Factors associated with immune responses to SARS-CoV-2 vaccination in individuals with autoimmune diseases.

Patients with autoimmune diseases are at higher risk for severe infection due to their underlying disease and immunosuppressive treatments. In this real-world observational study of 463 patients with autoimmune diseases, we examined risk factors for poor B and T cell responses to SARS-CoV-2 vaccination. We show a high frequency of inadequate anti-spike IgG responses to vaccination and boosting in the autoimmune population but minimal suppression of T cell responses. Low IgG responses in B cell-depleted patients with multiple sclerosis (MS) were associated with higher CD8 T cell responses. By contrast, patients taking mycophenolate mofetil (MMF) exhibited concordant suppression of B and T cell responses. Treatments with highest risk for low anti-spike IgG response included B cell depletion within the last year, fingolimod, and combination treatment with MMF and belimumab. Our data show that the mRNA-1273 (Moderna) vaccine is the most effective vaccine in the autoimmune population. There was minimal induction of either disease flares or autoantibodies by vaccination and no significant effect of preexisting anti-type I IFN antibodies on either vaccine response or breakthrough infections. The low frequency of breakthrough infections and lack of SARS-CoV-2-related deaths suggest that T cell immunity contributes to protection in autoimmune disease.

The differential role of SSa/SSb and Ro52 antibodies in defining clinical phenotypes in idiopathic inflammatory myopathies.

OBJECTIVE: In idiopathic inflammatory myopathies, anti-SSa/SSb and anti-Ro52 are associated with interstitial lung disease (ILD), yet few studies have compared their prognostic utility. Our study analyzes clinical phenotypes associated with anti-SSa/SSb and anti-Ro52 positivity in IIM and their association with ILD. METHODS: We performed a retrospective analysis of IIM patients >18-years-old, seen at Northwell Myositis Center 2007- 2018 who met 2017 EULAR/ACR criteria with available anti-SSa/SSb data. Patients who were anti-SSa/SSb(-) and anti-Ro52(+) were excluded from anti-SSa/SSb subgroup analysis but included in Ro52 subgroup analysis. Organ manifestations, pulmonary function tests (PFTs) and comorbidities were recorded. Statistical analyses included Chi-square, Fisher's Exact, Wilcoxon Rank Sum, McNemar's test. RESULTS: Of 94 patients included in the final analysis, 35% (33/94) were anti-SSa/SSb positive (+). Of 60 patients with anti-Ro52 data, 42% (25/60) were (+). ILD was more common in anti-SSa/SSb (+) versus anti-SSa/SSb negative patients and anti-Ro52(+) versus anti-Ro52 negative patients (58% vs 25%; p = 0.003 and 64% vs.26%; p = 0,004 respectively). Anti-SSa/SSb (+) was not associated with increased ILD severity based on PFTs. Anti-Ro52(+) group had lower DLCO than anti-Ro52(-) (47% vs 68%; p = 0.003). Anti-SSa/SSb positivity did not confer a difference in the frequency of other manifestations. Elevated rates of venous thromboembolism (VTE) (10%-12%) and osteoporosis (13-17%) were observed independent of anti-SSa/SSb or anti-Ro52 status. CONCLUSION: In IIM anti-SSa/SSb or anti-Ro52 positivity is associated with higher ILD rate. Both assays are useful to confer ILD risk, but anti-Ro52 is more predictive of severe ILD. High frequencies of osteoporosis and VTE were observed in all subgroups.

Belimumab treatment of adult idiopathic inflammatory myopathy.

OBJECTIVE: To evaluate belimumab addition to the standard of care in patents with refractory idiopathic inflammatory myopathy (IIM). METHODS: We conducted a 40-week multicentre, randomized, double-blind, placebo-controlled trial with 1:1 IV belimumab 10 mg/kg or placebo randomization and a 24-week open-label extension. Clinical responses were measured by the definition of improvement (DOI) and total improvement score (TIS). Flow cytometry analyses were performed on available samples before randomization, at 24 and 60-64 weeks. Descriptive statistics, t-test, Fisher's exact test and analysis of variance tests were used. RESULTS: A total of 17 patients were randomized, 15 received five or more doses of belimumab or placebo and were included in the intention-to-treat analysis. More belimumab patients vs placebo attained a TIS ≥40 [55.5% vs 33.3%; P = non-significant (NS)] and achieved the DOI (33.3% vs 16.7%; P = NS) at weeks 40 and 64; the mean TIS was similar among groups. Two patients achieved major responses (TIS = 72.5) after week 40 in the belimumab arm and none in the placebo arm. No improvement in the placebo arm after switching to the open-label phase was observed. There was no steroid-sparing effect. No new safety signals were detected. Although total B cells were not reduced, belimumab induced naïve B cell depletion while enhancing the number and frequency memory B cells. CONCLUSION: The study did not meet the primary endpoint and no statistically significant differences were observed in clinical responses between arms. More patients achieved sustained TIS ≥40 and reached the DOI. Most patients who received belimumab for >40 weeks had clinical improvement. Phenotypic changes in B cell populations were not associated with clinical responses. CLINICAL TRIAL REGISTRATION NUMBER: Clinicaltrials.gov (https://clinicaltrials.gov/), NCT02347891.

A Multianalyte Assay Panel With Cell-Bound Complement Activation Products Predicts Transition of Probable Lupus to American College of Rheumatology-Classified Lupus.

OBJECTIVE: To evaluate the usefulness of biomarkers to predict the evolution of patients suspected of systemic lupus erythematosus (SLE), designated as probable SLE (pSLE), into classifiable SLE according to the American College of Rheumatology (ACR) classification criteria. METHODS: Patients suspected of SLE were enrolled by lupus experts if they fulfilled three ACR criteria for SLE and were followed for approximately 1-3 years to evaluate transition into ACR-classifiable SLE. Individual cell-bound complement activation products (CB-CAPs), serum complement proteins (C3 and C4), and autoantibodies were measured by flow cytometry, turbidimetry, and enzyme-linked immunosorbent assay, respectively. Blood levels of hydroxychloroquine (HCQ) were measured by mass spectrometry. A multianalyte assay panel (MAP), which includes CB-CAPs, was also evaluated. A MAP of greater than 0.8 reflected the optimal cutoff for transition to SLE. Time to fulfillment of ACR criteria was evaluated by Kaplan-Meier analysis and Cox proportional hazards model. RESULTS: Of the 92 patients with pSLE enrolled, 74 had one or two follow-up visits 9-35 months after enrollment for a total of 128 follow-up visits. Overall, 28 patients with pSLE (30.4%) transitioned to ACR-classifiable SLE, including 16 (57%) in the first year and 12 (43%) afterwards. A MAP score of greater than 0.8 at enrollment predicted transition to classifiable SLE during the follow-up period (hazard ratio = 2.72; P = 0.012), whereas individual biomarkers or fulfillment of Systemic Lupus International Collaborating Clinics criteria did not. HCQ therapy was not associated with the prevention of transition to SLE. CONCLUSION: Approximately one-third of patients with pSLE transitioned within the study period. MAP of greater than 0.8 predicted disease evolution into classifiable SLE.

Cytokine-induced liver injury in coronavirus disease-2019 (COVID-19): untangling the knots.

Liver dysfunction manifesting as elevated aminotransferase levels has been a common feature of coronavirus disease-2019 (COVID-19) infection. The mechanism of liver injury in COVID-19 infection is unclear. However, it has been hypothesized to be a result of direct cytopathic effects of the virus, immune dysfunction and cytokine storm-related multiorgan damage, hypoxia-reperfusion injury and idiosyncratic drug-induced liver injury due to medications used in the management of COVID-19. The favored hypothesis regarding the pathophysiology of liver injury in the setting of COVID-19 is cytokine storm, an aberrant and unabated inflammatory response leading to hyperproduction of cytokines. In the current review, we have summarized the potential pathophysiologic mechanisms of cytokine-induced liver injury based on the reported literature.

The Longitudinal Immune Response to Coronavirus Disease 2019: Chasing the Cytokine Storm.

The clinical progression of the severe acute respiratory syndrome coronavirus 2 infection, coronavirus 2019 (COVID-19), to critical illness is associated with an exaggerated immune response, leading to magnified inflammation termed the "cytokine storm." This response is thought to contribute to the pathogenicity of severe COVID-19. There is an initial weak interferon response and macrophage activation that results in delayed neutrophil recruitment leading to impeded viral clearance. This causes prolonged immune stimulation and the release of proinflammatory cytokines. Elevated inflammatory markers in COVID-19 (e.g., d-dimer, C-reactive protein, lactate dehydrogenase, ferritin, and interleukin-6) are reminiscent of the cytokine storm seen in severe hyperinflammatory macrophage disorders. The dysfunctional immune response in COVID-19 also includes lymphopenia, reduced T cells, reduced natural killer cell maturation, and unmitigated plasmablast proliferation causing aberrant IgG levels. The progression to severe disease is accompanied by endotheliopathy, immunothrombosis, and hypercoagulability. Thus, both parts of the immune system-innate and adaptive-play a significant role in the cytokine storm, multiorgan dysfunction, and coagulopathy. This review highlights the importance of understanding the immunologic mechanisms of COVID-19 as they inform the clinical presentation and advise potential therapeutic targets.

Comparative Survival Analysis of Immunomodulatory Therapy for Coronavirus Disease 2019 Cytokine Storm.

BACKGROUND: Cytokine storm is a marker of coronavirus disease 2019 (COVID-19) illness severity and increased mortality. Immunomodulatory treatments have been repurposed to improve mortality outcomes. RESEARCH QUESTION: Do immunomodulatory therapies improve survival in patients with COVID-19 cytokine storm (CCS)? STUDY DESIGN AND METHODS: We conducted a retrospective analysis of electronic health records across the Northwell Health system. COVID-19 patients hospitalized between March 1, 2020, and April 24, 2020, were included. CCS was defined by inflammatory markers: ferritin, > 700 ng/mL; C-reactive protein (CRP), > 30 mg/dL; or lactate dehydrogenase (LDH), > 300 U/L. Patients were subdivided into six groups: no immunomodulatory treatment (standard of care) and five groups that received either corticosteroids, anti-IL-6 antibody (tocilizumab), or anti-IL-1 therapy (anakinra) alone or in combination with corticosteroids. The primary outcome was hospital mortality. RESULTS: Five thousand seven hundred seventy-six patients met the inclusion criteria. The most common comorbidities were hypertension (44%-59%), diabetes (32%-46%), and cardiovascular disease (5%-14%). Patients most frequently met criteria with high LDH (76.2%) alone or in combination, followed by ferritin (63.2%) and CRP (8.4%). More than 80% of patients showed an elevated D-dimer. Patients treated with corticosteroids and tocilizumab combination showed lower mortality compared with patients receiving standard-of-care (SoC) treatment (hazard ratio [HR], 0.44; 95% CI, 0.35-0.55; P < .0001) and with patients treated with corticosteroids alone (HR, 0.66; 95% CI, 0.53-0.83; P = .004) or in combination with anakinra (HR, 0.64; 95% CI, 0.50-0.81; P = .003). Corticosteroids when administered alone (HR, 0.66; 95% CI, 0.57-0.76; P < .0001) or in combination with tocilizumab (HR, 0.43; 95% CI, 0.35-0.55; P < .0001) or anakinra (HR, 0.68; 95% CI, 0.57-0.81; P < .0001) improved hospital survival compared with SoC treatment. INTERPRETATION: The combination of corticosteroids with tocilizumab showed superior survival outcome when compared with SoC treatment as well as treatment with corticosteroids alone or in combination with anakinra. Furthermore, corticosteroid use either alone or in combination with tocilizumab or anakinra was associated with reduced hospital mortality for patients with CCS compared with patients receiving SoC treatment.

Biologics in the treatment of Sjogren's syndrome, systemic lupus erythematosus, and lupus nephritis.

PURPOSE OF REVIEW: It is an understatement to say that drug approvals in systemic lupus erythematosus (SLE), lupus nephritis, and Sjogren's syndrome have lagged far behind those in other autoimmune diseases, such as rheumatoid arthritis and psoriatic arthritis. Reasons for this are multiple and include the molecular and clinical heterogeneity of these conditions; confounding by background medications, especially corticosteroids; and clinical trial endpoints. However, the tides are changing, and there have been several bright spots in our attempts to bring more efficacious drugs to our patients. RECENT FINDINGS: Several positive phase II and phase III trials in SLE and lupus nephritis with drugs such as anifrolumab, voclosporin, belimumab, and obinutuzumab will no doubt eventually generate regulatory approvals for most, if not all, of these drugs. Although early in development, the promising results in Sjogren's syndrome with iscalimab and ianalumab should make the Sjogren's syndrome community quite hopeful of future drug approvals. SUMMARY: In this review, we highlight recent study results in Sjogren's syndrome, SLE, and lupus nephritis, emphasizing investigational therapies in late stage development, but we also provide a glimpse into drugs of the future.

Cell-bound complement activation products associate with lupus severity in SLE.

OBJECTIVES: To evaluate the association between lupus severity and cell-bound complement activation products (CB-CAPs) or low complement proteins C3 and C4. METHODS: All subjects (n=495) fulfilled the American College of Rheumatology (ACR) classification criteria for SLE. Abnormal CB-CAPs (erythrocyte-bound C4d or B-lymphocyte-bound C4d levels >99th percentile of healthy) and complement proteins C3 and C4 were determined using flow cytometry and turbidimetry, respectively. Lupus severity was estimated using the Lupus Severity Index (LSI). Statistical analysis consisted of multivariable linear regression and groups comparisons. RESULTS: Abnormal CB-CAPs were more prevalent than low complement values irrespective of LSI levels (62% vs 38%, respectively, p<0.0001). LSI was low (median 5.44, IQR: 4.77-6.93) in patients with no complement abnormality, intermediate in patients with abnormal CB-CAPs (median 6.09, IQR: 5.31-8.20) and high in the group presenting with both abnormal CB-CAPs and low C3 and/or C4 (median 7.85, IQR: 5.51-8.37). Odds of immunosuppressant use was higher in subjects with LSI ≥5.95 compared with subjects with LSI <5.95 (1.60 vs 0.53, p<0.0001 for both). Multivariable regression analysis revealed that higher LSI scores associated with abnormal CB-CAPs-but not low C3/C4-after adjusting for younger age, race and longer disease duration (p=0.0001), which were also independent predictors of disease severity (global R2=0.145). CONCLUSION: Abnormalities in complement activation as measured by CB-CAPs are associated with increased LSI.

Complement Activation in Patients With Probable Systemic Lupus Erythematosus and Ability to Predict Progression to American College of Rheumatology-Classified Systemic Lupus Erythematosus.

OBJECTIVE: To evaluate the frequency of cell-bound complement activation products (CB-CAPs) as a marker of complement activation in patients with suspected systemic lupus erythematosus (SLE) and the usefulness of this biomarker as a predictor of the evolution of probable SLE into SLE as classified by the American College of Rheumatology (ACR) criteria. METHODS: Patients in whom SLE was suspected by lupus experts and who fulfilled 3 ACR classification criteria for SLE (probable SLE) were enrolled, along with patients with established SLE as classified by both the ACR and the Systemic Lupus International Collaborating Clinics (SLICC) criteria, patients with primary Sjögren's syndrome (SS), and patients with other rheumatic diseases. Individual CB-CAPs were measured by flow cytometry, and positivity rates were compared to those of commonly assessed biomarkers, including serum complement proteins (C3 and C4) and autoantibodies. The frequency of a positive multianalyte assay panel (MAP), which includes CB-CAPs, was also evaluated. Probable SLE cases were followed up prospectively. RESULTS: The 92 patients with probable SLE were diagnosed more recently than the 53 patients with established SLE, and their use of antirheumatic medications was lower. At the enrollment visit, more patients with probable SLE were positive for CB-CAPs (28%) or MAP (40%) than had low complement levels (9%) (P = 0.0001 for each). In probable SLE, MAP scores of >0.8 at enrollment predicted fulfillment of a fourth ACR criterion within 18 months (hazard ratio 3.11, P < 0.01). CONCLUSION: Complement activation occurs in some patients with probable SLE and can be detected with higher frequency by evaluating CB-CAPs and MAP than by assessing traditional serum complement protein levels. A MAP score above 0.8 predicts transition to classifiable SLE according to ACR criteria.

Randomised prospective trial to assess the clinical utility of multianalyte assay panel with complement activation products for the diagnosis of SLE.

OBJECTIVE: We compared the physician-assessed diagnostic likelihood of SLE resulting from standard diagnosis laboratory testing (SDLT) to that resulting from multianalyte assay panel (MAP) with cell-bound complement activation products (MAP/CB-CAPs), which reports a two-tiered index test result having 80% sensitivity and 86% specificity for SLE. METHODS: Patients (n=145) with a history of positive antinuclear antibody status were evaluated clinically by rheumatologists and randomised to SDLT arm (tests ordered at the discretion of the rheumatologists) or to MAP/CB-CAPs testing arm. The primary endpoint was based on the change in the physician likelihood of SLE on a five-point Likert scale collected before and after testing. Changes in pharmacological treatment based on laboratory results were assessed in both arms. Statistical analysis consisted of Wilcoxon and Fisher's exact tests. RESULTS: At enrolment, patients randomised to SDLT (n=73, age=48±2 years, 94% females) and MAP/CB-CAPs testing arms (n=72, 50±2 years, 93% females) presented with similar pretest likelihood of SLE (1.42±0.06 vs 1.46±0.06 points, respectively; p=0.68). Post-test likelihood of SLE resulting from randomisation in the MAP/CB-CAPs testing arm was significantly lower than that resulting from randomisation to SDLT arm on review of test results (-0.44±0.10 points vs -0.19±0.07 points) and at the 12-week follow-up visit (-0.61±0.10 points vs -0.31±0.10 points) (p<0.05). Among patients randomised to the MAP/CB-CAPs testing arm, two-tiered positive test results associated significantly with initiation of prednisone (p=0.034). CONCLUSION: Our data suggest that MAP/CB-CAPs testing has clinical utility in facilitating SLE diagnosis and treatment decisions.

An Interim Report of the Scleroderma Clinical Trials Consortium Working Groups.

The Scleroderma Clinical Trials Consortium (SCTC) represents many of the clinical researchers in the world who are interested in improving the efficiency of clinical trials in Systemic Sclerosis (SSc). The SCTC has established 11 working groups (WGs) to develop and validate better ways of measuring and recording multiple aspects of this heterogeneous disease. These include groups working on arthritis, disease damage, disease activity, cardiac disease, juvenile SSc, the gastrointestinal tract, vascular component, calcinosis, scleroderma renal crisis, interstitial lung disease, and skin measurement. Members of the SCTC may join any one or more of these groups. Some of the WGs have only recently started their work, some are nearing completion of their mandated tasks and others are in the midst of their projects. All these projects, which are described in this paper, will help to improve clinical trials and observational studies by improving or developing better, more sensitive ways of measuring various aspects of the disease. As Lord Kelvin stated, "To measure is to know. If you cannot measure it you cannot improve it." The SCTC is dedicated to improving the lives of patients with SSc and it is our hope that the contributions of the WGs will be one important step in this process.

Update on clinical trials in systemic lupus erythematosus.

PURPOSE OF REVIEW: With advancement in our understanding of pathogenic mechanisms in systemic lupus erythematosus (SLE), there is tremendous enthusiasm in examining drugs, old and new, to improve outcomes. This review highlights recent trials' successes and impasses that have come to fore. RECENT FINDINGS: Among B-cell therapies, belimumab continues its run of successes with sustained safety and tolerability documented in a long-term extension as well as the likely approval of a subcutaneous formulation in the near future. With greater antibody-dependent cytotoxicity and less immunogenicity, there is hope for obinituzumab to succeed where its anti-CD 20 predecessors have failed. Drugs targeting type I interferons - sifalimumab and anifrolumab - have been efficacious albeit with an increase in incidence of Herpes zoster infections. There is also renewed interest in evaluating the efficacy of calcineurin inhibitors, specifically tacrolimus in the induction and maintenance of lupus nephritis. Introspection into clinical trial designs have highlighted the effects of entry criteria, end points, background medications and geographical differences on study outcomes. SUMMARY: There are at least 50 drugs and targets being evaluated in SLE. In addition to developing new drugs to treat lupus, future trials have to focus on more effective study designs to improve chances of trial success.

Citrulline dependence of anti-cyclic citrullinated peptide antibodies in systemic lupus erythematosus as a marker of deforming/erosive arthritis.

OBJECTIVE: Anti-cyclic citrullinated peptide (CCP) antibodies are a serological marker for rheumatoid arthritis (RA); up to 10%-15% of patients with systemic lupus erythematosus (SLE) are also positive. While anti-CCP in RA is citrulline-dependent, anti-CCP in some other diseases is citrulline-independent and reacts with both CCP and the unmodified (arginine-containing) cyclic arginine peptide (CAP). We investigated the citrulline dependence of anti-CCP and its significance in the arthritis of SLE. METHODS: IgG anti-CCP was compared by ELISA to anti-CAP in sera from patients with SLE (n = 335) and RA (n = 47) and healthy controls (n = 35). SLE patients were divided into 5 groups based on their joint involvement: subset I: deforming/erosive arthritis (n = 20); II: arthritis fulfilling (or likely fulfilling) American College of Rheumatology criteria for RA but without erosions (n = 18); III: joint swelling but not fulfilling RA criteria (n = 39); IV: arthritis without documented joint swelling (n = 194); and V: no arthritis (n = 58). RESULTS: Anti-CCP (> 1.7 units) was found in 68% (32/47) of patients with RA and 17% (55/329) of those with SLE. It was more common in SLE patients with deforming/erosive arthritis (38%). High anti-CCP (> 10 units) was found in RA (26%) and deforming/erosive SLE (12%). High anti-CCP/CAP ratios (> 2, indicating a selectivity to CCP) were found in 91% of anti-CCP-positive RA and 50% of anti-CCP-positive SLE patients with deforming/erosive arthritis. Patients from subset II did not have high anti-CCP/CAP. CONCLUSION: Citrulline dependence or high levels (> 10) of anti-CCP were common in SLE patients with deforming/erosive arthritis, while most anti-CCP in SLE patients was citrulline-independent. This may be useful in identifying a subset of SLE patients with high risk for development of deforming/erosive arthritis.

Increased expression of FcgammaRI/CD64 on circulating monocytes parallels ongoing inflammation and nephritis in lupus.

INTRODUCTION: The high-affinity receptor for IgG Fcgamma/CD64 is critical for the development of lupus nephritis (LN). Cross-linking Fc receptor on recruited monocytes by IgG-containing immune complexes is a key step in immune-complex-mediated nephritis in systemic lupus erythematosus (SLE). The goal of this study was to determine whether expression of Fc receptor (FcgammaR) I on circulating monocytes is associated with systemic inflammation and renal disease in SLE patients. METHODS: We studied 205 SLE patients (132 with LN and 73 without LN) along with 74 healthy control individuals. Surface expression of CD14 (monocytes), FcgammaRI/CD64, FcgammaRII/CD32, and FcgammaRIII/CD16 was evaluated by flow cytometry. Monocyte function was assessed by determining the migratory capacity and the ability to produce CCL2 (monocyte chemotractic protein 1). High-sensitivity C-reactive protein, C3 and C4 were measured by nephelometry. RESULTS: There was little difference in the expression of FcgammaRIII/CD16 or FcgammaRIII/CD32 on circulating monocytes between patients with SLE and control individuals. In contrast, FcgammaRI/CD64 expression was significantly higher in SLE patients and even higher in patients with LN. FcgammaRI/CD64 expression was positively associated with serum creatinine and indicators of systemic inflammation. Monocytes from patients with high FcgammaRI/CD64 expression also exhibited increased chemotaxis and capacity to produce monocyte chemotractic protein 1. CONCLUSIONS: Increased FcgammaRI/CD64 expression on circulating monocytes parallels systemic inflammation and renal disease in SLE patients. We propose that circulating monocytes activated by immune complexes and/or proinflammatory mediators upregulate surface expression of FcgammaRI/CD64 in SLE. The enhanced chemotactic and inflammatory potential of the activated monocytes may participate in a vicious cycle of immune cell recruitment and renal injury in SLE.

In vivo adjuvant activity of the RNA component of the Sm/RNP lupus autoantigen.

OBJECTIVE: Many lupus autoantigens contain small, highly structured RNAs, and studies have shown that the RNA components of lupus autoantigens activate production of type I interferon by dendritic cells (DCs) in vitro via the Toll-like receptor (TLR)-myeloid differentiation factor 88 pathway. This study was undertaken to examine whether U1 RNA possesses adjuvant activity in vivo. METHODS: U1 RNA was affinity purified from K562 cells. C57BL/6 or OT-II mice were immunized with 4-hydroxy-3-nitrophenyl acetyl (NP)-conjugated keyhole limpet hemocyanin (NP-KLH) or ovalbumin(323-337) peptide, using either U1 RNA or aluminum hydroxide (alum) as the adjuvant. Activation of DCs and lymphocytes was measured using flow cytometry. NP-specific antibody responses were measured using enzyme-linked immunosorbent assay. Antigen-specific T cell proliferation was determined using 3H-thymidine incorporation. RESULTS: Similar to the results with the standard adjuvant, alum, U1 RNA coadministered with NP-KLH enhanced production of NP-specific IgM and IgG (on days 8 and 16 postinjection, respectively). Moreover, proliferation of antigen-specific CD4+ T cells was enhanced to comparable levels in the mice immunized with either U1 RNA or alum. Injection of U1 RNA into the footpad of mice resulted in DC recruitment to draining lymph nodes and induction of DC maturation. U1 RNA, at 24 hours' postinjection, also increased expression of the early activation marker CD69 in both B and T lymphocytes. Pretreatment of U1 RNA with RNase or coadministration with a TLR-7 antagonist inhibited the effects of this adjuvant. CONCLUSION: A small RNA of cellular origin can drive DC maturation, B and T cell activation/proliferation, and antibody responses to exogenous antigens. These results support the idea that U1 RNA is an endogenous adjuvant, helping to explain the striking predilection of lupus autoantibodies for RNA-protein complexes such as Sm/RNP.

Type I interferon as a novel risk factor for endothelial progenitor cell depletion and endothelial dysfunction in systemic lupus erythematosus.

OBJECTIVE: The premature atherosclerosis seen in patients with systemic lupus erythematosus (SLE) is not explained by traditional risk factors. SLE disease activity, such as renal involvement and presence of autoantibodies, is associated with elevated serum levels of type I interferon (IFN-I), a family of cytokines with potent antiviral and antiproliferative effects. This study was undertaken to test the hypothesis that elevated IFN-I levels could lead to endothelial dysfunction, a surrogate for cardiovascular disease, by causing a reduction in the number of endothelial progenitor cells (EPCs), bone marrow-derived cells that participate in endothelial repair. METHODS: EPCs were enumerated in the peripheral blood of SLE patients (n = 70) and healthy controls (n = 31), using a colony-forming assay. Serum IFN-I levels were quantified by real-time polymerase chain reaction measurement of the expression of the IFN-I-inducible gene MX1. Endothelial function was determined by peripheral arterial plethysmography. RESULTS: SLE patients had markedly reduced levels of EPC colony-forming units compared with controls (median 5.7/ml peripheral blood [interquartile range 1.9-12.8] versus 28.5/ml peripheral blood [14.7-47.3]; P < 0.0001), and the depletion of EPCs was more dramatic in patients with elevated levels of IFN-I. Stepwise multiple regression analysis showed that MX1 expression and serum levels of C-reactive protein were independently associated with the reduction of EPCs. Importantly, high IFN-I levels were associated with impaired endothelial function in patients with SLE. CONCLUSION: These data support the novel hypothesis that depletion of EPCs caused by excessive IFN-I may be linked to endothelial dysfunction and increased cardiovascular risk in SLE.