A Prospective Study on Complement Activation Distinguishes Focal Segmental Glomerulosclerosis from Minimal Change Disease.

Introduction: Minimal change disease (MCD) and focal segmental glomerulosclerosis (FSGS) are related podocytopathies with distinct kidney outcomes. Surprisingly, elevated urinary activation fragments have been found in FSGS despite little complement deposition on immunofluorescence (IF) staining. Whether complement activation distinguishes FSGS from MCD, participating in the development of segmental lesions, remains unknown. Methods: We performed an observational study in patients with MCD and FSGS, and proteinuria ≥1 g/g of creatinine. We included both primary and secondary or unknown causes. We compared urinary fragments of terminal pathway activation, sC5b9, and C5a expressed as creatinine ratios, between MCD and FSGS. Results: Patients with FSGS (n = 41) had a serum albumin of 31±10 g/l and proteinuria of 5.1 (2.6-9.1) g/g at sampling, whereas those with MCD (n = 15) had a lower serum albumin (22 ± 9 g/l; P = 0.002), and a proteinuria of 3.8 (1.9-7.7) g/g (P = 0.40). Urinary sC5b9 and C5a were 8.7 (1.7-52.3) and 1.26 (0.45-1.84) µg/mmol of creatinine, respectively in patients with FSGS; compared to 0.8 (0.0-1.5) and 0.06 (0.01-0.15) µg/mmol of creatinine in MCD (P < 0.001), respectively. We found no association between urinary complement fragments and age, estimated glomerular filtration rate (eGFR), or chronic kidney lesions. When analyzing samples with proteinuria ≥ 3 g/g, the c-statistics for urinary sC5b9 and C5a were 0.96 and 1.00, respectively, in differentiating FSGS from MCD. Conclusion: We found no urinary complement activation fragments in MCD, in comparison to FSGS, despite similar levels of proteinuria. This suggests a role for complement activation in the pathogenesis of FSGS and provides an additional tool for distinguishing these 2 entities.

Evidence from the large VALIGA cohort validates the subclassification of focal segmental glomerulosclerosis in IgA nephropathy.

Evidence from the Oxford IgA nephropathy (IgAN) cohort supports the clinical value of subclassifying focal segmental glomerulosclerosis lesions (S1). Using the larger Validation in IgA (VALIGA) study cohort, we investigated the association between podocytopathic changes and higher proteinuria, kidney outcome and response to immunosuppressive therapy. All biopsies were evaluated for glomeruli with segmental capillary occlusion by matrix ("not otherwise specified", NOS lesion), simple capsular adhesion without capillary occlusion (Adh), tip lesions, and podocyte hypertrophy (PH). S1 required a NOS lesion and/or Adh. A Chi-Squared Automatic Interaction Detection method was used to identify subgroups of FSGS lesions associated with distinctive proteinuria at biopsy. We assessed survival from a combined event (kidney failure or 50% decline in estimated glomerular filtration rate). Finally, we evaluated within each subgroup if immunosuppression was associated with a favorable outcome using propensity analysis. In 1147 patients, S1 was found in 70% of biopsies. Subclassification found NOS lesions in 44%, Adh in 59%, PH in 13%, and tip lesions in 3%, with much overlap. Four subgroups were identified with progressively higher proteinuria: from lowest, S1 without NOS, S1 with NOS but without Adh/PH, to highest, S1 with NOS and Adh but without PH, and S1 with NOS and PH. These four subgroups showed progressively worse kidney survival. Immunosuppression was associated with a better outcome only in the two highest proteinuria subgroups. Propensity analysis in these two groups, adjusted for clinical and pathological findings, found a significantly reduced time-dependent hazard of combined outcome with corticosteroids. Podocyte hypertrophy and glomeruli with simple adhesions appeared to reflect active lesions associated with a response to corticosteroids, while other S1 lesions defined chronicity. Thus, our findings support subclassifying S1 lesions in IgAN.

Focal Segmental Glomerulosclerosis: Assessing the Risk of Relapse.

Introduction: Kidney outcomes are improved in primary focal segmental glomerulosclerosis (FSGS) by maintaining a remission in proteinuria. However, characteristics associated with relapses are uncertain. We sought to identify these by analyzing each remission. Methods: We performed a retrospective study in patients with biopsy-proven lesions of FSGS, absent identifiable secondary cause, who had at least 1 remission from nephrotic-range proteinuria. In each patient, we identified every remission, every relapse, and their durations. Using a multilevel logistic regression to account for the clustering of multiple remissions within a patient, we tested which clinical characteristics were independently associated with relapses. Results: In 203 individuals, 312 remissions occurred, 177 with and 135 without relapse. A minority of remissions were atypical, defined by either absent hypoalbuminemia and/or no immunosuppression (IS), in contrast to the classic nephrotic syndrome that remits with IS. Atypical remission variants were just as likely to relapse as the classical presentation. Only 24% of remission events were on maintenance therapy at relapse. Independent characteristics associated with relapses were higher maximal proteinuria while nephrotic; and in remission, higher nadir proteinuria, lower serum albumin, and higher blood pressure. Using these variables, we created a tool estimating the 1-year risk of relapse ranging from 9% to 80%, well-calibrated to the observed data. Conclusion: In FSGS, relapses are frequent but predictable using independent clinical characteristics. We also provide evidence that atypical presentations remit and relapse following the same pattern as classic FSGS presentations. Treatment strategies to prolong remission duration should be addressed in future trials.

Quantifying the benefits of remission duration in focal and segmental glomerulosclerosis.

BACKGROUND: Although the clinical benefit of obtaining a remission in proteinuria in nephrotic patients with focal segmental glomerulosclerosis (FSGS) is recognized, the long-term value of maintaining it and the impact of relapses on outcome are not well described. METHODS: We examined the impact of remissions and relapses on either a 50% decline in kidney function or end-stage kidney disease (combined event) using time-dependent and landmark analyses in a retrospective study of all patients from the Toronto Glomerulonephritis Registry with biopsy-proven FSGS, established nephrotic-range proteinuria and at least one remission. RESULTS: In the 203 FSGS individuals with a remission, 89 never relapsed and 114 experienced at least one relapse. The first recurrence was often followed by a repeating pattern of remission and relapse. The 10-year survival from a combined event was 15% higher in those with no relapse versus those with any relapse. This smaller than anticipated difference was related to the favourable outcome in individuals whose relapses quickly remitted. Relapsers who ultimately ended in remission (n = 46) versus in relapse (n = 68) experienced a 91% and 32% 7-year event survival (P < .001), respectively. Using time-varying survival analyses that considered all periods of remission and relapse in every patient and adjusting for each period's initial estimated glomerular filtration rate, the state of relapse was associated with a 2.17 (95% confidence interval 1.32-3.58; P = .002) greater risk of experiencing a combined event even in this FSGS remission cohort. CONCLUSION: In FSGS, unless remissions are maintained and relapses avoided, long-term renal survival remains poor. Treatment strategies addressing remission duration remain poorly defined and should be an essential question in future trials.

Incidence of thrombotic microangiopathies in Quebec: insight from a laboratory centralizing ADAMTS-13 testing.

BACKGROUND: Thrombotic microangiopathies (TMA) are serious medical conditions requiring a prompt diagnosis to adapt treatment. The determination of ADAMTS-13 activity enables discriminating thrombotic thrombocytopenic purpura (TTP) from other forms of TMA. The purpose of this study was to provide an estimate of the incidence of TTP and TMA in the Canadian Quebec province using data collected from a laboratory centralizing ADAMTS-13 testing for the whole province. RESULTS: From 2012 to 2019, 846 patients were evaluated for plasma ADAMTS-13 activity due to a suspicion of TMA. TTP was identified in 147 patients. Of these, 118 patients with a median age of 51.5 years and a male-female ratio of 1:1.4 had their first episode of TTP during the study period. The number of ADAMTS-13 tests performed and the number of patients with suspected TMA increased annually by 19% and 21% respectively. While the incidence of non-TTP TMA increased annually, that for TTP remained unchanged. This averaged 10.2 (95% CI 5.9-14.4) per million persons per year for suspected non-TTP TMA and 1.8 (95% CI 1.3-2.4) for confirmed TTP. The incidence rate of TMA other than TTP was higher in the age group 70-79 years (21.8; 95% CI 5.4-38.1) for females and in the age group 80-89 years (24.4; 95% CI 7.2-41.7) for males compared to other age groups. The incidence rate of TTP was higher in the age group 40-49 years (4.0; 95% CI 2.0-5.9) for women and in the age group 60-69 years (3.4; 95% CI 1.1-5.6) for men compared to other age groups. CONCLUSION: The analysis of centralized data measuring ADAMTS-13 activity allowed us to adequately establish the incidence rate and demographic characteristics of TMA, particularly TTP, in Quebec. TTP incidence remained stable while suspected non-TTP TMA steadily increased from 2012 to 2019.

Comparison of Complement Pathway Activation in Autoimmune Glomerulonephritis.

Introduction: Studies on complement activation have implicated a combination of the classical pathway (CP), lectin pathway (LP), and alternative pathway (AP) in triggering the terminal pathway (TP) for each common autoimmune glomerulonephritis (GN). Evaluating different pathways simultaneously may help identify whether one is preferentially activated and, consequently, which is best to target for each disease. Methods: We followed 112 patients with focal segmental glomerular sclerosis (FSGS), membranous nephropathy (MN), IgA nephropathy (IgAN), lupus nephritis (LN), and antineutrophil cytoplasmic autoantibody-associated vasculitis (AAV) for a median duration of 22 (12-52) months. At the time of greatest clinical activity, we simultaneously evaluated urinary C3a (C3 convertase activity), C5a and sC5b-9 (TP), MASP-1 and MASP-2 (LP), C1q (CP), C4a (CP/LP), and Ba and Bb (AP). We evaluated the relation between activation fragments of the AP and CP/LP with the TP. Results: Urinary complement biomarkers for each pathway were associated with the severity of proteinuria. Fragments of the TP were higher among patients with FSGS and MN compared with patients with IgAN, LN, and AAV. For the AP, urinary Ba level was lower in those with IgAN and LN compared with those with FSGS. For the CP/LP, urinary C4a, MASP-1, and MASP-2 levels were similar between diseases whereas urinary C1q levels were lower in those with LN. For each GN, independent associations existed between the activation markers of the AP and CP/LP with the degree of TP activation, except for the AP in AAV, although perhaps underpowered. Conclusion: The AP and CP/LP contribute individually to the TP activation in autoimmune GN, and both seem to be valid potential therapeutic targets.

Meta-GWAS Reveals Novel Genetic Variants Associated with Urinary Excretion of Uromodulin.

BACKGROUND: Uromodulin, the most abundant protein excreted in normal urine, plays major roles in kidney physiology and disease. The mechanisms regulating the urinary excretion of uromodulin remain essentially unknown. METHODS: We conducted a meta-analysis of genome-wide association studies for raw (uUMOD) and indexed to creatinine (uUCR) urinary levels of uromodulin in 29,315 individuals of European ancestry from 13 cohorts. We tested the distribution of candidate genes in kidney segments and investigated the effects of keratin-40 (KRT40) on uromodulin processing. RESULTS: Two genome-wide significant signals were identified for uUMOD: a novel locus (P 1.24E-08) over the KRT40 gene coding for KRT40, a type 1 keratin expressed in the kidney, and the UMOD-PDILT locus (P 2.17E-88), with two independent sets of single nucleotide polymorphisms spread over UMOD and PDILT. Two genome-wide significant signals for uUCR were identified at the UMOD-PDILT locus and at the novel WDR72 locus previously associated with kidney function. The effect sizes for rs8067385, the index single nucleotide polymorphism in the KRT40 locus, were similar for both uUMOD and uUCR. KRT40 colocalized with uromodulin and modulating its expression in thick ascending limb (TAL) cells affected uromodulin processing and excretion. CONCLUSIONS: Common variants in KRT40, WDR72, UMOD, and PDILT associate with the levels of uromodulin in urine. The expression of KRT40 affects uromodulin processing in TAL cells. These results, although limited by lack of replication, provide insights into the biology of uromodulin, the role of keratins in the kidney, and the influence of the UMOD-PDILT locus on kidney function.

Activated Alpha 2-Macroglobulin Is a Novel Mediator of Mesangial Cell Profibrotic Signaling in Diabetic Kidney Disease.

Diabetic kidney disease (DKD) is caused by the overproduction of extracellular matrix proteins (ECM) by glomerular mesangial cells (MCs). We previously showed that high glucose (HG) induces cell surface translocation of GRP78 (csGRP78), mediating PI3K/Akt activation and downstream ECM production. Activated alpha 2-macroglobulin (α2M*) is a ligand known to initiate this signaling cascade. Importantly, increased α2M was observed in diabetic patients' serum, saliva, and glomeruli. Primary MCs were used to assess HG responses. The role of α2M* was assessed using siRNA, a neutralizing antibody and inhibitory peptide. Kidneys from type 1 diabetic Akita and CD1 mice and human DKD patients were stained for α2M/α2M*. α2M transcript and protein were significantly increased with HG in vitro and in vivo in diabetic kidneys. A similar increase in α2M* was seen in media and kidneys, where it localized to the mesangium. No appreciable α2M* was seen in normal kidneys. Knockdown or neutralization of α2M/α2M* inhibited HG-induced profibrotic signaling (Akt activation) and matrix/cytokine upregulation (collagen IV, fibronectin, CTGF, and TGFß1). In patients with established DKD, urinary α2M* and TGFß1 levels were correlated. These data reveal an important role for α2M* in the pathogenesis of DKD and support further investigation as a potential novel therapeutic target.

Clinical Use of Complement, Inflammation, and Fibrosis Biomarkers in Autoimmune Glomerulonephritis.

INTRODUCTION: Complement activation, inflammation, and fibrosis play central roles in the mechanisms of injury in autoimmune glomerulonephritis (GN) but they are seldom assessed in epidemiologic studies. The measurement of urinary biomarkers of these pathways of injury could parallel disease activity and add clinical value beyond proteinuria. METHODS: We performed a prospective cohort study of 100 patients with focal and segmental glomerulosclerosis (FSGS), membranous nephropathy (MN), IgA nephropathy (IgAN), lupus nephritis (LN), anti-neutrophil cytoplasmic autoantibody-associated vasculitis (AAV), and membranoproliferative GN (MPGN) followed for 33 (18-54) months. Repeated urinary samples were collected throughout their follow-up to determine proteinuria, urinary sC5b-9, monocyte chemoattractant protein-1 (MCP-1), and transforming growth factor-beta 1 (TGF-ß1), expressed as creatinine ratios. We identified 177 periods of active and inactive disease based on current remission definitions for each disease. RESULTS: Urinary sC5b-9, MCP-1, and TGF-ß1 were present in each disease. In periods leading to a remission, the reduction of urinary sC5b-9 was 91%, greater than for proteinuria with 76%. During inactive periods, those who did not experience a relapse maintained lower levels of biomarkers compared with those who relapsed. At that time, the increase in urinary sC5b-9 was significantly greater than the rise in proteinuria (8.5-fold increase compared with 3.2-fold) and urinary MCP-1 and TGF-ß1. Using current remission definitions for each disease, thresholds for each biomarker were determined using receiver operating characteristic curves. Individuals who averaged levels below these cutoffs during their follow-up had better renal outcomes. CONCLUSION: In autoimmune glomerular diseases, urinary sC5b-9, MCP-1, and TGF-ß1 are present and parallel disease activity and outcomes. Urinary sC5b-9 appears to be a more discerning marker of immunologic remissions and relapses.

Clinicopathologic predictors of renal outcomes in light chain cast nephropathy: a multicenter retrospective study.

Light chain cast nephropathy (LCCN) in multiple myeloma often leads to severe and poorly reversible acute kidney injury. Severe renal impairment influences the allocation of chemotherapy and its tolerability; it also affects patient survival. Whether renal biopsy findings add to the clinical assessment in predicting renal and patient outcomes in LCCN is uncertain. We retrospectively reviewed clinical presentation, chemotherapy regimens, hematologic response, and renal and patient outcomes in 178 patients with biopsy-proven LCCN from 10 centers in Europe and North America. A detailed pathology review, including assessment of the extent of cast formation, was performed to study correlations with initial presentation and outcomes. Patients presented with a mean estimated glomerular filtration rate (eGFR) of 13 ± 11 mL/min/1.73 m2, and 82% had stage 3 acute kidney injury. The mean number of casts was 3.2/mm2 in the cortex. Tubulointerstitial lesions were frequent: acute tubular injury (94%), tubulitis (82%), tubular rupture (62%), giant cell reaction (60%), and cortical and medullary inflammation (95% and 75%, respectively). Medullary inflammation, giant cell reaction, and the extent of cast formation correlated with eGFR value at LCCN diagnosis. During a median follow-up of 22 months, mean eGFR increased to 43 ± 30 mL/min/1.73 m2. Age, ß2-microglobulin, best hematologic response, number of cortical casts per square millimeter, and degree of interstitial fibrosis/tubular atrophy (IFTA) were independently associated with a higher eGFR during follow-up. This eGFR value correlated with overall survival, independently of the hematologic response. This study shows that extent of cast formation and IFTA in LCCN predicts the quality of renal response, which, in turn, is associated with overall survival.

Impact of updated recommendations on acetylsalicylic acid use for primary prevention of cardiovascular disease in Canada: a population-based survey.

BACKGROUND: The debate over acetylsalicylic acid (ASA) therapy for primary prevention of cardiovascular disease (CVD) has recently resurfaced, but scarce data are available on prophylactic ASA use in Canada for this purpose. This study aimed to evaluate the prevalence and factors associated with ASA use, and the potential impact of implementing the most recent (2016) US Preventive Services Task Force recommendations for primary CVD prevention in a Canadian setting. METHODS: We performed a cross-sectional analysis using data from the CARTaGENE study, which included a representative sample (n = 20 004) of the 2018 general population of the province of Quebec. We assessed eligibility for ASA treatment using US Preventive Services Task Force criteria (age 50-69 yr, no past history of myocardial infarction or stroke, and 10-year risk of CVD of at least 10%). We extrapolated to the entire 2018 Quebec population the number of people who would need to start ASA treatment. RESULTS: A total of 6231 respondents in the CARTaGENE study (54.2% of those aged 50-69 yr with no prior history of CVD) were found to be potentially eligible for ASA use for primary CVD prevention. Of the 6231, 1379 (22.1%) were receiving prophylactic ASA treatment. Factors found to be related to ASA use included age, male sex, regular medical visits, lower education level, obesity, hypertension, diabetes and dyslipidemia. Income and smoking status were not found to be significantly associated with ASA use. Our results indicate that 885 261 people would potentially have started ASA treatment if the US Preventive Services Task Force recommendations had been implemented in Quebec in 2018. INTERPRETATION: Prevalent ASA use for primary CVD prevention was low. Implementation of the 2016 US Preventive Services Task Force recommendations would require initiating ASA treatment in a substantial proportion of people, with undetermined potential benefits.

Increased urinary excretion of hedgehog interacting protein (uHhip) in early diabetic kidney disease.

Glomerular endothelial cell (GEC) dysfunction occurs in diabetic kidney disease (DKD) and generally precedes albuminuria. We recently reported that hedgehog interacting protein (Hhip), highly expressed in GECs, contributes to DKD development in diabetic mice. Here, we hypothesized that urinary Hhip (uHhip) could identify early DKD; we tested uHhip in mice and humans with diabetes (DM). In both type 1 (Akita) and type 2 (db/db) DM mice, uHhip is elevated prior to the development of albuminuria, while non-DM controls excrete minimal amount of uHhip. In 87 type 2 DM patients and 39 healthy controls, the uHhip/creatinine (Cr) ratio provides a significant discrimination between non-DM and DM groups; 0 [0-69.5] in non-DM, 9.9 [1.7-39.5] in normoalbuminuric DM, 167.7 [95.7-558.7] in microalbuminuric DM, and 207.9 [0-957.2] in macroalbuminuric DM (median [IQR] ng/mmol, P < 0.0001). The log-uHhip/Cr is positively correlated with urine albumin/Cr ratio (UACR) (spearman correlation coefficient 0.47, P < 0.0001). The log-uHhip/Cr is also associated with eGFR, pulse pressure, and urinary cytokines (IL-1ß, IL-6, IL-8, and TGFß1) independent of UACR. By immunostaining, Hhip is localized in glomeruli and tubules, and is increased in human DM kidneys compared with non-DM kidneys. TGFß1 shares the similar staining pattern as Hhip in human DM kidneys. Thus, uHhip appears to be a novel indicator of diabetic GEC injury and is elevated in early DKD before the development of microalbuminuria in mice and humans. Clinical value for detecting early DKD warrants further investigation.

Clinical Value of Complement Activation Biomarkers in Overt Diabetic Nephropathy.

BACKGROUND: Experimental studies support a role of complement activation in diabetic nephropathy (DN), yet few clinical correlates exist. We evaluated urinary levels of sC5b-9 membrane attack complex (MAC) in patients with overt DN, and examined its association with the glomerular filtration rate (GFR) decline, proteinuria, and inflammatory biomarkers. We explored different complement pathways and compared our findings to autoimmune glomerulonephritis. METHODS: We prospectively followed 83 patients with DN and obtained repeated measurements of proteinuria, complement fragments (sC5b-9, C4a, C1q, mannose-binding lectin-associated serine protease [MASP]-1, and factor Bb), monocyte chemoattractant protein-1 (MCP-1), and transforming growth factor (TGF)-ß1. We assessed independence and interactions using general linear models and repeated measures analyses and compared levels with subjects with active focal and segmental glomerulosclerosis, ANCA-associated vasculitis, and membranous and IgA nephropathies (n = 63). RESULTS: The diabetic cohort had an initial GFR of 25 ± 9 ml/min per 1.73 m2 and a renal function decline of 2.9 ± 3.0 ml/min per 1.73 m2 per year. All complement biomarkers were strongly intercorrelated and associated with biomarker inflammation and fibrosis, proteinuria, and the rate of renal function decline. There was a significant interaction (P = 0.03) between the level of proteinuria and urinary sC5b-9: in individuals with higher levels of urinary MAC, the relationship between proteinuria and the rate of renal function decline was more pronounced than in those with low urinary MAC. Finally, patients with DN had levels of urinary sC5b-9 comparable to autoimmune glomerulonephritis, when stratified by the level of proteinuria. CONCLUSION: Urinary MAC is present in patients with overt DN at levels comparable to autoimmune glomerulonephritis and correlates with the GFR decline, supporting that complement activation and its measurement are clinically relevant in DN.

Estimating the Change in Renal Function During the First Year of Therapy in ANCA-Associated Vasculitis.

INTRODUCTION: Studies in antineutrophil cytoplasmic autoantibody (ANCA)-associated vasculitis (AAV) consistently show that the months following diagnosis have the greatest impact on the long-term renal function. Yet, it remains uncertain how much early gain should be expected with treatment. We sought to determine the factors associated with the change in glomerular filtration rate (GFR) throughout the first year. METHODS: We retrospectively reviewed patients from 3 university hospitals who received treatments. We assessed the proportions of glomeruli with crescents, with global sclerosis, the AAV glomerulonephritis classification, the severity of chronic vascular and tubulo-interstitial disease, and the presence of acute tubular injury (ATI). We used repeated-measures analyses of variance (ANOVAs) to determine factors associated with the change in GFR throughout the first year. RESULTS: There were 162 individuals with AAV identified, 96 with a valid renal biopsy and 82 with at least 12 months of follow-up. The initial GFR of 30 ± 25 ml/min per 1.73 m2 rose by 15 ± 20 during the first year. The severity of pathology findings, myeloperoxidase positivity, and those with kidney- and lung-limited disease presented with a lower GFR. Younger patients with a lower initial GFR and the presence of ATI correlated with a greater increase in GFR by 12 months. A higher proportion of crescents did not predict the change in GFR, contrary to global glomerulosclerosis, where each 10% increase added a loss of 2.7 ± 1.3 ml/min per 1.73 m2 per year (P = 0.03). These factors remained independent of each other. CONCLUSION: Multiple factors influence renal recovery during the first year of therapy. Estimating the change in GFR early on will help identify and reassess outliers.