Infection as a trigger of acute, transient glomerular deposition of clonal immunoglobulins.

Glomerular deposition of monoclonal IgM, frequently in the form of intracapillary pseudothrombi, can be seen in Waldenström macroglobulinemia (WM) and type I cryoglobulinemia (CG). They are typically associated with plasma cell or B-lymphoid neoplasms, particularly lymphoplasmacytic lymphoma (LPL). While infection is a frequent trigger of mixed (type II and III) CG, its association with type I CG is uncommon. We report two cases in which striking lambda-chain-restricted IgM deposits and acute kidney injury (AKI) occurred in the setting of known or suspected systemic infections, with prompt resolution on treatment of the infection.

Renal biopsy in systemic infections: expect the unexpected.

Infection-related glomerulonephritis is well recognized in patients with ongoing infections. It can be missed, however, if the infection is unusual or undetected. We present three cases where the renal biopsy findings prompted the identification or treatment of systemic infections.Case 1: A 84-year-old male presented with acute kidney injury (AKI) and IgA vasculitis on skin biopsy. A renal biopsy showed active glomerulonephritis with abundant neutrophils and predominantly mesangial immune complex deposits containing IgA. The findings prompted an infectious workup which was positive for COVID-19, suggesting exacerbation of IgA nephropathy by recent COVID-19 infection. Case 2: A 31-year-old female status post kidney transplant for granulomatosis with polyangiitis (GPA) had recent pregnancy with preterm delivery, disseminated herpes simplex virus (HSV) infection with HSV hepatitis, E. coli on urine culture, and AKI. A renal biopsy showed proliferative glomerulonephritis with subendothelial and mesangial immune complex deposits containing IgG and C3. The findings were most consistent with infection-related immune complex glomerulonephritis, most likely HSV-related. Case 3: A 78-year-old female presented with AKI, proteinuria, hematuria, and positive p-ANCA. Clinically, ANCA vasculitis was suspected, and renal biopsy did show focal, segmental, necrotizing glomerulonephritis. However, immunofluorescence and electron microscopy showed IgM-rich deposits in the mesangium. The unusual presentation prompted an infectious workup including a Bartonella antibody panel which showed very high titers, suggesting Bartonella endocarditis.Infection-related glomerulonephritis has a wide variety of presentations histologically and clinically. The three cases we present here emphasize the importance of recognizing these entities to help guide treatment and improve patient care.

BK virus nephropathy in non-renal solid organ transplant recipients: Are we looking hard enough?

We retrospectively examined the clinical characteristics, pathological features, and outcomes of BK viremia and nephropathy in a population of non-renal solid organ transplant patients (NRSOT) referred for outpatient nephrology consultation over a period of 5 years. In the entire cohort of liver, heart, and lung transplant recipients referred to this clinic, 14% percent were found to have BK viremia with a median peak serum BK viral load of 35 500 copies/ml (range 250 to 21 100 000 copies/ml). BK viremia resolved in six of the seventeen patients (35%). Four out of five patients biopsied showed BK virus (BKV) nephropathy. Eleven out of seventeen patients with BK viremia developed advanced (stage 4 or 5) chronic kidney disease. Four patients developed rejection of their solid organ transplant within the first year post detection of BK viremia after immunosuppression reduction. We conclude that a multi-center study is required to evaluate whether implementation of a systematic BK screening program would be effective in early detection and management of this problem in the NRSOT population.

Electron microscopy in renal pathology: overall applications and guidelines for tissue, collection, preparation, and stains.

Electron microscopy is a mainstay in the analysis of renal biopsies, where it is typically employed in a correlative fashion along with light and immunofluorescence microscopy. Despite the development of a growing armamentarium of molecular and biochemical analytic methods as well as new immunostains with a widening panel of immunoreactants, electron microscopy remains crucial to the diagnosis of a number of disorders involving the renal glomerulus, vasculature, and tubulointerstitial compartment. The number of renal biopsies continues to grow and the indications for these biopsies continue expanding together with our understanding of disease processes. Proper collection of biopsies and careful analysis of data emanating from diagnostic modalities, clinical information, imaging, gross and microscopic tissue analysis, including a wide range of ancillary studies, represent the essential paradigm for generating detailed diagnoses with clinical significance. This communication offers a guide to the pre-analytic and analytic process for renal biopsy examination, discusses diagnostic keys and pitfalls for an important category of renal diseases (immune complex disorders), and provides an introduction to a useful adjunct diagnostic method (ultrastructural immunolabeling). Renal pathologists should render expert diagnoses that guide patient management, provide prognostic information and lead to targeted new therapeutic interventions that are currently available.

Parenteral administration of oral medications in lung transplant recipients: An underrecognized problem.

Microcrystalline cellulose (MCC) is an insoluble material commonly used as a binder and filler in oral medications. Identification of pulmonary intravascular deposition of MCC in transbronchial biopsies from lung transplant (LT) recipients following parenteral injection of oral medications has only been reported once. A search of our surgical pathology electronic database was performed from January 1, 2000 to November 1, 2017 using the text "transplant transbronchial." The diagnosis field for all cases retrieved was then searched for the text "cellulose." These cases were queried for patient demographics and outcomes. Between January 1, 2000 and November 1, 2017, 1558 lung transplants were performed in 1476 individual patients at our institution; 12 were identified to have MCC in their lung tissue. Patients with MCC identified on biopsies were more likely to be transplanted for cystic fibrosis versus other indications and younger versus older. MCC identified in 2 of our cases was favored to be donor derived. Of the 12 patients, 6 (50%) are deceased. MCC within the pulmonary vasculature may be an indicator of increased complications, mortality, or shortened survival in LT recipients. Detecting intravascular MCC and distinguishing it from aspirated foreign material can be challenging. Awareness of the differential diagnosis for pulmonary foreign material is of paramount importance for the pathologist.

In vivo Modeling Implicates APOL1 in Nephropathy: Evidence for Dominant Negative Effects and Epistasis under Anemic Stress.

African Americans have a disproportionate risk for developing nephropathy. This disparity has been attributed to coding variants (G1 and G2) in apolipoprotein L1 (APOL1); however, there is little functional evidence supporting the role of this protein in renal function. Here, we combined genetics and in vivo modeling to examine the role of apol1 in glomerular development and pronephric filtration and to test the pathogenic potential of APOL1 G1 and G2. Translational suppression or CRISPR/Cas9 genome editing of apol1 in zebrafish embryos results in podocyte loss and glomerular filtration defects. Complementation of apol1 morphants with wild-type human APOL1 mRNA rescues these defects. However, the APOL1 G1 risk allele does not ameliorate defects caused by apol1 suppression and the pathogenicity is conferred by the cis effect of both individual variants of the G1 risk haplotype (I384M/S342G). In vivo complementation studies of the G2 risk allele also indicate that the variant is deleterious to protein function. Moreover, APOL1 G2, but not G1, expression alone promotes developmental kidney defects, suggesting a possible dominant-negative effect of the altered protein. In sickle cell disease (SCD) patients, we reported previously a genetic interaction between APOL1 and MYH9. Testing this interaction in vivo by co-suppressing both transcripts yielded no additive effects. However, upon genetic or chemical induction of anemia, we observed a significantly exacerbated nephropathy phenotype. Furthermore, concordant with the genetic interaction observed in SCD patients, APOL1 G2 reduces myh9 expression in vivo, suggesting a possible interaction between the altered APOL1 and myh9. Our data indicate a critical role for APOL1 in renal function that is compromised by nephropathy-risk encoding variants. Moreover, our interaction studies indicate that the MYH9 locus is also relevant to the phenotype in a stressed microenvironment and suggest that consideration of the context-dependent functions of both proteins will be required to develop therapeutic paradigms.

Renal biopsy in patients with systemic lupus erythematosus: Not just lupus glomerulonephritis!

Kidney biopsy is a mainstay in the diagnosis and management of renal disease in patients with systemic lupus erythematosus. Though biopsies from patients with lupus typically show various forms of immune complex glomerulonephritis, other pathologies are occasionally encountered, including unusual lupus-related nephropathies, other forms of autoimmune disease, and occasional renal disorders without any direct connection with lupus or autoimmunity. Electron microscopy is a powerful tool for detecting and classifying these unusual conditions, which frequently have important therapeutic and prognostic implications.

A novel missense mutation of Wilms' Tumor 1 causes autosomal dominant FSGS.

FSGS is a clinical disorder characterized by focal scarring of the glomerular capillary tuft, podocyte injury, and nephrotic syndrome. Although idiopathic forms of FSGS predominate, recent insights into the molecular and genetic causes of FSGS have enhanced our understanding of disease pathogenesis. Here, we report a novel missense mutation of the transcriptional regulator Wilms' Tumor 1 (WT1) as the cause of nonsyndromic, autosomal dominant FSGS in two Northern European kindreds from the United States. We performed sequential genome-wide linkage analysis and whole-exome sequencing to evaluate participants from family DUK6524. Subsequently, whole-exome sequencing and direct sequencing were performed on proband DNA from family DUK6975. We identified multiple suggestive loci on chromosomes 6, 11, and 13 in family DUK6524 and identified a segregating missense mutation (R458Q) in WT1 isoform D as the cause of FSGS in this family. The identical mutation was found in family DUK6975. The R458Q mutation was not found in 1600 control chromosomes and was predicted as damaging by in silico simulation. We depleted wt1a in zebrafish embryos and observed glomerular injury and filtration defects, both of which were rescued with wild-type but not mutant human WT1D mRNA. Finally, we explored the subcellular mechanism of the mutation in vitro. WT1(R458Q) overexpression significantly downregulated nephrin and synaptopodin expression, promoted apoptosis in HEK293 cells and impaired focal contact formation in podocytes. Taken together, these data suggest that the WT1(R458Q) mutation alters the regulation of podocyte homeostasis and causes nonsyndromic FSGS.

Rare hereditary COL4A3/COL4A4 variants may be mistaken for familial focal segmental glomerulosclerosis.

Focal segmental glomerulosclerosis (FSGS) is a histological lesion with many causes, including inherited genetic defects, with significant proteinuria being the predominant clinical finding at presentation. Mutations in COL4A3 and COL4A4 are known to cause Alport syndrome (AS), thin basement membrane nephropathy, and to result in pathognomonic glomerular basement membrane (GBM) findings. Secondary FSGS is known to develop in classic AS at later stages of the disease. Here, we present seven families with rare or novel variants in COL4A3 or COL4A4 (six with single and one with two heterozygous variants) from a cohort of 70 families with a diagnosis of hereditary FSGS. The predominant clinical finding at diagnosis was proteinuria associated with hematuria. In all seven families, there were individuals with nephrotic-range proteinuria with histologic features of FSGS by light microscopy. In one family, electron microscopy showed thin GBM, but four other families had variable findings inconsistent with classical Alport nephritis. There was no recurrence of disease after kidney transplantation. Families with COL4A3 and COL4A4 variants that segregated with disease represent 10% of our cohort. Thus, COL4A3 and COL4A4 variants should be considered in the interpretation of next-generation sequencing data from such patients. Furthermore, this study illustrates the power of molecular genetic diagnostics in the clarification of renal phenotypes.

A novel role for type 1 angiotensin receptors on T lymphocytes to limit target organ damage in hypertension.

RATIONALE: Human clinical trials using type 1 angiotensin (AT(1)) receptor antagonists indicate that angiotensin II is a critical mediator of cardiovascular and renal disease. However, recent studies have suggested that individual tissue pools of AT(1) receptors may have divergent effects on target organ damage in hypertension. OBJECTIVE: We examined the role of AT(1) receptors on T lymphocytes in the pathogenesis of hypertension and its complications. METHODS AND RESULTS: Deficiency of AT(1) receptors on T cells potentiated kidney injury during hypertension with exaggerated renal expression of chemokines and enhanced accumulation of T cells in the kidney. Kidneys and purified CD4(+) T cells from "T cell knockout" mice lacking AT(1) receptors on T lymphocytes had augmented expression of Th1-associated cytokines including interferon-γ and tumor necrosis factor-α. Within T lymphocytes, the transcription factors T-bet and GATA-3 promote differentiation toward the Th1 and Th2 lineages, respectively, and AT(1) receptor-deficient CD4(+) T cells had enhanced T-bet/GATA-3 expression ratios favoring induction of the Th1 response. Inversely, mice that were unable to mount a Th1 response due to T-bet deficiency were protected from kidney injury in our hypertension model. CONCLUSIONS: The current studies identify an unexpected role for AT(1) receptors on T lymphocytes to protect the kidney in the setting of hypertension by favorably modulating CD4(+) T helper cell differentiation.

Subnormothermic Oxygenated Machine Perfusion (24 h) in DCD Kidney Transplantation.

Background: Ex vivo kidney perfusion is an evolving platform that demonstrates promise in preserving and rehabilitating the kidney grafts. Despite this, there is little consensus on the optimal perfusion conditions. Hypothermic perfusion offers limited functional assessment, whereas normothermic perfusion requires a more complex mechanical system and perfusate. Subnormothermic machine perfusion (SNMP) has the potential to combine the advantages of both approaches but has undergone limited investigation. Therefore, the present study sought to determine the suitability of SNMP for extended kidney preservation. Methods: SNMP at 22-25 °C was performed on a portable device for 24 h with porcine kidneys. Graft assessment included measurement of mechanical parameters and biochemical analysis of the perfusate using point-of-care tests. To investigate the viability of kidneys preserved by SNMP, porcine kidney autotransplants were performed in a donation after circulatory death (DCD) model. SNMP was also compared with static cold storage (SCS). Finally, follow-up experiments were conducted in a subset of human kidneys to test the translational significance of findings in porcine kidneys. Results: In the perfusion-only cohort, porcine kidneys all displayed successful perfusion for 24 h by SNMP, evidenced by stable mechanical parameters and biological markers of graft function. Furthermore, in the transplant cohort, DCD grafts with 30 min of warm ischemic injury demonstrated superior posttransplant graft function when preserved by SNMP in comparison with SCS. Finally, human kidneys that underwent 24-h perfusion exhibited stable functional and biological parameters consistent with observations in porcine organs. Conclusions: These observations demonstrate the suitability and cross-species generalizability of subnormothermic machine perfusion to maintain stable kidney perfusion and provide foundational evidence for improved posttransplant graft function of DCD kidneys after SNMP compared with SCS.

Detection of BK polyomavirus after kidney transplantation: a comparison of urine electron microscopy with plasma polymerase chain reaction.

BK polyomavirus (BKV) infection continues to be a significant source of allograft dysfunction in kidney transplant recipients. The optimal screening method to detect BKV remains undetermined. In this retrospective analysis of 347 consecutive kidney transplant recipients, we compare the diagnostic and screening performance of urine electron microscopy (EM) with plasma polymerase chain reaction (PCR) in testing for BKV, using biopsy-proved polyomavirus-associated nephropathy (PVAN) as the gold standard. Sixty-nine of 347 recipients had a positive screening test for BKV infection. Twenty-nine patients underwent biopsy, and 11 were diagnosed with PVAN. Sensitivity rates of urine EM and plasma PCR were 88% and 100%, respectively. Specificity rates of urine EM and plasma PCR were 91% and 78%. There was no statistical difference in the operating characteristics of the two tests. The majority of both plasma PCR and urine EM tests were positive in the six months prior to a diagnostic biopsy confirming PVAN. In those patients who had evidence of BKV infection but did not have PVAN, the percentage of positive screening tests decreased with aggressive lowering of immunosuppression. We conclude that urine EM and plasma PCR both function well in screening for BKV infection and in the diagnosis of PVAN. There is an opportunity to detect viral replication, lower immunosuppression, and to prevent PVAN in this population.

The continued vital role of electron microscopy in the diagnosis of renal disease/dysfunction.

Ultrastructural examination remains a crucial diagnostic tool for analysis of renal biopsies, along with light and immunofluorescence microscopy. Several advantages of electron microscopy are the ability to detect most forms of glomerular disease/dysfunction; the ability to examine small, suboptimal, or unusual specimens; and superior resolution that allows the investigator to hunt without preconceived ideas of what he/she is looking for or will find. A few shortcomings must be noted for completeness, including inability to detect disorders without specific ultrastructural signature, biochemical heterogeneity of deposits with similar or identical ultrastructural appearances, and occasional false-negative results in diseases with focal manifestations.

Mechanisms of the proteinuria induced by Rho GTPases.

Podocytes are highly differentiated cells that play an important role in maintaining glomerular filtration barrier integrity; a function regulated by small GTPase proteins of the Rho family. To investigate the role of Rho A in podocyte biology, we created transgenic mice expressing doxycycline-inducible constitutively active (V14 Rho) or dominant-negative Rho A (N19 Rho) in podocytes. Specific induction of either Rho A construct in podocytes caused albuminuria and foot process effacement along with disruption of the actin cytoskeleton as evidenced by decreased expression of the actin-associated protein synaptopodin. The mechanisms of these adverse effects, however, appeared to be different. Active V14 Rho enhanced actin polymerization, caused a reduction in nephrin mRNA and protein levels, promoted podocyte apoptosis, and decreased endogenous Rho A levels. In contrast, the dominant-negative N19 Rho caused a loss of podocyte stress fibers, did not alter the expression of either nephrin or Rho A, and did not cause podocyte apoptosis. Thus, our findings suggest that Rho A plays an important role in maintaining the integrity of the glomerular filtration barrier under basal conditions, but enhancement of Rho A activity above basal levels promotes podocyte injury.

Case Report: Unusual Aggregation of Different Glomerulopathies in a Family Resolved by Genetic Testing and Reverse Phenotyping.

Glomerular diseases (GDs) are a major cause of chronic kidney disease in children. The conventional approach to diagnosis of GDs includes clinical evaluation and, in most cases, kidney biopsy to make a definitive diagnosis. However, in many cases, clinical presentations of different GDs can overlap, leading to uncertainty in diagnosis and management even after renal biopsy. In this report, we identify a family with clinical diagnoses of postinfectious glomerulonephritis and IgA nephropathy in a parent and two children. Renal biopsies were initially inconclusive; however, genetic testing showed that the two individuals diagnosed at different points with IgA nephropathy carried novel segregating pathogenic variants in COL4A5 gene. We were only able to make the final diagnoses in each of the family members after genetic testing and reverse phenotyping. This case highlights the utility of genetic testing and reverse phenotyping in resolving clinical diagnosis in families with unusual constellations of different glomerulopathies. We propose that clustering of different glomerular disease phenotypes in a family should be an indication for genetic testing followed by reverse phenotyping.

Steroid-sensitive nephrotic syndrome candidate gene CLVS1 regulates podocyte oxidative stress and endocytosis.

We performed next-generation sequencing in patients with familial steroid-sensitive nephrotic syndrome (SSNS) and identified a homozygous segregating variant (p.H310Y) in the gene encoding clavesin-1 (CLVS1) in a consanguineous family with 3 affected individuals. Knockdown of the clavesin gene in zebrafish (clvs2) produced edema phenotypes due to disruption of podocyte structure and loss of glomerular filtration barrier integrity that could be rescued by WT CLVS1 but not the p.H310Y variant. Analysis of cultured human podocytes with CRISPR/Cas9-mediated CLVS1 knockout or homozygous H310Y knockin revealed deficits in clathrin-mediated endocytosis and increased susceptibility to apoptosis that could be rescued with corticosteroid treatment, mimicking the steroid responsiveness observed in patients with SSNS. The p.H310Y variant also disrupted binding of clavesin-1 to α-tocopherol transfer protein, resulting in increased reactive oxygen species (ROS) accumulation in CLVS1-deficient podocytes. Treatment of CLVS1-knockout or homozygous H310Y-knockin podocytes with pharmacological ROS inhibitors restored viability to control levels. Taken together, these data identify CLVS1 as a candidate gene for SSNS, provide insight into therapeutic effects of corticosteroids on podocyte cellular dynamics, and add to the growing evidence of the importance of endocytosis and oxidative stress regulation to podocyte function.

Acute cellular rejection and humoral sensitization in lung transplant recipients.

Despite the recent development of many new immunosuppressive agents for use in transplantation, acute cellular and humoral rejection represent extremely prevalent and serious complications after lung transplantation. Acute cellular rejection, defined as perivascular or bronchiolar mononuclear inflammation, affects over 50% of lung transplant recipients within the first year. Furthermore, the frequency and severity of acute rejections are the most important risk factors for the subsequent development of bronchiolitis obliterans syndrome (BOS), a condition of progressive airflow obstruction that severely limits survival after lung transplantation. Treatment options for cellular rejection include high-dose methylprednisolone, antithymocyte globulin, or alemtuzumab. Emerging evidence also suggests that humoral rejection occurs in lung transplantation, characterized by local complement activation or the presence of antibody to donor human leukocyte antigens and is associated with an increased risk for BOS. Treatment options for humoral rejection include intravenous immunoglobulin, plasmapheresis, or rituximab. Herein, we review the clinical presentation, diagnosis, mechanisms, and treatment of cellular and humoral rejection after lung transplantation.

Metastatic Urothelial Carcinoma from Transplanted Kidney with Complete Response to an Immune Checkpoint Inhibitor.

BACKGROUND: Donor-derived malignancy is a rare complication in patients who undergo organ transplant. Approaches to treatment have largely been individualized based on clinical circumstances given the lack of evidence-based guidelines, with therapeutic options ranging from discontinuation of immunosuppression and transplantectomy to the addition of chemotherapy or radiotherapy. Case Presentation. Herein, we describe a 60-year-old woman with metastatic donor-derived upper tract urothelial carcinoma (UTUC) discovered nine years postrenal transplant. Molecular diagnostic studies using polymerase chain reaction amplification of short tandem repeat alleles and HLA tissue typing proved that the urothelial carcinoma originated from donor tissue. She achieved sustained complete remission with transplant nephroureterectomy, retroperitoneal lymphadenectomy, immunosuppression withdrawal, and immunotherapy with pembrolizumab. Routine radiologic surveillance has demonstrated 15-month progression-free survival to date off pembrolizumab, and she is now under consideration for retransplantation. CONCLUSIONS: Immunotherapy using checkpoint inhibitors can serve as a novel treatment option for patients in the clinical predicament of having a solid organ transplant and simultaneous metastatic malignancy. In this report, we also discuss the oncogenic potential of BK virus, the use of checkpoint inhibitors in urothelial carcinoma, and the feasibility of retransplant for this patient population.

Serving as a Temporary Pathology Chair: "Boon" or "Boondoggle"?

The 2019 Association of Pathology Chairs Annual Meeting included a discussion group sponsored by the Senior Fellows Group (former chairs of academic departments of pathology who have remained active in Association of Pathology Chairs) that was focused on serving as temporary pathology chair. Such positions include "acting chair" (service while the permanent chair is on leave or temporarily indisposed), "interim chair" (service after departure of the prior chair and before a new chair is appointed), "term-limited chair" (usually one nonrenewable term of less than 5 years), and "terminal chair" (permanent chair being asked to stay until a successor is appointed). Discussion group panelists represented each of these positions and included the perspective of 3 former deans about the rationale for making such appointments. The potential benefits and risks of serving in these roles were discussed. Issues addressed included acting as "caretaker manager" or "change-agent leader"; whether such service and experience would enhance or harm one's chances to become a permanent chair of that or another department; the effect of such service on academic productivity; the influence of department and institutional factors on the position; the range of authority provided, particularly in addressing significant problems affecting the department's future; and the impact of time served in these various positions. The "lame-duck" effect of prolonged service as "terminal chair" was also discussed. The observations and advice provided by the panelists and audience discussion are reported and may be useful for those considering service as temporary chair in pathology as well as other academic leadership positions.