Post-renal Biopsy Retroperitoneal Haematoma Accompanied by Decreased Coagulation Factor XIII Levels in Immunoglobulin A Nephropathy.

Post-biopsy bleeding is the primary complication of renal biopsy. Retroperitoneal haematoma is a rare but severe bleeding complication; it commonly occurs among patients who have risk factors or vascular lesions. The bleeding risks in patients with immunoglobulin A (IgA) nephropathy (IgAN) have been discussed in the literature, but clinical data are lacking. Here, we report a case of a post-biopsy retroperitoneal haematoma accompanied by decreased coagulation factor XIII (FXIII) in a patient with IgAN. A 14-year-old male patient with haematuria and proteinuria but no bleeding or family history of bleeding underwent pre-renal biopsy evaluation that showed no coagulation abnormalities. He underwent percutaneous renal biopsy, and the histopathological diagnosis was IgAN. Five days after the biopsy, he presented with delayed bleeding from a retroperitoneal haematoma. During the workup for undiagnosed haemorrhagic diatheses, a mildly decreased FXIII level was discovered. This result suggested the possibility of bleeding complications associated with decreased FXIII. Some bleeding diatheses, including FXIII deficiency, cannot be evaluated in routine pre-biopsy coagulation tests. Mild FXIII deficiency can increase the risk of post-biopsy bleeding complications. Therefore, physicians should consider unevaluated haemorrhagic diatheses when a patient presents with major bleeding complications or delayed bleeding following renal biopsy without any known risk factors or vascular lesions.

Urinary extracellular vesicles in childhood kidney diseases.

Most biological fluids contain extracellular vesicles (EVs). EVs are surrounded by a lipid bilayer and contain biological macromolecules such as proteins, lipids, RNA, and DNA. They lack a functioning nucleus and are incapable of replicating. The physiological characteristics and molecular composition of EVs in body fluids provide valuable information about the status of originating cells. Consequently, they could be effectively utilized for diagnostic and prognostic applications. Urine contains a heterogeneous population of EVs. To date, these urinary extracellular vesicles (uEVs) have been ignored in the standard urinalysis. In recent years, knowledge has accumulated on how uEVs should be separated and analyzed. It has become clear how uEVs reflect the expression of each molecule in cells in nephron segments and how they are altered in disease states such as glomerular/tubular disorders, rare congenital diseases, acute kidney injury (AKI), and chronic kidney disease (CKD). Significant promise exists for the molecular expression signature of uEVs detected by simple techniques such as enzyme-linked immunosorbent assay (ELISA), making them more applicable in clinical settings. This review presents the current understanding regarding uEVs, emphasizing the potential for non-invasive diagnostics, especially for childhood kidney diseases.

Enzyme-linked immunosorbent assay to detect surface marker proteins of extracellular vesicles purified from human urine.

The molecular profile of extracellular vesicles released in urine reflects the pathophysiological processes occurring within originating cells located in diverse nephron segments. Here, we present an enzyme-linked immunosorbent assay for quantitative membrane protein detection in extracellular vesicles in human urine samples. We describe steps for preparing urine samples, biotinylated antibodies, and microtiter plates to purify extracellular vesicles and detect membrane-bound biomarkers. The specificity of signals and the limited variability by freeze-thaw cycles or cryopreservation have been verified. For complete details on the use and execution of this protocol, please refer to Takizawa et al. (2022).1.

Comparison of bleeding complications after pediatric kidney biopsy between intravenous sedation and general anesthesia: a nationwide cohort study.

BACKGROUND: An increasing number of studies are evaluating the safety of intravenous sedation compared with that of general anesthesia; however, data on bleeding complications after pediatric percutaneous renal biopsy performed under intravenous sedation or general anesthesia are lacking. We aimed to examine differences in bleeding complications between intravenous sedation and general anesthesia in pediatric patients. METHODS: Data of pediatric patients aged ≤ 15 years undergoing percutaneous kidney biopsy for kidney disease between July 2007 and March 2019 were retrieved from a national inpatient database in Japan. We examined differences in bleeding complications after renal biopsy performed under intravenous sedation, defined by the absence of the record of general anesthesia with intubation but by the presence of intravenous sedation during biopsy, and general anesthesia, defined by the presence of the record of general anesthesia with intubation during biopsy, among pediatric patients admitted for percutaneous renal biopsy. We performed binomial regression using overlap weights based on propensity scores for patients receiving intravenous sedation. Analyses stratified by age or sex, a sensitivity analysis using generalized estimating equations considering cluster effects by hospital among a propensity score-matched cohort, and another sensitivity analysis using the instrumental variable method were performed to confirm the robustness of the results. RESULTS: We identified 6,560 biopsies performed in 5,999 children aged 1-15 years from 328 hospitals and 178 events. Only three severe complications and no death were observed. No significant difference in the proportion of bleeding complications was observed between procedures performed under intravenous sedation and those performed under general anesthesia (unadjusted proportions, 2.8% and 2.3%; adjusted proportions, 2.5% and 2.2%), with an unadjusted relative risk of 1.21 (95% confidence interval, 0.80-1.81) and adjusted relative risk of 1.13 (95% confidence interval, 0.74-1.73). Both age- and sex-stratified analyses yielded similar results. The analysis using generalized estimating equation and the instrumental variable method showed relative risks of 0.95 (95% confidence interval, 0.48-1.88) and 1.18 (95% confidence interval, 0.74-1.89), respectively. CONCLUSION: This retrospective cohort study using a national database revealed that the risk of biopsy-related bleeding was comparable between intravenous sedation and general anesthesia during pediatric percutaneous kidney biopsy, suggesting that intravenous sedation alone and general anesthesia may have a similar bleeding risk in pediatric percutaneous kidney biopsies.

Precise clinicopathologic findings for application of genetic testing in pediatric kidney transplant recipients with focal segmental glomerulosclerosis/steroid-resistant nephrotic syndrome.

BACKGROUND: Establishing a molecular genetic diagnosis of focal segmental glomerulosclerosis (FSGS)/steroid-resistant nephrotic syndrome (SRNS) can be useful for predicting post-transplant recurrence. Monogenic causes are reportedly present in approximately 20-30% of patients with FSGS/SRNS. However, the characteristics of patients who are likely to have a monogenic cause remain to be determined. METHODS: Pediatric recipients with SRNS and/or biopsy-proven FSGS who underwent their first kidney transplantation at our center between 1999 and 2019 were analyzed. Patients with secondary FSGS/SRNS were excluded. The recipients were divided into three groups: familial/syndromic, presumed primary, and undetermined FSGS/SRNS. Patients who met all of the following criteria were categorized as having presumed primary FSGS/SRNS: (i) nephrotic syndrome, (ii) complete or partial remission with initial steroid therapy and/or additional immunosuppressive therapies, and (iii) diffuse foot process effacement on electron microscopy in the native kidney biopsy. All patients underwent genetic testing using next-generation sequencing. RESULTS: Twenty-four patients from 23 families were analyzed in this study. Pathogenic or likely pathogenic variants in FSGS/SRNS-related genes were identified in four of four families, zero of eight families, and 10 of 11 families with familial/syndromic, presumed primary, and undetermined FSGS/SRNS, respectively. Post-transplant recurrence only occurred in patients with presumed primary FSGS/SRNS. CONCLUSIONS: Our systematic approach based on precise clinicopathological findings including nephrotic syndrome, treatment responses, and diffuse foot process effacement might be useful to differentiate pediatric kidney transplant recipients with FSGS/SRNS who are likely to have a monogenic cause from patients who are not, and to predict post-transplant recurrence. A higher resolution version of the Graphical abstract is available as Supplementary information.

A case of right hypodysplastic kidney and ectopic ureter associated with bicornuate uterus in a prepubertal girl.

Congenital anomalies of the kidney and urinary tract (CAKUT) are frequently associated with Mullerian anomalies. This can be explained by the fact that Mullerian duct elongation depends on the preformed Wolffian duct during embryogenesis. While CAKUT such as unilateral renal agenesis and multicystic dysplastic kidney are commonly identified prenatally by routine ultrasound, the diagnosis of Mullerian anomalies is often delayed, increasing the risk of complications such as endometriosis or pelvic inflammatory disease. Herein, we report a case of a premenarchal girl who had initially been diagnosed with right multicystic dysplastic kidney. She presented with continuous urinary incontinence at 4 years old and further evaluation by contrast-enhanced computed tomography, cystoscopy, colposcopy, ureterography, and hysterosalpingography led to the final diagnosis of right hypodysplastic kidney and ectopic ureter associated with bicornuate uterus. A strong family history of uterine malformations prompted the examination of the uterus. Genetic testing was suggested but the family declined. She is planned to be referred to a gynecologist at puberty for further assessment. The recognition and screening rate of concurrent Mullerian anomalies in CAKUT patients varies between institutions. Screening for Mullerian anomalies in prediagnosed CAKUT girls may enable to provide timely counseling and to prevent gynecological complications.

Urinary extracellular vesicles signature for diagnosis of kidney disease.

Congenital disorders characterized by the quantitative and qualitative reduction in the number of functional nephrons are the primary cause of chronic kidney disease (CKD) in children. We aimed to describe the alteration of urinary extracellular vesicles (uEVs) associated with decreased renal function during childhood. By nanoparticle tracking analysis and quantitative proteomics, we identified differentially expressed proteins in uEVs in bilateral renal hypoplasia, which is characterized by a congenitally reduced number of nephrons. This expression signature of uEVs reflected decreased renal function in CKD patients by congenital anomalies of the kidney and urinary tract or ciliopathy. As a proof-of-concept, we constructed a prototype ELISA system that enabled the isolation of uEVs and quantitation of expression of molecules representing the signature. The system identified decreased renal function even in its early stage. The uEVs signature could pave the way for non-invasive methods that can complement existing testing methods for diagnosing kidney diseases.

A novel de novo truncating TRIM8 variant associated with childhood-onset focal segmental glomerulosclerosis without epileptic encephalopathy: a case report.

BACKGROUND: Heterozygous truncating variants in the Tripartite motif containing 8 (TRIM8) gene have been reported to cause epileptic encephalopathy, both with and without proteinuria. A recent study showed a lack of TRIM8 protein expression, with suppressor of cytokine signaling 1 (SOCS1) overexpression, in podocytes and tubules from a patient with a TRIM8 variant, who presented with epileptic encephalopathy and focal segmental glomerulosclerosis (FSGS). To date, no patients with TRIM8 variants who presented with nephrotic syndrome but without neurological manifestations have been described. CASE PRESENTATION: An 8-year-old girl presented with nephrotic syndrome, without epilepsy or developmental delay. Her kidney biopsy specimens showed FSGS and cystic dilatations of the distal tubules. Whole-exome sequencing identified a novel de novo heterozygous variant in the C-terminal encoding portion of TRIM8 (c.1461C > A), resulting in a premature stop codon (p.Tyr487*). Reverse transcription-polymerase chain reaction using peripheral blood mononuclear cells identified the mRNA sequence of the mutant allele, which confirmed an escape from nonsense-mediated mRNA decay. Immunofluorescence studies showed a lack of TRIM8 expression in glomerular and tubular cells and cystic dilatation of distal tubules. Immunohistochemical studies showed overexpression of SOCS1 in glomerular and tubular cells. CONCLUSIONS: We reported a patient with FSGS, associated with a de novo heterozygous TRIM8 variant, without any neurological manifestations. Our results expanded the clinical phenotypic spectrum of TRIM8 variants.

An infant with X-linked anhidrotic ectodermal dysplasia with immunodeficiency presenting with Pneumocystis pneumonia: A case report.

Pneumocystis jirovecii pneumonia associated with primary immunodeficiency should be considered in infants with slowly progressing cyanosis, even without fever or respiratory symptoms. Genetic counseling is crucial for incontinentia pigmenti families in advance of pregnancy because lethal infections can occur before the diagnosis of X-linked anhidrotic ectodermal dysplasia with immunodeficiency.

Degree of foot process effacement in patients with genetic focal segmental glomerulosclerosis: a single-center analysis and review of the literature.

Determining the cause of focal segmental glomerulosclerosis (FSGS) has crucial implications for evaluating the risk of posttransplant recurrence. The degree of foot process effacement (FPE) on electron micrographs (EM) of native kidney biopsies can reportedly differentiate primary FSGS from secondary FSGS. However, no systematic evaluation of FPE in genetic FSGS has been performed. In this study, percentage of FPE and foot process width (FPW) in native kidney biopsies were analyzed in eight genetic FSGS patients and nine primary FSGS patients. All genetic FSGS patients showed segmental FPE up to 38% and FPW below 2000 nm, while all primary FSGS patients showed diffuse FPE above 88% and FPW above 3000 nm. We reviewed the literature which described the degree of FPE in genetic FSGS patients and identified 38 patients with a description of the degree of FPE. The degree of FPE in patients with mutations in the genes encoding proteins associated with slit diaphragm and cytoskeletal proteins was varied, while almost all patients with mutations in other FSGS genes showed segmental FPE. In conclusion, the present study suggests that the degree of FPE in native kidney biopsies may be useful for differentiating some genetic FSGS patients from primary FSGS patients.

A case of 17q12 deletion syndrome that presented antenatally with markedly enlarged kidneys and clinically mimicked autosomal recessive polycystic kidney disease.

The gene encoding hepatocyte nuclear factor 1ß (HNF1B), a transcription factor involved in the development of the kidney and other organs, is located on chromosome 17q12. Heterozygous deletions of chromosome 17q12, which involve 15 genes including HNF1B, are known as 17q12 deletion syndrome and are a common cause of congenital anomalies of the kidneys and urinary tract (CAKUT) and may also present as a multisystem disorder. Autosomal recessive polycystic kidney disease (ARPKD), on the other hand, is a severe form of polycystic kidney disease caused by mutations in PKHD1 (polycystic kidney and hepatic disease 1). It is important to differentiate between these two diseases because they differ significantly in inheritance patterns, renal prognosis, and extrarenal manifestations. Here we report a case of 17q12 deletion syndrome that clinically mimicked ARPKD in which genetic testing was essential for appropriate genetic counseling and monitoring of possible extrarenal manifestations. The patient presented antenatally with markedly enlarged kidneys and showed bilaterally hyperechoic kidneys with poor corticomedullary differentiation and multiple cysts on ultrasonography. There was no family history of renal disease. ARPKD was clinically suspected and genetic testing was performed to confirm diagnosis, resulting in an unexpected finding of 17q12 deletion including HNF1B. While some research has been done to identify patients that should be tested for HNF1B anomalies, this case illustrates the difficulty of recognizing HNF1B-related disease and the importance of genetic testing in appropriately managing CAKUT cases.

Laminin ß2 variants associated with isolated nephropathy that impact matrix regulation.

Mutations in LAMB2, encoding laminin ß2, cause Pierson syndrome and occasionally milder nephropathy without extrarenal abnormalities. The most deleterious missense mutations that have been identified affect primarily the N-terminus of laminin ß2. On the other hand, those associated with isolated nephropathy are distributed across the entire molecule, and variants in the ß2 LEa-LF-LEb domains are exclusively found in cases with isolated nephropathy. Here we report the clinical features of mild isolated nephropathy associated with 3 LAMB2 variants in the LEa-LF-LEb domains (p.R469Q, p.G699R, and p.R1078C) and their biochemical characterization. Although Pierson syndrome missense mutations often inhibit laminin ß2 secretion, the 3 recombinant variants were secreted as efficiently as WT. However, the ß2 variants lost pH dependency for heparin binding, resulting in aberrant binding under physiologic conditions. This suggests that the binding of laminin ß2 to negatively charged molecules is involved in glomerular basement membrane (GBM) permselectivity. Moreover, the excessive binding of the ß2 variants to other laminins appears to lead to their increased deposition in the GBM. Laminin ß2 also serves as a potentially novel cell-adhesive ligand for integrin α4ß1. Our findings define biochemical functions of laminin ß2 variants influencing glomerular filtration that may underlie the pathogenesis of isolated nephropathy caused by LAMB2 abnormalities.

Development of an exon skipping therapy for X-linked Alport syndrome with truncating variants in COL4A5.

Currently, there are no treatments for Alport syndrome, which is the second most commonly inherited kidney disease. Here we report the development of an exon-skipping therapy using an antisense-oligonucleotide (ASO) for severe male X-linked Alport syndrome (XLAS). We targeted truncating variants in exon 21 of the COL4A5 gene and conducted a type IV collagen α3/α4/α5 chain triple helix formation assay, and in vitro and in vivo treatment efficacy evaluation. We show that exon skipping enabled trimer formation, leading to remarkable clinical and pathological improvements including expression of the α5 chain on glomerular and the tubular basement membrane. In addition, the survival period was clearly prolonged in the ASO treated mice group. This data suggests that exon skipping may represent a promising therapeutic approach for treating severe male XLAS cases.

A cellular model of albumin endocytosis uncovers a link between membrane and nuclear proteins.

Cubilin (CUBN) and amnionless (AMN), expressed in kidney and intestine, form a multiligand receptor complex called CUBAM that plays a crucial role in albumin absorption. To date, the mechanism of albumin endocytosis mediated by CUBAM remains to be elucidated. Here, we describe a quantitative assay to evaluate albumin uptake by CUBAM using cells expressing full-length CUBN and elucidate the crucial roles of the C-terminal part of CUBN and the endocytosis signal motifs of AMN in albumin endocytosis. We also demonstrate that nuclear valosin-containing protein-like 2 (NVL2), an interacting protein of AMN, is involved in this process. Although NVL2 was mainly localized in the nucleolus in cells without AMN expression, it was translocated to the extranuclear compartment when coexpressed with AMN. NVL2 knockdown significantly impaired internalization of the CUBN-albumin complex in cultured cells, demonstrating an involvement of NVL2 in endocytic regulation. These findings uncover a link between membrane and nucleolar proteins that is involved in endocytic processes.

Incomplete cryptic splicing by an intronic mutation of OCRL in patients with partial phenotypes of Lowe syndrome.

Mutations of OCRL cause Lowe syndrome, which is characterised by congenital cataracts, infantile hypotonia with mental retardation, and renal tubular dysfunction and Dent-2 disease, which only affects the kidney. While few patients with an intermediate phenotype between these diseases have been reported, the mechanism underlying variability in the phenotype is unclear. We identified an intronic mutation, c.2257-5G>A, in intron 20 of OCRL in an older brother with atypical Lowe syndrome without eye involvement and a younger brother with renal phenotype alone. This mutation created a splice acceptor motif that was accompanied by a cryptic premature termination codon at the junction of exons 20 and 21. The mutation caused incomplete alternative splicing, which created a small amount of wild-type transcript and a relatively large amount of alternatively spliced transcript with a premature termination codon. In the patients' cells, the alternatively spliced transcript was degraded by nonsense-mediated decay and the wild-type transcript was significantly decreased, but not completely depleted. These findings imply that an intronic mutation creating an incomplete alternative splicing acceptor site results in a relatively low level of wild-type OCRL mRNA expression, leading to partial phenotypes of Lowe syndrome.

Renal hypoplasia can be the cause of membranous nephropathy-like lesions.

BACKGROUND: Renal hypoplasia (RH) is the most common cause of chronic kidney disease in children. In cases of RH, proteinuria is often induced by glomerular hypertrophy and hyperfiltration that is commonly associated with focal segmental glomerulosclerosis. This study reports the first case series of a possible association between RH and membranous nephropathy (MN). METHODS: Of the 168 children with RH who visited our department between 1999 and 2017, five with overt proteinuria (≥ 1 g/gCr) underwent renal biopsy. We retrospectively reviewed the medical charts and analyzed biopsy specimens using light microscopy (LM), immunofluorescence (IF), and electron microscopy. RESULTS: The five children (four boys and one girl) had a median age of 5.5 years at the time of renal biopsy. The median proteinuria was 4.23 g/gCr (range 1.46-14.25), median serum albumin, 2.9 g/dL (range 2.3-3.7), and median estimated glomerular filtration rate, 59.7 mL/min/1.73 m2 (range 36.7-103.6). LM showed segmental spike formation and mesangial hypercellularity and IF study showed segmental granular immunoglobulin G (IgG) staining (IgG1 and IgG3 dominant) along the capillary loops in all five patients. Electron-dense deposits were observed in the subepithelial and mesangial areas. Thus, the pathological studies showed MN-like lesions in all patients. CONCLUSION: Our study suggests that RH can be the cause of MN-like lesions.

Clinical and genetic characterization of nephropathy in patients with nail-patella syndrome.

Nail-patella syndrome (NPS) is a multi-system disorder characterized by hypoplastic nails, hypoplastic patella, skeletal deformities, and iliac horns, which is caused by heterozygous variants of LMX1B. Nephropathy ranging from mild urinary abnormality to end-stage renal disease occurs in some individuals with NPS. Because of the low prevalence of NPS and the lack of longitudinal studies of its kidney involvement, the clinical, pathological, and genetic features characterizing severe nephropathy remain unclear. We conducted a Japanese survey of NPS with nephropathy, and analyzed their clinical course, pathological features, and factors associated with severe renal phenotype. LMX1B gene analysis and luciferase reporter assay were also performed. Among 13 NPS nephropathy cases with genetic validation, 5 patients who had moderate-to-massive proteinuria progressed to advanced chronic kidney disease or end-stage renal disease. Pathological findings in the early phase did not necessarily correlate with renal prognosis. Variants associated with deteriorated renal function including a novel variants were confined to the N-terminal region of the LIM domain and a short sequence in the LMX1B homeodomain, which were distinct from reported variants found in isolated nephropathy without extrarenal manifestation (LMX1B-associated nephropathy). Luciferase reporter analysis demonstrated that variants in patients with severe renal phenotype caused haploinsufficiency, but no dominant-negative effects on promoter activation. A distinct proportion of NPS nephropathy patients progressed to end-stage renal disease in adolescence or young adulthood. Patients with moderate or severe proteinuria, especially those with variants in specific regions of LMX1B, should be monitored for potential deterioration of renal function.

Relationship between post-IVIG IgG levels and clinical outcomes in Kawasaki disease patients: new insight into the mechanism of action of IVIG.

INTRODUCTION/OBJECTIVES: The dosing of intravenous immunoglobulin (IVIG) therapy for Kawasaki disease (KD) has been a matter of debate for decades, with recent studies implicating that larger doses lead to better outcomes. Despite this, few have investigated post-IVIG infusion immunoglobulin G (IgG) levels in relation to outcomes of KD such as response to IVIG and development of coronary artery abnormalities (CAAs). The present study investigated how varying levels of post-infusion IgG affected these outcomes. METHOD: We collected demographic and laboratory data, including post-infusion IgG, from children with KD who were admitted to six hospitals in Japan between 2006 and 2012. We conducted multivariate analyses to examine the relationship between independent variables and non-response to IVIG and development of CAAs. We used random forest, a decision tree-based machine learning tool, to investigate the marginal effect of varying post-infusion IgG levels on non-response to IVIG and development of CAAs. RESULTS: Of 456 patients included in the study, 130 (28.5%) were non-responders and 38 (8.3%) developed CAAs. Sodium, post-infusion IgG, and AST were significantly associated with non-response. Post-infusion IgG and sodium were significantly associated with CAA development. The random forest plots revealed a decrease in non-response and CAA rates with increasing post-infusion IgG until post-infusion IgG was near the median (2821 mg/dL), after which the non-response and CAA rates leveled off. CONCLUSIONS: Greater post-infusion IgG is associated with better response to IVIG and decreased CAA development in KD patients, but this effect levels off at post-infusion IgG levels greater than the median. Key points • Though previous studies have shown that post-intravenous immunoglobulin (IVIG) infusion immunoglobulin G (IgG) is associated with non-response to IVIG therapy and coronary artery abnormality (CAA) development in Kawasaki disease (KD) patients, no study has investigated the relationship between varying levels of post-infusion IgG and these clinical outcomes. • Our study showed that non-response to IVIG therapy and CAA development in Kawasaki disease patients follow a decreasing trend with increasing post-infusion IgG at post-infusion IgG levels below the median. • At values of post-infusion IgG greater than the median, non-response and CAA development rates remain relatively constant with increasing post-infusion IgG. • Our study suggests that when post-infusion IgG is greater than the median, IgG may have fully bound to the therapeutic targets of KD, and in these patients, there may be limited benefit in administering additional IVIG.