Control of dermatomyositis skin disease activity lags behind control of muscles disease activity during the early treatment stages of classic dermatomyositis: A retrospective, single-centre study.

This study aimed to retrospectively identify differences in relative control of cutaneous and muscular disease activity in adult dermatomyositis (DM) patients at a single, tertiary care centre following initiation of diagnosis and treatment. Our results demonstrated a significantly lower complete treatment response rate of skin disease compared to muscular disease at 6-months and persistent but not statistically significant lower skin disease response at 12 months. These results suggest DM skin disease activity may be more refractory to treatment than muscle disease activity, especially in the early disease phase.

Ablation of SigH+ pDCs in B6.Nba2 mice prevents lupus-like disease development only if started before disease is fully established.

Systemic lupus erythematosus is characterized by hyper-activation of the immune system, multi-organ inflammation, and end-organ damage. Type I interferons (IFN-I) have been strongly implicated a role in disease etiology as has the main IFN-I-producing cell subset, the plasmacytoid dendritic cell (pDC). The B6.Nba2 mouse model develops a lupus-like disease characterized by elevated IFN-I levels and pDC pathogenicity. We have previously shown that pDC ablation prior to disease development in B6.Nba2 mice effectively prevents disease; however, it remains unclear if a similar protection can be seen if pDC ablation is initiated during later disease stages. This is important as Systemic lupus erythematosus patients are rarely diagnosed until disease is well-established and thus preventative treatment is unlikely to take place. Here we show that ablation of pDCs in the B6.Nba2 mouse model must be initiated early in order to effectively block disease development and that sustained reduction in pDC numbers is necessary for sustained effects. Finally, targeting of pDCs have been hypothesized to affect immunity towards infectious agents, in particular virus and intracellular bacteria. We show here that pDC ablation in B6.Nba2 mice does not affect the anti-viral response to encephalomyocarditic virus or a model T-dependent antigen. In summary, pDC ablation does not affect general immunity, but needs to happen early and be sustained to prevent lupus-like disease development in B6.Nba2 mice.

Lack of Association Between Sex Hormones, MDSCs, LDGs and pDCs in Males and Females With Systemic Lupus Erythematosus.

Plasmacytoid dendritic cells (pDCs) and low-density granulocytes (LDGs) are interferon-alpha producing cells that create a pro-inflammatory response in Systemic Lupus Erythematosus (SLE) leading to auto antibody production and organ damage. Both pDCs and LDGs have been shown to be dysfunctional in patients with active SLE. Myeloid-derived suppressor cells (MDSCs) have the capacity to control T and B cell activation and differentiation, and have recently been identified as cells of interest in SLE as well. While not fully understood, previous studies have suggested that pDCs are regulated in part by both X chromosome inactivation and estradiol. Whether sex chromosomes or sex hormones regulate MDSCs and LDGs remain to be determined. We aimed to explore the relative role of sex and sex hormones on pDC, MDSC and LDG frequency and function in SLE patients. We recruited patients with SLE as defined by ACR or SLICC classification criteria and healthy controls in conjunction with the Cleveland Clinic Lupus Cohort and Clinical Research Unit. We analyzed serum sex hormone levels by ELISA, and frequencies of pDCs, MDSCs, and LDGs among PBMCs and serum cytokine levels by flow cytometry. PBMCs were further analyzed for expression of genes involved in or induced by toll-like receptor (TLR)7 or TLR9 stimulation. In all SLE patients, the serum estradiol/testosterone ratio and levels of granulocytic MDSCs and LDGs were increased, while levels of pDCs were decreased. Furthermore, pDCs from active SLE patients expressed lower levels of TLR7 and TLR9 and showed diminished production of TLR9-induced IFNα and TNFα as compared to healthy controls. LDGs from healthy controls and SLE patients expressed very low levels of TLR7 and TLR9 and largely failed to respond to TLR9 stimulation. Thus, regardless of sex and sex-hormone levels, frequencies of pDCs, MDSCs and LDGs, TLR7 and TLR9 expression, and TLR9-driven cytokine production were similarly altered in male and female SLE patients.

Spontaneous CD4+ T Cell Activation and Differentiation in Lupus-Prone B6.Nba2 Mice Is IFNAR-Independent.

Systemic lupus erythematosus (SLE) is an autoimmune disorder characterized by dysregulated T and B lymphocytes. Type I interferons (IFN-I) have been shown to play important pathogenic roles in both SLE patients and mouse models of lupus. Recent studies have shown that B cell intrinsic responses to IFN-I are enough to drive B cell differentiation into autoantibody-secreting memory B cells and plasma cells, although lower levels of residual auto-reactive cells remain present. We speculated that IFN-I stimulation of T cells would similarly drive specific T-cell associated lupus phenotypes including the upregulation of T follicular helper cells and Th17, thereby affecting autoantibody production and the development of glomerulonephritis. Using the B6.Nba2 mouse model of lupus, we evaluated disease parameters in T cell specific IFN-I receptor (IFNAR)-deficient mice (cKO). Surprisingly, all measured CD4+ T cell abnormalities and associated intra-splenic cytokine levels (IFNγ, IL-6, IL-10, IL-17, IL-21) were unchanged and thus independent of IFN-I. In contrast B6.Nba2 cKO mice displayed reduced levels of effector CD8+ T cells and increased levels of Foxp3+ CD8+ regulatory T cells, suggesting that IFN-I induced signaling specifically affecting CD8+ T cells. These data suggest a role for both pathogenic and immunosuppressive CD8+ T cells in Nba2-driven autoimmunity, providing a model to further evaluate the role of these cell subsets during lupus-like disease development in vivo.

S100a9 Protects Male Lupus-Prone NZBWF1 Mice From Disease Development.

Systemic lupus erythematosus (SLE) is an autoimmune disorder disproportionally affecting women. A similar sex difference exists in the murine New Zealand Black/White hybrid model (NZBWF1) of SLE with all females, but only 30-40% of males, developing disease within the first year of life. Myeloid-derived suppressor cells (MDSCs) are prominent in NZBWF1 males and while depletion of these cells in males, but not females, promotes disease development, the mechanism of suppression remains unknown. S100a9, expressed by neutrophils and MDSCs, has previously been shown to exert immunosuppressive functions in cancer and inflammation. Here we investigated if S100a9 exerts immunosuppressive functions in NZBWF1 male and female mice. S100a9+/+, S100a9+/- and S100a9-/- NZBWF1 mice were followed for disease development for up to 8 months of age. Serum autoantibody levels, splenomegaly, lymphocyte activation, glomerulonephritis and proteinuria were measured longitudinally or at the time of harvest. In accordance with an immunosuppressive function of MDSCs in male mice, S100a9-deficient male NZBWF1 mice developed accelerated autoimmunity as indicated by increased numbers of differentiated effector B and T cells, elevated serum autoantibody levels, increased immune-complex deposition and renal inflammation, and accelerated development of proteinuria. In contrast, female mice showed either no response to S100a9-deficiency or even a slight reduction in disease symptoms. Furthermore, male, but not female, S100a9-/- NZBWF1 mice displayed an elevated type I interferon-induced gene signature, suggesting that S100a9 may dampen a pathogenic type I interferon signal in male mice. Taken together, S100a9 exerts an immunosuppressive function in male NZBWF1 mice effectively moderating lupus-like disease development via inhibition of type I interferon production, lymphocyte activation, autoantibody production and the development of renal disease.

Lupus-prone B6.Nba2 male and female mice display anti-DWEYS reactivity and a neuropsychiatric phenotype.

OBJECTIVE: Neuropsychiatric lupus (NPSLE), a manifestation of the autoimmune disease systemic lupus erythematosus (SLE), is characterized by psychiatric symptoms including anxiety and depression and upregulated autoantibodies. The B6.Nba2 spontaneous mouse model develops SLE, but has not previously been tested for NPSLE. METHODS: We investigated the NPSLE phenotype in male and female B6.Nba2 mice (n = 12 each) and age- and sex-matched B6 controls (n = 10 each) via behavioral assessments for anxiety, depression, and memory deficits. Serum anti-dsDNA, anti-nRNP, anti-DWEYS peptide reactive IgG autoantibody levels and soluble TWEAK levels were determined by ELISA. Hippocampal regions were stained for activated microglia and neurons. RESULTS: Both male and female B6.Nba2 mice showed elevated anti-dsDNA IgG, anti-nRNP IgG and anti-DWEYS reactive antibodies, elevated serum soluble TWEAK levels, and a strong anxiety and depression phenotype (p < 0.05-0.0001). Male B6.Nba2 mice developed this phenotype at a slightly older age than females. Female B6.Nba2 mice displayed reduced numbers of neurons in the hippocampal region compared to female B6 controls (p < 0.05). CONCLUSION: The B6.Nba2 mouse model recapitulates many known NPSLE phenotypes, making it a promising model to investigate the development of NPSLE in the context of SLE.

Androgen-Mediated Anti-inflammatory Cellular Processes as Therapeutic Targets in Lupus.

Systemic Lupus Erythematosus (SLE), among many other auto-immune diseases, is known to be more prevalent in females than in males. This observation has served as the foundation for studies into how sex hormones may interact with the immune system to either drive or inhibit immune activation. Early studies using castration in lupus mouse models showed the potential protective effect of testosterone against lupus development. These studies were later corroborated by observational studies in lupus patients, who upon treatment with testosterone therapy, displayed decreased disease burden. However, there are numerous limitations to treating (especially female) lupus patients with testosterone. Thus, identification of testosterone-targeted cellular and molecular mechanisms affecting immune activation is an attractive target for lupus treatment in the future. Recent studies have examined the effects of androgens on the activation of anti-inflammatory processes. As such, immunoregulatory cell types including myeloid-derived suppressor cells (MDSCs) and regulatory T and B cells have been shown to be susceptible to manipulation by sex hormones. Here, we review studies of SLE and lupus-like disease in which testosterone or testosterone-derivatives were used to skew an ongoing immune reaction toward an anti-inflammatory state. Via evaluation of both clinical studies and immunologic models we propose new areas for research with the goal of identifying testosterone-driven anti-inflammatory mediators suitable for therapeutic targeting in patients with lupus and other autoimmune diseases.

Low Levels of Vitamin D Promote Memory B Cells in Lupus.

Background: Vitamin D deficiency is a known risk factor for Systemic Lupus Erythematosus (SLE), yet clinical trials have not demonstrated efficacy and few studies have utilized lupus models to understand the mechanism underlying this relationship. The Act1-/- mouse is a spontaneous model of lupus and Sjögren's syndrome, characterized by increased Th17 cells and peripheral B cell expansion. Vitamin D3 has anti-inflammatory properties, reduces Th17 cells and impairs B cell differentiation/activation. Therefore, we assessed how varying amounts of vitamin D3 affected lupus-like disease in the Act1-/- mouse. Methods: Act1-/- mice were fed either low/restricted (0 IU/kg), normal (2 IU/kg), or high/supplemented (10 IU/kg) vitamin D3 chow for 9 weeks, after which lupus-like features were analyzed. Results: While we found no differences in Th17 cells between vitamin D3 groups, vitamin D3 restriction specifically promoted memory B cell development, accompanied by elevated levels of serum IgM, IgG1, IgG3, and anti-dsDNA IgG. A similar significant negative association between serum vitamin D and memory B cells was confirmed in a cohort of SLE patients. Conclusion: Low levels of vitamin D3 are associated with elevated levels of memory B cells in an animal model of lupus and well-controlled SLE patients.

Partial Protection From Lupus-Like Disease by B-Cell Specific Type I Interferon Receptor Deficiency.

Systemic lupus erythematosus (SLE) is an autoimmune disease that can present with many different permutations of symptom presentation. A large subset of SLE patients have been shown to present with elevated interferon stimulated gene (ISG) expression, and Type I IFNs (IFNαß) have been shown to drive disease in murine models through global IFNα Receptor (IFNAR) knockouts. However, the disease contribution of distinct immune cell subsets in response to constitutively increased levels of IFNαß is not fully understood. We utilized a B-cell specific IFNAR knockout (BΔIFNAR) on the B6.Nba2 spontaneous-lupus background to determine the contribution of IFNαß stimulated B cells in disease. We found that IFNαß signaling in B cells is driving increased splenomegaly, increased populations of activated B cells, and increased populations of germinal center (GC) B cells, memory B cells, and plasma blasts/cells, but did not affect the development of glomerulonephritis and immune-complex deposition. IFNAR expression by B cells also drove production of anti-chromatin IgG, and anti-dsDNA and -nRNP IgG and IgG2C auto-antibody levels, as well as increased Bcl2 expression, affecting GC B cell survival in B6.Nba2 mice.

Limited Effect of Indolamine 2,3-Dioxygenase Expression and Enzymatic Activity on Lupus-Like Disease in B6.Nba2 Mice.

B6.Nba2 mice spontaneously develop a lupus-like disease characterized by elevated levels of serum anti-nuclear autoantibody (ANA) immune complexes and constitutive type I interferon (IFNα) production. During disease progression, both plasmacytoid dendritic cells (pDCs) and antibody secreting plasma cells accumulate in spleens of B6.Nba2 mice. Indoleamine 2,3-dioxygenase (IDO) has been suggested to play a role in several autoimmune diseases including in the MRL/lpr model of mouse lupus-like disease; however, it remains unknown if IDO is involved in disease development and/or progression in other spontaneous models. We show here that IDO1 protein and total IDO enzymatic activity are significantly elevated in lupus-prone B6.Nba2 mice relative to B6 controls. IDO1 expression was restricted to PCs and SignR1+ macrophages in both strains, while significantly increased in B6.Nba2-derived SiglecH+ (SigH+) pDCs. Despite this unique expression pattern, neither pharmacologic inhibition of total IDO nor IDO1 gene ablation altered serum autoantibody levels, splenic immune cell activation pattern, or renal inflammation in B6.Nba2 mice. Interestingly, IDO pharmacologic inhibition, but not IDO1 deficiency, resulted in diminished complement factor C'3 fixation to kidney glomeruli, suggesting a possible therapeutic benefit of IDO inhibition in SLE patients with renal involvement.

Immunological effects of vitamin D and their relations to autoimmunity.

Vitamin D deficiency is an established risk factor for many autoimmune diseases and the anti-inflammatory properties of vitamin D underscore its potential therapeutic value for these diseases. However, results of vitamin D3 supplementation clinical trials have been varied. To understand the clinical heterogeneity, we reviewed the pre-clinical data on vitamin D activity in four common autoimmune diseases: multiple sclerosis (MS), rheumatoid arthritis (RA), systemic lupus erythematosus (SLE), and inflammatory bowel disease (IBD), in which patients are commonly maintained on oral vitamin D3 supplementation. In contrast, many pre-clinical studies utilize other methods of manipulation (i.e. genetic, injection). Given the many actions of vitamin D3 and data supporting a vitamin D-independent role of the Vitamin D receptor (VDR), a more detailed mechanistic understanding of vitamin D3 activity is needed to properly translate pre-clinical findings into the clinic. Therefore, we assessed studies based on route of vitamin D3 administration, and identified where discrepancies in results exist and where more research is needed to establish the benefit of vitamin D supplementation.

Relationships Between Vitamin D, Gut Microbiome, and Systemic Autoimmunity.

There is increasing recognition of the role the microbiome plays in states of health and disease. Microbiome studies in systemic autoimmune diseases demonstrate unique microbial patterns in Inflammatory Bowel Disease, Rheumatoid Arthritis, and Systemic Lupus Erythematosus to a lesser extent, whereas there is no single bug or pattern that characterizes Multiple Sclerosis. Autoimmune diseases tend to share a predisposition for vitamin D deficiency, which alters the microbiome and integrity of the gut epithelial barrier. In this review, we summarize the influence of intestinal bacteria on the immune system, explore the microbial patterns that have emerged from studies on autoimmune diseases, and discuss how vitamin D deficiency may contribute to autoimmunity via its effects on the intestinal barrier function, microbiome composition, and/or direct effects on immune responses.

Act1 is a negative regulator in T and B cells via direct inhibition of STAT3.

Although Act1 (adaptor for IL-17 receptors) is necessary for IL-17-mediated inflammatory responses, Act1- (but not Il17ra-, Il17rc-, or Il17rb-) deficient mice develop spontaneous SLE- and Sjögren's-like diseases. Here, we show that Act1 functions as a negative regulator in T and B cells via direct inhibition of STAT3. Mass spectrometry analysis detected an Act1-STAT3 complex, deficiency of Act1 (but not Il17ra-, Il17rc-, or Il17rb) results in hyper IL-23- and IL-21-induced STAT3 activation in T and B cells, respectively. IL-23R deletion or blockade of IL-21 ameliorates SLE- and Sjögren's-like diseases in Act1-/- mice. Act1 deficiency results in hyperactivated follicular Th17 cells with elevated IL-21 expression, which promotes T-B cell interaction for B cell expansion and antibody production. Moreover, anti-IL-21 ameliorates the SLE- and Sjögren's-like diseases in Act1-deficient mice. Thus, IL-21 blocking antibody might be an effective therapy for treating SLE- and Sjögren's-like syndrome in patients containing Act1 mutation.

Androgen-Induced Immunosuppression.

In addition to determining biological sex, sex hormones are known to influence health and disease via regulation of immune cell activities and modulation of target-organ susceptibility to immune-mediated damage. Systemic autoimmune disorders, such as systemic lupus erythematosus, rheumatoid arthritis, and multiple sclerosis are more prevalent in females, while cancer shows the opposite pattern. Sex hormones have been repeatedly suggested to play a part in these biases. In this review, we will discuss how androgens and the expression of functional androgen receptor affect immune cells and how this may dampen or alter immune response(s) and affect autoimmune disease incidences and progression.

Super-resolution microscopy reveals functional organization of dopamine transporters into cholesterol and neuronal activity-dependent nanodomains.

Dopamine regulates reward, cognition, and locomotor functions. By mediating rapid reuptake of extracellular dopamine, the dopamine transporter is critical for spatiotemporal control of dopaminergic neurotransmission. Here, we use super-resolution imaging to show that the dopamine transporter is dynamically sequestrated into cholesterol-dependent nanodomains in the plasma membrane of presynaptic varicosities and neuronal projections of dopaminergic neurons. Stochastic optical reconstruction microscopy reveals irregular dopamine transporter nanodomains (∼70 nm mean diameter) that were highly sensitive to cholesterol depletion. Live photoactivated localization microscopy shows a similar dopamine transporter membrane organization in live heterologous cells. In neurons, dual-color dSTORM shows that tyrosine hydroxylase and vesicular monoamine transporter-2 are distinctively localized adjacent to, but not overlapping with, the dopamine transporter nanodomains. The molecular organization of the dopamine transporter in nanodomains is reversibly reduced by short-term activation of NMDA-type ionotropic glutamate receptors, implicating dopamine transporter nanodomain distribution as a potential mechanism to modulate dopaminergic neurotransmission in response to excitatory input.The dopamine transporter (DAT) has a crucial role in the regulation of neurotransmission. Here, the authors use super-resolution imaging to show that DAT clusters into cholesterol-dependent membrane regions that are reversibly regulated by ionotropic glutamate receptors activation.

New Treatments for Systemic Lupus Erythematosus on the Horizon: Targeting Plasmacytoid Dendritic Cells to Inhibit Cytokine Production.

Patients with systemic lupus erythematosus (SLE) often have elevated levels of type I interferon (IFN, particularly IFNα), a cytokine that can drive many of the symptoms associated with this autoimmune disorder. Additionally, the presence of autoantibody-secreting plasma cells contributes to the systemic inflammation observed in SLE and IFNα supports the survival of these cells. Current therapies for SLE are limited to broad immunosuppression or B cell-targeting antibody-mediated depletion strategies, which do not eliminate autoantibody-secreting plasma cells. Recent clinical trials testing the efficacy of IFNα neutralization in SLE have delivered disappointing results, with primary endpoints not being met or with minimal improvements, while studies evaluating antibody therapy targeting the type I IFN receptor was more successful and is currently being tested in phase III clinical studies. As many studies have supported the idea that plasmacytoid dendritic cells (pDCs) are the main source of IFNα in SLE, specifically targeting pDCs in SLE represents a new therapeutic option. Murine models suggest pDC ablation effectively ameliorates or reduces lupus-like disease development in spontaneous models of lupus and pre-clinical and phase I clinical trials support the safety of such a therapy in humans. Here we review animal studies and the current status of clinical trials targeting IFNα, type I interferon receptor and pDCs in SLE.

A kinome wide screen identifies novel kinases involved in regulation of monoamine transporter function.

The high affinity transporters for the monoamine neurotransmitters, dopamine, norepinephrine, and serotonin, play a key role in controlling monoaminergic neurotransmission. It is believed that the transporters (DAT, NET and SERT, respectively) are subject to tight regulation by the cellular signaling machinery to maintain monoaminergic homeostasis. Kinases constitute a pivotal role in cellular signaling, however, the regulation of monoamine transporters by the entire ensemble of kinases is unknown. Here, we perform a whole human kinome RNA interference screen to identify novel kinases involved in regulation of monoamine transporter function and surface expression. A primary screen in HEK 293 cells stably expressing DAT or SERT with siRNAs against 573 human kinases revealed 93 kinases putatively regulating transporter function. All 93 hits, which also included kinases previously implicated in monoamine transporter regulation, such as Protein kinase B (Akt) and mitogen-activated protein kinases (MAPK), were validated with a new set of siRNAs in a secondary screen. In this screen we assessed both changes in uptake and surface expression leading to selection of 11 kinases for further evaluation in HEK 293 cells transiently expressing DAT, SERT or NET. Subsequently, three kinases; salt inducible kinase 3 (SIK3), cAMP-dependent protein kinase catalytic subunit alpha (PKA C-α) and protein kinase X-linked (PrKX); were selected for additional exploration in catecholaminergic CATH.a differentiated cells (CAD) and rat chromocytoma (PC12) cells. Whereas SIK3 likely transcriptionally regulated expression of the three transfected transporters, depletion of PKA C-α was shown to decrease SERT function. Depletion of PrKX caused decreased surface expression and function of DAT without changing protein levels, suggesting that PrKX stabilizes the transporter at the cell surface. Summarized, our data provide novel insight into kinome regulation of the monoamine transporters and identifies PrKX as a yet unappreciated possible regulator of monoamine transporter function.