non-LN Patients Research Articles

P0359CROSS-TISSUE AND MURINE-HUMAN COMPARATIVE TRANSCRIPTOME ANALYSES IDENTIFY TARGETABLE GENES FOR HUMAN SYSTEMIC LUPUS ERYTHEMATOUS AND LUPUS NEPHRITIS

Abstract Background and Aims Systemic Lupus Erythematosus (SLE) is a complex disease associated with non-synchronous multi-tissue dysfunction of varying severity. Involvement of major organs, such as kidneys, contributes significantly to morbidity and mortality. We sought to unravel new pathways to be used as potential biomarkers and therapeutic targets. To this end, we compared the patterns of gene transcription -as determined by the use of RNA-sequencing- across tissues between healthy and lupus-prone mice at different stages of the disease, and explored their implications for human disease and prediction of kidney involvement, in our whole blood RNA sequencing dataset comprised of 120 SLE [55 lupus nephritis (LN), 65 non-LN] patients and 58 healthy individuals (HI). Method NZB/W-F1 lupus-prone mice were sacrificed at the pre-puberty, pre-autoimmunity and nephritic stage. Age-matched C57BL/6 mice were used as controls. An “effector” tissue (spleen) and major end-organ tissues (kidneys, brain) were collected. Total RNA was isolated, and mRNA-sequencing was performed. Differential expression and time-series analyses were performed using DESeq2. Differentially expressed genes (DEGs) were hierarchically clustered and functionally interpreted using gProfiler enrichment analysis. Human orthologous genes of mouse common DEGs in each tissue and across all disease stages, were compared to human DEGs. Using machine learning techniques, human orthologs identified in the common DEGs across all stages in the kidneys of lupus-prone vs healthy mice were used to predict kidney involvement in the human dataset, which was split in training and validation sets. Results: Gene signatures: the common cross-tissue signature was identified by the comparison of DEGs between tissues of lupus-prone vs healthy mice at each stage of the disease. A total 134 genes (including C4A, LYRM7 and HDDC3) were found, suggesting their involvement in a common pathogenic mechanism across “effector” and end-organ tissues. Tissue-specific signatures showed enrichment of FCERI mediated NF-kB activation pathway in the spleen, steroid hormone biosynthesis pathway in the kidney and phosphatidylcholine metabolic process in the brain, suggesting distinct pathways implicated in end-organ injury. Comparative murine-human transcriptome analysis: 76 human orthologs (including CCL5, IFIT and HLA genes) identified in the murine spleen signature were also differentially expressed in SLE patients vs HI, suggesting their involvement in systemic autoimmunity. A total of 68 human orthologs (including FCGR2A, C1R and JAK1) identified in the mouse kidney-signature and 25 human orthologs (including APOA2) identified in the mouse brain-signature, were differentially expressed in LN patients vs HI and neuropsychiatric SLE patients vs HI, respectively. Kidney involvement prediction in human SLE: using a neural network model, 193 human orthologs predicted LN patients vs HI with high accuracy (accuracy=0.86, sensitivity=0.82, specificity=0.91 in the validation set). Using a support vector machine model, 30 human orthologs and age and gender were the best predictors of LN vs non-LN SLE patients (accuracy=0.71, sensitivity=0.73, specificity=0.69 in the validation set). Conclusion Murine RNA-sequencing uncovered both shared cross-tissue and tissue-specific gene signatures that could be potentially targeted in SLE. Murine-human comparative transcriptome analysis revealed common gene signatures suggesting that similar biological processes and pathways are disturbed across species, with murine kidney lupus signature predicting kidney involvement in human SLE. Validation in other datasets is ongoing.

PSY29 CHARACTERISTICS AND DISEASE MANAGEMENT IN A LARGE REAL WORLD LUPUS NEPHRITIS COHORT IN THE US

Lupus nephritis (LN) is a common but heterogenous serious manifestation of systemic lupus erythematosus (SLE) associated with increased morbidity and mortality. Some evidence suggests men with SLE more frequently develop LN and with greater severity. The heterogeneity of LN complicates development of new treatment strategies. The purpose of this study was to characterize LN patients compared to non-LN SLE patients in a large, longitudinal, diverse dataset. The OM1 SLE Registry (OM1, Boston, MA) follows more than 37,000 patients longitudinally with deep clinical data, including laboratory, patient-reported and disease activity information, starting from 2013. Registry patients were identified as having new onset or prevalent LN based on relevant diagnosis codes (e.g., glomerular disease in SLE, glomerulonephritis, chronic kidney disease, proteinuria; ≥ 2 codes at least 30 days apart) or documented dialysis or kidney transplant. As of November 2019, a total of 7267 (19.3%) of SLE patients met applied criteria for LN. Median age at SLE follow-up start was 51 years (IQR 38-63) with a median follow-up duration of 6 years. At least one eGFR value was available for 71% of patients and 82% had clinical notes available. A higher proportion of LN patients compared to non-LN patients were male (12.3% vs. 7.3%) and of black race (28.7% vs. 16.5%). Treatments included hydroxychloroquine (74.5%), mycophenolate mofetil (31.7%), azathioprine (14.3%) and cyclophosphamide (1.8%). Cardiovascular (CVD) risk factors were more common in LN patients such as hypertension (71.5% vs 37.1%), hyperlipidemia (41.9% vs. 25.4%), and type 2 diabetes (23.5% vs. 11.1%), and LN patients were more likely to have coronary artery disease (15.9% vs. 6.5%) and cerebrovascular disease (10.5% vs. 4.9%). Over 8% of LN patients had end stage renal disease. Understanding the heterogeneous characteristics and course of LN patients is critical to refining clinical trial populations and improving renal and CVD outcomes.

Choroidal thickness in lupus nephritis.

This study aimed to compare choroidal thickness between patients with systemic lupus erythematosus (SLE) and lupus nephritis (LN) in complete renal remission to that of patients with SLE without LN. This was a retrospective case-control study of 23 SLE patients meeting either the American College of Rheumatology or Systemic Lupus International Collaborating Clinics classification criteria and followed at Washington University School of Medicine Rheumatology or Nephrology, and Ophthalmology outpatient clinics. The diagnosis of LN was based on renal pathology, and complete renal remission was defined as proteinuria <500 mg/daily and serum creatinine at baseline. Extra-renal flare status was determined using modified Fortin criteria. Choroidal thickness was measured using spectral-domain optical coherence tomography and read by blinded reviewers. In SLE patients without extra-renal flare, choroidal thickness of LN patients was 281 ± 78 µm compared to 288 ± 70 µm in non-LN SLE patients (p = 0.766) at the fovea. Choroidal thickness was not different in patients with LN in remission compared to non-LN SLE patients in remission. Additional studies are needed to examine choroidal thickness in patients with SLE with active LN.

The importance of serum neutrophil gelatinase-associated lipocalin level in patients with lupus nephritis

Introduction: The neutrophil gelatinase-associated lipocalin (NGAL) has emerged as a biomarker of renal damage. Objectives: The aim of this study was to assess the serum levels of NGAL (sNGAL) as a marker of disease activity in individuals with lupus nephritis (LN). Patients and Methods: This study contained 50 systemic lupus erythematosus (SLE) individuals with (n = 25) and without (n = 25) nephritis, and 39 healthy controls. The sNGAL levels were measured by ELISA. Renal function test, urinary parameters, lupus serology activity, and also calculated SLE disease activity index (SLEDAI) were analyzed to determine their associations with sNGAL. Results: The results revealed that the SLE individuals with or without nephritis had a raised serum NGAL levels as compared to control subjects (P&lt;0.001). Additionally, sNGAL levels in LN individuals were meaningfully higher compared to those in non-LN patients (P&lt;0.001). Serum NGAL showed a significant correlation with the SLEDAI, serum creatinine, and 24-h urinary protein (P&lt;0.05). More importantly, sNGAL had a significant positive correlation with the activity index of LN (r = 0.616, P=0.001). In the ROC curve analysis, the measurement of sNGAL level showed a good diagnostic performance for distinguishing individuals with LN from SLE patients without renal involvement with AUC=0.902 (P&lt;0.001), 72% sensitivity, and 99% specificity. Moreover, sNGAL could identify all of SLE patients from controls with high accuracy, AUC= 0.99, P&lt;0.001, with 99% sensitivity, and 97% specificity. Conclusion: Serum NGAL had an association with clinical parameters and could discriminate LN from SLE patients without renal involvement. Our result suggests that serum NGAL can be used for early diagnosis of LN and identifying active LN.

Polyomavirus BK, BKV microRNA, and urinary neutrophil gelatinase-associated lipocalin can be used as potential biomarkers of lupus nephritis.

ObjectiveLupus nephritis (LN) frequently progresses to end-stage renal disease. Finding a biomarker for LN and a predictor for the development of chronic kidney disease (CKD) is important for patients with systemic lupus erythematosus (SLE).MethodsNinety patients with SLE were divided into biopsy-proven LN (n = 54) and no kidney involvement (non-LN) (n = 36) groups and followed up for 54 months.ResultsOf 36 patients with LN, 3 (5.6%) had class II disease, 3 (5.6%) had class III, 35 (64.8%) had class IV, 10 (18.5%) had class V, and 3 (5.6%) had class VI (advanced sclerosis). Compared to the non-LN group, patients in the LN group had higher autoimmunity evidenced by a higher proportion of low C3 and C4 levels, positive anti-double-stranded DNA antibody levels, and lower estimated glomerular filtration rates (eGFR). Urinary neutrophil gelatinase-associated lipocalin (uNGAL) levels were significantly higher in the LN group (LN vs non-LN, 670 vs 33 ng/mL, respectively). The patients with LN had a higher urinary polyomavirus BK (BKV) load (3.6 vs 3.0 log copies/mL) and a lower urinary BKV miRNA (miR-B1) 5p level (0.29 vs 0.55 log copies/mL, p = 0.025), while there was no significant difference in the level of miR-B1-3p. Urinary miR-B1-5p level but not urinary BKV load was negatively correlated with uNGAL level (r = -0.22, p = 0.004). At the cutoff value of 80 ng/mL, the receiver operating characteristic curve analysis showed that uNGAL level as a predictor of the presence of LN had a high sensitivity (98%) and specificity (100%) (area under the curve [AUC], 0.997; p < 0.001). During the 54-month follow-up period, 14 (7%) patients with LN and none of the non-LN patients developed CKD. Multivariate Cox regression analysis revealed that baseline uNGAL level was the only predictive factor for CKD development, while baseline serum creatinine level and eGFR were not.ConclusionAn elevated urinary BKV viral load with a decreased level of miR-B1 implies the presence of LN. In addition, an increased uNGAL level is a good biomarker not only in predicting the presence of LN but also for prediction of CKD development in patients with SLE.

Phosphatidylserine-Mediated Procoagulant Activity of Microparticles and Their Originating Cells Contributes to Hypercoagulability in Lupus Nephritis

Introduction: Lupus nephritis (LN) is associated with a hypercoagulable state and an increased risk for thrombosis. Glomerular microthrombosis is detected in approximately 33% of LN patients. The incidence of thrombotic disease in patients with LN is 1.35 to 6.2 times higher than in other systemic lupus erythematosus (SLE) patients. While the immune complex is thought to elicit most forms of injury in LN, thromboembolic complications may be another important cause of renal injury and kidney dysfunction. However, the mechanisms specific to LN that promote a hypercoagulable state have not yet been identified. Antiphospholipid antibodies (aPLs) have been found to severely affect the pathophysiology of LN. But several authors have reported that the presence of aPLs is not sufficient for thrombus formation in vivo. Despite the clinical efficacy of the anti-inflammatory and immunosuppressive properties of glucocorticoid (GC) in treating LN, increased risk of thrombosis has been observed in GC users. Therefore, further studies should aim at finding other appropriate targets and effective treatments to correct coagulation abnormalities in LN. Recent studies have shown that phosphatidylserine (PS), a membrane constituent, plays an important role in thrombosis. However, the procoagulant role of PS in LN is not fully understood. Our objective was to elucidate the effects of PS exposure on microparticles (MPs) and their originating cells in LN.Methods: LN patients (n = 40) were divided into two groups, defined as active LN (aLN) and inactive LN (iLN), and compared with non-LN SLE patients (n = 20) and controls (n = 20). Patients with aLN were those with impaired kidney function with proteinuria (>0.5 g/day), an active urine sediment, or kidney biopsy with active glomerulonephritis. Inactive LN patients had decreased proteinuria levels (<0.5 g/day) and inactive urinary sediment with stable kidney function. PS exposure on MPs and MP-origin cells was quantified by lactadherin, and resulting procoagulant activity (PCA) was assessed with coagulation function assays. Fibrin production was determined by turbidity. Phosphatidylserine exposure and fibrin strands were observed using confocal microscopy. To analyze the role of glucocorticoids in MPs generation, pooled MPs suspensions were prepared from paired samples of each patient group before and after glucocorticoid treatment.Results:We found that LN patients had high levels of PS+ MPs, blood cells (BCs) and cultured endothelial cells (ECs) by flow cytometry. Circulating levels of all MP subsets were significantly higher in aLN, with the largest increases seen in MPs derived from platelets (CD41a+, PMPs), T lymphocytes (CD3+, TLMPs) and neutrophils (CD66b+, NMPs) compared to controls (all P<0.01, Figure 1A-H). Furthermore, the aLN patients had greatly elevated PS+ MPs compared with iLN. In a paired sampling study, total PS+ MPs increased significantly after glucocorticoid treatment (all P<0.05, Figure 1I). The percentage of PS+ BCs was also markedly increased in aLN, primarily in lymphocytes (32.1 ± 2.6%) followed by neutrophils (26.4 ± 1.2%), monocytes (12.5 ± 0.8%), platelets (11.3 ± 0.4%) and erythrocytes (6.2 ± 0.3%). In addition, circulating PS+ MPs and MP-origin cells in LN, especially in active stage, dramatically shortened coagulation time and increased FXa/thrombin generation and fibrin formation. Blocking PS with lactadherin prolonged coagulation time, inhibited the PCA over 80% and reduced fibrin formation. More importantly, confocal microscopy images showed fibrin strands formed on MPs and ECs in the same regions that bound lactadherin, and showed FVa/Xa costaining. Lastly, the percentage of PS+ cells significantly correlated with disease severities (proteinuria, serum creatinine, SLEDAI and urinary albumin) of LN patients.Conclusions:The current findings suggested that these activated or injured cells may contribute to the hypercoagulability of LN through PS exposure and MPs release. Our results enable us to better understand the developing trend of the thrombosis in LN. Blockade of PS prior to thrombus formation might be a novel therapeutic approach in these patients. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Two-Parametric Immunological Score Development for Assessing Renal Involvement and Disease Activity in Systemic Lupus Erythematosus.

Objective Anti-double-stranded (ds) DNA and anti-C1q autoantibodies are useful tools in the assessment of disease activity and nephritis in systemic lupus erythematosus (SLE) patients. This study aimed to explore the utility of these antibodies along with anti-Ku antibodies in an oligoparametric model approach for the assessment of disease activity and lupus nephritis. Methods Samples from 261 well-characterized SLE patients were tested using chemiluminescent immunoassays (CIA) for anti-dsDNA and anti-Ku antibodies as well as by anti-C1q antibody ELISA (Inova Diagnostics, USA). Of these SLE patients, 26.4% had lupus nephritis (LN) at the time of blood draw or had a history of LN, and modified SLE disease activity index-2K (SLEDAI) scores were used to assess disease activity. Results All three antibodies demonstrated higher prevalence and higher antibody levels in active versus inactive SLE patients and in LN versus non-LN patients. When oligoparametric analysis was performed, the likelihood of LN and patients with active disease increased with dual and triple positivity. Conclusions Anti-dsDNA and anti-C1q antibodies are useful tools to identify disease activity and/or renal involvement in SLE patients. In addition, the combination of those antibodies in a two-parametric score might improve the clinical utility of those markers.

Circulating Angiogenic T Cells Are Increased in Lupus Nephritis Patients.

BackgroundPatients with systemic lupus erythematosus (SLE), especially with lupus nephritis (LN), undergo vascular damage and repair during the course of the disease. Since the recently identified angiogenic T cells (Tang) are involved in endothelial repair coupled with endothelial progenitor cells (EPCs), this study investigated the circulating Tang cells in LN patients and their potential correlations with disease features.Material/MethodsCirculating Tang cells and EPCs were assessed by flow cytometry in peripheral blood samples from 67 SLE patients; of these, 32 had LN and 30 were matched healthy controls (HCs). The plasma levels of interleukin IL-17, IL-8, and vascular endothelial growth factor (VEGF) were quantified by immunoassays.ResultsThe percentage of circulating Tang cells in LN patients was significantly increased as compared to the non-LN patients and HCs, and they were positively correlated with the level of EPC and VEGF. Additionally, circulating Tang cell percentages were positively correlated with the extent of proteinuria in LN patients.ConclusionsThe increased levels of circulating Tang cells in LN patients might play a role in the balance of endothelium dysfunction in these patients.

Combined utilization of untimed single urine of MCP-1 and TWEAK as a potential indicator for proteinuria in lupus nephritis: A case-control study.

The aim of this study was to determine whether combined utilization of untimed single urine monocyte chemoattractant protein 1 (uMCP-1) and tumor necrosis factor (TNF)-like weak inducer of apoptosis (uTWEAK) could serve as a screening test for proteinuria in patients with lupus nephritis (LN).A case-control study that contained 39 biopsy-proven LN patients, 20 non-LN systemic lupus erythematosus (SLE) patients, and 10 healthy controls (HCs) were carried out. Correlations between uMCP-1, uTWEAK, and traditional clinical markers were analyzed by Spearman correlation test. Diagnostic values of uMCP-1, uTWEAK, and urine albumin/creatinine ratio (uACR) in the assessment of proteinuria were investigated by receiver operating characteristic (ROC) curves.Biopsy-proven LN patients showed higher levels of uMCP-1 and uTWEAK than non-LN patients. uMCP-1 and uTWEAK were elevated in renal active patients (rSLEDAI ≥4). Both uMCP-1 and uTWEAK showed significant correlation with patients' rSLEDAI, 24-hour urine proteinuria (24hr UP), and anti-double-stranded DNA (anti-dsDNA) antibodies. No correlations of these 2 biomarkers between cystatin C (Cys-C), creatinine (Cr), and blood urea nitrogen (BUN) were observed. An algorithm combining the moderate sensitivity of uMCP-1 and high specificity of uTWEAK displayed great specificity and sensitivity for proteinuria screening.Both uMCP-1 and uTWEAK were positively correlated with the impairments of LN, and the combined utility of untimed single uMCP-1 and uTWEAK might be used as potential predictors for proteinuria in LN.

Association of Tumor Necrosis Factor Alpha-Induced Protein 3 Interacting Protein 1 (TNIP1) Gene Polymorphism (rs7708392) with Lupus Nephritis in Egyptian Patients.

Lupus nephritis (LN) is a major cause of morbidity and mortality in systemic lupus erythematosus (SLE). Previous studies suggest that mutant A20 binding inhibitor of NF-κB 1 (ABIN1) protein encoded by tumor necrosis factor alpha-induced protein 3 interacting protein 1 (TNIP1) gene is associated with LN via NF-κB dysregulation. The aim of the current study was to evaluate the association of TNIP1 gene SNP rs7708392 with SLE and LN in Egyptian patients. 5' nuclease Allelic discrimination was used to evaluate the frequency of TNIP1 SNP rs7708392 in 53 patients with LN, 57 SLE patients without nephritis and 85 healthy controls. The genotyping analysis revealed that the CC genotype was more frequent in controls than SLE patients, while GC and GG genotypes were more common in SLE patients. Moreover, the GG genotype and the G allele were significantly more prevalent among LN patient than non-LN patients (P < 0.001). In LN patients, the most common genotype was GG (56.6%), while among the non-LN patients; the CG genotype was the most common (59.6%). Regression analysis demonstrated that SLE patients carrying only one G allele had a 3.4 folds increased risk for LN. Our results suggested that TNIP1 SNP (rs7708392) might be associated with the LN in Egyptian SLE patients. TNIP1 SNP (rs7708392) might be used to identify patients at risk of developing LN, which could help in early detection and treatment before progression to end-stage renal disease, improving patients' outcome and quality of life.

Urinary neutrophil gelatinase-associated lipocalin as a marker for disease activity in lupus nephritis

The neutrophil gelatinase-associated lipocalin (NGAL) has been emerging as a novel biomarker of acute kidney injury while its value in lupus nephritis is uncertain. The aim of this study was to assess urinary NGAL levels as a marker for disease activity in patients with lupus nephritis.This study included 70 systemic lupus erythematosus (SLE) patients; 50 with active lupus nephritis (LN) and 20 without as well as 20 matched controls. The neutrophil gelatinase-associated lipocalin (NGAL) in both serum and urine samples was measured by enzyme-linked immunosorbent assay (ELISA). Patients with active LN received standard treatment then assessed for response as well as the value of urinary NGAL (uNGAL). Our results revealed that, The SLE patients with or without LN had an elevated urinary NGAL as compared to controls (p < 0.000) and the mean of uNGAL was (20.67 ± 5.34),(10.63 ± 3.53),(5.65 ± 2.49) respectively. Furthermore,Urinary NGAL levels in LN patients were significantly higher than those in non-LN patients (P < 0.0001). In the ROC curve analysis , the diagnostic performance of uNGAL for discriminating patients with nephritis from those without nephritis showed that the best cutoff value was 13.66 ng/ml ,sensitivity 92%,specificity 75%,area undercurve (0.959) and (P < 0.0001). Measurement of urinary NGAL levels showed an excellent diagnostic performance for discriminating patients with LN from SLE without nephritis.

Low level of circulating basophil counts in biopsy-proven active lupus nephritis.

Basophils have been shown to be important players in promoting lupus nephritis (LN). However, the relationship between circulating basophil counts and renal pathology activity of LN remains unclear. In this retrospective study, 159 clinical and pathology samples from patients with biopsy-proven LN were analyzed. The renal activity and classification were evaluated according to renal pathology. The correlations between circulating basophil counts and renal pathology activity index were assessed. Overall, circulating basophil counts correlated with total systemic lupus erythematosus disease activity index (SLEDAI) score (r=-0.31), renal SLEDAI score (r=-0.35), activity index (AI) score(r=-0.40), and renal histologic activity parameters (p<0.05, respectively). Compared with systemic lupus erythematosus (SLE) non-LN patients, the LN group had lower basophil counts (0.007±0.007 vs. 0.011±0.010×109/L, p=0.04). Subgroup analyses revealed that the circulating basophil counts in group B (AI>8) were significantly lower than that in group A (AI≤8) (0.004±0.006 vs. 0.009±0.009×109/L, p<0.001). The difference was still significant when eliminating the influence of SLEDAI. Significant differences were found in circulating basophil counts among LN pathology classification groups (p<0.01). Groups of classes III, IV, and V were more likely to have lower circulating basophil counts when compared with group of class I/II (p<0.05). These findings suggest a potential role of circulating basophil counts as a convenient and helpful marker for renal activity of LN.

A functional SNP MCP-1 (−2518A/G) predispose to renal disorder in Indian Systemic Lupus Erythematosus patients

Systemic Lupus Erythematosus (SLE) is a clinically heterogeneous chronic, inflammatory autoimmune disorder that affects multiple organs where exact etiology of the disease is not yet clearly understood. Various evidences suggest that genetic polymorphisms in inflammatory mediators like cytokines and chemokines may influence development of the disease. Here, we investigated whether functional polymorphism at the Monocyte Chemoattractant Protein-1 (MCP-1) regulatory region associates with disease phenotype in Indian SLE patients. This case control study included 200 SLE patients and 201 ethnically matched healthy controls. Genotyping of MCP-1 (−2518 A/G) polymorphism was performed using PCR-RFLP method. Serum MCP-1 levels were detected by bead-based multiplex immunoassay. Serum MCP-1 levels were found to be higher in patients compared with healthy individuals (p<0.0001). A significant difference for MCP-1G allele frequency (OR=1.9, 95%CI=1.4–2.6, p<0.0001) was observed among SLE patients against healthy individuals. A significant difference in the distribution of MCP-1 −2518GG (OR=3.0, 95%CI=1.4–6.7, p=0.0041) and AG+GG genotypes (OR=2.0, 95%CI=1.4–3.0, p=0.0005) was also noted among SLE patients when compared with healthy individuals. A significant association was observed between A/G and G/G versus A/A genotypes with renal manifestations (p<0.0001, Pc<0.001). Serum MCP-1 levels in active LN patients were found to be significantly higher than inactive LN (p=0.0059), mild LN (p=0.0061) as well as non-LN patients (p=0.0001). These findings suggest that −2518G allele of MCP-1 −2518 A/G polymorphism is associated with renal disorders and may influence MCP-1 gene expression among Indian SLE patients.

Pentraxin-3 and its Association with C1q-CIC, hsCRP and Pro-Inflammatory Cytokines (TNF-Α and IL-1β) among SLE Patients from India

Introduction: Systemic Lupus Erythematosus (SLE) is a prototype autoimmune disease with alternating periods of flares and remission. Disease pathogenesis involves vast inflammatory responses, characterized by impaired cell signaling and T-cell dysfunction with interplay of cytokines and a wide network of protein cascades. Aim of this study is to understand the correlation between PTX-3 and pro-inflammatory cytokines (TNF-α and IL-1β) and their role in disease pathogenesis of SLE. Materials and Methods: Sixty-three SLE patients classified as per ACR 1997 criteria were included, of which 36 had renal involvement (LN). Autoantibodies were detected by IFA and ANA BLOT techniques. Serum Complement levels and hsCRP (by nephelometer), Pentraxin-3 and C1q-CIC (by ELISA), TNF-α and IL-1β levels (by Multiplex immunoassay) were assessed. Results: ANA was present in 90.5% patients, anti-dsDNA antibodies were present in 87.3% patients. Reduced C3 ( 5 mg/L) in 49.2% patients. C1q-CIC levels were high (>50 μg/ml) in 50.8% patients. Serum PTX-3 levels and TNF-α levels were significantly higher in SLE patients than healthy controls (p<0.0001 and p<0.0001 respectively). It was observed that PTX-3 and IL-1β levels were significantly higher in LN patients as compared with nonLN patients (p=0.0107; p=0.0022 respectively). PTX-3 positively correlates with C1q-CIC and negatively correlates with hsCRP and IL-1β levels among SLE patients. Conclusion: This study suggests that serum PTX-3 though do not induce an immediate inflammatory response, its positive correlation with C1q-CIC levels, suggested its possible role in classical complement pathway activation. Further studies in this regard are needed to understand the role of PTX-3 in pathogenesis of SLE.

The Level of IgA Antibodies to Endothelial Cells Correlates with Histological Evidence of Disease Activity in Patients with Lupus Nephritis.

Anti-endothelial cell antibodies (AECA) are frequently detected in patients with systemic lupus erythematosus (SLE), but their pathological role remains unclear. We recently developed a solubilized cell surface protein capture enzyme-linked immunosorbent assay (CSP-ELISA) to detect antibodies against membrane proteins involved in autoimmune reactions. In this study, sera from 51 patients with biopsy-proven lupus nephritis (LN), 25 with SLE without renal involvement (non-LN SLE), 42 disease control (DC) subjects, and 80 healthy control (HC) subjects were tested for IgG- and IgA-AECA for human umbilical vein endothelial cells (HUVEC) and human glomerular EC (HGEC) by using CSP-ELISA. IgG- and IgA-AECA titers were significantly higher in LN and non-LN SLE patients than in the DC or HC (P < 0.001) groups. IgG- and IgA-AECA titers for HUVEC corresponded well with those for HGEC. The IgA-AECA level correlated with the SLE disease activity index and with histological evidence of active lesions (cellular proliferations, hyaline thrombi and wire loops, leukocytic infiltration, and fibrinoid necrosis) in LN patients (P < 0.001). The sensitivity of IgA-AECA as a diagnostic test for histological evidence of active lesions in LN patients was 0.92, with a specificity of 0.70. The significant correlation of IgA-AECA with glomerular hypercellularity indicates that IgA-AECA are associated with endothelial damage in LN.

A discrete cluster of urinary biomarkers discriminates between active systemic lupus erythematosus patients with and without glomerulonephritis.

BackgroundManagement of lupus nephritis (LN) would be greatly aided by the discovery of biomarkers that accurately reflect changes in disease activity. Here, we used a proteomics approach to identify potential urinary biomarkers associated with LN.MethodsUrine was obtained from 60 LN patients with paired renal biopsies, 25 active non-LN SLE patients, and 24 healthy controls. Using Luminex, 128 analytes were quantified and normalized to urinary creatinine levels. Data were analyzed by linear modeling and non-parametric statistics, with corrections for multiple comparisons. A second cohort of 33 active LN, 16 active non-LN, and 30 remission LN SLE patients was used to validate the results.ResultsForty-four analytes were identified that were significantly increased in active LN as compared to active non-LN. This included a number of unique proteins (e.g., TIMP-1, PAI-1, PF4, vWF, and IL-15) as well as known candidate LN biomarkers (e.g., adiponectin, sVCAM-1, and IL-6), that differed markedly (>4-fold) between active LN and non-LN, all of which were confirmed in the validation cohort and normalized in remission LN patients. These proteins demonstrated an enhanced ability to discriminate between active LN and non-LN patients over several previously reported biomarkers. Ten proteins were found to significantly correlate with the activity score on renal biopsy, eight of which strongly discriminated between active proliferative and non-proliferative/chronic renal lesions.ConclusionsA number of promising urinary biomarkers that correlate with the presence of active renal disease and/or renal biopsy changes were identified and appear to outperform many of the existing proposed biomarkers.Electronic supplementary materialThe online version of this article (doi:10.1186/s13075-016-1120-0) contains supplementary material, which is available to authorized users.

International validation of a urinary biomarker panel for identification of active lupus nephritis in children

BackgroundConventional markers of juvenile-onset systemic lupus erythematosus (JSLE) disease activity fail to adequately identify lupus nephritis (LN). While individual novel urine biomarkers are good at detecting LN flares, biomarker panels may improve diagnostic accuracy. The aim of this study was to assess the performance of a biomarker panel to identify active LN in two international JSLE cohorts.MethodsNovel urinary biomarkers, namely vascular cell adhesion molecule-1 (VCAM-1), monocyte chemoattractant protein 1 (MCP-1), lipocalin-like prostaglandin D synthase (LPGDS), transferrin (TF), ceruloplasmin, alpha-1-acid glycoprotein (AGP) and neutrophil gelatinase-associated lipocalin (NGAL), were quantified in a cross-sectional study that included participants of the UK JSLE Cohort Study (Cohort 1) and validated within the Einstein Lupus Cohort (Cohort 2). Binary logistic regression modelling and receiver operating characteristic curve analysis [area under the curve (AUC)] were used to identify and assess combinations of biomarkers for diagnostic accuracy.ResultsA total of 91 JSLE patients were recruited across both cohorts, of whom 31 (34 %) had active LN and 60 (66 %) had no LN. Urinary AGP, ceruloplasmin, VCAM-1, MCP-1 and LPGDS levels were significantly higher in those patients with active LN than in non-LN patients [all corrected p values (pc) < 0.05] across both cohorts. Urinary TF also differed between patient groups in Cohort 2 (pc = 0.001). Within Cohort 1, the optimal biomarker panel included AGP, ceruloplasmin, LPGDS and TF (AUC 0.920 for active LN identification). These results were validated in Cohort 2, with the same markers resulting in the optimal urine biomarker panel (AUC 0.991).ConclusionIn two international JSLE cohorts, urinary AGP, ceruloplasmin, LPGDS and TF demonstrate an ‘excellent’ ability for accurately identifying active LN in children.

Immune cells and type 1 IFN in urine of SLE patients correlate with immunopathology in the kidney

The immunopathological events in the kidneys of lupus nephritis (LN) patients are poorly understood due in part to the difficulty in acquiring serial biopsies and the inherent limitations in their analysis. To identify a means to circumvent these limitations, we investigated whether immune cells of kidney origin are present in patient urine and whether they correlate with kidney pathology. Flow cytometry analysis was performed on peripheral blood and urine cells of 69 SLE patients, of whom 41 were LN patients. In addition, type I IFN (IFNα/β) levels were determined in plasma and urine by bioassay. Approximately 60% of non-LN patients had urine lymphocytes. In these patients, T cells were always present and predominantly CD8+, while B cells were either absent or a mixture of naïve and memory B cells. In contrast, >90% of LN patients had urine lymphocytes. In half, the B and T cells resembled those in non-LN patient urine; however, in the remaining patients, the B cells were exclusively Ig-secreting plasmablasts or plasma cells (PB/PCs) and the T cells were predominantly CD4+. In addition, pDCs and IFNα/β frequently accompanied PB/PCs. The majority of patients with urine PB/PCs presented with proliferative nephritis and a significant loss of kidney function, which in some cases had progressed to end stage renal disease (ESRD). In conclusion, urine can provide access to cells of kidney resident populations for phenotypic and functional characterization. Analysis of these cells provides insight into the kidney immunopathology and may serve as biomarkers to identify patients at risk for developing LN and progressing to ESRD.

Genetic Polymorphism (rs329498) in the Pellino-1 Gene as Possible Predisposal Factor for Systemic Lupus Erythematosus in a Chinese Population

ABSTRACTObjective: The purpose of this study was to analyze the association of two single nucleotide polymorphisms (SNPs) in Peli-1 gene with systemic lupus erythematosus (SLE) in a Chinese population.Methods: We conducted a case–control study and a total of 738 SLE patients and 827 healthy controls were finally recruited. Peli-1 rs329498 and rs10496105 polymorphisms were specified from genomic DNA using TaqMan genotyping assay on Fluidigm 192.24 system.Results: Allele contrast showed the minor allele C was associated with decreased risk for SLE when compared with the A allele (OR = 0.851, 95% CI = 0.737–0.983, p = 0.028). Significant difference was observed in genotype distribution of rs329498 polymorphism between lupus nephritis (LN) patients and non-LN patients (χ2 = 8.18, p = 0.017). Furthermore, we also found a decreased frequency of the minor allele C in LN patients (29.2%) than in non-LN patients (37.7%) (χ2 = 8.67, p = 0.003). Moreover, a significant difference was also detected under a dominant model with regard to the distribution of genotype frequencies between LN patients and non-LN patients (CC + AC vs. AA: OR = 0.632, 95% CI = 0.451–0.884, p = 0.007). Clinical features analysis showed a significant difference in the distribution of genotypic frequencies between patients with malar rash and patients without this feature (χ2 = 6.63, p = 0.036). Unfortunately, we failed to find any significant results between Peli-1 gene rs10496105 and SLE susceptibility.Conclusions: Our observations suggested that Peli-1 gene polymorphism rs329498 might contribute to SLE susceptibility in Chinese Han Population. Likewise, the rs329498 SNP was also associated with the clinical features LN and malar rash in SLE patients.

Diagnostic value of progranulin in patients with lupus nephritis and its correlation with disease activity

The aim of this study was to explore whether progranulin (PGRN) can be a useful marker not only for accurate diagnosis of patients with active lupus nephritis (LN), but also for prediction of the disease activity in this population. A total of 154 LN patients were enrolled in this study, 76 of which were diagnosed as having active LN and 78 as having stable LN. Additionally, 71 age-matched non-LN patients were enrolled as controls. The serum and urine PGRN levels of each study population were measured using the enzyme-linked immunosorbent assay method. The diagnostic performance of both indicators and their correlation with the disease activity of LN were systematically investigated using receiver operating characteristic (ROC) analysis and correlation analysis. The active LN population had significantly higher serum and urine PGRN levels than the other two populations. ROC analysis further demonstrated that these two indicators, particularly in combination, appear to have a strong performance in discriminating active LN patients from the rest of the LN population. In the active LN population, serum and urine PGRN levels were not only significantly correlated with SLEDAI score, rSLEDAI score, and activity index, but also had a considerable association with several key markers reflecting the disease activity of LN, including serum levels of complement component 3 and ds-DNA. Nevertheless, neither of the two indicators were correlated with the pathological classification of LN, chronicity index, serum creatinine level, and 24-h urine protein levels. Our findings demonstrate that PGRN may have great potential as a diagnostic factor for active LN and as a predictor for its disease activity.