Renal Pathological Activity Research Articles

V-set immunoglobulin domain-containing protein 4 (VSIG4) as a potential non-invasive biomarker of lupus nephritis

Abstract Objective: This study aimed to identify a non-invasive biomarker of lupus nephritis (LN) for diagnosis, disease monitoring or flare prediction of LN. Methods: Both the VSIG4 free form and immune complex (ICx) bound form in the urine of LN patients (N = 97) were examined using ELISA, to compare with chronic kidney diseases (CKD) controls and healthy controls. Immunohistochemistry was used to confirm the elevated tissue expression of VSIG4 in the renal biopsy of LN. Immunofluorescence was used to examine the VSIG4-mediated immune complex deposition in the kidney of LN. Results: The urinary levels of both VSIG4 free form (P = 0.01) and VSIG4-ICx (P = 6.3 x 10-14) and were significantly elevated in LN patients, particularly in renal active LN (P = 4.8 x 10-16) compared to healthy controls. VSIG4-ICx effectively discriminated HC from LN (AUC = 0.84, P< 0.01) and active LN from inactive LN (AUC = 0.91, P< 0.01). The urinary immune complex form of VSIG4 was positively correlated with renal pathology activity index (AI). Immunohistochemistry highlighted the significantly elevated VSIG4 expression in both glomeruli and tubules in LN patients, compared to healthy controls. Conclusion: Our findings underscore the diagnostic potential of VSIG4 as a non-invasive urinary biomarker for LN. With its correlation to disease activity and renal pathology, VSIG4 presents a promising avenue for enhanced diagnosis and monitoring of lupus nephritis, paving the way for improved patient care.

Risk factors and prognosis of pediatric silent lupus nephritis with class Ⅲ to Ⅴ

To study the risk factors and prognostic characteristics of pediatric silent lupus nephritis (SLN) with class Ⅲ to V. A retrospective study was conducted to collect clinical data from 30 children diagnosed with SLN at the Department of Pediatrics, Second Xiangya Hospital, Central South University, from May 2007 to April 2023. Based on renal pathological classification, the patients were divided into a class Ⅱ group (12 cases) and a class Ⅲ to Ⅴ group (18 cases). The risk factors for the occurrence of class Ⅲ to Ⅴ SLN were analyzed, and the prognostic characteristics were summarized. Among the 30 SLN patients, the median follow-up time was 61.50 months. There were no statistically significant differences in the proportions of patients who discontinued glucocorticoids or achieved low disease activity status, nor in the annual decline rate of estimated glomerular filtration rate (eGFR) between the class Ⅱ and class Ⅲ to V groups (P>0.05). However, three patients in the class Ⅱ group progressed to stage 1 chronic kidney disease (CKD), while eight patients in the class III to V group reached stage 1 CKD, and four patients reached stage 2 CKD. Among the 26 female SLN patients, serum complement C3 levels in the class III to V group were lower than those in the class Ⅱ group (P<0.05). Serum C3 levels in SLN patients, as well as in female SLN patients, were negatively correlated with the fluorescence intensity of IgA, IgG, and C3 immune complexes in the kidneys (P<0.05). Additionally, serum C3 levels in female SLN patients were negatively correlated with the renal pathological activity index (P<0.05). Binary logistic regression analysis indicated that being female and having low serum complement C3 levels were risk factors for the occurrence of class Ⅲ to V SLN in children (P<0.05). Class Ⅲ to V SLN is not uncommon among SLN children, and there remains a risk of long-term renal function progression. Being female and having low serum complement C3 levels are identified as risk factors for class Ⅲ to V SLN in children.

Association of Mitochondrial Pyruvate Carrier with the Clinical and Histological Features in Lupus Nephritis.

Mounting evidence suggests that mitochondrial dysfunction contributes to lupus nephritis (LN) pathogenesis. Mitochondrial pyruvate carrier 1 (MPC1) and mitochondrial pyruvate carrier 2 (MPC2) mediating pyruvate transport from the cytoplasm to the mitochondrial matrix, determines the cell survival and cellular energy supply. Here, we aimed to investigate the association of mitochondrial pyruvate carrier expression with the clinical and histological features in LN. Patients with biopsy-proven proliferative LN (class III and class IV, n=18) and membranous LN (class V, n=18) were included. Expression of MPC1 and MPC2 were examined by immunohistochemistry. MPC protein levels in the two groups were evaluated by the Student's t-test. Correlation analysis between MPC levels and clinicopathological features was performed by Spearman's rank correlation. Both MPC1 and MPC2 were exclusively expressed in renal tubules of enrolled LN. Significantly lower MPC1 and MPC2 were observed in patients with proliferative LN compared to membranous LN. In addition, the MPC1 and MPC2 were negatively correlated with SLEDAI-2K score, renal function, and renal pathology activity index. Both MPC1 and MPC2 were localized in renal tubules, and decreased MPC content was more pronounced in proliferative LN than membranous LN. MPC levels were significantly correlated with renal functions and renal pathology activity.

Symposium 1

Proteomics has been a very informative tool in uncovering several biomarkers that may have clinical utility in managing patients with lupus nephritis (LN). Both non-targeted (mass spectrometry-based) and targeted (ligand-based) proteomics point to promising biomarkers in the urine of LN patients. Aptamer-based interrogation of 1129 proteins using the Somascan platform, followed by ELISA-validation in two independent cohorts has indicated that the urine proteins that best distinguish active LN from controls were ALCAM, PF-4, properdin, and VCAM-1 among African Americans, E-selectin, VCAM-1, BFL-1 and Hemopexin among Caucasians, and ALCAM, VCAM-1, TFPI and PF-4 among Asians (PMID: 32366845). Most of these urine proteins correlate significantly with disease activity indices and surpass conventional metrics in identifying active LN. Several elevated urinary molecules are also expressed within LN kidneys, based on single-cell RNA-seq analysis. Of these, ALCAM has been validated in a large multi-ethnic, multi-national cohort of LN subjects, and has also been shown to be a pathogenic driver and therapeutic target in LN (PMID: 34981775). Antibody array based screening of 1000 proteins using the Raybiotech Kiloplex platform has identified urine L-selectin, Angptl4, TGF[Formula: see text] 1, TPP1, TSP-1, FOLR2, PDGFR[Formula: see text], and PRX2, as markers of active renal lupus, outperforming conventional metrics, with L-selectin, Angptl4 and TGF[Formula: see text] 1 tracking well with concurrent or pending disease flares in LN (PMID: 32651195). Of these, urine L-selectin has been validated in large multi-ethnic/national cohorts of LN subjects (under review). Proteomic screens of LN urine using an electrochemiluminescence platform (Mesoscale/MSD) has identified IL-7, IL-12p40, IL-15, IP-10, and TARC as markers of active renal lupus that correlated significantly with disease activity (PMID: 30618193). Very recent global proteomic screens of LN urine using a proximity ligation assay platform from Olink has added several additional biomarker candidates that have been successfully ELISA-validated (under review). Given the plethora of urine biomarkers that have been identified using proteomics, it now becomes important to validate the clinical utility of these urine proteins, using multiple well-annotated patient cohorts, across ethnicities. Among the completed studies, we now know that some urine proteins are diagnostic of concurrent renal pathology LN Class or Activity Index or Chronicity index (e.g. CD163: PMID: 32351512; VCAM-1: PMID: 29076253; Angiostatin: PMID: 29076253; Hemopexin, Kim-1 and Lipoxcalin-2/NGAL; Renin and SAP; PMID: 20065116), while others exhibit prognostic value (e.g., ALCAM and VCAM-1; PMID: 31722419). Ongoing studies also show that most of the urine proteins interrogated in LN are not elevated in circulation (PMID: 36091001), but are expressed within LN kidneys, as ascertained by immunohistochemistry (Clinical Immunology, 2022) and Imaging Mass Cytometry (Tissue CyTOF). To facilitate monitoring of urinary biomarkers in outpatient clinics or to enable self-monitoring of disease status by the patient from the comfort of one’s home, we have recently engineered a smartphone-readable nanophosphor-based lateral flow assay for point of care monitoring of urine ALCAM (Frontiers in Immunology, 2022). The long-term goal is to decipher the clinical utility of different urine biomarkers in managing lupus and LN in the clinics, to empower LN patients with easy-to-use tools to monitor their own disease status, and to understand the mechanistic significance of the uncovered molecules in renal disease pathogenesis. Ongoing research on this front is summarized at: https://mohanlab.bme.uh.edu/ .

The ratio of neutrophil to lymphocyte as a potential marker of clinicopathological activity for lupus nephritis.

The ratio of neutrophil to lymphocyte (NLR) is a novel inflammatory factor that is elevated in systemic lupus erythematosus (SLE). However, the relationship between NLR and renal pathological manifestations in patients with lupus nephritis (LN) has not been investigated. A retrospective study included 240 SLE patients, in which 186 patients with renal involvement and 124 LN patients underwent renal biopsy, 125 healthy volunteers and 125 chronic kidney disease (CKD) controls. Patients with SLE disease activity 2000 (SLEDAI-2K) > 9 and ≤ 9 were defined as severely active and mildly active, respectively. Clinical parameters and renal pathological data were collected from medical records. The correlations between NLR and clinicopathological features were analyzed. The NLR of SLE group was significantly higher than that of the sex-age matched control groups. Patients with nephritis had higher NLR levels than those without nephritis (P = 0.044). Increased NLR was observed in severely active group compared to mildly active group (P = 0.020). NLR was significantly positively related with SLEDAI score, Renal SLEDAI score, C-reactive protein (CRP), 24-h urine protein, renal activity index (AI), cellular crescents and tubular atrophy, and negatively correlated with serum albumin. NLR was significantly decreased after treatment. Based on the receiver operating characteristic (ROC) curve, the best NLR cut-off value to predict severe activity of SLE and cellular crescents in renal pathology was 2.19 and 3.16, respectively. NLR may be a non-invasive and potential inflammatory marker in evaluating clinical and renal pathological activity in LN patients.

V-Set Immunoglobulin Domain-Containing Protein 4 as a Novel Serum Biomarker of Lupus Nephritis and Renal Pathology Activity.

To discover novel serum biomarkers that have diagnostic or predictive value in lupus nephritis (LN). Using a quantitative protein microarray, we screened for high-abundant proteome expression in the serum of patients with LN compared to healthy controls. Top candidates from this screening were validated using a larger cohort of patients with LN compared to a disease control cohort (subjects with other chronic kidney diseases) and a healthy control cohort. Promising markers were then selected using a machine-learning model and further validated with a larger patient cohort. The corresponding autoantibodies and immune complexes containing these proteins were also examined. In total, 13 proteins were found to be significantly elevated in LN patient serum in the screening, among which 8 proteins were validated by enzyme-linked immunosorbent assay using 81 serum samples from LN patients and control subjects. Three serum markers with LN diagnostic potential were identified using feature importance analysis and further validated using 155 serum samples from LN patients and control subjects. V-set immunoglobulin domain-containing protein 4 (VSIG4) appeared to be the most promising marker in distinguishing LN from healthy controls, with an area under the curve of 0.93. VSIG4 could also discriminate active LN from inactive LN. Furthermore, serum VSIG4 levels were positively correlated with all of the following clinical parameters: the Systemic Lupus Erythematosus Disease Activity Index (SLEDAI) score (Spearman's rank correlation rs = 0.42, P < 0.001), the renal domain score of the SLEDAI (rs = 0.46, P < 0.001), the urinary protein-to-creatinine ratio (rs = 0.56, P < 0.001), and the serum creatinine level (rs = 0.41, P < 0.001). Importantly, we found that serum VSIG4 levels tracked with LN disease activity longitudinally, and that serum VSIG4 levels reflected the renal pathology activity index (AI), particularly the AI components of crescent formation and hyaline deposits. VSIG4 may be a promising novel serum biomarker and therapeutic target in patients with LN.

Renal C4d is a potential biomarker of disease activity and severity in pediatric lupus nephritis patients.

Systemic lupus erythematosus (SLE), a multisystemic autoimmune disease, is very aggressive in pediatric-onset patients as they are prone to develop lupus nephritis (LN). Although renal C4d positivity is correlated with the activity of renal disease and SLE in adult-onset LN patients, available information for pediatric-onset patients is limited. To evaluate the potential diagnostic significance of renal C4d staining in pediatric LN patients, we retrospectively detected C4d staining by immunohistochemistry on renal biopsy specimens from 58 pediatric LN patients. The clinical and laboratory data at the time of the kidney biopsy and the renal disease activity of histological injury were analyzed according to the C4d staining status. Glomerular C4d (G-C4d)-positive staining was detected in all 58 cases of LN. Patients with a G-C4d score of 2 displayed more severe proteinuria than those with a G-C4d score of 1 (24-h urinary protein: 3.40 ± 3.55 g vs. 1.36 ± 1.24 g, P < 0.05). Peritubular capillary C4d (PTC-C4d) positivity was found in 34 of 58 LN patients (58.62%). The PTC-C4d-positive patient groups (patients with a PTC-C4d score of 1 or 2) had higher serum creatinine and blood urea nitrogen levels as well as renal pathological activity index (AI) and SLE disease activity index (SLEDAI) scores; however, they had lower serum complement C3 and C4 levels compared to PTC-C4d-negative patients (P < 0.05). In addition, there was positive tubular basement membrane C4d (TBM-C4d) staining in 11 of 58 LN patients (18.96%), and a higher proportion of TBM-C4d-positive patients than TBM-C4d-negative patients (63.63% vs. 21.27%) had hypertension. Our study revealed that G-C4d, PTC-C4d, and TMB-C4d were positively correlated with proteinuria, disease activity and severity, and hypertension, respectively, in pediatric LN patients. These data suggest that renal C4d is a potential biomarker for disease activity and severity in pediatric LN patients, providing insights into the development of novel identification and therapeutic approaches for pediatric-onset SLE with LN.

Risk factors of childhood systemic lupus erythematosus with thyroid dysfunction

Objective: To investigate the risk factors of childhood systemic lupus erythematosus (SLE) with thyroid dysfunction and to explore the relationship between thyroid hormone and kidney injury of lupus nephritis (LN). Methods: In this retrospective study, 253 patients who were diagnosed with childhood SLE and hospitalized in the First Affiliated Hospital of Zhengzhou University from January 2019 to January 2021 were enrolled in the case group, and 70 healthy children were the control cases. The patients in the case group were divided into the normal thyroid group and the thyroid dysfunction group. Independent t-test, χ2 test, and Mann-Whitney U test were used for comparison between the groups, Logistic regression analysis was used for multivariate analysis, and Spearman correlation. Results: A total of 253 patients, there were 44 males and 209 females in the case group, and the age of onset was 14 (12, 16) years; a total of 70 patients, 24 males and 46 females were in the control group, and the age of onset was 13 (10, 13) years. The incidence of thyroid dysfunction in the case group was higher than that in the control group (48.2% (122/253) vs. 8.6% (6/70), χ²=36.03, P<0.05). Of the 131 patients, there were 17 males and 114 females in the normal thyroid group, and the age of onset was 14 (12, 16) years. Of the 122 patients in the thyroid dysfunction group, 28 males and 94 females were in the thyroid dysfunction group, and the age of onset was 14 (12, 16) years. Of the 122 had thyroid dysfunction, including 51 cases (41.8%) with euthyroid sick syndrome, 25 cases (20.5%) with subclinical hypothyroidism, 18 cases (14.8%) patients with sub-hyperthyroidism, 12 cases (9.8%) with hypothyroidism, 10 cases (8.2%) with Hashimoto's thyroiditis, 4 cases (3.3%) with hyperthyroidism, and 2 cases (1.6%) with Graves disease. Compared to patients with normal thyroid function, the serum level of triglyceride, total cholesterol, urine white blood cell, urine red blood cell, 24 h urine protein, D-dimer, and fibrinogen, ferritin and systemic lupus erythematosus disease activity Index-2000 (SLEDAI-2K) score were higher in patients with thyroid dysfunction (Z=3.07, 3.07, 2.48, 3.16, 2.40, 3.99, 2.68, 2.55, 2.80, all P<0.05), while the serum level of free thyroxine and C3 were lower in thyroid disfunction patients (10.6 (9.1, 12.7) vs. 11.3 (10.0, 12.9) pmol/L, and 0.46 (0.27, 0.74) vs. 0.57 (0.37, 0.82) g/L, Z=2.18, 2.42, both P<0.05). The higher level of triglyceride and D-dimer were the independent risk factors for childhood SLE with thyroid dysfunction (OR=1.40 and 1.35, 95%CI 1.03-1.89 and 1.00-1.81, respectively, both P<0.05). There were 161 patients with LN in the case group, all of which were conducted with renal biopsies, including 11 cases (6.8%) with types Ⅰ LN, 11 cases (6.8%) with typesⅡLN, 31 cases (19.3%) with types Ⅲ LN, 92 cases (57.1%) with types Ⅳ LN, and 16 cases (9.9%) with types Ⅴ LN. There were significant differences in the level of free triiodothyronine and thyroid stimulating hormone among different types of kidney pathology (both P<0.05); compared with types I LN, the serum level of free triiodothyronine was lower in types Ⅳ LN (3.4 (2.8, 3.9) vs. 4.3 (3.7, 5.5) pmol/L, Z=3.75, P<0.05). The serum level of free triiodothyronine was negatively correlated with the acute activity index score of lupus nephritis (r=-0.228, P<0.05), while the serum level of thyroid stimulating hormone was positively correlated with the renal pathological acute activity index score of lupus nephritis (r=0.257, P<0.05). Conclusions: There is a high incidence of thyroid dysfunction in childhood SLE patients. The higher SLEDAI and more severe renal damage were found in SLE patients with thyroid dysfunction compared to these with normal thyroid functions. The risk factors of childhood SLE with thyroid dysfunction are the higher level of triglyceride and D-dimer. The serum level of thyroid hormone is possibly related to the kidney injury of LN.

Renal NLRP3 Inflammasome activation is associated with disease activity in lupus nephritis

The current study was initiated to comprehensively evaluate renal NLRP3 inflammasome pathway activation in lupus nephritis (LN) patients and their clinicopathological significances based on a Chinese LN cohort. We found that the expressions of NLRP3, ASC, caspase-1, IL-1β and IL-18 were all significantly higher in the kidneys of LN patients and were predominantly expressed in glomerular mesangial cells, podocytes, renal tubular epithelial cells and macrophages. The expressions of NLRP3, ASC, caspase-1 and IL-1β were positively correlated to SLEDAI scores and several renal pathological activity indices, while the expression of NLRP3 was negatively associated with chronicity scores. Moreover, the foot process width was positively correlated with glomerular caspase-1 levels, and several podocyte injury markers were decreased significantly in LN patients with higher caspase-1 expression compared with those with lower expression. Our findings indicated that renal NLRP3 inflammasome was activated in LN patients and correlated with disease activity, which needs further explorations.

A novel technology for home monitoring of lupus nephritis that tracks the pathogenic urine biomarker ALCAM.

The gold standard for diagnosis of active lupus nephritis (ALN), a kidney biopsy, is invasive with attendant morbidity and cannot be serially repeated. Urinary ALCAM (uALCAM) has shown high diagnostic accuracy for renal pathology activity in ALN patients. Lateral flow assays (LFA) for assaying uALCAM were engineered using persistent luminescent nanoparticles, read by a smartphone. The stability and reproducibility of the assembled LFA strips and freeze-dried conjugated nanoparticles were verified, as was analyte specificity. The LFA tests for both un-normalized uALCAM (AUC=0.93) and urine normalizer (HVEM)-normalized uALCAM (AUC=0.91) exhibited excellent accuracies in distinguishing ALN from healthy controls. The accuracies for distinguishing ALN from all other lupus patients were 0.86 and 0.74, respectively. Periodic monitoring of uALCAM using this easy-to-use LFA test by the patient at home could potentially accelerate early detection of renal involvement or disease flares in lupus patients, and hence reduce morbidity and mortality.

Type I interferon and neutrophil transcripts in lupus nephritis renal biopsies: clinical and histopathological associations.

To investigate the expression of type I IFN (IFN-I) and neutrophil transcripts in kidney tissue from patients with different classes of LN and their association with distinct clinical and histopathological features. Quantitation of IFN-I, defensin-α3 and formyl peptide receptor-like 1 (FPRL-1) transcripts was performed in kidney biopsy tissue from 24 patients with various classes of LN (6 class III, 14 class IV, 4 class V) and 3 control samples. Patient demographics, glomerular filtration rate (eGFR) and histopathological characteristics, including activity and chronicity indices, were analysed. IFNα2 and IFNβ transcripts were overexpressed in renal tissues from patients with proliferative forms of LN (III/IV) compared with patients with membranous nephritis and control kidneys. Patients with LN and impaired renal function, attested by eGFR, displayed higher relative expression of IFNα2 transcripts in renal tissues compared with those with normal renal function (23.0 ± 16.2 vs 12.0 ± 14.8, P = 0.04). Defensin-α3, but not FPRL-1, transcripts were overexpressed in LN tissues, particularly those with segmental necrotizing lesions, and were correlated with higher renal pathological activity indices (r = 0.61, P = 0.02), urinary protein levels (r = 0.44, P = 0.048) and IFNα2 expression (r = 0.50, P = 0.01). IFN-I transcripts are expressed locally in kidneys from patients with proliferative LN and are associated with impaired renal function. Elevated defensin-α3 transcripts, a neutrophil product associated with neutrophil extracellular traps, may identify a driver of local IFN-I expression. These findings provide insights into the mechanisms of proliferative LN and may inform therapeutic decisions regarding selection of IFN-I pathway inhibitors.

Exploring urine:serum fractional excretion ratios as potential biomarkers for lupus nephritis.

ObjectivesThe goal of this exploratory study is to determine if urine:serum fractional excretion ratios can outperform the corresponding urinary biomarker proteins in identifying active renal disease in systemic lupus erythematosus (SLE).MethodsThirty-six adult SLE patients and twelve healthy controls were examined for serum and urine levels of 8 protein markers, namely ALCAM, calpastatin, hemopexin, peroxiredoxin 6 (PRDX6), platelet factor 4 (PF4), properdin, TFPI and VCAM-1, by ELISA. Fractional excretion of analyzed biomarkers was calculated after normalizing both the urine and serum biomarker levels against creatinine. A further validation cohort of fifty SLE patients was included to validate the initial findings.ResultsThe FE ratios of all 8 proteins interrogated outperformed conventional disease activity markers such as anti-dsDNA, C3 and C4 in identifying renal disease activity. All but VCAM-1FE were superior to the corresponding urine biomarkers levels in differentiating LN activity, exhibiting positive correlation with renal SLEDAI. ALCAMFE, PF4FE and properdinFE ratios exhibited the highest accuracy (AUC>0.9) in distinguishing active LN from inactive SLE. Four of the FE ratios exhibited perfect sensitivity (calpastatin, PRDX6, PF4 and properdin), while ALCAMFE, PF4FE and properdinFE exhibited the highest specificity values for active LN. In addition, several of these novel biomarkers were associated with higher renal pathology activity indices. In the validation cohort ALCAMFE, PF4FE and properdinFE once again exhibited higher accuracy metrics, surpassing corresponding urine and serum biomarkers levels, with ALCAMFE exhibiting 95% accuracy in distinguishing active LN from inactive SLE.ConclusionsWith most of the tested proteins, urine:serum fractional excretion ratios outperformed corresponding urine and serum protein measurements in identifying active renal involvement in SLE. Hence, this novel class of biomarkers in SLE ought to be systemically evaluated in larger independent cohorts for their diagnostic utility in LN assessment.

Urinary CD163 is a marker of active kidney disease in childhood-onset lupus nephritis.

The objective of this study was to evaluate the utility of urine CD163 for detecting disease activity in childhood-onset SLE (cSLE) patients. Sixty consecutive pediatric patients fulfilling four or more ACR criteria for SLE and 20 healthy controls were recruited for testing of urinary CD163 using ELISA. SLE disease activity was assessed using the SLEDAI-2K. Urine CD163 was significantly higher in patients with active LN than inactive SLE patients and healthy controls, with receiver operating characteristics area under the curve values ranging from 0.93 to 0.96. LN was ascertained by kidney biopsy. Levels of CD163 significantly correlated with the SLEDAI, renal SLEDAI, urinary protein excretion and C3 complement levels. Urine CD163 was also associated with high renal pathology activity index and chronicity index, correlating strongly with interstitial inflammation and interstitial fibrosis based on the examination of concurrent kidney biopsies. Urine CD163 emerges as a promising marker for identifying cSLE patients with active kidney disease. Longitudinal studies are warranted to validate the clinical utility of urine CD163 in tracking kidney disease activity in children with lupus.

Anti-DFS70 Antibodies Are Associated With Proliferative Lupus Nephritis and Renal Pathological Activity.

ObjectiveThe significance of anti-dense fine speckles 70 (DFS70) antibodies in systemic lupus erythematosus (SLE) is still unclear, especially in lupus nephritis (LN) patients. We investigated the prevalence, clinical and pathological relevance of anti-DFS70 antibodies in LN patients.MethodsAnti-DFS70 antibodies were measured using enzyme-linked immunosorbent assays in 377 biopsy-proven LN patients, 268 non-LN SLE patients, 232 chronic kidney disease (CKD) patients, and 78 healthy individuals (HI). Demographic, clinical, and pathological parameters were compared between LN patients with and without anti-DFS70 antibodies. Stepwise multivariable logistic regression was performed to identify covariates associated with anti-DFS70 antibodies.ResultsThe prevalence of anti-DFS70 antibodies in LN (19.6%) was comparable to non-LN SLE patients (19.8%, P=0.9630), but was significantly higher than CKD patients (13.4%, P=0.0468) and HI (9.0%, P=0.0252). Using multivariable logistic regression analysis, the titer of anti-double-stranded DNA (dsDNA) antibodies (adjusted odds ratio=1.002, 95% confidence interval 1.001-1.003, P=0.004) was associated with positive anti-DFS70 antibodies in LN patients. In addition, anti-DFS70 antibodies were more prevalent in proliferative LN (22.0%, 68/309) compared to membrane LN patients (10.2%, 6/59, P=0.0376). Furthermore, LN patients with positive anti-DFS70 antibodies had significantly higher activity index (AI) compared to patients who were negative (8.0 vs 6.0, P=0.0131). However, the chronicity index was similar between the groups (3.0 vs 3.0, P=0.8412).ConclusionAnti-DFS70 antibodies were not associated with LN development in SLE patients but were associated with anti-dsDNA antibodies, proliferative LN, and renal AI. This suggests their potential to serve as a non-histological biomarker for LN subclass and activity status.

MO277THE RATIO OF NEUTROPHIL TO LYMPHOCYTE AS A POTENTIAL MARKER OF CLINICOPATHOLOGICAL ACTIVITY FOR SYSTEMIC LUPUS ERYTHEMATOSUS

Abstract Background and Aims The ratio of neutrophils to lymphocytes (NLR) is a novel inflammatory factor that is elevated in systemic lupus erythematosus (SLE) and related to disease activity. However, the relationship between NLR and renal pathological manifestations in patients with lupus nephritis (LN) has not been studied. Method In this retrospective study, 240 SLE patients were recruited. 186 patients with renal involvement and 124 LN patients underwent renal biopsy. In the meanwhile, control groups included 125 chronic kidney disease (CKD) patients and 125 healthy volunteers. Patients with SLE disease activity 2000 (SLEDAI-2K) &amp;gt;9 and ≤ 9 were defined as severely active and mildly active, respectively. Clinical parameters and pathological data were collected. The correlations between NLR and clinicopathological features were analyzed. Results The NLR of SLE group was significantly higher than that of the sex-age matched control groups. Patients with nephritis had higher NLR levels than those without nephritis [2.88(1.81,4.32) vs. 2.43(1.55,3.90), P=0.044; Figure 1]. Increased NLR was observed in severely active group compared to mildly active group [3.00(1.84,4.28) vs.2.36(1.61,3.51), P=0.020; Figure 1]. NLR was significantly positively related with SLEDAI score (r=0.131, P=0.043), Renal SLEDAI score (r=0.173, P=0.023), C-reactive protein (CRP; r=0.213, P=0.002), 24-hour urine protein (r=0.274, P&amp;lt;0.001; Figure 2A), renal activity index (AI; r=0.192, P=0.033), cellular crescents (r=0.274, P=0.006) and tubular atrophy (r=0.226, P=0.011), and negatively correlated with serum albumin (r=-0.187, P=0.004; Figure 2A). Based on the receiver operating characteristic (ROC) curve, the best NLR cut-off value to predict severe activity and cellular crescents was 2.19 and 3.16, respectively. The ability of NLR to differentiate severely active from mildly active SLE was stronger than CRP and weaker than C3 (Figure 3A). The ability of NLR to predict cellular crescents was better than C3, but not superior to SLEDAI and RSLEDAI. Interestingly, the ROC fitted by NLR and RSLEDAI had a higher AUC and sensitivity [AUC=0.75(0.66,0.84), sensitivity=82.6%, specificity=63.6%; Figure 3B]. Conclusion NLR was a non-invasive and potential inflammatory factor to evaluate clinical and renal pathological activity in patients with SLE.

Association of antiphospholipid antibodies with clinical activity and renal pathological activity in patients with lupus nephritis.

This study aimed to investigate the association of antiphospholipid antibodies (aPL) with clinical activity and renal pathological activity in patients with lupus nephritis (LN). Levels of anticardiolipin () antibodies, anti-β2-glycoprotein I (anti-β2-GPI) antibodies and lupus anticoagulant (LAC) were measured, and other clinical and pathological data were also obtained during the same period before renal biopsy. A total of 83 patients with LN were included in this study, 40 patients (48.2%) in the s positive group and 43 patients in the aPL negative group. LN patients with positive aPL had significantly higher SLEDAI (p = 0.012), more hematuria (p = 0.043), lower serum C3 (p = 0.003) and C4 (p = 0.014), and a higher pathological activity index (p = 0.012), more micro-thrombosis (p = 0.046) and more C3 deposits (p = 0.038) in the glomerulus than patients with negative aPL The level of IgG- was significantly correlated with SLEDAI and serum level of C3 (r = 0.44, p < 0.001; r = -0.39, p = 0.003, respectively). The level of IgM- was significantly correlated with SLEDAI, and serum levels of C3 and C4 (r = 0.27, p = 0.014; r = -0.22, p = 0.041; r = -0.23, p = 0.035, respectively). Our work suggests that aPL, especially, are correlated with both clinical activity and renal pathological activity in patients with LN.

Relationship of MDR1 gene polymorphism and P-glycoprotein expression in Chinese refractory lupus nephritis

To evaluate the association of multidrug resistance 1 (MDR1) polymorphism and the expression of P-glycoprotein (Pgp) in Chinese refractory lupus nephritis (LN) patients. Polymerase chain reaction-direct sequencing was used to analyze MDR1 polymorphism. The genotype distribution of MDR1 polymorphism in 132 SLE (systemic lupus erythematosus) patients was evaluated. ELISA was used to measure the expression of Pgp. Relationship among Pgp expression, MDR1 polymorphism, SLEDAI (SLE disease activity index), and kidney pathological score was analyzed by using One-way ANOVA and Pearson linear correlation. The frequency distribution of the MDR1 gene was consistent with the Hardy-Weinberg equilibrium. Compared with CT and CC, patients with T/T homozygote in MDR1 C3435T had significantly increased Pgp expression in the refractory group (p < 0.05). Additionally, SLEDAI score was positively correlated with Pgp expression (r = 0.481, p < 0.05). Also, Pgp expression was positively correlated with renal pathological activity index (r = 0.76, p < 0.05). MDR1 C3435T polymorphism is significantly associated with Pgp expression in patients with refractory LN. Pgp expression is closely related to SLEDAI and renal pathological score. Thus, Pgp may be useful in evaluation of the prognosis of patients with refractory LN.

Urinary activated leukocyte cell adhesion molecule as a novel biomarker of lupus nephritis histology

BackgroundLupus nephritis (LN) is one of the most severe complications of SLE patients. We aim to validate urinary ALCAM as a biomarker in predicting renal disease histpathology in a Chinese lupus cohort.MethodsIn this cross-sectional study, a total of 256 patients and controls were recruited. Urinary levels of ALCAM were determined by ELISA. Renal histopathology was reviewed by an experienced renal pathologist.ResultsUrinary ALCAM levels were significantly increased in active LN patients when compared to active SLE patients without renal involvement (p < 0.001), inactive LN patients (p = 0.023), inactive SLE patients without renal involvement (p < 0.001), and healthy controls (p < 0.001). Correlation analysis revealed a positive correlation between urinary ALCAM and general disease activity—SLEDAI score (r = 0.487, p < 0.001), as well as renal disease activity—rSLEDAI (r = 0.552, p < 0.001) and SLICC RAS (r = 0.584, p < 0.001). Urinary ALCAM also correlated with lab parameters including 24-h urine protein, hemoglobin, and complement 3. Moreover, urinary ALCAM levels were significantly increased in class III and IV (proliferative) LN as compared to those in class V (membranous) LN. It outperformed conventional biomarkers (anti-dsDNA antibody, C3, C4, proteinuria) in discriminating the two groups of LN. On renal histopathology, urinary ALCAM levels correlated positively with activity index (r = 0.405, p < 0.001) but not chronicity index (r = 0.079, p = 0.448).ConclusionUrinary ALCAM is a potential biomarker for predicting renal pathology activity in LN and may serve as a valuable surrogate marker of renal histopathology.

Soluble TNF-R1, VEGF and other cytokines as markers of disease activity in systemic lupus erythematosus and lupus nephritis.

Current non-invasive methods of assessing disease activity in systemic lupus erythematosus (SLE) are of limited sensitivity and specificity. Testing includes acute phase markers, autoantibodies and complement levels. Although measurements of dsDNA antibodies and complement C3/C4 levels are routine, they remain of limited value. Improved blood and urine markers may help in early detection of flare, distinction between flare and chronic damage, and monitoring response to therapy. A total of 87 patients with SLE were tested for the following cytokines in serum and urine: monocyte chemoattractant protein 1 (MCP-1), regulated upon activation, normal T cell expressed and secreted (RANTES), soluble tumour necrosis factor receptor 1 (sTNF-R1), interferon-inducible protein 10 (IP-10), monocyte inhibitory protein 1α (MIP-1α) and vascular endothelial growth factor (VEGF). Patients attending the Lupus Unit at St Thomas' Hospital, London, UK were divided into active lupus nephritis (LN), inactive LN and non-renal SLE groups based on their renal pathology and SLE disease activity index (SLEDAI). Cytokine testing was performed using the FIDIS multiplex bead assay. The mean level of serum sTNF-R1 was higher in the active LN group compared with both inactive LN and non-renal SLE groups ( p < 0.001). For urine measurements there were significant differences between active LN and non-renal SLE for VEGF ( p = 0.016), after statistical correction for multiple testing. Both urinary and serum sTNF-R1 and IP-10 levels correlated with SLEDAI scores ( p < 0.001), while serum VEGF correlated weakly with SLEDAI ( p = 0.025). The optimum combination for differentiating active from inactive LN patients was serum VEGF, sTNF-R1, MCP-1 and glomerular filtration rate plus urinary sTNF-R1 and protein-creatinine ratio. These results indicate that for active LN, sTNF-R1 could be a useful serum cytokine marker, with potential for VEGF in the urine. This study has confirmed the ability of the multiplex bead technique to detect cytokines in a good analytical range, including very low and high levels, in both serum and urine. Combining serum and urine markers provided additional sensitivity in distinguishing active from inactive LN.

Correlation analysis of TLR9 and IL-23 expression in patients with lupus nephritis.

The aim of this study was to investigate the expression levels of toll-like receptor 9 (TLR9) and interleukin-23 (IL-23) in renal tissue and serum of patients with lupus nephritis (LN), and to explore their clinical correlation. LN patients and healthy controls were enrolled in the experimental group and the control group, respectively. Blood samples, serum, and peripheral blood mononuclear cells (PBMCs) were collected. Renal lesion tissues and adjacent normal tissues of LN patients were harvested from a renal tissue biopsy. Serum level of IL-23 was detected by enzyme-linked immunosorbent assay (ELISA). Expression of IL-23 in PBMCs was determined by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) and Western blot, respectively. Meanwhile, TLR9 expression in renal tissues was accessed by immunohistochemistry staining. Subsequently, 24-h protein urine, renal tubular pathological activity index, erythrocyte sedimentation rate (ESR), serum complement C3 level, and blood albumin level of LN patients were recorded. Also, the correlation between TLR9 expression and these pathological indexes was measured by correlation analysis. Serum level of IL-23 in LN patients was significantly higher than that of healthy controls. Similarly, the mRNA and protein expressions of IL-23 in PBMCs of LN patients were markedly higher than those of healthy controls. IL-23 expression was positively correlated with renal tubular pathological activity index of LN patients. Meanwhile, TLR9 was highly expressed in renal tissues of LN patients. Furthermore, TLR9 expression was positively correlated with 24-h protein urine, renal tubular pathological activity index and ESR, whereas negatively correlated with serum complement C3 level and blood albumin level of LN patients. IL-23 is highly expressed in the serum of LN patients, and its expression is closely related to the occurrence of LN. Also, the expression of TLR9 is up-regulated in the tubulointerstitium of LN patients, which is correlated with relevant clinical indexes.