Ch Muñoz

Potential Involvement of Platelet-Derived Microparticles and Microparticles Forming Immune Complexes during Monocyte Activation in Patients with Systemic Lupus Erythematosus

Microparticles (MPs) are vesicles derived from the plasma membrane of different cells, are considered a source of circulating autoantigens, and can form immune complexes (MPs-ICs). The number of MPs and MPs-ICs increases in patients with systemic lupus erythematosus (SLE). MPs activate myeloid cells by inducing IL-6 and TNF-α in both SLE and other diseases. Therefore, we propose that the recognition of MPs-ICs by monocytes rather that MPs may define their phenotype and contribute to the inflammatory process in patients with SLE. Thus, the aims of this study were to evaluate the association among circulating MPs-ICs from different cell sources, alterations observed in monocyte subsets, and disease activity in patients with SLE and to establish whether monocytes bind and respond to MPs-ICs in vitro. Circulating MPs and monocyte subsets were characterized in 60 patients with SLE and 60 healthy controls (HCs) using multiparametric flow cytometry. Patients had higher MP counts and frequencies of MPs-CD41a + (platelet-derived) compared with HCs, regardless of disease activity. MPs from patients with SLE were C1q + and formed ICs with IgM and IgG. MPs-IgG + were positively correlated with active SLE (aSLE), whereas MPs-IgM + were negatively correlated. Most of the circulating total ICs-IgG + were located on MPs. The proportion and number of non-classical monocytes were significantly decreased in patients with SLE compared with HCs and in patients with aSLE compared with patients with the inactive disease. Non-classical monocytes obtained from patients with SLE exhibited increased levels of CD64 associated with MPs-IgG +, MPs-C1q +, total circulating ICs-IgG +, and disease activity. The direct effects of MPs and MPs-IgG + on monocytes were evaluated in cell culture. Monocytes from both HCs and patients bound to and internalized MPs and MPs-IgG + independent of CD64. These vesicles derived from platelets (PMPs), mainly PMPs-IgG +, activated monocytes in vitro and increased the expression of CD69, CD64, and pro-inflammatory cytokines such as IL-1β, TNF-α, and IFN-α. Therefore, MPs are one of the most representative sources of the total amount of circulating ICs-IgG + in patients with SLE. MPs-IgG + are associated with SLE activity, and PMPs-IgG + stimulate monocytes, changing their phenotype and promoting pro-inflammatory responses related to disease activity.

Infiltrating CD16+ Are Associated with a Reduction in Peripheral CD14+CD16++ Monocytes and Severe Forms of Lupus Nephritis

Our aim was to characterize glomerular monocytes (Mo) infiltration and to correlate them with peripheral circulating Mo subsets and severity of lupus nephritis (LN). Methods. We evaluated 48 LN biopsy samples from a referral hospital. Recognition of Mo cells was done using microscopic view and immunohistochemistry stain with CD14 and CD16. Based on the number of cells, we classified LN samples as low degree of diffuse infiltration (<5 cells) and high degree of diffuse infiltration (≥5 cells). Immunophenotyping of peripheral Mo subsets was done using flow cytometry. Results. Mean age was 34.0 ± 11.7 years and the mean SLEDAI was 17.5 ± 6.9. The most common SLE manifestations were proteinuria (91%) and hypocomplementemia (75%). Severe LN was found in 70% of patients (Class III, 27%; Class IV, 43%). Severe LN patients and patients with higher grade of CD16+ infiltration had lower levels of nonclassical (CD14+CD16++) Mo in peripheral blood. Conclusions. Our results might suggest that those patients with more severe forms of LN had a higher grade of CD14+CD16+ infiltration and lower peripheral levels of nonclassical (CD14+CD16++) Mo and might reflect a recruitment process in renal tissues. However, given the small sample, our results must be interpreted carefully.

Transferrina y ceruloplasmina en orina de pacientes con lupus eritematoso sistémico. ¿Son útiles para diferenciar pacientes con nefritis lúpica?

BACKGROUND AND OBJECTIVE Diagnosis of lupus nephritis (LN) is usually based on renal biopsy, which is an invasive technique that involves multiple risks. Therefore, different biomarkers have emerged as alternatives for the diagnosis of LN. Nonetheless, studies regarding urinary biomarkers in Latin American patients are limited. The objective of this study was to assess the diagnostic value of urinary transferrin and ceruloplasmin to differentiate patients who have renal involvement from those who do not. MATERIALS AND METHODS Systemic lupus erythematosus (SLE) patients that met the revised American College of Rheumatology (ACR) classification criteria were recruited. Patients with another autoimmune disease, active infection (urinary tract or systemic infection), renal replacement therapy, human immunodeficiency virus infection or pregnancy were excluded. A urine sample was collected from each patient. LN was diagnosed according to ACR criteria. The activity and chronicity of LN were measured using the Austin indices. Urinary transferrin and ceruloplasmin levels were measured using commercial enzyme-linked immunosorbent assay (ELISA) kits. Mann-Whitney U test and Students t-test were used to compare data. Spearmans rank correlation was used to determine associations. Lastly, receiver operating characteristic (ROC) curves were created. RESULTS The study involved 120 SLE patients. In all, 85% were female, 76% mestizo, the mean age was 32.8±12.1years and mean systemic lupus erythematosus disease activity index (SLEDAI) was 8.4±8.9; 64% had renal involvement. Urinary levels of the two biomarkers were significantly higher in patients with LN compared to those without LN. Similarly, urinary levels of both biomarkers were significantly higher in patients with active LN compared to those with inactive LN. Furthermore, urinary transferrin levels were significantly higher in Afro-Latin American patients. On the other hand, urinary transferrin levels correlated with SLEDAI and proteinuria, and transferrin and ceruloplasmin levels correlated with each other. The diagnostic value of ROC curves for these urinary biomarkers for LN were good. CONCLUSIONS In our cohort of SLE patients, we found that transferrin and ceruloplasmin were potential biomarkers for LN, and can even differentiate active LN.

Tuberculous sacroiliitis in a patient with systemic lupus erythematosus: a case report and literature review:

Systemic lupus erythematosus (SLE) patients are at higher risk of developing opportunistic infections such as tuberculosis (TB), especially extrapulmonary forms like osteoarticular TB, compared to the general population. However, tuberculous sacroiliitis has been scarcely reported in these patients. We present a 34-year-old woman with SLE who developed articular tuberculosis simultaneously affecting the right sacroiliac joint and the left knee. The patient was successfully treated with antituberculosis therapy for nine months. In this case, in addition to the immunological abnormalities of lupus, the long-term glucocorticoid therapy at high dosages was the main risk factor for the development of osteoarticular tuberculosis.

Urinary neutrophil gelatinase-associated lipocalin and monocyte chemoattractant protein 1 as biomarkers for lupus nephritis in Colombian SLE patients

Background Information regarding urinary biomarkers in Mestizo and Afro-Latin-American patients is very limited. We investigated whether levels of urinary neutrophil gelatinase-associated lipocalin (NGAL), and monocyte chemoattractant protein 1 (MCP-1) are good biomarkers to differentiate patients with lupus nephritis among Latin-American systemic lupus erythematosus (SLE) patients. Methods SLE patients meeting the revised American College of Rheumatology classification criteria for SLE were recruited. Urinary levels of NGAL and MCP-1 were measured using a commercial ELISA kit. Serum anti-C1q antibodies were measured by ELISA. SLE activity was measured with the systemic lupus erythematosus disease activity index (SLEDAI). Mann–Whitney tests were used to compare data and Spearman’s rank correlations were used to examine associations between continuous variables. In addition, receiver operating characteristic curves were performed. Results One hundred and twenty SLE patients were recruited (87% women) with a median age of 32.8 ± 12.1 years and median disease duration of 7.3 ± 6.9 years. Afro-Latin-Americans had a significantly higher prevalence of lupus nephritis and higher SLEDAI scores than Mestizos. The three biomarkers were significantly higher in patients with lupus nephritis than in patients without lupus nephritis. In addition, urinary NGAL and MCP-1 were significantly higher in patients with active lupus nephritis than in inactive lupus nephritis. Urinary NGAL levels were significantly higher in Afro-Latin-American patients. A receiver operating characteristic curve for urinary biomarkers for lupus nephritis in all SLE patients showed a good level of sensitivity and specificity. Conclusion In our cohort of SLE patients, we found that urinary NGAL and MCP-1 in addition to anti-C1q antibodies were useful biomarkers for the identification of renal involvement and discrimination of active lupus nephritis among patients with renal disease.

Anti C1q antibodies. A promising biomarker for cocaine-levamisole induced vasculitis

During the 1960s, Levamisole was used as an anthelmintic agent. It was subsequently found to have immunomodulatory effects and was used to treat several inflammatory disorders. During the last decade, attention has been focused on the use of levamisole as a cutting agent for cocaine and on several cases of severe agranulocytosis associated with cocaine use.1 Cocaine/levamisole-induced vasculitis – LIVEN – is a heterogeneous vasculopathy characterized by skin necrosis concentrated in acral areas [ears, cheeks, genitals and digital necrosis], retiform purpura, general symptoms and cytopenias.2 Histopathological samples reveal pauci-inflammatory thrombotic diathesis accompanied with intravascular monocytes and evidence of complement activation.3 In 23 consecutive patients with systemic vasculitis from Hospital Universitario San Vicente Fundación, Medellín, Colombia, we analyzed the presence of Anti C1q antibodies, including antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis, (N = 13), polyarteritis nodosa (N = 4), LIVEN (N = 4) and Takayasu arteritis (N = 2). Serum Anti-C1q were measured by ELISA techniques (QUANTA Lite, Inova, USA). This study was approved by the institutional review board at our institution. From this small pilot study, we found that anti C1q antibodies were more prevalent in LIVEN patients in comparison with other vasculitis (75% vs. 5.3%, p < 0.001). Mean anti C1q titers were significantly higher in patients with LIVEN than in other vasculitis (51.0 ± 25.3 vs. 10.4 ± 7.1 IU, p = 0.04). Main clinical and serological characteristics including serum complement levels are summarized in Table 1.

273 Urinary tweak levels as biomarker of lupus nephritis in colombian sle patients

Background and aims TNF-like WEAK inducer of apoptosis (TWEAK), a TNF ligand superfamily is mainly produced by monocytes/macrophages, and is widely expressed at the RNA level in tissues including kidneys. The usefulness of urinary TWEAK (uTWEAK) to identify renal involvement in Mestizo and African-Latin American (ALA) SLE patients has not been examined yet. Methods Patients meeting the revised ACR criteria for SLE were recruited from 2 different centres at Medellín and Baranquilla, Colombia. uTWEAK were measured using an ELISA kit (R and D system, USA) Results 158 SLE patients were recruited (89% female) with median age of 32.8±12.1 years and median disease duration of 7.27±6.6 years. Mestizo (77%) and ALA (20%) were majority. 64% of patients had lupus nephritis (LN). 50 out of 71 biopsy proven LN had proliferative forms. Mean SLEDAI score was 8.5±8.7. LN patients (2803±6086 vs 672±1042, p=0.013) (Fig 1A) and ALA patients (3995±9656 vs 1618±2653, p=0.002) had significant higher levels of uTWEAK. uTWEAK levels were higher in patients with active LN and in Class V LN (Fig. 1B). uTWEAK levels were significantly correlated with 24 hours proteinuria, SLEDAI (Fig. 1C) and serum anti-C1q titers. An ROC curve constructed showed a good level of sensitivity and specificity (Fig. 1D) Abstract 273 Figure 1 Conclusions In our cohort of Colombian SLE patients, uTWEAK levels were 4 and 2 times higher in LN patients and ALA respectively. uTWEAK were significantly higher in active LN and were correlated with disease activity, proteinuria and anti-C1q antibodies.

274 Prevalence of anti-dfs70 antibodies in a colombian cohort: a case-control study

Background and aims Anti-dense fine speckled 70 (anti-DFS70) antibodies were initially identified as an ANA IIF pattern from patients with interstitial cystitis; however, some recent studies showed that anti-DFS70 antibodies are common among ANA positive individuals with no evidence of systemic autoimmune disease (SAD)(Mahler M. 2012). Information of anti-DFS70 in Latin-American countries is very limited. We determined the prevalence of Anti-DFS70 antibodies in a Colombian cohort. Methods We evaluated individuals≥18 years old, including 100 SLE patients, 102 SADs, 200 healthy controls, and 56 subjects suspected of having autoimmune disease with ANA positive and negative anti ds-DNA antibodies. The presence of anti-DFS70 antibodies was determined by QUANTA Flash by chemiluminescent techniques (Inova/Werfen, San Diego) Results Our final cohort included 458 samples. The mean age of SLE patients was 33±12 years, for SADs was 41±19 and for healthy controls was 36±10 years. The main diagnoses of SAD were: Vasculitis (n=28), RA (n=21), Systemic sclerosis (n=12), primary antiphospholipid syndrome (n=11), dermatomyositis (n=10) among others. Racial/ethnic breakdown was: 76% Mestizo and 20% Afro-latin Americans. Anti-DFS70 antibodies were positive in 1.8% of subjects with ANAs positive/anti DNA negative, in 1% of SLE patients, 0.9% of patients with other SADs and in 0.5% of healthy controls. Given the low prevalence of anti-DFS70 antibodies, no clinical correlations were possible. Conclusions Despite anti-DFS70 antibodies are a good diagnostic tool for discrimination among healthy individuals and SADs (including SLE), we found a very low prevalence of anti-DFS70 antibodies in our Colombian cohort.

Síndrome de seudotumor cerebral en una mujer embarazada y con lupus eritematoso sistémico

Systemic lupus erythematous is a chronic multi-systemic autoimmune disease that affects multiple organ systems, including the central nervous system. Pseudotumor cerebri is a disorder associated with increased intracranial pressure in the absence of a space-occupying lesion or other identifiable cause that affects young and obese women.We present the case of a pregnant woman with both pseudotumor cerebri and a new diagnosis of active systemic lupus erythematous.

AB0538 Urinary levels of vcam-1 and tweak as biomarkers of lupus nephritis

Background TNF-like WEAK inducer of apoptosis (TWEAK) is able to increase the expression of various molecules involved in the inflammatory response with relevant effects also in angiogenesis. The pathological functions of TWEAK are primarily attributed to its ability to induce the expression of several pro-inflammatory cytokines, chemokines and cell adhesion molecules, including vascular cell adhesion molecule-1 (VCAM-1) (1). Some previous studies (most of them in Caucasic, Afro-American and Asiatic cohorts) have demonstrated clinical utility of VCAM-1 and TWEAK as biomarkers in patients with systemic lupus erythematosus SLE (2). Objectives To evaluate the diagnostic value of urinary VCAM-1 and TWEAK in a cohort of Latin-American patients with SLE. Methods Patients meeting the revised ACR criteria for SLE were recruited from 2 different centers at Medellín and Baranquilla, Colombia. Urinary levels of VCAM-1 (uVCAM-1) and TWEAK (uTWEAK) were measured using an ELISA kit (R&D system, USA). SLE activity was measured with SLEDAI. Inactive LN was defined by the presence of: 24 hours proteinuria ≤500 mg/dl, inactive urine sediments and stable serum creatinine. Mann-Whitney tests were used to compare data and Spearmans rank correlations were used to examine associations. Results One hundred and fifty eight SLE patients were recruited (89% female) with median age of 32.8±12.1 years and median disease duration of 7.27±6.6 years. Mestizo (77%) and African Latin-American patients (20%) were majority. Mean SLEDAI score was 8.5±8.7. One hundred and four patients (64%) had lupus nephritis (LN). 76 out of 104 patients had biopsy proven LN, in 62% of cases with proliferative forms. uVCAM-1 and uTWEAK were significantly higher in patients with LN than without LN. At the same time, uVCAM-1 and uTWEAK were significantly higher in patients with active vs inactive LN (Figure). uVCAM-1 (581±1197 vs 189±256 ng/ml, p<0.001) and uTWEAK levels (3202±3778 vs 1123±1873 pg/ml, p=0.038) were significantly higher in patients with Class V LN in comparison with other LN classes. uVCAM-1 and uTWEAK levels had a mild positive correlation with SLEDAI (r=0.22 and r=0.16, respectively). In addition, uTWEAK correlated with 24 hours proteinuria (r=0.28). No significant correlation was found between uVCAM-1 and uTWEAK.Figure 1. Urinary levels of VCAM-1 and TWEAK in patients with and without LN (A and B) and in patients with active vs inactive LN (C and D). Conclusions uVCAM-1 and uTWEAK are useful biomarkers in Latin-American patients with SLE for the identification of patients with LN and active LN. In addition, urinary levels of VCAM-1 and TWEAK were significantly more elevated in patient with membranous LN. References González-Sánchez DA, Άlvarez CM, Vásquez G, Gόmez-Puerta JA. Role of TWEAK/Fn14 signalling pathway in lupus nephritis and other clinical settings. Nefrologia. 2016. pii: S0211–6995(16)30084–4. Skeoch S, Haque S, Pemberton P, Bruce IN. Cell adhesion molecules as potential biomarkers of nephritis, damage and accelerated atherosclerosis in patients with SLE. Lupus. 2014;23:819–24. Acknowledgements JA Gόmez-Puerta was supported by Colciencias (conv. 656 de 2014). Disclosure of Interest None declared