Faculty Bibliography

At birth, the human immune system already contains substantial levels of polymeric IgM, that include autoantibodies to neo-epitopes on apoptotic cells (ACs) that are proposed to play homeostatic and anti-inflammatory roles. Yet the biologic origins and developmental regulation of these naturally arising antibodies remain poorly understood. Herein, we report that levels of IgM-antibodies to malondialdehyde (MDA) protein adducts, a common type of in vivo generated oxidative stress-related neoepitope, directly correlate with the relative binding of neonatal-IgM to ACs. Levels of IgM to phosphorylcholine (PC), a natural antibody prevalent in adults, were relatively scant in cord blood, while there was significantly greater relative representation of IgM anti-MDA antibodies in newborns compared to adults. To investigate the potential interrelationships between neonatal IgM with pathogenic IgG-autoantibodies, we studied 103 newborns born to autoimmune mothers with IgG anti-Ro (i.e., 70 with neonatal lupus and 33 without neonatal lupus). In these subjects the mean levels of IgM anti-Ro60 were significantly higher than in the newborns from non-autoimmune mothers. In contrast, levels of IgM anti-MDA in IgG anti-Ro exposed neonates were significantly lower than in neonates from non-autoimmune mothers. The presence or absence of neonatal lupus did not appear to influence the total levels of IgM in the anti-Ro exposed newborns. Taken together, our studies provide evidence that the immune development of the natural IgM-repertoire may be affected, and become imprinted by, the transfer of maternal IgG into the fetus.

Background/Objective: We and others have suggested that complement activation can serve as an initiating signal that increases the thrombosis risk in SLE patients with antiphospholipid antibodies (aPL). Generation of complement activation products can result in proinflammatory and/or prothrombotic responses providing a permissive environment for the pathological effects of antibodies to negatively charged phospholipid protein complexes. Methods: We analyzed the NYU SLE SAMPLE Biorepository initiated in September 2013, which consists of 599 patients fulfilling ACR and/or SLICC criteria for systemic lupus erythematosus. We identified 98 patients whose criteria included the presence of one or more for the following aPL: lupus anticoagulant, IgG or IgM anti-beta2- glycoprotein-I, or IgG or IgM anticardiolipin antibodies and determined if these patients received SLEDAI points for hypocomplementemia during any encounter. We then reviewed each patient's medical record to identify the prevalence of thrombosis defined as DVT, PE, CVA, arterial occlusion with gangrene or amputation, and obstetric events as well as noncriteria manifestation of thrombocytopenia or valvulitis. We then compared the prevalence of these APLS manifestations in the SLE patients with and without evidence of complement activation. Results: The NYU SLE SAMPLE biorepository includes 599 patients (90% female, mean age 43.0+/-.9, and 10 % men, mean age 41.0+/-.3, 54% Caucasian, 31% African American, 15% Asian, 30% Hispanic White, and 5% Hispanic Black). 98 of the 599 SLE patients had aPL, 86 female and 12 male (mean age 43.0+/-.2, 56% Caucasian, 33% African American, and 11% Asian). 24 % were Hispanic white and 4% Hispanic Black. The total number of patients with adverse events was 54 of 98 (55%) with 33/50 (66%) in the SLE patients with aPL and evidence of hypocomplementemia and 21/48 (43%) (p=0.04) in the patients without evidence of hypocomplementemia. The most common thrombotic event was DVT followed by CVA. Conclusions: The prevalence of aPL as a criteria in the NYU SLE registry is 98 of 599, and adverse events were more common in the patients with evidence of hypocomplementemia (33/50, 66%) as compared to the patients without complement activation (21/48, 43%). These findings can inform decisions regarding which patient subsets that may benefit from the prophylactic use of low-dose aspirin for primary prevention in asymptomatic lupus patients. Moreover, future clinical trials should be stratified on the basis of complement consumption to be sure that equal numbers of these patients appear in both the experimental and comparator treatment arms. Finally, future prospective studies should explore the interaction between complement activation products, platelets, neutrophils, mononuclear cells, endothelial cells, coagulation cascade in the adverse events that constitute antiphospholipid syndrome

Background/Objective: Thrombocytopenia is a common feature of both SLE and antiphospholipid syndrome (APS) and in former most frequently results from antiplatelet antibodies (ie AITP) or antiphospholipid antibodies (aPL). Patients with AITP paradoxically have an increased risk of thrombosis and it has been speculated that this can result from co-presence of aPL. Primary APS is associated with both thrombosis and thrombocytopenia. We assessed whether there was an association between a history of thrombocytopenia and prevalence of a thrombotic event in a large, multiethnic cohort of SLE patients. Methods: We analyzed the NYU SLE SAMPLE Registry, consisting of patients fulfilling ACR and/or SLICC criteria for SLE. We identified 105 patients whose SLE criteria included the presence of one or more of the following APLS antibodies: lupus anticoagulant; IgG or IgM anti-beta2-glycoprotein-I; IgG or IgM anticardiolipin antibodies, and determined whether these patients had thrombocytopenia among their SLE classification criteria (<100,000/muL). We reviewed each patient's medical record to identify the prevalence of thrombosis (defined as DVT, PE, CVA, arterial occlusion with gangrene or amputation), and/or obstetric events. We compared the prevalence of thrombotic events in SLE patients with and without history of thrombocytopenia. Results The NYU SLE SAMPLE currently includes 612 patients (90% female, mean age 43.0+/-0.9 years, and 10% men, mean age 41.0+/-0.3 years). 54% of subjects were white, 31% African American, and 15% Asian. 30% were Hispanic White, and 5% Hispanic Black. 17% had aPL, of whom 89% were female and 11% male (mean age 43.0+/-0.2 years, 56% Caucasian, 33% African American, and 11% Asian). 24% were Hispanic white and 4% Hispanic Black. The total numbers of patients with thrombotic events were 45 (43%), with 5/21 (23%) in the SLE patients with aPL and prior history of thrombocytopenia and 40/ 84 (47%) (p=0.042) in the patients without thrombocytopenia. The most common thrombotic event was DVT followed by CVA. Conclusion The prevalence of aPL in the NYU SLE registry was 17%, and adverse thrombotic events were less common in the patients with prior history of thrombocytopenia (5/21, 23%) as compared to those without (40/84, 47%). This unexpected finding could be explained by protective benefit of antiplatelet antibodies when co-occur with aPL in SLE or less thrombogenic aPL in SLE when produces thrombocytopenia. Additionally, our data suggest that there might be different consequences of aPL between primary APS patients and SLE patients with aPL

Background Several studies have evaluated mortality and shortterm morbidity in neonatal lupus (NL), however there is minimaldata on long term outcomes in children exposed to maternalanti-Ro antibodies in utero. A previous pilot study utilising theResearch Registry for Neonatal Lupus (RRNL) raised concernregarding the development of autoimmune disease in childhood, however the numbers evaluated were small and the patientsstudied were young. This study was initiated to ascertain the current prevalence of autoimmune disease in NL children and theirunaffected siblings, and to evaluate whether fetal or maternal factors associated with the development of future autoimmunity.Materials and methods A retrospective cohort of family membersfrom the RRNL were contacted to evaluate for autoimmune disease. Follow-up questionnaires were completed which included35 items describing symptoms and diagnoses associated withautoimmunity in 138 cardiac NL children, 74 cutaneous NL children, and 134 unaffected siblings. Medical records were obtainedand evaluated from the patient's physicians to confirm diagnoses.Maternal diagnosis of systemic lupus and/or Sjogren's syndromeand fetal cardiac disease severity based on a previously describedseverity score were associated with postnatal autoimmune diseases using chi square and Mann-Whitney analyses.Results Seventeen (8.0%) of NL affected children developed anautoimmune disease at the time of follow up (mean age 11.6+/-9.0 years). These included 3 patients with SLE, 1 with JIA, 3with thyroid disease, 5 with psoriasis, 1 with IBD, 1 with uveitis,1 with UAS and 2 with type 1 DM. Six (4.5%) unaffected siblingsdeveloped an autoimmune disease (mean age 10.6+/-7.1 years),which included 1 with JIA, 1 with Sarcoidosis/ITP, 1 with Myasthenia Gravis/Celiac disease, 1 with psoriasis, 1 with UAS and1 with type 1 DM,. There was a significant association ofbetween having an autoimmune disease and having advancedheart block (11.0% vs. 4.2%, p = 0.03) and a trend towards anassociation with cardiac NL disease severity score (4.43+/-4.89 vs.2.58+/-4.24, p = 0.06). Mother's diagnosis of SLE or Sjogren'sdid not associate with the children's development of autoimmunedisease (p = 0.828).Conclusions Fetuses that develop advanced congenital heartblock as a manifestation of NL may be at greater risk for developing autoimmune diseases later in life. This could potentially relateto a genetic component that makes a Ro exposed fetus both moreprone to inflammatory effects of passive immunity and predisposes to future autoimmunity, independent of the mother's rheumatic disease status

Background SLE is an archetypical systemic autoimmune disease,which has been attributed to interactions between genetic andenvironmental factors that are currently not well understood. Yetrecent reports have begun to elucidate how intestinal bacteriainfluence the development of physiologic B-cell/T-cell responses,and can affect the pathogenesis of inflammatory and autoimmuneconditions. Our studies are designed to shed light on the potential roles of the gut microbiome in SLE pathogenesis.Methods We have assembled and characterised a cohort of 60female SLE patients and 20 healthy controls. DNA from theunfractionated bacteria in faecal samples, and from the sortedendogenous IgA-coated and non-coated bacterial fractions, wasthen extracted. 16 S bacterial rRNA genes were then barcodedand amplified, and over 20,000 reads were determined per sample using illumina NGS technology. Faecal and serum total Ig andautoantibodies were measured by ELISA.Results Our analysis showed less microbiome diversity in SLEthan healthy controls (p = 0.002). This dysbiosis was treatmentindependent, with more severe intestinal dysbiosis and decreasedbacterial diversity in patients with high disease activity, based onSLEDAI. In addition, SLE patients had increased representationof certain bacterial families, genus's and species, based on 16 SrRNA assignments of operational taxonomic units (OTUs).Patients with active disease displayed contractions of bacterialtaxa with reported protective properties and reciprocal expansions of taxa with putative pathobiont properties. We alsoassessed IgA, which is the most prevalent antibody isotype madeby the human body, and found evidence of exuberant levels inboth intestinal and blood samples of SLE patients. While only aminority of bacterial taxa are specifically coated by endogenousintestinal IgA, IgA-coated bacteria in SLE patients had differentialrepresentation with recurrent taxa-specific expansion in SLEpatients. Strikingly, Prevotella copri, which has recently beenlinked to new-onset RA, was significantly over-representedamong the IgA-coated taxa only in SLE patients with high diseaseactivity, and was not detected in healthy controls.Conclusion Our studies provide the first evidence that SLE isassociated with gut microbiome dysbiosis with expansions of specific bacterial taxa that may contribute to immune dysregulation.This imbalance was more significant in patients with high diseaseactivity Characterisation of in vivo IgA-coated bacteria demonstrated that certain microbes taxa/species are preferentially recognised by the adaptive immune system of SLE patients, and thesediffer significantly from healthy adults. We are now studyingthese candidate pathobionts in longitudinal studies to addresswhether the microbiome predicts and/or tracks flares

Background When found in the absence of antibodies toextractable nuclear antigens (ENA) or anti-double-stranded DNA(dsDNA) (i.e., monospecific), autoantibodies to the nuclear autoantigen dense fine speckles 70 (DFS70) are purported to ruleout SLE. The reported frequency of anti-DFS70 by chemiluminescence (CIA) in SLE is low compared to healthy individuals(0-5.7% vs. 1.3-23.2%), while the frequency of monospecificanti-DFS70 in SLE is even lower at 0-0.4%. There are no studies examining the frequency of anti-DFS70 in an early inceptionSLE cohort. This study determined the prevalence of antiDFS70 in a multi-national, multi-ethnic early inception SLEcohort and examined demographic, clinical, and autoantibodyassociations.Materials and methods Patients fulfilling ACR Classification Criteria for SLE were enrolled in the Systemic Lupus InternationalCollaborating Clinics (SLICC) inception cohort within 15months of diagnosis. Demographic and clinical data were collected at enrollment. ANAs were detected by indirect immunofluorescence on HEp-2 cells (ImmunoConcepts, Sacramento)and ENAs and dsDNA by an addressable laser bead immunoassay (FIDIS Connective13, TheraDiag, Paris). Anti-DFS70 antibodies were measured by CIA (Inova Diagnostics, San Diego).The association between anti-DFS70 and baseline demographic,clinical, and autoantibody profiles was assessed using univariateand multivariate logistic regression. For the most informativemodel, only the remaining statistically significant predictors atthe 95% CI: were included, after eliminating other potentialpredictors individually, starting with the least likely to be associated with the outcome.Results 1137 patients were included; 89.9% were female and93.8% were ANA positive (Table 1). The frequency of antiDFS70 was 7.1% [95% CI: 5.7-8.8%]. 13 of 1137 (1.1%)[95% CI: 0.6-1.9%] were positive for anti-DFS70 only (monospecific). In univariate analysis, patients with musculoskeletalactivity (based on SLEDAI items) or anti-b-2 glycoprotein-1(anti-b2GP1) were more likely to have anti-DFS70, whereasthose with anti-dsDNA, anti-SSA/Ro60, anti-SSB/La, or antiU1RNP were less likely to have anti-DFS70. In multivariateanalysis, patients with musculoskeletal activity (Odd Ratio (OR)1.25 [95% CI: 1.10, 1.41]) or anti-b2GP1 (OR 2.15, 95% CI:1.21, 3.84) were more likely to have anti-DFS70, while thosewith anti-dsDNA (OR 0.53, 95% CI: 0.31, 0.92) or anti-SSB/La(OR 0.25, 95% CI:0.08, 0.82) were less likely to have antiDFS70.Conclusions The prevalence of anti-DFS70 in newly diagnosedSLE patients was at the high end of the range previouslypublished for SLE (7.1% vs. 0-5.7%) and was associated withmusculoskeletal activity and anti-b2GP1. However, 'monospecific' anti-DFS70 was rare (1.1%) and is potentially useful to discriminate between ANA positive healthy individuals and SLE

Background Progress in the management of pregnancy in patientswith lupus and asymptomatic women with anti-Ro antibodies hasbeen made on several fronts in the last year.Results Data from the PROMISSE study (Predictors of PRegnancy Outcome: Bio Markers In Antiphospholipid Syndrome andSystemic Lupus Erythematosus) under the leadership of Dr. JaneSalmon which enrolled 389 women with or without anti-phospholipid antibodies, identified several important and potentiallyactionable baseline predictors of adverse pregnancy outcomes(APO). These include: taking hypertensive medications, having aplatelet count less than 100 k, being positive for the lupus anticoagulant (LAC), and having a physician global assessment of lupusactivity at >1. The absence of a rise in C3 during the second trimester also constituted a risk factor. Being non-Hispanic whitewas protective. Both mild/moderate and severe flares were infrequent, 13% and 3%, respectively. De novo renal disease was rare,occurring in 3/265 patients who never met ACR renal criteria.Overall, in patients with no risk factors at baseline, the APO ratewas 7.8%; fetal/neonatal mortality 3.9%. In contrast, in patientswho are either LAC positive, or LAC negative but non-White andtreated with antihypertensives, the APO rate was 58%; fetal/neonatal mortality 22%. With regard to clinical and basic translational work in neonatal lupus, there have been several advances.Based on review of 156 cases of heart block absent any extranodal involvement the use of fluorinated steroids did not reversethe block, influence the cumulative probability of extranodal disease, the cumulative probability of survival, or the cumulativeprobability of pacemaker implantation. Accordingly, these datado not support the use of dexamethasone in isolated block forthe sole purpose of preventing more progressive disease. Evaluation of umbilical cord blood from 139 anti-Ro exposed neonateswith and without heart block suggests innate and parenchymalimmune cell activation in the affected fetus. Specifically, cordCRP, NT-proBNP, MMP-2, uPA, uPAR, and plasminogen levelswere higher in affected fetuses than in unaffected cases, independent of maternal rheumatic disease, season at highest risk ofheart block, and medications taken during pregnancy. These biomarkers were positively associated with a disease severity scorederived from known risk factors for mortality. Based on the consistent demonstration of fibrosis of the atrioventricular node surrounded by macrophages and multinucleated giant cells in antiRo antibody exposed fetuses dying with heart block, investigational studies focused on macrophage signalling stimulated byssRNA associated (hY3) with the Ro60 protein and the impact ofantagonising innate cell drivers such as TLR7/8. Ligation ofTLR7/8 resulted in increased histone methylation as measured byincreased H3K4me2, a requirement for binding of NF-kB at certain promoters, specifically the kB1 region in the TNF promoterwhich was significantly decreased by hydroxychloroquine.Conclusion Translation of these finding to the bedside suggeststhat hydroxychloroquine may be efficacious in preventing heartblock. An ongoing study is currently addressing secondaryprevention

OBJECTIVE: To measure antiphospholipid antibody (aPL) variance during pregnancy; to determine if variation affects outcomes. METHODS: We used data from PROMISSE, a multicenter prospective study of pregnant women with aPL and/or SLE. APL was present if any of the following was positive: anticardiolipin (aCL), anti-beta2glycoprotein I (abeta2GPI) titers >/=40 GPL or MPL units, and/or lupus anticoagulant (LAC). APL were measured every trimester and post-partum. Adverse pregnancy outcomes (APOs) were defined as: fetal/neonatal death, preterm delivery <36 weeks due to preeclampsia or placental insufficiency; or growth restriction. RESULTS: One hundred and fifty-two aPL-positive patients were studied: 57% with clinical APS and 36% with SLE. aPL IgG levels were significantly lower during 2nd and 3rd trimesters compared to screening, but IgG aCL and abeta2GPI remained high-positive through pregnancy in 93% and 85% of patients, respectively. APL IgM titers were negative in the majority of patients and fell modestly during pregnancy. LAC frequency also decreased, but 75% remained positive through the 2nd trimester. Only 4% of patients with aPL at baseline did not have aPL at either 2nd or 3rd trimester. Changes in aPL levels or aPL status were not associated with APOs. LAC was the only aPL associated with APOs. CONCLUSION: APL levels decreased marginally during pregnancy, and changes were not associated with pregnancy outcome. Our findings suggest that measurement of aPL early is sufficient to assess risk . Repeat aPL testing through pregnancy is unnecessary

Antibodies to DNA and chromatin drive autoimmunity in systemic lupus erythematosus (SLE). Null mutations and hypomorphic variants of the secreted deoxyribonuclease DNASE1L3 are linked to familial and sporadic SLE, respectively. We report that DNASE1L3-deficient mice rapidly develop autoantibodies to DNA and chromatin, followed by an SLE-like disease. Circulating DNASE1L3 is produced by dendritic cells and macrophages, and its levels inversely correlate with anti-DNA antibody response. DNASE1L3 is uniquely capable of digesting chromatin in microparticles released from apoptotic cells. Accordingly, DNASE1L3-deficient mice and human patients have elevated DNA levels in plasma, particularly in circulating microparticles. Murine and human autoantibody clones and serum antibodies from human SLE patients bind to DNASE1L3-sensitive chromatin on the surface of microparticles. Thus, extracellular microparticle-associated chromatin is a potential self-antigen normally digested by circulating DNASE1L3. The loss of this tolerance mechanism can contribute to SLE, and its restoration may represent a therapeutic opportunity in the disease.