Delphi: A Democratic and Cost-Effective Method of Consensus Generation in Transplantation.

The Thrombotic Microangiopathy Banff Working Group (TMA-BWG) was formed in 2015 to survey current practices and develop minimum diagnostic criteria (MDC) for renal transplant TMA (Tx-TMA). To generate consensus among pathologists and nephrologists, the TMA BWG designed a 3-Phase study. Phase I of the study is presented here. Using the Delphi methodology, 23 panelists with >3 years of diagnostic experience with Tx-TMA pathology listed their MDC suggesting light, immunofluorescence, and electron microscopy lesions, clinical and laboratory information, and differential diagnoses. Nine rounds (R) of consensus resulted in MDC validated during two Rs using online evaluation of whole slide digital images of 37 biopsies (28 TMA, 9 non-TMA). Starting with 338 criteria the process resulted in 24 criteria and 8 differential diagnoses including 18 pathologic, 2 clinical, and 4 laboratory criteria. Results show that 3/4 of the panelists agreed on the diagnosis of 3/4 of cases. The process also allowed definition refinement for 4 light and 4 electron microscopy lesions. For the first time in Banff classification, the Delphi methodology was used to generate consensus. The study shows that Delphi is a democratic and cost-effective method allowing rapid consensus generation among numerous physicians dealing with large number of criteria in transplantation.

Thrombotic Microangiopathy in the Renal Allograft: Results of the TMA Banff Working Group Consensus on Pathologic Diagnostic Criteria.

The Banff community summoned the TMA Banff Working Group to develop minimum diagnostic criteria (MDC) and recommendations for renal transplant TMA (Tx-TMA) diagnosis, which currently lacks standardized criteria. Using the Delphi method for consensus generation, 23 nephropathologists (panelists) with >3 years of diagnostic experience with Tx-TMA were asked to list light, immunofluorescence, and electron microscopic, clinical and laboratory criteria and differential diagnoses for Tx-TMA. Delphi was modified to include 2 validations rounds with histological evaluation of whole slide images of 37 transplant biopsies (28 TMA and 9 non-TMA). Starting with 338 criteria in R1, MDC were narrowed down to 24 in R8 generating 18 pathological, 2 clinical, 4 laboratory criteria, and 8 differential diagnoses. The panelists reached a good level of agreement (70%) on 76% of the validated cases. For the first time in Banff classification, Delphi was used to reach consensus on MDC for Tx-TMA. Phase I of the study (pathology phase) will be used as a model for Phase II (nephrology phase) for consensus regarding clinical and laboratory criteria. Eventually in Phase III (consensus of the consensus groups) and the final MDC for Tx-TMA will be reported to the transplantation community.

Immune-mediated diseases after coronavirus disease 2019 vaccination: rare but important complication.

Since the beginning of mass vaccination against coronavirus disease 2019 (COVID-19), vaccine-linked immune-mediated diseases have been increasingly reported. The development of these diseases after COVID-19 vaccination may be attributed to the mechanisms of molecular mimicry and cross-reactivity between the viral spike protein and self-antigens. The most frequent vaccine-linked glomerular disease is immunoglobulin A nephropathy (IgAN). Cutaneous vasculitis has also been reported after COVID-19 vaccination. In both diseases, deposition of immune complexes activates the inflammatory response with end-organ damage. We report on a case of de novo IgAN in a young man and a case of severe cutaneous vasculitis in a 68-year-old woman, both after the second dose of Pfizer-BioNTech COVID-19 vaccine. Neither of the patients had a history of autoimmunity or adverse reactions to vaccines. The temporal association between vaccination and disease development in the absence of other possible intercurrent inciting events suggests a causal mechanism, although coincidental co-occurrence cannot be excluded. In both cases, immunosuppressive treatment was warranted to stop disease progression and to partially or completely resolve the disease. A timely reaction is needed if new-onset signs of an immune-mediated disease appear after vaccination.

PULMONARY HEMORRHAGE AND CRESCENTIC GLOMERULONEPHRITIS IN A PATIENT WITH SEROPOSITIVE ANTI-GLOMERULAR BASEMENT MEMBRANE DISEASE AND ANTI-NEUTROPHIL CYTOPLASMIC ANTIBODIES.

Anti-glomerular basement membrane (anti-GBM) disease is an acute and life-threatening systemic autoimmune disorder. The coexistence of circulating anti-neutrophil cytoplasmic antibodies (ANCA) and anti-GBM disease, the so-called double-positive disease (DPD), is exceptionally rare. We report a unique case of DPD manifesting as pulmonary-renal syndrome (PRS) in a 46-year-old woman who first presented with clinical and radiological suspicion of pneumonia. Chest computed tomography scan later revealed bilateral alveolar hemorrhage. Kidney biopsy showed necrotizing crescentic (100% glomeruli) glomerulonephritis. On immunofluorescence microscopy, glomeruli were global linear positive for IgG, confirming anti-GBM disease. Double positivity was detected for circulating anti-myeloperoxidase ANCA (p-ANCA) and anti-GBM antibodies. Acute renal failure evolved rapidly. Therapeutic plasma exchange (TPE) and hemodialysis (HD) were initiated early in combination with intravenous pulse corticosteroid therapy followed by oral methylprednisolone and cyclophosphamide. Pulmonary hemorrhage resolved, but renal function could not be preserved. The patient remains HD dependent. This case report highlights that pulmonary symptomatology may be the leading clinical presentation of PRS, with initially normal renal function at DPD onset. Early recognition and diagnosis are therefore crucial to timely clinical intervention. The role of prompt kidney biopsy and initiation of TPE and HD in PRS must not be underestimated.

Clinical and pathohistological characteristics of Alport spectrum disorder caused by COL4A4 mutation c.193-2A>C: a case series.

AIM: To present the pathohistological and clinical characteristics of five Croatian families with Alport spectrum disorders caused by splice acceptor pathogenic variant c.193-2A>C in COL4A4 at the genomic position chr2:227985866. METHODS: The study enrolled five probands with kidney biopsy analysis and five family members. Mutation screening was performed with Illumina MiSeq platform. The pathogenic variant was confirmed with standard dye-terminator sequencing. RESULTS: The only homozygous patient, aged two, had proteinuria and hematuria with preserved kidney function and no extrarenal manifestations. This patient had changes characteristic for Alport syndrome observed on electron microscopy of the kidney biopsy. In the heterozygous group, six patients had hematuria, four biopsied probands had proteinuria, and only one had moderately reduced kidney function. Heterozygous probands had variable kidney biopsy findings. Three patients had thin glomerular basement membrane nephropathy visible on electron microscopy and focal segmental glomerulosclerosis on light microscopy, two of them with focal lamellation on electron microscopy. One heterozygous patient had changes characteristic for Alport syndrome on electron microscopy without focal segmental glomerulosclerosis. CONCLUSION: The homozygous patient had hematuria and proteinuria with preserved kidney function. The heterozygous patients presented with reasonably mild clinical phenotype and variable pathohistological findings.

Expression Pattern of α-Tubulin, Inversin and Its Target Dishevelled-1 and Morphology of Primary Cilia in Normal Human Kidney Development and Diseases.

The spatiotemporal expression of α-tubulin, inversin and dishevelled-1 (DVL-1) proteins associated with the Wnt-signaling pathway, and primary cilia morphology were analyzed in developing kidneys (14th-38th developmental weeks), healthy postnatal (1.5- and 7-years old) and pathologically changed human kidneys, including multicystic dysplastic kidneys (MCDK), focal segmental glomerulosclerosis (FSGS) and nephrotic syndrome of the Finnish type (CNF). The analysis was performed by double immunofluorescence, electron microscopy, semiquantitative and statistical methods. Cytoplasmic co-expression of α-tubulin, inversin and DVL-1 was observed in the proximal convoluted tubules (pct), distal convoluted tubules (dct) and glomeruli (g) of analyzed tissues. During kidney development, the overall expression of α-tubulin, inversin and DVL-1 decreased, while in the postnatal period slightly increased. The highest expressions of α-tubulin and inversin characterized dct and g, while high DVL-1 characterized pct. α-tubulin, inversin and DVL-1 expression pattern in MCDK, FSGS and CNF kidneys significantly differed from the healthy control. Compared to healthy kidneys, pathologically changed kidneys had dysmorphic primary cilia. Different expression dynamics of α-tubulin, inversin and DVL-1 during kidney development could indicate that switch between the canonical and noncanonical Wnt-signaling is essential for normal kidney morphogenesis. In contrast, their disturbed expression in pathological kidneys might be associated with abnormal primary cilia, leading to chronic kidney diseases.

The Onset of Systemic Oxidative Stress Associated with the Accumulation of Lipid Peroxidation Product Acrolein in the Skin of Patients with Small-Vessel Vasculitis.

Small-vessel vasculitis (SVV) is the inflammation of the vessel wall that can result in hemorrhage and/or ischemia. Among the histological findings in SVV are increased infiltrating neutrophils, which, due to their oxidative burst and myeloperoxidase activity, release excessive reactive oxygen species, triggering a chain reaction of lipid peroxidation and yielding reactive aldehydes such as acrolein. The implication of oxidative stress in the pathogenesis of SVV was studied, focusing on acrolein immunohistochemistry in the affected skin vessels and systemic stress response. Samples from SVV patients and healthy subjects were collected and analyzed for total serum peroxides, total antioxidant capacity, inflammatory and immunological parameters, as well as for the presence of acrolein-protein adducts in the skin tissue specimens. The obtained data showed that systemic redox homeostasis and iron metabolism are altered in SVV patients. Possible biomarkers in the evaluation of oxidative status, disease activity and prevalence were indicated. Furthermore, a strong correlation between the accumulation of acrolein-protein adducts in the skin and the progression of the disease was revealed. Thus, the results of this study demonstrate that SVV is not only associated with systemic oxidative stress but also with tissue-specific oxidative stress that promotes acrolein formation and protein modification correlating with the severity of cutaneous vasculitis.

A CHILD WITH DENSE DEPOSIT DISEASE AND DECREASED CLASSIC COMPLEMENT PATHWAY ACTIVITY.

We report a rare case of nephritic syndrome underlying dense deposit disease (DDD) with alternative complement pathway dysfunction explained with both C3 nephritic factor (C3NeF) antibodies and DDD associated polymorphism of factor H. An 8-year-old boy presented with macroscopic hematuria, hypertension and periorbital edema followed by persistently low C3 during the 8-week follow-up. Positive C3 staining on immunofluorescence microscopy, supported by dense deposits within the glomerular basement membrane on electron microscopy, confirmed the diagnosis of DDD. Preliminary tests for complement activation showed decreased classic pathway and deficient alternative complement pathway, as well as slightly positive C3NeF, supporting the diagnosis of DDD. Genetic analysis revealed a polymorphism of the complement factor H gene with an increased risk of developing DDD. Supportive therapy led to satisfactory recovery of renal function and normalization of C3. Given the poor prognosis of the disease, proper approach to such specific glomerulopathy is important to avoid or at least slow down progression to end-stage renal disease.

Reproducibility of the Oxford classification of immunoglobulin A nephropathy, impact of biopsy scoring on treatment allocation and clinical relevance of disagreements: evidence from the VALidation of IGA study cohort.

BACKGROUND: The VALidation of IGA (VALIGA) study investigated the utility of the Oxford Classification of immunoglobulin A nephropathy (IgAN) in 1147 patients from 13 European countries. Methods. Biopsies were scored by local pathologists followed by central review in Oxford. We had two distinct objectives: to assess how closely pathology findings were associated with the decision to give corticosteroid/immunosuppressive (CS/IS) treatments, and to determine the impact of differences in MEST-C scoring between central and local pathologists on the clinical value of the Oxford Classification. We tested for each lesion the associations between the type of agreement (local and central pathologists scoring absent, local present and central absent, local absent and central present, both scoring present) with the initial clinical assessment, as well as long-term outcomes in those patients who did not receive CS/IS. RESULTS: All glomerular lesions (M, E, C and S) assessed by local pathologists were independently associated with the decision to administer CS/IS therapy, while the severity of tubulointerstitial lesions was not. Reproducibility between local and central pathologists was moderate for S (segmental sclerosis) and T (tubular atrophy/interstitial fibrosis), and poor for M (mesangial hypercellularity), E (endocapillary hypercellularity) and C (crescents). Local pathologists found statistically more of each lesion, except for the S lesion, which was more frequent with central review. Disagreements were more likely to occur when the proportion of glomeruli affected was low. The M lesion, assessed by central pathologists, correlated better with the severity of the disease at presentation and discriminated better with outcomes. In contrast, the E lesion, evaluated by local pathologists, correlated better with the clinical presentation and outcomes when compared with central review. Both C and S lesions, when discordant between local and central pathologists, had a clinical phenotype intermediate to double absent lesions (milder disease) and double present (more severe). CONCLUSION: We conclude that differences in the scoring of MEST-C criteria between local pathologists and a central reviewer have a significant impact on the prognostic value of the Oxford Classification. Since the decision to offer immunosuppressive therapy in this cohort was intimately associated with the MEST-C score, this study indicates a need for a more detailed guidance for pathologists in the scoring of IgAN biopsies.

Autosomal recessive Alport syndrome caused by a novel COL4A4 splice site mutation: a case report.

Alport syndrome (AS) is a genetically heterogenic, structural disorder of the glomerular basement membrane (GBM) due to the mutation of COL4A3, COL4A4, or COL4A5 genes, which clinically presents as progressive hematuric nephritis with ultrastructural changes of the GBM, high tone sensorineural hearing loss, and ocular lesions. About 15% of AS cases have autosomal mutations of COL4A3 and COL4A4 genes, including homozygous and compound heterozygous mutations. Here, we present a case of a two-year-old boy with autosomal recessive Alport syndrome (ARAS) caused by a novel c.193-2A>C COL4A4 mutation. The patient had a delayed motor and sensory development coupled with speech and language delay, megalencephaly, hematuria and proteinuria, and normal tonal audiogram and ophthalmology exam. Extensive genetic, metabolic, and neurologic workup performed at the age of 10 months was unremarkable and patient's megalencephaly was described as familial benign megalencephaly. Kidney biopsy analysis showed characteristic signs of AS. Mutations screening with use of Illumina MiSeq platform revealed that the patient was homozygous for a newly discovered splice acceptor pathogenic variant c.193-2A>C found in COL4A4 at the genomic position chr2:227985866 and both parents were heterozygous carriers. The genetic heterogeneity of AS makes the diagnostic process challenging. Although renal biopsy provides information about the characteristic GBM changes and the degree of renal parenchyma damage (interstitial fibrosis and tubular atrophy ratio), genetic testing is a more sensitive and specific method that also gives insight into potential disease severity and clinical course, and provides the basis for genetic counseling.

IgM as a novel predictor of disease progression in secondary focal segmental glomerulosclerosis.

AIM: To determine the role of immunoglobulin M (IgM) deposits in clinical manifestations, disease outcome, and treatment response of idiopathic and secondary focal segmental glomerulosclerosis (FSGS). METHODS: Kidney biopsy specimens of 171 patients diagnosed with FSGS (primary and secondary) and 50 control patients were retrospectively included in the study. For each patient, clinical and outcome data were obtained and compared to morphological parameters, including immunofluorescence analysis of mesangial IgM and complement 3 (C3) deposits analyzed on kidney biopsy samples. RESULTS: There were significant positive correlations between IgM and C3 deposition in secondary FSGS (P<0.001) and between IgM and mesangial deposits detected by electron microscopy in secondary FSGS (P=0.015), which indicated that higher IgM deposition correlated with higher C3 deposition and mesangial deposits only in secondary FSGS. Patients with secondary FSGS and the deposition of IgM showed inferior renal outcomes at earlier time points in comparison with patients with negative IgM expression (P=0.022). CONCLUSIONS: We detected a positive correlation between IgM and C3 in secondary FSGS. The association between IgM deposition and worse renal outcome in secondary FSGS indicates that IgM may play a role in the progression of this disease.

C4d Is an Independent Predictor of the Kidney Failure in Primary IgA Nephropathy.

Background: C4d deposits are present in a substantial proportion of patients with IgA nephropathy (IgAN), indicating the activation of the lectin pathway (LP) of the complement system. It seems that patients with activated LP have worse renal prognosis. The aim of this study was to investigate the prevalence and prognostic significance of C4d in our cohort of patients with primary IgA nephropathy (pIgAN). Methods: Patients with pIgAN were recruited from a hospital register of kidney biopsies of the Department of Nephrology and Dialysis, Dubrava University Hospital, Zagreb. Additional immunohistochemistry staining for C4d was performed on paraffin-embedded kidney tissue, and patients were stratified into being C4d positive or C4d negative. The clinical and histologic features of patients were analyzed and compared regarding C4d positivity. The primary outcome was defined as kidney failure (KF), and predictor variables of KF and renal survival were analyzed. Results: Of a total of 95 patients with pIgAN included in the study, C4d was present in 43 (45.3%). C4d-positive patients had a higher value of systolic (p = 0.039) and diastolic (p = 0.006) blood pressure at diagnosis as well as higher 24 h proteinuria (p = 0.018), serum urate (p = 0.033), and lower eGFR (p < 0.001). C4d-positive patients had worse renal survival (p < 0.001), higher rates of disease progression to KF (p < 0.001), and higher proteinuria (p < 0.001) and lower eGFR (p < 0.001) at the last follow-up. Glomerular C4d was an independent predictor of disease progression to KF (HR = 5.87 [0.95 CI 1.06-32.44], p = 0.032). Conclusions: C4d is an independent predictor of disease progression in patients with pIgAN. C4d may be used as an additional marker of progressive disease course in IgAN. The therapeutic implications of C4d status in IgAN, particularly in terms of complement inhibitors application, are not yet known.

Crescentic Glomerulonephritis Due to Enterococcal Endocarditis.

Glomerulonephritis following an enterococcal endocarditis is an extremely rare and life-threatening condition. We present the case of a 71-year-old patient with rapidly progressive glomerulonephritis following enterococcal endocarditis after surgical replacement of the aortic valve. The combination of antibiotic therapy, corticosteroid therapy and haemodialysis led to an improvement in renal function; however, the severity of cardiac deterioration resulted in a fatal outcome.

Clinical and histopathological characteristics of COL4A3 c.2881+1G>A variant causing Alport spectrum disorders in Croatian population.

Alport syndrome (AS) and thin basement membrane nephropathy (TBMN) are part of the spectrum of kidney disorders caused by pathogenic variants in α3, α4, or α5 chains of the collagen type IV, the major structural component of the glomerular basement membrane (GBM). Using targeted next-generation sequencing (NGS), 34 AS/TBMN patients (58.8% male) from 12 unrelated families were found positive for heterozygous c.2881+1G>A variant of the COL4A3gene, that is considered disease-causing. All patients were from the continental or island part of Croatia. Clinical, laboratory, and histopathological data collected from the medical records were analyzed and compared to understand the clinical course and prognosis of the affected patients. At the time of biopsy or first clinical evaluation, the mean age was 31 years (median: 35 years; range: 1 - 72 years). Hematuria was present in 33 patients (97.1%) and 19 (55.9%) patients had proteinuria. There were 6 (17.6%) patients with hearing loss, 4 (11.8%) with ocular lesions, and 11 (32.4%) with hypertension. Twenty-three (67.6%) patients had proteinuria at follow-up, and 5 (14.7%) patients with the median age of 48 years (range: 27-55) progressed to kidney failure, started dialysis, or underwent kidney transplantation. Of the 13 patients who underwent kidney biopsy, 4 (30.8%) developed focal segmental glomerulosclerosis (FSGS), and 8 (66.7%) showed lamellation of the GBM, including all patients with FSGS. It is essential to conduct a detailed analysis of each collagen type IV genetic variant to optimize the prognosis and therapeutic approach for affected patients.

Resistant and Relapsing Collapsing Glomerulopathy Successfully Treated with Rituximab-A Case Report.

Collapsing glomerulopathy (CG) or collapsing focal segmental glomerulosclerosis (cFSGS) is an aggressive disease with a high tendency of progression to end-stage renal disease due to common resistance to conventional immunosuppressants. Rituximab (RTX), a monoclonal antibody against CD20 B cells, showed some benefit in the treatment of CG. We are reporting about female patients with an idiopathic form of CG presenting with nephrotic syndrome (NS) and renal insufficiency resistant to several immunosuppressive agents such as steroids (ST), calcineurin inhibitors (CNI), and cyclophosphamide (CYC). This multidrug-resistant disease responded to RTX with complete remission. Forty-four months after initial RTX administration, a relapse of CG with severe NS and acute renal insufficiency occurred. Repeated application of RTX led to complete remission again. To the best of our knowledge, we are reporting the first case of the relapsing multidrug-resistant form of CG, which responded to RTX. Current data about the treatment of CG with RTX is lacking and is based on rare case reports and small case series. Thus, our report can contribute to determining the role of RTX in the treatment of CG.

Loss of decay-accelerating factor triggers podocyte injury and glomerulosclerosis.

Kidney glomerulosclerosis commonly progresses to end-stage kidney failure, but pathogenic mechanisms are still poorly understood. Here, we show that podocyte expression of decay-accelerating factor (DAF/CD55), a complement C3 convertase regulator, crucially controls disease in murine models of adriamycin (ADR)-induced focal and segmental glomerulosclerosis (FSGS) and streptozotocin (STZ)-induced diabetic glomerulosclerosis. ADR induces enzymatic cleavage of DAF from podocyte surfaces, leading to complement activation. C3 deficiency or prevention of C3a receptor (C3aR) signaling abrogates disease despite DAF deficiency, confirming complement dependence. Mechanistic studies show that C3a/C3aR ligations on podocytes initiate an autocrine IL-1ß/IL-1R1 signaling loop that reduces nephrin expression, causing actin cytoskeleton rearrangement. Uncoupling IL-1ß/IL-1R1 signaling prevents disease, providing a causal link. Glomeruli of patients with FSGS lack DAF and stain positive for C3d, and urinary C3a positively correlates with the degree of proteinuria. Together, our data indicate that the development and progression of glomerulosclerosis involve loss of podocyte DAF, triggering local, complement-dependent, IL-1ß-induced podocyte injury, potentially identifying new therapeutic targets.

Clinical, serological and histological determinants of patient and renal outcome in ANCA-associated vasculitis with renal involvement: an analysis from a referral centre.

PURPOSE: To evaluate significance of clinical and histopathological prognostic factors for renal and patient outcome in AAV patient cohort. METHODS: Retrospective study included consecutive patients diagnosed with pauci-immune crescentic glomerulonephritis from January 2003 to December 2013. Primary outcome was combined endpoint patient death or progression to end-stage renal disease (ESRD). Secondary outcomes were patient survival and progression to ESRD (renal survival) singularly and disease relapse. Kaplan-Meyer survival analysis and multivariate Cox proportional hazard regression analysis were used to explore difference between phenotypes and finding significant predictors regarding outcomes. RESULTS: Out of 81 patients, 40.7% patients reached primary endpoint, 22.2% died, 29.6% reached ESRD and 16% relapsed during follow-up. Multivariate Cox proportional hazards regression-adjusted analysis found higher BVAS (HR 1.08, 95% CI 1.01-1.17, p = 0.042), higher baseline maximal serum creatinine (HR 1.02, 95% CI 1.01-1.03, p = 0.04) and lower haemoglobin (HR 0.97, 95% CI 0.95-0.99, p = 0.011) significantly associated with primary endpoint. Higher BVAS (HR 1.25, 95% CI 1.01-1.43, p = 0.001) and lower haemoglobin (HR 0.95, 95% CI 0.91-0.99, p = 0.008) were significantly associated with patient survival, while for renal survival, lower haemoglobin (HR 0.97, 95% CI 0.94-0.99, p = 0.041) and the need for acute haemodialysis (HR 3.15, 95% CI 1.20-8.26, p = 0.02) were significant predictors. On multivariate-adjusted analysis, no significant predictors for disease relapse were found. Kaplan-Meier survival analysis found no difference between clinical, serological and pathohistological phenotypes for all of the endpoints. CONCLUSIONS: Renal function at presentation, anaemia and BVAS should be included in prediction models for the outcomes for the AAV patients.