. Interferon gamma is an inducible cytokine with a crucial role in the innate host response to intracellular bacteria. Toll-like receptors are trans-membrane structures responsible for coordination of innate and adaptive immune responses. The solute carrier family 11 member 1 (SLC11A1, formerly NRAMP1) gene plays an important role in innate immunity, preventing bacterial growth in macrophages during the initial stages of infection. The objective of this candidate gene case-control study was to characterize the distribution of polymorphisms in three candidate genes related to the immune function; interferon gamma (BoIFNG), toll-like receptor 4 (TLR4), and SLC11A1 genes and to test their role as potential risk factors for paratuberculosis infection in cattle. The statistical analysis demonstrated significant differences in allelic frequencies between cases and controls for BoIFNG-SNP(1)2781 and SLC11A1 microsatellites, indicating a significant association between infection and variant alleles. In the analysis of genotypes, a significant association was also found between infection status and BoIFNG-SNP(1)2781 and SLC11A1-275-279-281 microsatellites. However, when variables such as breed and age were included in the multivariate logistic regression analysis, a tendency toward statistical significance for the effect of polymorphisms in the odds of infection was only found for alleles SLC11A1-275 and 279.
Paratuberculosis represents a major problem in farmed ruminants and at the present is considered a potential zoonosis. The disease is caused by Mycobacterium avium subsp. paratuberculosis, and susceptibility to infection is suspected to have a genetic component. Caspase recruitment domain 15 (CARD15) gene encodes for a cytosolic protein implicated in bacterial recognition during innate immunity. Crohns disease (CD) is an idiopathic inflammatory bowel disease in humans comparable in many features to bovine paratuberculosis involving an abnormal mucosal immune response. The association between mutations in the CARD15 gene and increased risk of Crohns disease has been described. The objective of this candidate gene case-control study was to characterize the distribution of three polymorphisms in the bovine CARD15 gene and test their association with paratuberculosis infection in cattle. Three previously reported single nucleotide polymorphisms (E2[-32] intron 1; 2197/C733R and 3020/Q1007L) were screened for the study population (431 adult cows). The statistical analysis resulted in significant differences in allelic frequencies between cases and controls for SNP2197/C733R (P<0.001), indicating a significant association between infection and variant allele. In the analysis of genotypes, a significant association was also found between SNP2197/C733R and infection status (P<0.0001); cows with the heterozygous genotype were 3.35 times more likely to be infected than cows with the reference genotype (P=0.01). Results suggest a role for CARD15 gene in the susceptibility of cattle to paratuberculosis infection. These data contribute to the understanding of paratuberculosis, suggest new similarities with Crohns disease and provide new information for the control of bovine paratuberculosis.
The lupus-prone NZM2410 mice present an expanded B1a cell population that we have mapped to the Sle2c1 lupus susceptibility locus. The expression of Cdkn2c, a gene encoding for cyclin-dependent kinase inhibitor p18(Ink4c) and located within Sle2c1, is significantly lower in B6.Sle2c1 B cells than in B6 B cells. To test the hypothesis that the B1a cell expansion in B6.Sle2c1 mice was due to a defective p18 expression, we analyzed the B1a cell phenotypes of p18-deficient C57BL/6 mice. We found a dose-dependent negative correlation between the number of B1a cells and p18 expression in B cells, with p18-deficient mice showing an early expansion of the peritoneal B1a cell pool. p18 deficiency enhanced the homeostatic expansion of B1a cells but not of splenic conventional B cells, and the elevated number of B6.Sle2c1 B1a cells was normalized by cyclin D2 deficiency. These data demonstrated that p18 is a key regulator of the size of the B1a cell pool. B6.p18(-/-) mice produced significant amounts of anti-DNA IgM and IgG, indicating that p18 deficiency contributes to humoral autoimmunity. Finally, we have shown that Sle2c1 increases lpr-associated lymphadenopathy and T cell-mediated pathology. B6.p18(-/-).lpr mice showed a greater lymphadenopathy than B6.Sle2c1.lpr mice, but their renal pathology was intermediate between that of B6.lpr and B6.Sle2c1.lpr mice. This indicated that p18-deficiency synergizes, at least partially, with lpr-mediated pathology. These results show that Cdkn2c contributes to lupus susceptibility by regulating the size of the B1a cell compartment and hence their contribution to autoimmunity.
Sle1c is a sublocus of the NZM2410-derived Sle1 major lupus susceptibility locus. We have shown previously that Sle1c contributes to lupus pathogenesis by conferring increased CD4(+) T cell activation and increased susceptibility to chronic graft-versus-host disease (cGVHD), which mapped to the centromeric portion of the locus. In this study, we have refined the centromeric sublocus to a 675-kb interval, termed Sle1c2. Mice from recombinant congenic strains expressing Sle1c2 exhibited increased CD4(+) T cell intrinsic activation and cGVHD susceptibility, similar to mice with the parental Sle1c. In addition, B6.Sle1c2 mice displayed a robust expansion of IFN-?-expressing T cells. NZB complementation studies showed that Sle1c2 expression exacerbated B cell activation, autoantibody production, and renal pathology, verifying that Sle1c2 contributes to lupus pathogenesis. The Sle1c2 interval contains two genes, only one of which, Esrrg, is expressed in T cells. B6.Sle1c2 CD4(+) T cells expressed less Esrrg than B6 CD4(+) T cells, and Esrrg expression was correlated negatively with CD4(+) T cell activation. Esrrg encodes an orphan nuclear receptor that regulates oxidative metabolism and mitochondrial functions. In accordance with reduced Esrrg expression, B6.Sle1c2 CD4(+) T cells present reduced mitochondrial mass and altered mitochondrial functions as well as altered metabolic pathway utilization when compared with B6 CD4(+) T cells. Taken together, we propose Esrrg as a novel lupus susceptibility gene regulating CD4(+) T cell function through their mitochondrial metabolism.
Considerable efforts have been deployed over the years to decipher the genetic basis of systemic lupus erythematosus (SLE). The NZM2410 strain is murine model in which the genetic analysis of SLE is the most advanced. NZM2410 studies have shown that, as in SLE patients, lupus susceptibility is achieved by the coexpression of many susceptibility alleles, each of which with a small contribution to the overall disease phenotype. This mouse model has also revealed the critical role played by gene-gene interactions, which are believed to be an essential contribution to human SLE heritability, although it has been much more difficult to characterize. We have now reached a phase in which NZM2410 susceptibility genes have been identified, all them novel in their association with lupus or even with immune functions. Ongoing studies geared at understanding how these genes impact immune tolerance and interact with each other in the mouse, and their impact on the human immune system or target organs, will undoubtedly lead to important discovery for a better understanding on the disease and potential identification of therapeutic targets.
We have previously shown that a novel -74 C-to-T mutation in the promoter of the cyclin-dependent kinase inhibitor p18(Ink4c) (p18) gene was associated with a reduced p18 expression in B cells from mice carrying the Sle2c1 lupus susceptibility locus. To determine the function of the -74 C/T single nucleotide polymorphism, we have characterized the proximal promoter of the mouse p18 gene. Functional analysis of the 5 flanking region by sequential deletions revealed crucial elements between -300 and +1, confirming the in silico prediction that the -74 T allele created a novel Yin-Yang 1 (YY-1) binding site adjacent to an existing one common to both alleles. Moreover, we found that YY-1, E2F1, and Sp-1 can synergistically enhance the activity of the p18 promoter. Mutational inactivation revealed that YY-1 binding regulates the p18 activity in an allele-dependent fashion. EMSAs with splenic B cell extracts directly demonstrated that YY-1 binds to the p18 promoter with differences between the C and the T alleles. We also determined in vivo by chromatin immunoprecipitation that the T allele resulted in increased YY-1 and decreased Nrf-2 binding to the p18 promoter as compared with the C allele in B cells. Thus, YY-1 is a direct regulator of p18 gene expression in an allele-dependent fashion that is consistent with the lupus-associated T allele, inducing a lower p18 transcriptional activity by increasing YY-1 binding. These results establish the p18 -74 C/T mutation as the leading causal variant for the B1a cell expansion that characterizes the NZB and NZM2410 lupus-prone strains.
CSF-1, required for macrophage (Mø) survival, proliferation, and activation, is upregulated in the tubular epithelial cells (TECs) during kidney inflammation. CSF-1 mediates Mø-dependent destruction in lupus-susceptible mice with nephritis and, paradoxically, Mø-dependent renal repair in lupus-resistant mice after transient ischemia/reperfusion injury (I/R). We now report that I/R leads to defective renal repair, nonresolving inflammation, and, in turn, early-onset lupus nephritis in preclinical MRL/MpJ-Faslpr/Fas(lpr) mice (MRL-Fas(lpr) mice). Moreover, defective renal repair is not unique to MRL-Fas(lpr) mice, as flawed healing is a feature of other lupus-susceptible mice (Sle 123) and MRL mice without the Fas(lpr) mutation. Increasing CSF-1 hastens renal healing after I/R in lupus-resistant mice but hinders healing, exacerbates nonresolving inflammation, and triggers more severe early-onset lupus nephritis in MRL-Fas(lpr) mice. Probing further, the time-related balance of M1 "destroyer" Mø shifts toward the M2 "healer" phenotype in lupus-resistant mice after I/R, but M1 Mø continue to dominate in MRL-Fas(lpr) mice. Moreover, hypoxic TECs release mediators, including CSF-1, that are responsible for stimulating the expansion of M1 Mø inherently poised to destroy the kidney in MRL-Fas(lpr) mice. In conclusion, I/R induces CSF-1 in injured TECs that expands aberrant Mø (M1 phenotype), mediating defective renal repair and nonresolving inflammation, and thereby hastens the onset of lupus nephritis.
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