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Systemic lupus erythematosus (SLE) is a multiorgan autoimmune disease associated with impaired immune system regulation. The exact mechanisms of SLE development remain to be elucidated. TAM receptor tyrosine kinases (RTKs) are important for apoptotic cell clearance, immune homeostasis, and resolution of immune responses. TAM deficiency leads to lupus-like autoimmune diseases. Activation of TAM receptors leads to proteolytic cleavage of the receptors, generating soluble forms of TAM. Circulating TAM receptors have an immunoregulatory function and may also serve as biomarkers for disease prognosis. Here, we review the biological function and signaling of TAM RTKs in the development and pathogenesis of lupus and lupus nephritis. Targeting Gas6/TAM pathways may be of therapeutic benefit. A discussion of potential TAM activation and inhibition in the treatment of lupus and lupus nephritis is included.

The unique cytokine signature of COVID-19 might provide clues to disease mechanisms and possible future therapies. Here, we propose a pathogenic model in which the alarmin cytokine, interleukin (IL)-33, is a key player in driving all stages of COVID-19 disease (ie, asymptomatic, mild–moderate, severe–critical, and chronic–fibrotic). In susceptible individuals, IL-33 release by damaged lower respiratory cells might induce dysregulated GATA-binding factor 3-expressing regulatory T cells, thereby breaking immune tolerance and eliciting severe acute respiratory syndrome coronavirus 2-induced autoinflammatory lung disease. Such disease might be initially sustained by IL-33-differentiated type-2 innate lymphoid cells and locally expanded γδ T cells. In severe COVID-19 cases, the IL-33–ST2 axis might act to expand the number of pathogenic granulocyte–macrophage colony-stimulating factor-expressing T cells, dampen antiviral interferon responses, elicit hyperinflammation, and favour thromboses. In patients who survive severe COVID-19, IL-33 might drive pulmonary fibrosis by inducing myofibroblasts and epithelial–mesenchymal transition. We discuss the therapeutic implications of these hypothetical pathways, including use of therapies that target IL-33 (eg, anti-ST2), T helper 17-like γδ T cells, immune cell homing, and cytokine balance.

Autoantibodies in systemic autoimmunity are directed against only ~5% of the proteome. The purpose of this study was to assess whether the properties of assemblages (also known as Membraneless Organelles and Biological Condensates) and their protein constituents partly explain the immunological selectivity of autoimmunity. Assemblages arise from phase separation of their protein components, akin to partitioning of oil droplets in water. We obtained from a prediction algorithm (Vernon et al., elife 7, 2018) the propensity scores (PScores), i.e., likelihood, for phase separation of autoantigens and non-autoantigens. We then compared autoantigens with the highest PScores to identify shared structural properties. The mean PScores for autoantigens (n = 1050) and the entire human proteome of non-autoantigens (n = 17,532) were 1.46 and 1.09 (p = 1.2E-08). To varying extents, the 25 autoantigens with the highest phase separation propensities shared additional features such as compositional bias, repeated domains, coiled coil regions, nucleic acid binding, and disorder. Most of these properties were present with greater frequencies than their frequencies in the non-autoantigens. We conclude that, on average, autoantigens have a higher predisposition to undergo phase separation, thus, they are more likely to exist in assemblages compared with the average non-autoantigen. We suggest that assemblage formation and the greater than average presence of certain structural features are key factors in selection of a portion of the autoimmune repertoire. Other properties of assemblage proteins, such as high concentration and tendency to form novel complexes with other proteins, may partially explain why assemblages are potent sources of autoantigens.

Programmed cell death is a complex biological process involving multiple signaling pathways. Autoimmunity can result from defects in the multiple stages of the process of apoptosis in immunocompetent cells, ranging from mutant death receptors and ligands to biochemical alterations in death-inducing signaling. Ineffective recognition and clearance of apoptotic cells leading to increased numbers of apoptotic and necrotic cells can also cause autoimmunity. In this chapter, we review the mechanisms of apoptosis and the means by which abnormalities in this process can lead to self-immunization and to the development of autoimmunity. The induction of autoimmunity by receptors that recognize the nucleic acids derived from dead and dying cells is a critical pathway in self-immunization, and the immune complexes containing apoptotic cells may amplify and perpetuate existing autoimmunity.

Mer receptor tyrosine kinase (MerTK) is key for efficient phagocytosis of apoptotic neutrophils (ANs) and homeostasis of IL-10 production by human anti-inflammatory M2c monocytes/macrophages. We asked whether stimulation of M2c surface receptors contributes in turn to MerTK activation. For this purpose, human monocytes/macrophages were differentiated under M1, M2a, and M2c polarizing conditions. The effects of antibody-mediated cross-linking of M2c receptors (i.e., CD14, CD16, CD32, CD163, CD204) on MerTK phosphorylation and phagocytosis of ANs were tested. MerTK expression was also studied by flow cytometry and western blot in the presence of LPS and in M2c-derived microvesicles (MVs). Antibody cross-linking of either CD14 or CD32/FcγRII led to Syk activation and MerTK phosphorylation in its two distinct glycoforms (175–205 and 135–155 kDa). Cross-linked CD14 enhanced efferocytosis by M2c macrophages and enabled M1 and M2a cells to clear ANs efficiently. In M1 conditions, LPS abolished surface MerTK expression on CD14bright cell subsets, so disrupting the anti-inflammatory pathway. In M2c cells, instead, MerTK was diffusely and brightly co-expressed with CD14, and was also detected in M2c macrophage-derived MVs; in these conditions, LPS only partially downregulated MerTK on cell surfaces, while the smaller MerTK glycoform contained in MVs remained intact. Altogether, cooperation between CD14 and MerTK may foster the clearance of ANs by human monocytes/macrophages. CD14 stands between M1-related LPS co-receptor activity and M2c-related MerTK-dependent response. MerTK interaction with CD32/FcγRII, its detection in M2c MVs, and the differential localization and LPS susceptibility of MerTK glycoforms add further new elements to the complexity of the MerTK network.

Sjögren's Syndrome (SjS) is a chronic, systemic autoimmune disease causing xerostomia, xerophthalmia, and systemic symptoms. The principal pathological finding in SjS is the accumulation of lymphocytes in exocrine glandular tissue and elsewhere, leading to secretory dysfunction and other abnormalities. A rational therapeutic approach might be to interfere with lymphocyte migration to the periphery from central lymphoid tissues. We thus examined in an animal model of SjS the effects of Fingolimod (FTY720, Gilenya™), which interferes with migration of lymphocytes to peripheral sites. Fingolimod induces sequestration of lymphocytes in lymphoid organs by altering lymphocyte expression of sphingosine-1-phosphate receptors. In the C57Bl/6. NOD.Aec1Aec2 (AEC) model of SjS, Fingolimod reduced circulating T and B cell numbers. Treatment of AEC mice with Fingolimod increased salivary output and decreased the size of salivary gland infiltrates. Oral Fingolimod thus merits further consideration in the management of SjS in humans.

Background Mer receptor tyrosine kinase (MerTK) is key for efficient phagocytosis of apoptotic neutrophils (ANs) and homeostasis of IL-10 production by human anti-inflammatory M2c monocytes/macrophages. We asked whether stimulation of certain M2c surface receptors contribute in turn to MerTK activation. Methods Human monocytes/macrophages were differentiated under M1, M2a and M2c polarizing conditions. We tested the effects of antibody-mediated cross-linking of M2c receptors (i.e., CD14, CD16, CD32, CD163, CD204) on MerTK phosphorylation and phagocytosis of ANs. MerTK expression was also studied by flow cytometry and western blot in the presence of LPS and in M2c-derived microvesicles (MVs). Results Antibody cross-linking of either CD14 or CD32/FcγRII led to Syk activation and MerTK phosphorylation in its two distinct glycoforms (175–205 and 135–155 KDa). Cross-linked CD14 enhanced efferocytosis by M2c macrophages and enabled M1 and M2a cells to clear ANs efficiently. In M1 conditions, LPS abolished surface MerTK expression on CD14bright cell subsets, so disrupting the anti-inflammatory pathway. In M2c cells, instead, MerTK was diffusely and brightly co-expressed with CD14, and was also detected in M2c macrophage-derived MVs; in these conditions, LPS only partially down-regulated MerTK on the cell surface, while the smaller MerTK glycoform contained in MVs remained intact. Conclusions Cooperation between CD14 and MerTK fosters tethering and engulfment of ANs by human monocytes/macrophages. CD14 stands between M1-related LPS co-receptor activity and M2c-related MerTK-dependent responses. MerTK interaction with CD32/FcγRII, its detection in M2c MVs, and the differential localization and LPS susceptibility of MerTK glycoforms add further new elements to the complexity of the MerTK network. Acknowledgements NIAID and Lupus Research Alliance

Systemic lupus erythematosus (SLE) is a complex multisystem autoimmune disease, characterized by a spectrum of autoantibodies that target multiple cellular components. Glomerulonephritis is a major cause of morbidity in patients with SLE. Little is known about the pathogenesis of SLE renal damage and compromised renal function. Activation of both Stat1 and Stat3 has been reported in lupus and lupus nephritis. The reciprocal activation of these two transcription factors may have a major impact on renal inflammation. To study the role of Stat1 in a lupus model, we induced lupus-like chronic graft-versus-host disease (cGVHD) in Stat1-knockout (KO) and wild-type (WT) mice by i.p. injection of class II–disparate bm12 splenocytes. WT recipients of these alloreactive cells developed anti-dsDNA autoantibodies starting at week 2 as expected, with a decline after week 4. In contrast, Stat1-KO hosts exhibited a prolonged and significant increase of anti-dsDNA autoantibody responses compared with WT mice (week 4 to week 8). Increased autoantibody titers were accompanied by increased proteinuria and mortality in the cGVHD host mice lacking Stat1. Further analysis revealed expression and activation of Stat3 in the glomeruli of Stat1-KO host mice but not WT mice with cGVHD. Glomerular Stat3 activity in the Stat1-KO mice was associated with increased IL-6 and IFN-γ secretion and macrophage infiltration. Interactions between Stat1 and Stat3 thus appear to be crucial in determining the severity of lupus-like disease in the cGVHD model.

The Mertk receptor tyrosine kinase facilitates macrophage and DC apoptotic-cell clearance and regulates immune tolerance. Mertk may also contribute to B-cell activation, because Mertk-KO mice fail to develop autoantibodies when allo-activated by T cells. We investigated this possibility with a well-characterized model in which injection of mice with goat anti-IgD antibody causes membrane IgD cross-linking that induces T-independent B cell activation and antigen presentation to T cells. Goat anti-mouse IgD antibody-injected C57BL/6 Mertk-KO mice had normal initial B cell activation and proliferation, but significantly lower T cell activation and proliferation, as well as lower IgE and IgG anti-goat IgG responses, as compared to C57BL/6 WT controls. B cell antigen processing, analyzed by evaluating B cell fluorescence following injection of monoclonal anti-IgD antibody labeled with biotin or FITC, was comparable between Mertk-KO mice and WT mice. IgD Ab primed B cells from Mertk-KO mice exhibited significantly lower ability in activating memory T cells isolated from WT mice injected with the same antigen 10 days before. These observations suggest that Mertk expression is required for optimal B-cell antigen presentation, which is, in turn, required in this model for optimal T cell activation and subsequent T cell-dependent B cell differentiation.

Review question/objective The overall research question for this systematic review is: what is the cost effectiveness of Mohs micrographic surgery for non-melanoma skin cancer? More specifically, the objectives are to: Examine the evidence on the cost effectiveness, cost benefit, cost minimization, and cost utility of Mohs micrographic surgery, compared to other surgical/ablative interventions such as excisional surgery and radiation therapy, as well as non-surgical/chemotherapeutic interventions such as topical 5-fluorouracil and imiquimod immunotherapy, for non-melanoma skin cancer clinical care outcomes; Determine the cost effectiveness, cost benefit, cost minimization and cost utility of Mohs micrographic surgery from the patient, provider, healthcare system and societal perspectives in order to elucidate the economic and clinical benefits of the procedure at these various levels; Provide a comprehensive assessment of the relative economic and clinical effectiveness of this procedure for patients with non-melanoma skin cancer in different contexts globally. Inclusion criteria Types of participants This review will consider studies that include participants with an established diagnosis of non-melanoma skin cancer. For the purposes of this review, an established diagnosis of non-melanoma skin cancer will consist of either squamous cell carcinoma or basal cell carcinoma , confirmed histologically or clinically or both, by a healthcare provider trained to do so. Types of intervention(s)/phenomena of interest This review will consider studies that evaluate Mohs micrographic surgery as a treatment for non-melanoma skin cancer. Types of outcomes This review will consider studies that measured any clinical care outcomes related to non-melanoma skin cancer, including but not limited to mortality, quality of life, quality-adjusted life years , incidence and recurrence of non-melanoma skin cancer lesions, and frequency of adverse events. However, to be considered for inclusion, studies must also provide an economic evaluation of the relative costs of using Mohs micrographic surgery as a therapeutic modality when compared to other known treatment modalities.

Exposure of human monocytes/macrophages to anti-inflammatory agents, such as IL-10 or glucocorticoids, can lead to two separate fates: either Fas/CD95-mediated apoptosis or differentiation into regulatory and efferocytic M2c (CD14brightCD16+CD163+Mer tyrosine kinase+) macrophages. We found that the prevalent effect depends on the type of Th cytokine environment and on the stage of monocyte-to-macrophage differentiation. In particular, the presence of IFN-γ (Th1 inflammation) or the prolonged exposure to IL-4 (chronic Th2 inflammation) promotes apoptosis of monocytes/macrophages and causes resistance to M2c differentiation, thus provoking impaired clearance of apoptotic neutrophils, uncontrolled accumulation of apoptotic cells, and persistent inflammation. In contrast, the presence of IL-17 (Th17 environment) prevents monocyte/macrophage apoptosis and elicits intense M2c differentiation, thus ensuring efficient clearance of apoptotic neutrophils and restoration of anti-inflammatory conditions. Additionally, the Th environment affects the expression of two distinct Mer tyrosine kinase isoforms: IL-4 downregulates the membrane isoform but induces an intracellular and Gas6-dependent isoform, whereas IFN-γ downregulates both and IL-17 upregulates both. Our data support an unexpected role for IL-17 in orchestrating resolution of innate inflammation, whereas IFN-γ and IL-4 emerge as major determinants of IL-10 and glucocorticoid resistance.

Objectives Hyperplasia of synovial fibroblasts, infiltration with lymphocytes and tissue hypoxia are major characteristics of rheumatoid arthritis (RA). Extensive data support a key role for toll-like receptors (TLRs) in RA. Little is known regarding the impact of hypoxia on TLR-induced inflammation in RA. The aim of this study was to reveal the effects of hypoxia and its regulator, hypoxia-inducible factor-1α (HIF-1α), on the inflammatory response of RA synovial fibroblasts (RASF) to TLR ligands. Methods Hypoxia was induced in RASF by incubation with Na 2 S 2 O 4 . TLR3 ligand polyIC, TLR2 ligand peptidoglycan, TLR4 ligand LPS and TLR9 ligand CpG were used to stimulate the cells. Effects of hypoxia on TLR-induced inflammatory mediators were determined by RT-PCR, qPCR and ELISA. Overexpression of HIF-1α as well as knocking-down its expression was used to reveal its fundamental role. RASF-induced inflammatory T cell expansion was determined by flow cytometry analysis of T helper (Th)1/Th17 cells, and IFN-γ/IL-17 production by ELISA after RASF/T cell coculture. Results Hypoxia potentiated the expression of inflammatory cytokines, metalloproteinases and VEGF in RASF stimulated by different TLR ligands, especially polyIC, a synthetic mimic of dsRNA from viruses or apoptotic cells. HIF-1α played a fundamental role in this synergy. Moreover, HIF-1α overexpression enhanced RASF-mediated expansion of inflammatory Th1 and Th17 cells, leading to proinflammatory IFN-γ and IL-17 production. Conclusions Our findings suggest that hypoxia and HIF-1α may function in conjunction with TLR-stimulated innate immune responses to drive inflammation in RA. This pathway may serve as a therapeutic target for the disease.

Objective To study the safety and clinical efficacy of rituximab therapy for primary Sjogren's syndrome, as well as to investigate its mechanisms. Methods Patients with primary Sjogren's syndrome were enrolled in an open-label trial, were given rituximab (1 gm) infusions on days 1 and 15, and were monitored through week 52. The primary end point was safety, with secondary end points evaluating clinical and biologic efficacy. Blood was obtained for enumeration of lymphocyte subsets, measurement of serum autoantibody and BAFF levels, and analysis of gene expression. Results Twelve female patients with primary Sjogren's syndrome were administered rituximab. They had a median age of 51 years (range 34–69 years) and a median disease duration of 8.0 years (range 2–18 years). We observed no unexpected toxicities from the rituximab therapy. Modest improvements were observed at week 26 in patient-reported symptoms of fatigue and oral dryness, with no significant improvement in the objective measures of lacrimal and salivary gland function. The recovery of blood B cells following the nadir from rituximab therapy was characterized by a predominance of transitional B cells and a lack of memory B cells. While blood B cell depletion was associated with an increase in serum BAFF levels, no significant changes were observed in the levels of serum anti-Ro/SSA, anti-La/SSB, and anti–type 3 muscarinic acetylcholine receptor autoantibodies or in the blood interferon signature. Conclusion In patients with primary Sjogren's syndrome, a single treatment course of rituximab was not associated with any unexpected toxicities and led to only modest clinical benefits despite effective depletion of blood B cells.

Activation and migration of marginal zone B (MZB) cells into follicular (FO) regions of the spleen has been proposed as one of the mechanisms that regulate the development of autoreactive B cells. The mer receptor tyrosine kinase (Mertk) mediates apoptotic cell clearance and regulates activation and cytokine secretion. In the well-studied class II chronic GVH model of bm12 cells into B6 hosts, we observed that Mertk deficient B6 mice did not generate autoantibodies in response to this allogeneic stimulus. We posited that Mertk is important in MHC-II-mediated B cell signaling. In the present study, we show that B cells from Mertk(-/-) mice but not WT B6 mice exhibited decreased calcium mobilization and tyrosine phosphorylation when stimulated by MHC-II cross-linking. The finding that Mertk was important for class II signaling in B cells was further supported by the preponderance of a-allotype autoantibodies in cGVH in RAG-KO mice reconstituted with a mixture of bone marrow from Mertk(-/-) mice (b-allotype) and C20 mice (a-allotype). MZB cells from Mertk(-/-) mice were unable to down regulate surface CD1d expression and subsequent inclusion in the MZ, associated with significantly lower germinal center responses compared to MZB cells from WT. Moreover, Mertk(-/-) mice treated with an anti-CD1d down regulating antibody responded significantly to bm12 cells, while no response was observed in Mertk(-/-) mice treated with control antibodies. Taken together, these findings extend the role of Mertk to include CD1d down regulation on MZB cells, a potential mechanism limiting B cell activation in cGVH.

Objective Increased levels of type I interferon (IFN) and type I IFN–regulated genes are found in patients with systemic lupus erythematosus (SLE) and may be central to its pathogenesis. Mitochondrial antiviral signaling protein (MAVS) is a key regulator of type I IFN that undergoes a dramatic prion-like aggregation and self propagates the activation signal from viral RNA to amplify downstream IFN production. We undertook this study to determine whether such MAVS aggregates might play a role in the sustained increased production of type I IFN in SLE. Methods Peripheral blood mononuclear cells were isolated and mitochondrial extracts were prepared. MAVS aggregation was detected by semidenatured agarose gel electrophoresis and confirmed by immunofluorescence staining. MAVS-associated signaling proteins were analyzed by Western blotting. MAVS aggregation–associated gene expression signature was analyzed by microarray. Results In blood cells from 22 of 67 SLE patients, essentially all MAVS was in a high molecular weight aggregated form. None of 6 rheumatoid arthritis patients and only 3 of 33 healthy controls had abnormal MAVS. Compared to MAVS aggregate–negative patients, MAVS aggregate–positive SLE patients had significantly higher serum levels of IFNβ and significantly increased levels of autoantibodies against Sm and U1 RNP. Gene array data revealed a characteristic gene expression pattern in these patients, with altered expression of genes involved in IFN signaling and membrane trafficking. Conclusion Persistent MAVS aggregates may lead to increased type I IFN production and result in unmitigated signals leading to autoimmunity.

Mer tyrosine kinase (MerTK) is a major macrophage apoptotic cell (AC) receptor. Its functional impairment promotes autoimmunity and atherosclerosis, whereas overexpression correlates with poor prognosis in cancer. However, little is known about mechanisms regulating MerTK expression in humans. We found that MerTK expression is heterogenous among macrophage subsets, being mostly restricted to anti-inflammatory M2c (CD14+CD16+CD163+CD204+CD206+CD209−) cells, differentiated by M-CSF or glucocorticoids. Small numbers of MerTK+ “M2c-like” cells are also detectable among circulating CD14brightCD16+ monocytes. MerTK expression levels adapt to changing immunologic environment, being suppressed in M1 and M2a macrophages and in dendritic cells. Remarkably, although glucocorticoid-induced differentiation is IL-10 independent, M-CSF–driven M2c polarization and related MerTK upregulation require IL-10. However, neither IL-10 alone nor TGF-β are sufficient to fully differentiate M2c (CD16+CD163+MerTK+) macrophages. M-CSF and IL-10, both released by T lymphocytes, may thus be required together to promote regulatory T cell–mediated induction of anti-inflammatory monocytes-macrophages. MerTK enables M2c macrophages to clear early ACs more efficiently than other macrophage subsets, and it mediates AC clearance by CD14brightCD16+ monocytes. Moreover, M2c cells release Gas6, which in turn amplifies IL-10 secretion via MerTK. IL-10–dependent induction of the Gas6/MerTK pathway may, therefore, constitute a positive loop for M2c macrophage homeostasis and a critical checkpoint for maintenance of anti-inflammatory conditions. Our findings give new insight into human macrophage polarization and favor a central role for MerTK in regulation of macrophage functions. Eliciting M2c polarization can have therapeutic utility for diseases such as lupus, in which a defective AC clearance contributes to initiate and perpetuate the pathological process.

Sjogren's syndrome (SS) is characterized by infiltration of exocrine glands with T and B lymphocytes, leading to glandular dysfunction and frequently accompanied by hypergammaglobulinemia and autoantibodies. The role of T cells, which predominate in the lesions, has attracted much interest. CD4 T cells seem to be responding to autoantigens on apoptotic cells, such as the Ro and La antigens, or to the cytoskeletal antigen α-fodrin. Physical injury to ocular surfaces may also lead to T cell mediated responses to self antigens and perpetuate disease. Within the salivary glands, T cell responsiveness is further promoted by the special capacity of salivary epithelial tissue to provide costimulation and enhanced antigen presentation. Cytokines are key mediators of the T cell contribution to pathology, with roles attributed both to Th1 and Th2 cells. Recently, striking data implicate Th17 cells in the stimulation of B cells, and a role for the related cytokine IL-21 produced by follicular T helper cells is now appreciated. Dysfunction of T regulatory cells has been shown to have a role in the exuberant production of cytokines by Th17 cells. Beyond their role in provoking B cell hyperactivity and immunoglobulin secretion, T cells are directly involved in destruction of glands through Fas and perforin-mediated cytotoxicity. Animal models of SS have confirmed the role of T cell derived cytokines in disease and support a role for effector-memory cells in pathogenesis. Further elucidation of the role of T cells will open avenues for better treatment of SS, whose current management is still mainly supportive.

Primary Sjögren's syndrome (SjS) is an autoimmune disorder characterized by lymphocytic infiltration of the salivary and lacrimal glands, B-cell clonal expansions and an increased risk of lymphoma. In order to understand the role of B cells in this disorder, the antibody repertoire and B-cell maturation were studied in a mouse model of SjS called B6.Aec1/2.B6.Aec1/2 serum was analyzed for antibodies by ELISA and immunoprecipitation, B-cell development by flow cytometry, and antibody gene rearrangements by CDR3 spectratyping and quantitative PCR. In order to test the functional consequences of the observed defects, B6.Aec1/2 mice were crossed with anti-dsDNA antibody heavy chain knock-in mice (B6.56R).B6.Aec1/2 mice exhibit B-cell clonal expansions, have altered serum immunoglobulin levels and spontaneously produce multireactive autoantibodies. B6.Aec1/2 mice also have decreased numbers of bone marrow pre-B cells and decreased frequencies of kappa light chain gene deletion. These findings suggest that B6.Aec1/2 mice have a defective early B-cell tolerance checkpoint. B6.56R.Aec1/2 mice unexpectedly had lower anti-dsDNA antibody levels than B6.56R mice and less salivary gland infiltration than B6.Aec1/2 mice.These data suggest that the early tolerance checkpoint defect in B6.Aec1/2 mice is not sufficient to promulgate disease in mice with pre-formed autoantibodies, such as B6.56R. Rather, B6.Aec1/2 mice may require a diverse B-cell repertoire for efficient T-B-cell collaboration and disease propagation. These findings imply that therapies aimed at reducing B-cell diversity or T-B interactions may be helpful in treating SjS.

a b s t r a c t A potential link between obesity, circulating leptin levels and autoimmune disease symptoms sug- gests that targeting the leptin receptor (ObR) might be a viable novel strategy to combat rheumatoid arthritis (RA). However, studies in animal models and evaluation of clinical cases did not pro- vide clear view on leptin's involvement in RA. To validate ObR as RA target, we used our peptide -based ObR agonists and antagonist in different in vitro and in vivo models of the disease. In human peripheral blood mononuclear cells, leptin and its agonist fragment, desI2-E1/Aca, moderately induced constitutive activation of a major proinflammatory transcription factor, NF-B, while the ObR antago- nist peptide Allo-aca inhibited the process. Leptin administration itself did not induce arthritis in rats, but worsened the clinical condition of mice given K/BxN serum transfer arthritis. Simultaneous admin- istration of Allo-aca reduced leptin-dependent increase in disease severity by more than 50%, but the antagonist was ineffective when injected with a 3-day delay. In rats inflicted with mild adjuvant-induced arthritis, both leptin and Allo-aca reduced the extent of joint swelling and the number of arthritic joints. In a more aggressive disease stage, Allo-aca decreased the number of arthritic joints in a dose-dependent manner but did not affect other arthritis markers. In summary, leptin exerts diverse effects on RA depend- ing on the experimental model. This might reflect the heterogeneous character of RA, which is differently impacted by leptin and is unmasked by ObR antagonism. Nevertheless, the results suggest that ObR antagonists might become useful therapeutics in leptin-sensitive early stages of RA.

The Mer receptor tyrosine kinase is strongly expressed in the glomerulus. We wondered if this molecule might modify immune-mediated glomerular disease through its functions as a receptor for apoptotic cells and immunoregulatory molecule. Mer-knockout (KO) mice showed decreased survival rate and greatly increased proteinuria and serum urea levels compared to wild type (WT) mice by day 3 after injection of NTS. Their glomeruli were hyperplastic and later became necrotic. While in the glomerulus of WT mice, a significant increase of Mer expression was observed. Apoptotic bodies were evident in NTS-treated Mer-KO kidneys, but not in normal controls. NTS-treated Mer-KO mice had massive neutrophil infiltration and inflammatory cytokine expression. Mer thus has a critical role in attenuating renal inflammation, both as a receptor for apoptotic cells and as a molecule that down regulates inflammation.

Systemic lupus erythematosus is characterized by loss of tolerance to DNA and other nuclear Ags. To understand the role of T cells in the breaking of tolerance, an anti-DNA site-specific transgenic model of spontaneous lupus, B6.56R, was studied. T cells were eliminated by crossing B6.56R with CD4−/− or TCRβ−/−δ−/− mice, and the effects on anti-dsDNA serum levels, numbers of anti-dsDNA Ab-secreting cells, and isotypes of anti-dsDNA were analyzed. In addition, the development and activation of B cells in these mice were examined. Surprisingly, the presence of T cells made little difference in the development and character of the serum anti-dsDNA Ab in B6.56R mice. At 1 mo of age, anti-dsDNA Abs were somewhat lower in mice deficient in αβ and γδ T cells. Levels of Abs later were not affected by T cells, nor was autoantibody class switching. B cell activation was somewhat diminished in T cell-deficient mice. Thus, in the B6 background, the presence of an anti-dsDNA transgene led the production of autoantibodies with a specificity and isotype characteristic of murine systemic lupus erythematosus with little influence from T cells. TLR9 also did not appear to play a role. Although we do not yet understand the mechanism of this failure of immunoregulation, these results suggest that similar processes may influence autoimmunity associated with clinical disease.

The Mer receptor tyrosine kinase mediates apoptotic cell phagocytosis and modulates macrophage cytokine production. Mer−/− mice have defective clearance of apoptotic debris and develop a systemic lupus erythematosus-like autoimmune syndrome. It was surprising then that B6-Mer−/− recipients of bm12 spleen cells failed to develop anti-dsDNA and anti-chromatin autoantibodies, whereas B6 hosts produced the expected autoimmune chronic graft-vs-host (cGVH) reaction. The lack of autoantibody formation in cGVH was not due to the failure of Mer-deficient hosts to provoke alloreactivity, because Mer−/− spleen cells were recognized by bm12 T cells in MLR. Cell transfer experiments in Rag-knockout mice indicated that the lack of autoantibody production in Mer−/− cGVH disease hosts was due to an intrinsic B cell defect. This defect did not cause a global inability to produce autoantibodies, because in vivo exposure to LPS stimulated production of autoantibodies in both B6 and Mer−/− mice. We further observed that wild-type B6 B cells up-regulated Mer upon activation in cGVH, and that B cells from mice lacking Mer showed a decreased up-regulation of activation-associated cell surface markers. These findings indicate that Mer serves an important role in the activation of self-reactive B cells in systemic autoimmunity.

Chronic graft-vs-host (cGVH) disease is a well-characterized systemic lupus erythematosus (SLE) model. Induction of cGVH in anti-DNA H chain knockin (3H9KI) transgenic mice results in specific activation of anti-dsDNA B cells. In this study, we show that B cells from 3H9KI mice were activated by cGVH even when adoptively transferred into irradiated JHT−/− recipients that lack endogenous B cells. This process of activation was reflected by high autoantibody titers and changes in phenotypic markers. We have used this system to characterize the particular B cell subsets that were responsible for secreting autoantibodies during cGVH response. We isolated splenic B cell subsets based on their expression of specific cell surface markers and used them in our adoptive transfer studies. We found that mature B cells were the most vulnerable to the allostimulus and were the major source of autoantibodies compared with immature B cells. The greater susceptibility of mature B cells to become activated and thereby lose tolerance was unanticipated and has implications for maintenance of peripheral tolerance and for the development of autoimmunity. Furthermore, of the mature B cells, marginal zone B cells were particularly responsible for mounting the initial response to the cGVH stimulus. This observation underscores the critical role of marginal zone B cells in activation and production of autoantibodies.

I read with interest the article by M.S. Kim and T.S. Kim ([1][1]), in which the authors demonstrate that AIMP1 activates B cells and promotes Ag-specific production of Ab. Certain patients suffer from antisynthetase syndrome, an illness characterized by muscle and skin inflammation accompanied by

Chronic graft-vs-host (cGVH) disease is induced in nonautoimmune mice by the transfer of alloreactive T cells that recognize foreign MHC class II. It closely resembles systemic lupus erythematosus, with antinuclear Abs and immune-mediated nephritis. Recent work has implicated TLRs, particularly TLR9, in the recognition of certain autoantigens in vitro and in vivo. To explore further the role of TLR9 in systemic autoimmunity, we induced cGVH disease in C57BL/6 (B6) mice lacking TLR9, including B6 mice expressing the anti-DNA-encoding IgH transgenes 3H9 or 56R (B6.3H9.TLR9−/−, B6.56R.TLR9−/−). We found that cGVH disease caused breakdown of B cell tolerance to chromatin and DNA in TLR9−/− recipients of alloreactive cells, yet that nephritis was less severe and that some autoantibody titers were lower compared with B6-cGVH controls. Spleen lymphocyte analysis showed that cGVH disease strikingly depleted marginal zone B cells in B6 mice, but did not influence T cell subsets in either B6 or B6-TLR9−/− hosts. B6.56R.TLR9−/− mice had less spontaneous production of autoantibodies than B6.56R mice, but there were no significant differences between B6.56R and B6.56R.TLR9−/− postinduction of cGVH disease. Taken together, these results suggested that TLR9 may worsen some aspects of systemic autoimmunity while alleviating others.

Programmed cell death and the disposal of cell corpses by phagocytic cells are highly regulated ongoing processes essential for the survival and well-being of higher organisms. Abnormalities in the susceptibility of certain cells to receptor-induced death are known to lead to certain human diseases (e.g., autoimmune lymphoproliferative syndrome) and may contribute to the pathogenesis of systemic lupus erythematosus. Impaired clearance of apoptotic cells is also likely to be an important factor in lupus pathogenesis, though the biological basis of such a defect remains elusive. Finally, the process of apoptosis has been shown to contribute to lupus disease effector mechanisms. A better understanding of the role of apoptosis in lupus very likely will lead to improved diagnosis and therapy.

Systemic lupus erythematosus is characterized by production of autoantibodies and glomerulonephritis. The murine chronic graft-vs-host (cGVH) model of systemic lupus erythematosus is induced by allorecognition of foreign MHC class II determinants. Previous studies have shown that cGVH could not be induced in CD4 knockout (CD4KO) mice. We have further explored the role of host CD4 T cells in this model. Our studies now show that B cells in CD4KO mice have intrinsic defects that prevent them from responding to allohelp. In addition, B cells in CD4KO mice showed phenotypic differences compared with congeneic C57BL/6 B cells, indicating some degree of in vivo activation and increased numbers of cells bearing a marginal zone B cell phenotype. The transfer of syngeneic CD4 T cells at the time of initiation of cGVH did not correct these B cell abnormalities; however, if CD4 T cells were transferred during the development and maturation of B cells, then the B cells from CD4KO mice acquire the ability to respond in cGVH. These studies clearly indicate that B cells need to coexist with CD4 T cells early in their development to develop full susceptibility to alloactivation signals.

The nuclear self-Ags targeted in systemic lupus erythematosus translocate to the cell membrane of UV-irradiated apoptotic keratinocytes and may represent an important source of self-immunization. It is hard to understand how the noninflammatory milieu accompanying most apoptosis might provoke an immunogenic response leading to autoantibodies. We have found that the precise amount of keratinocyte UV exposure is crucial in determining the rate of apoptosis, the amount of inflammatory cytokine production, and the degree of autoantigen translocation. Low doses of UVB (≤15 mJ/cm2) promptly induced a normal, caspase-dependent apoptosis, while intermediate doses of UV-B (35 mJ/cm2) caused apoptosis with altered morphology, slower DNA fragmentation, and poly(ADP-ribose) polymerase degradation accompanied by increased Bcl-2. High doses of UVB (80 mJ/cm2) induced instead necrosis. We observed IL-1 production upon intermediate and high UVB doses. Nuclear Ag redistribution was also markedly UV dose dependent: at low doses, Sm, Ku, and DNA translocated to the surfaces of early apoptotic cells. At intermediate doses, these Ags concentrated on the cell membrane when the nucleus was still visible. At high doses, these autoantigens diffused into the cytoplasm and were released into the supernatant. Taken together, the results show that low-dose UVB induces prompt noninflammatory apoptosis. In contrast, intermediate and high doses of UVB induce proinflammatory apoptosis and necrosis, where the production of inflammatory cytokines is accompanied by exposure and release of autoantigens. The key importance of the UV dose on the fate of apoptotic keratinocytes and on their potential immunogenicity should help clarify the role of UVB in inducing systemic lupus erythematosus autoimmunity.

The epitopes recognized by pathogenic T cells in systemic autoimmune disease remain poorly defined. Certain MHC class II-bound self peptides from autoimmune MRL/lpr mice are not found in eluates from class II molecules of MHC-identical C3H mice. Eleven of 16 such peptides elicited lymph node cell and spleen cell T cell proliferation in both MRL/lpr (stimulation index = 2.03–5.01) and C3H mice (stimulation index = 2.03–3.75). IL-2 and IFN-γ production were detected, but not IL-4. In contrast to what was seen after immunization, four self peptides induced spleen cell proliferation of T cells from naive MRL/lpr, but not from C3H and C57BL/6.H2k, mice. These peptides were derived from RNA splicing factor SRp20, histone H2A, β2-microglobulin, and MHC class II I-Akβ. The first three peptides were isolated from I-Ek molecules and the last peptide was bound to I-Ak. T cell responses, evident as early as 1 mo of age, depended on MHC class II binding motifs and were inhibited by anti-MHC class II Abs. Thus, although immunization can evoke peripheral self-reactive T cells in normal mice, the presence in MRL/lpr mice of spontaneous T cells reactive to certain MHC-bound self peptides suggests that these T cells actively participate in systemic autoimmunity. Peptides eluted from self MHC class II molecules may yield important clues to T cell epitopes in systemic autoimmunity.

Systemic autoimmunity is an important clinical problem, offering a window into fundamental questions of self-nonself tolerance. We have used cellular immunology, serology, and immunopathology to approach several spontaneous mouse models. Although much remains to be done, a picture is emerging of pathological antigen-driven immune responses to self nuclear antigens, highly dependent on multiple genes, and susceptible to abnormalities of apoptosis.

Objective Crystalline silica may act as an immune adjuvant to increase inflammation and antibody production, and findings of occupational cohort studies suggest that silica exposure may be a risk factor for systemic lupus erythematosus (SLE). We undertook this population-based study to examine the association between occupational silica exposure and SLE in the southeastern US. Methods SLE patients (n = 265; diagnosed between January 1, 1995 and July 31, 1999) were recruited from 4 university rheumatology practices and 30 community-based rheumatologists in 60 contiguous counties. Controls (n = 355), frequency-matched to patients by age, sex, and state of residence, were randomly selected from driver's license registries. The mean age of the patients at diagnosis was 39 years; 91% were women and 60% were African American. Detailed occupational and farming histories were collected by in-person interviews. Silica exposure was determined through blinded assessment of job histories by 3 industrial hygienists, and potential medium- or high-level exposures were confirmed through followup telephone interviews. Odds ratios (ORs) and 95% confidence intervals (95% CIs) were estimated by logistic regression. Results More patients (19%) than controls (8%) had a history of medium- or high-level silica exposure from farming or trades. We observed an association between silica and SLE (medium exposure OR 2.1 [95% CI 1.1–4.0], high exposure OR 4.6 [95% CI 1.4–15.4]) that was seen in separate analyses by sex, race, and at different levels of education. Conclusion These results suggest that crystalline silica exposure may promote the development of SLE in some individuals. Additional research is recommended in other populations, using study designs that minimize potential selection bias and maximize the quality of exposure assessment.

Introduction Patients with systemic lupus erythematosus (SLE) have an abnormal population of neutrophils, called low-density granulocytes (LDGs), that express the surface markers of mature neutrophils, yet their nuclear morphology resembles an immature cell. Because a similar discrepancy in maturation status is observed in myelodysplasias, and disruption of neutrophil development is frequently associated with genomic alterations, genomic DNA isolated from autologous pairs of LDGs and normal-density neutrophils was compared for genomic changes. Methods Alterations in copy number and losses of heterozygosity (LOH) were detected by cytogenetic microarray analysis. Microsatellite instability (MSI) was detected by capillary gel electrophoresis of fluorescently labeled PCR products. Results Control neutrophils and normal-density SLE neutrophils had similar levels of copy number variations, while the autologous SLE LDGs had an over twofold greater number of copy number alterations per genome. The additional copy number alterations found in LDGs were prevalent in six of the thirteen SLE patients, and occurred preferentially on chromosome 19, 17, 8, and X. These same SLE patients also displayed an increase in LOH. Several SLE patients had a common LOH on chromosome 5q that includes several cytokine genes and a DNA repair enzyme. In addition, three SLE patients displayed MSI. Two patients displayed MSI in greater than one marker, and one patient had MSI and increased copy number alterations. No correlations between genomic instability and immunosuppressive drugs, disease activity or disease manifestations were apparent. Conclusions The increased level of copy number alterations and LOH in the LDG samples relative to autologous normal-density SLE neutrophils suggests somatic alterations that are consistent with DNA strand break repair, while MSI suggests a replication error-prone status. Thus, the LDGs isolated have elevated levels of somatic alterations that are consistent with genetic damage or genomic instability. This suggests that the LDGs in adult SLE patients are derived from cell progenitors that are distinct from the autologous normal-density neutrophils, and may reflect a role for genomic instability in the disease. Electronic supplementary material The online version of this article (doi:10.1186/ar4681) contains supplementary material, which is available to authorized users.

Programmed cell death is a complex biological process involving multiple signaling pathways. Autoimmunity can result from defects in multiple stages of apoptosis, ranging from mutant death receptors and ligands to biochemical alterations in death-inducing signaling. While defects in apoptosis of immunocompetent cells can lead to systemic autoimmunity, increased levels of apoptotic cells are also associated with autoimmunity. Ineffective recognition and clearance of apoptotic cells is also a cause of autoimmunity. In this chapter, we review the mechanisms of apoptosis, and the means by which abnormalities in this process can lead to self-immunization and to the development of autoimmunity. The role of necrotic cells in inducing autoimmunity is also discussed, as well as the role of immune complexes containing apoptotic cells in amplifying and perpetuating autoimmunization.

The immunologic basis of systemic lupus erythematosus (SLE) is multifactorial and still elusive. Recent advances in the field of apoptosis have suggested new paradigms for the development of lupus autoimmunity. In the present studies we examined the possibility that individual populations of T and B cells are abnormally resistant to apoptosis or that they stand out in over- or underexpressing Fas. Fas was generally overexpressed in cells freshly isolated from SLE patients but the apoptotic response to FasL was normal. We did not find increased spontaneous ongoing apoptosis in SLE lymphocytes. Normal cleavage of PARP similarly implied that the final biochemical pathway of apoptosis is relatively intact in SLE. Finally we placed special emphasis on the response of SLE patient cells to UV irradiation, especially cells from photosensitive patients, and found no difference in Fas expression. In conclusion our results indicate that SLE patients do not suffer from a major apoptotic abnormality. The results also raise questions concerning the dynamic expression of Fas and the significance of ongoing apoptosis as a risk for autoimmune disease.

We have previously shown that the gld autoimmune syndrome is suppressed in lethally irradiated gld mice reconstituted with a mixture of normal and gld bone marrow (BM). Furthermore, in vivo depletion of normal Thy-1+ cells restores lymphoproliferation and autoantibody production in such chimeras, suggesting that T cells bearing Fas ligand are responsible for correcting the gld defect. In this study, mixed-BM chimeras lacking either normal CD4+ (B6CD4KO-B6gld) or normal CD8+ T cells (B6CD8KO-B6gld) were generated to determine the contribution of the normal T cell subsets to disease suppression. Lymphoproliferation was completely suppressed in B6CD4KO-B6gld chimeras but only modestly in B6CD8KO-B6gld chimeras. On the other hand, both types of mixed-BM chimeras had incomplete effects on the suppression of serum autoantibodies when compared with B6gld reconstituted with isologous BM. These results suggest that both T cell subsets provide Fas ligand to suppress immune cells responsible for autoantibody production; however, CD8+ T cells are mainly responsible for preventing lymphoproliferation.

Defective Fas-mediated apoptosis in mice, caused by the gld mutation in the fas ligand gene, results in the development of lupus-like autoantibodies and severe lymphoproliferation. We previously demonstrated ectopic expression of the costimulatory molecule B7-1 (CD80) on T lymphocytes in B6/gld mice. This report extends these observations by demonstrating similar results in B6/lpr mice, which possess a mutation in the gene encoding Fas. Additionally, we demonstrate that this phenomenon is age-dependent and occurs on multiple subsets of B6/gld T lymphocytes. B7-1 upregulation is observed on T cells from both conventionally housed and specific-pathogen-free B6/gld mice, suggesting that this is not a consequence of infection by pathogen. T cells from lpr and gld mice show increased binding of CTLA4-Ig fusion protein, suggesting that the upregulated B7-1 is functional. CD28, a receptor for B7-1 which activates T cells, is upregulated in B6/lpr and B6/gld mice, while CTLA4, a negative regulator of T cells which binds B7-1, is not. Our results suggest that ectopic expression of B7-1 on T cells of lpr and gld mice may be playing a role in exacerbation of lymphoproliferation and/or autoimmunity.

It has been proposed that the “normal” stimulation of the immune system that occurs from interactions with environmental stimuli, whether infectious or dietary, is necessary for the initiation and/or continuation of autoimmunity. We tested this hypothesis by deriving a group of MRL-lpr mice into a germfree (GF) environment. At 5 mo of age, no differences between GF and conventional MRL-lpr mice were noted in lymphoproliferation, flow cytometric analysis of lymph node cells (LN), or histologic analysis of the kidneys. Autoantibody levels were comparably elevated in both groups. A second experiment tested the role of residual environmental stimuli by contrasting GF mice fed either a low m.w., ultrafiltered Ag-free (GF-AF) diet or an autoclaved natural ingredient diet (GF-NI). At 4 mo of age, both groups showed extensive lymphoproliferation and aberrant T cell formation, although the GF-AF mice had ∼50% smaller LNs compared with sex-matched GF-NI controls. Autoantibody formation was present in both groups. Histologic analysis of the kidneys revealed that GF-AF mice had much lower levels of nephritis, while immunofluorescence analysis demonstrated no difference in Ig deposits but did reveal a paucity of C3 deposition in the kidneys of GF-AF mice.These data do not support a role for infectious agents in the induction of lymphoproliferation and B cell autoimmunity in MRL-lpr mice. Furthermore, they suggest that autoantibodies do not originate from B cells that were initially committed to exogenous Ags. They do suggest a possible contributory role for dietary exposure in the extent of lymphoproliferation and development of nephritis in this strain.

Objective— To study the role of Mer receptor tyrosine kinase (mertk) in atherosclerosis. Methods and Results— We irradiated and reconstituted atherosclerosis-susceptible C57Bl/6 low-density lipoprotein receptor-deficient female mice ( ldlr −/− ) with either a mertk +/+ or mertk −/− (tyrosine kinase-defective mertk) bone marrow. The mice were put on high-fat diet for either 8 or 15 weeks. Mertk deficiency led to increased accumulation of apoptotic cells within the lesions, promoted a proinflammatory immune response, and accelerated lesion development. Conclusions— Mertk expression by bone marrow-derived cells is required for the disposal of apoptotic cells and controls lesion development and inflammation.

Fas (CD95) is a cell surface protein that mediates apoptosis. lpr is a mutation of the Fas gene caused by a retroviral insertion resulting in premature termination of transcription and aberrant splicing of Fas mRNA. Mice homozygous for the lpr gene develop lymphoproliferation and produce autoantibodies closely resembling those of human systemic lupus erythematosus. While lpr mice have been reported to express low levels of normally spliced Fas mRNA, it is unknown whether they express functional Fas protein. Here we show that splenocytes from lpr mice that have been damaged by γ-irradiation expressed Fas protein. Fas was up-regulated on irradiated B6 cells and could be detected on B6/lpr cells undergoing apoptosis following in vitro culture. Detection of Fas on live lpr cells was demonstrable when apoptosis was blocked by zinc. In a short term chimera system, Fas was shown to play a role, in vivo, in the disposition of radiation-injured cells from both normal and lpr mice. The addition of anti-Fas Ab to in vitro cultures resulted in an increase in apoptosis in both B6 and B6/lpr cells. Detection of intact Fas message and low levels of Fas protein in lpr mice has led to the consideration of lpr as a leaky mutation. This study demonstrates that lpr mice can produce functional Fas protein. This system is also appropriate for identifying the in vivo role of Fas/FasL in apoptosis following other cell manipulations.

The CD40/CD40L (CD40 ligand) axis regulates several interactions between T cells and B cells. Blocking of CD40 engagement by CD40L inhibits Ig class switch by B cells as well as diminishes T cell response to an immunizing Ag. For these reasons, disruption of CD40/CD40L interactions by anti-CD40L administration or by genetic disruption of CD40L has ameliorated a variety of autoimmune conditions. More recent findings suggest that a direct signal can be transmitted to T cells via their expressed CD40L, which can costimulate proliferation with CD3 or promote germinal center formation. It is therefore possible that treatment with anti-CD40L Ab might produce a different outcome than observed in genetically CD40L-deficient mice. In this regard, we observe that in contrast to the genetic deletion of CD40L in MRL-lpr mice, which diminishes autoimmune disease but has little effect on adenopathy, administration of anti-CD40L to MRL-lpr mice accelerates both of these parameters. This difference appears to result from anti-CD40L actively delivering a signal that inhibits T cell apoptosis in lpr mice. This was confirmed by in vitro studies demonstrating that CD40L cross-linking on lpr thymocytes inhibited apoptosis and surface TCR down-modulation induced by CD3 ligation.

We wondered whether the apoptosis known to occur after UV-B irradiation might involve the Fas/Fas ligand (FasL) signaling pathway. We exposed PBLs from normal individuals, and also the Jurkat (E6-1) and U937 cell lines, to graded doses of UV-B irradiation and observed a prompt and marked increase in Fas expression at doses as low as 0.5 mJ/cm2. Increased Fas expression did not require new protein synthesis, since cycloheximide-treated cells also showed an increase in Fas after UV-B. UV-B-irradiated cells cultured in the presence of zinc showed inhibition of apoptosis coincident with a marked increase in Fas+ cells, apparently indicating the accumulation of Fas-bearing cells unable to undergo apoptosis. After UV-B irradiation, PBLs showed increased expression of Fas ligand; the E6-1 lymphocytic cell line also released soluble FasL. UV-B induced apoptosis could be partially blocked by neutralizing FasL Abs, and a FasL-resistant variant of E6-1 cell line showed reduced apoptosis after UV-B irradiation, implying that the increase in Fas expression signified a role for Fas in UV-induced apoptosis. UV-induced Fas expression may serve to target stress-injured cells for removal by FasL-bearing cells or by FasL produced by the cells themselves in response to the stimuli, and may represent a general function of the Fas/FasL pathway in facilitating the apoptosis and elimination of undesirable or harmful cells.

lpr, a murine mutation of the Fas apoptosis receptor, causes lymphadenopathy and autoantibody production, with lymphadenopathy primarily due to a population of CD4−CD8−B220+ T cells. Previous in vivo experiments, in which lpr and normal bone marrow cells were coinfused into lpr hosts, have demonstrated that only T cells of lpr origin accumulated abnormally and only B cells of lpr origin produced autoantibodies. Moreover, in these chimeras, B cells of normal origin were unable to respond to conventional, T cell-dependent exogenous Ag. To address the role of lpr B cells in regulation of lpr autoimmunity, we have prepared lpr-+ mixed chimeras and selectively eliminated lpr B cells using allele-specific, mAb treatment, thus allowing normal B cells to develop in an environment with lpr T cells. From these data, we arrived at four major conclusions: 1) Compared with control-treated chimeric mice, lpr B cell-depleted mice had greatly reduced total lymph node cell counts; 2) the T cells were derived equally from normal and lpr donors, and the percentage of lpr-derived CD4−CD8− T cells was greatly reduced; 3) despite the presence of the remaining lpr T cells, no autoantibodies were produced by the normal derived B cells; and 4) lpr T cells without lpr B cells were unable to prevent a normal B cell response to conventional Ag. These data demonstrate that B cells can play a critical and expansive regulatory role, not only for T cells, but for other B cells as well.

We have identified a novel activation related B-cell gene (bca) through differential hybridization screening of a murine B cell cDNA library. The deduced amino acid sequence predicted a protein of 482 amino acids with strong sequence similarity to the SH2 and SH3 domains present within the non-catalytic regions of several protein tyrosine kinases. Northern analysis of RNA from several murine B-cell lines revealed a transcript of 1.8 kb, which was not detected in T-cell and non-lymphoid cell lines. bca was transcribed at low levels in resting spleen cells from a variety of normal mouse strains and was strongly expressed in kidney RNA. bca expression was markedly increased in RNA prepared from mitogen activated B cells, and in freshly isolated spleen and lymph node cells of MRL/lpr and NZB autoimmune strains. The unique sequence of bca, which bears no obvious similarity to any specific class of proteins containing SH2 and SH3 domains, suggests that this gene encodes a novel protein potentially involved in B-cell signal transduction.

Tuberous sclerosis is an autosomal dominant disorder characterized by involvement of skin, nervous system, kidneys, and lungs. It results from mutations in 1 of 2 genes: TSC1 (encoding hamartin) or TSC2 (encoding tuberin), leading to dysregulation and activation of the mammalian target of rapamycin (mTOR) pathway. Constitutive activation of mTOR signaling has recently been reported in systemic lupus erythematosus (SLE), and inhibition of this pathway may benefit patients with SLE nephritis. We report a case of a young woman with tuberous sclerosis who developed fulminant SLE, with lower extremity edema, massive proteinuria, and class IV lupus glomerulonephritis. She died despite treatment with high-dose steroids, plasmapheresis, and cyclophosphamide. Although there are no prior reports of coexistence of these 2 rare diseases, this case is of considerable interest because of the possibility that activation of mTOR by the TSC mutations may have led to activation of the immune system and the development of unusually severe SLE.

Mice homozygous for the mutant Fas gene lpr develop generalized lymphoproliferation and produce autoantibodies resembling those found in human SLE. We have previously shown that these autoantibodies are produced by B2 cells rather than B1 cells and that the autoantibody-producing B cells are intrinsically abnormal. We investigated further the lpr B cell with a large panel of antibodies to B-cell surface markers to identify phenotypic abnormalities. B cells from spleen and bone marrow of age-matched congenic mice differing only at the lpr locus were examined by flow cytometry. Two consistent phenotypic differences were identified. First, spleen cells from older lpr mice had an increase in the number and percentage of IgM + B cells expressing low levels of CD23. Second, lpr bone marrow had decreased numbers of B220 h1 IgM + -syndecan-1 + CD23 + B cells. All other markers tested, except the previously identified modest increase of Ia on lpr spleen cells, showed no consistent differences. B cells from gld mice showed the same phenotypic abnormalities as those from lpr. Compared to T cells, the relative paucity of cell surface marker differences between lpr and +/+ B cells suggests that B cells may have fewer regulatory mechanisms to silence autoreactive specificities. The phenotypic differences identified may provide clues to the mechanism of autoantibody production in lpr mice, while the overwhelming phenotypic similarity between lpr and +/+ B cells suggests that the major abnormality of lpr B cells may lie in their specificity, that is, in their inability to delete autoreactive subsets.