C3 glomerulopathy in a patient with a history of post-infectious glomerulonephritis.

Post-infectious glomerulonephritis (PIGN) is an immune complex mediated glomerular injury occurring because of an infection, most commonly with group A beta-hemolytic streptococcus in children. C3 glomerulopathy (C3G) is a distinct clinicopathological entity occurring secondary to dysregulation of alternate complement pathway encompassing both C3 glomerulonephritis (C3GN) and dense deposit disease (DDD). While most patients with PIGN attain complete remission with normalized complement levels by 6-8 weeks after presentation, patients with C3G continue to have hypocomplementemia with high rates of progressive kidney disease. Here, we report a patient diagnosed with dense deposit disease after his initial presentation with PIGN three years prior. While current literature continues to explore the overlapping and distinguishing features of PIGN and C3G, including how underlying defects in the alternate complement pathway may commonly contribute to both diseases, this case further exemplifies the importance of recognizing the clinico-pathogenic features of PIGN and C3G in pediatric patients with glomerulonephritis.

Acetaminophen induced high anion gap metabolic acidosis: a potentially under-recognized consequence from a common medication.

While metabolic acidosis is one of the most common complications in patients with chronic kidney disease (CKD), there are several uncommon etiologies that are challenging to diagnose. Here, we describe a patient on peritoneal dialysis who developed high anion gap metabolic acidosis secondary to acquired 5-oxoprolinemia from acetaminophen use. While CKD is a known risk factor for developing this potentially serious complication, this case further highlights how 5-oxoproline accumulation can occur, even with therapeutic dosing of acetaminophen.

Stromal ß-catenin activation impacts nephron progenitor differentiation in the developing kidney and may contribute to Wilms tumor.

Wilms' tumor (WT) morphologically resembles the embryonic kidney, consisting of blastema, epithelial and stromal components, suggesting tumors arise from the dysregulation of normal development. ß-Catenin activation is observed in a significant proportion of WTs; however, much remains to be understood about how it contributes to tumorigenesis. Although activating ß-catenin mutations are observed in both blastema and stromal components of WT, current models assume that activation in the blastemal lineage is causal. Paradoxically, studies performed in mice suggest that activation of ß-catenin in the nephrogenic lineage results in loss of nephron progenitor cell (NPC) renewal, a phenotype opposite to WT. Here, we show that activation of ß-catenin in the stromal lineage non-autonomously prevents the differentiation of NPCs. Comparisons of the transcriptomes of kidneys expressing an activated allele of ß-catenin in the stromal or nephron progenitor cells reveals that human WT more closely resembles the stromal-lineage mutants. These findings suggest that stromal ß-catenin activation results in histological and molecular features of human WT, providing insights into how alterations in the stromal microenvironment may play an active role in tumorigenesis.

Identification and characterization of cellular heterogeneity within the developing renal interstitium.

Kidney formation requires the coordinated growth of multiple cell types including the collecting ducts, nephrons, vasculature and interstitium. There is a long-held belief that interactions between progenitors of the collecting ducts and nephrons are primarily responsible for kidney development. However, over the last several years, it has become increasingly clear that multiple aspects of kidney development require signaling from the interstitium. How the interstitium orchestrates these various roles is poorly understood. Here, we show that during development the interstitium is a highly heterogeneous patterned population of cells that occupies distinct positions correlated to the adjacent parenchyma. Our analysis indicates that the heterogeneity is not a mere reflection of different stages in a linear developmental trajectory but instead represents several novel differentiated cell states. Further, we find that ß-catenin has a cell autonomous role in the development of a medullary subset of the interstitium and that this non-autonomously affects the development of the adjacent epithelia. These findings suggest the intriguing possibility that the different interstitial subtypes may create microenvironments that play unique roles in development of the adjacent epithelia and endothelia.

Transcription Factors YAP/TAZ and SRF Cooperate To Specify Renal Myofibroblasts in the Developing Mouse Kidney.

BACKGROUND: The embryonic renal stroma consists of multiple molecularly distinct cell subpopulations, the functional significance of which is largely unknown. Previous work has demonstrated that the transcription factors YAP and TAZ play roles in the development and morphogenesis of the nephrons, collecting ducts, and nephron progenitor cells. METHODS: In embryonic mouse kidneys, we identified a subpopulation of stromal cells with enriched activity in YAP and TAZ. To evaluate the function of these cell types, we genetically ablated both Yap and Taz from the stromal progenitor population and examined how gene activity and development of YAP/TAZ mutant kidneys are affected over a developmental time course. RESULTS: We found that YAP and TAZ are active in a subset of renal interstitium and that stromal-specific coablation of YAP/TAZ disrupts cortical fibroblast, pericyte, and myofibroblast development, with secondary effects on peritubular capillary differentiation. We also demonstrated that the transcription factor SRF cooperates with YAP/TAZ to drive expression of at least a subset of renal myofibroblast target genes and to specify myofibroblasts but not cortical fibroblasts or pericytes. CONCLUSIONS: These findings reveal a critical role for YAP/TAZ in specific embryonic stromal cells and suggest that interaction with cofactors, such as SRF, influence the expression of cell type-specific target genes, thus driving stromal heterogeneity. Further, this work reveals functional roles for renal stroma heterogeneity in creating unique microenvironments that influence the differentiation and maintenance of the renal parenchyma.

Single-Cell Profiling of AKI in a Murine Model Reveals Novel Transcriptional Signatures, Profibrotic Phenotype, and Epithelial-to-Stromal Crosstalk.

BACKGROUND: Current management of AKI, a potentially fatal disorder that can also initiate or exacerbate CKD, is merely supportive. Therefore, deeper understanding of the molecular pathways perturbed in AKI is needed to identify targets with potential to lead to improved treatment. METHODS: We performed single-cell RNA sequencing (scRNA-seq) with the clinically relevant unilateral ischemia-reperfusion murine model of AKI at days 1, 2, 4, 7, 11, and 14 after AKI onset. Using real-time quantitative PCR, immunofluorescence, Western blotting, and both chromogenic and single-molecule in situ hybridizations, we validated AKI signatures in multiple experiments. RESULTS: Our findings show the time course of changing gene expression patterns for multiple AKI stages and all renal cell types. We observed elevated expression of crucial injury response factors-including kidney injury molecule-1 (Kim1), lipocalin 2 (Lcn2), and keratin 8 (Krt8)-and of several novel genes (Ahnak, Sh3bgrl3, and Col18a1) not previously examined in kidney pathologies. AKI induced proximal tubule dedifferentiation, with a pronounced nephrogenic signature represented by Sox4 and Cd24a. Moreover, AKI caused the formation of "mixed-identity cells" (expressing markers of different renal cell types) that are normally seen only during early kidney development. The injured tubules acquired a proinflammatory and profibrotic phenotype; moreover, AKI dramatically modified ligand-receptor crosstalk, with potential pathologic epithelial-to-stromal interactions. Advancing age in AKI onset was associated with maladaptive response and kidney fibrosis. CONCLUSIONS: The scRNA-seq, comprehensive, cell-specific profiles provide a valuable resource for examining molecular pathways that are perturbed in AKI. The results fully define AKI-associated dedifferentiation programs, potential pathologic ligand-receptor crosstalk, novel genes, and the improved injury response in younger mice, and highlight potential targets of kidney injury.

Clinicopathological features of C3 glomerulopathy in children: a single-center experience.

BACKGROUND: C3 glomerulopathy (C3G) is defined by dominant glomerular deposition of C3 and minimal or no immunoglobulin, with two subtypes-dense deposit disease (DDD) and C3 glomerulonephritis (C3GN)-distinguished by features on electron microscopy (EM). Given that this rare disease has generally unfavorable yet highly variable outcomes, we sought out to review the histopathology, complement/genetic studies, and renal outcomes of pediatric patients with C3G at our institution. METHODS: All native kidney biopsies performed in a single pediatric hospital over a 10-year period were reviewed for features of C3G. Of 589 biopsy reports, we identified 9 patients fulfilling the diagnostic criteria for C3G and retrospectively reviewed their clinical chart and renal biopsy findings. RESULTS: We identified 4 patients with DDD, 4 with C3GN, and 1 indeterminate case, with features of both C3GN and DDD. Five patients were positive for one or more nephritic factors (C3NeF, C4NeF, C5NeF) with 1 patient additionally positive for complement factor H (CFH) autoantibody. Genetic testing done in 5 of the 9 patients failed to identify any causative mutations. Three patients showed progressive renal dysfunction over a mean follow-up period of 33 months. CONCLUSIONS: Complement and genetic studies are now routinely recommended for patients with a histopathological diagnosis of C3G. Careful interpretation of these studies and their prognostic and therapeutic implications in conjunction with biopsy findings is needed to further understand the pathophysiology of this rare disease in children.

Unusual presentations of BK virus infections in pediatric renal transplant recipients.

BKV has emerged as a significant pathogen in the field of transplantation, predominantly causing BKV nephropathy in renal transplant recipients and hemorrhagic cystitis in HSCT recipients. However, case reports describe more diverse complications, and we too present three unusual cases of BKV infections in pediatric renal transplant recipients. First, we describe a case of biopsy-proven renal damage secondary to BKV prior to the onset of viremia, demonstrating that BKV nephropathy can occur without preceding viremia. We also present two renal transplant recipients with persistent BK viruria, one with BKV-associated hemorrhagic cystitis and the other with microscopic hematuria. Therefore, we conclude that BKV manifestations may be more diverse than previously thought and suggest clinical utility in urine BKV qPCR testing in specific transplant recipients.

Integration of Multiple, Diverse Methods to Identify Biologically Significant Marker Genes.

Identification of genes that reliably mark distinct cell types is key to leveraging single-cell RNA sequencing to better understand organismal biology. Such genes are usually chosen by measurement of differential expression between groups of cells and selecting those with the greatest magnitude or most statistically significant change. Many methods have been developed for performing such analyses, but no single, best method has emerged. Validating the results of these analyses is costly in terms of time, effort and resources. We demonstrate that applying an ensemble of such methods robustly identifies genes that mark cells that cluster together and that show restricted expression assessed by antisense mRNA in situ and immunofluorescence. This technique is easily extensible to any number of differential expression methods and the inclusion of additional methods is expected to result in further improvement in performance.

Case Report: Atypical HUS Presenting With Acute Rhabdomyolysis Highlights the Need for Individualized Eculizumab Dosing.

Background: Atypical hemolytic uremic syndrome (aHUS) is an ultra-rare orphan disease caused by dysregulated complement activation resulting in thrombotic microangiopathy. Although complement-mediated endothelial injury predominantly affects the renal microvasculature, extra-renal manifestations are present in a significant proportion of patients. While eculizumab has significantly improved the morbidity and mortality of this rare disease, optimizing therapeutic regimens of this highly expensive drug remains an active area of research in the treatment of aHUS. Case Presentation: This report describes the case of a previously healthy 4 year-old male who presented with rhabdomyolysis preceding the development of aHUS with anuric kidney injury requiring dialysis. Clinical stabilization required increased and more frequent eculizumab doses compared with the standardized weight-based guidelines. In the maintenance phase of his disease, pharmacokinetic analysis indicated adequate eculizumab levels could be maintained with an individualized dosing regimen every 3 weeks, as opposed to standard 2 week dosing, confirmed in this patient over a 4 year follow up period. Cost analyses show that weight-based maintenance dosing costs $312,000 per year, while extending the dosing interval to every 3 weeks would cost $208,000, a savings of $104,000 per year, relative to the cost of $72,000 from more frequent eculizumab dosing during his initial hospitalization to suppress his acute disease. Conclusion: This case exemplifies the potential of severe, multisystem involvement of aHUS presenting with extra-renal manifestations, including rhabdomyolysis as in this case, and highlights the possibility for improved clinical outcomes and higher value care with individualized eculizumab dosing in patients over the course of their disease.

Iron deficiency and renal development in the newborn rat.

Iron is essential for fetal organ development, but the effect of isolated iron deficiency on nephrogenesis is unknown. Human premature infants are at risk for disrupted nephrogenesis because glomerular development is incomplete until 36-wk gestation. We modeled the effects of iron on postnatal glomerulogenesis in four groups of immature rats from P4 to P12: dam fed controls (DF), dam fed with sham gastrostomy surgery (DF + SS), iron-deficiency anemia (IDA), fed iron-deficient formula through gastrostomy apart from the dam, and IDA plus simultaneous enteral iron rescue (IDA+Fe). Hematocrit, plasma ferritin, and body and kidney tissue iron contents were measured. Tissue was examined. Rats grew similarly, but IDA rats exhibited lower hematocrit, plasma ferritin, and body and kidney iron contents than DF, DF + SS, or IDA + Fe. IDA exhibited 1.7 fewer radial glomerular counts (RGCs), 26% reduced glomerular density, and 29% less planar glomerular surface area than DF, with partial improvement in IDA + Fe. Compared with DF or DF + SS, we observed elevated plasma CRP levels and tubulointerstitial fibrosis in the IDA and IDA + Fe groups. IDA reduced glomerular density, glomerular surface area, and promoted fibrosis. Iron substantially rescued renal growth and development, supporting the critical role of iron in late nephrogenesis.

Methods for renal lineage tracing: In vivo and beyond.

Lineage tracing has resulted in fundamental discoveries in kidney development and disease and remains a powerful technique to study mechanisms of organogenesis, homeostasis, and repair/regeneration. Following decades of research on the cellular and molecular regulation of renal organogenesis, the kidney has become one of the most well-characterized organs, resulting in exciting advancements in pluripotent stem cell differentiation, tissue bioengineering, and the potential for developing novel regenerative therapies for kidney disease. Lineage tracing, or the labeling of progeny cells arising from a single cell or group of cells, allows for spatial and temporal analyses of dynamic in vivo and in vitro processes. As lineage tracing techniques expand across disciplines of developmental biology, stem cell biology, and regenerative medicine, careful experimental design and interpretation, along with an understanding of the basic principles and technical limitations, are essential for utilizing genetically complex lineage tracing models to further understand kidney development and disease.

Disruption of Hox9,10,11 function results in cellular level lineage infidelity in the kidney.

Hox genes are important regulators of development. The 39 mammalian Hox genes have considerable functional overlap, greatly confounding their study. In this report, we generated mice with multiple combinations of paralogous and flanking Abd-B Hox gene mutations to investigate functional redundancies in kidney development. The resulting mice developed a number of kidney abnormalities, including hypoplasia, agenesis, and severe cysts, with distinct Hox functions observed in early metanephric kidney formation and nephron progenitor maintenance. Most surprising, however, was that extensive removal of Hox shared function in these kidneys resulted in cellular level lineage infidelity. Strikingly, mutant nephron tubules consisted of intermixed cells with proximal tubule, loop of Henle, and collecting duct identities, with some single cells expressing markers associated with more than one nephron segment. These results indicate that Hox genes are required for proper lineage selection/maintenance and full repression of genes involved in cell fate restriction in the developing kidney.