Development and Validation of a Multiclass Model Defining Molecular Archetypes of Kidney Transplant Rejection: A Large Cohort Study of the Banff Human Organ Transplant Gene Expression Panel.

Gene expression profiling from formalin-fixed paraffin-embedded (FFPE) renal allograft biopsies is a promising approach for feasibly providing a molecular diagnosis of rejection. However, large-scale studies evaluating the performance of models using NanoString platform data to define molecular archetypes of rejection are lacking. We tested a diverse retrospective cohort of over 1400 FFPE biopsy specimens, rescored according to Banff 2019 criteria and representing 10 of 11 United Network of Organ Sharing regions, using the Banff Human Organ Transplant panel from NanoString and developed a multiclass model from the gene expression data to assign relative probabilities of 4 molecular archetypes: No Rejection, Antibody-Mediated Rejection, T Cell-Mediated Rejection, and Mixed Rejection. Using Least Absolute Shrinkage and Selection Operator regularized regression with 10-fold cross-validation fitted to 1050 biopsies in the discovery cohort and technically validated on an additional 345 biopsies, our model achieved overall accuracy of 85% in the discovery cohort and 80% in the validation cohort, with ≥75% positive predictive value for each class, except for the Mixed Rejection class in the validation cohort (positive predictive value, 53%). This study represents the technical validation of the first model built from a large and diverse sample of diagnostic FFPE biopsy specimens to define and classify molecular archetypes of histologically defined diagnoses as derived from Banff Human Organ Transplant panel gene expression profiling data.

Discovery of seven novel putative antigens in membranous nephropathy and membranous lupus nephritis identified by mass spectrometry.

Multiple autoantigens have been identified in membranous nephropathy (MN) by tissue-based proteomics. However, antigenic targets of disease are unknown for over 10% of patients with MN and over half of those with membranous lupus nephritis (MLN). Here, we identified multiple new targets in PLA2R-/THSD7A-/EXT-/NELL1-quadruple negative MN biopsies through mass spectrometry of immune complexes recovered from biopsy tissue of patients with MN. Patients with MN negative for these four antigens were identified from Arkana Laboratories case archives. Protein G immunoprecipitation recovered immune complexes from frozen biopsy tissue from 142 quadruple-negative cases and 278 cases of known antigen type, followed by interrogation by mass spectrometry. Potential putative antigens were confirmed through paraffin immunofluorescence and co-localization with IgG within immune deposits. Consecutive series of 165 cases of PLA2R-negative MN and 142 MLN biopsies were screened to determine the frequency for each potential antigen. Seven putative antigens were discovered within immune complexes from biopsies of patients with MN including FCN3, CD206, EEA1, SEZ6L2, NPR3, MST1, and VASN. Peptides from these proteins were not enriched in the 278 cases of known antigen type. Between three to 30 unique peptides were detected for each new target. Frequencies of each biomarker, determined by staining consecutive case series, ranged from under 1 to 4.9%. NPR3 and CD206 were only positive in index cases. All cases showed co-localization of IgG within the immune deposits. Thus, seven putative antigens were newly identified in MN and MLN. Due to the number of antigens identified, it is becoming impractical to type PLA2R-negative MN or MLN cases through immunostaining alone. A multiplex approach is needed for subtyping of these diseases.

Proprotein convertase subtilisin/kexin type 6 (PCSK6) is a likely antigenic target in membranous nephropathy and nonsteroidal anti-inflammatory drug use.

Drugs are an important secondary cause of membranous nephropathy (MN) with the most common drugs associated with MN being nonsteroidal anti-inflammatory drugs (NSAIDs). Since the target antigen in NSAID-associated MN is not known, we performed laser microdissection of glomeruli followed by mass spectrometry (MS/MS) in 250 cases of PLA2R-negative MN to identify novel antigenic targets. This was followed by immunohistochemistry to localize the target antigen along the glomerular basement membrane and western blot analyses of eluates of frozen biopsy tissue to detect binding of IgG to the novel antigenic target. MS/MS studies revealed high total spectral counts of a novel protein Proprotein Convertase Subtilisin/Kexin Type 6 (PCSK 6) in five of the 250 cases in the discovery cohort. A validation cohort using protein G immunoprecipitation, MS/MS, and immunofluorescence detected PCSK6 in eight additional cases. All cases were negative for known antigens. Ten of 13 cases had a history of heavy NSAID use with no history available in one case. The mean serum creatinine and proteinuria at kidney biopsy were 0.93 ± 0.47 mg/dL and 6.5 ± 3.3 gms/day, respectively. Immunohistochemistry/immunofluorescence showed granular staining for PCSK6 along the glomerular basement membrane, and confocal microscopy showed co-localization of IgG and PCSK6. IgG subclass analysis in three cases revealed codominance of IgG1 and IgG4. Western blot analysis using eluates from frozen tissue showed IgG binding to PCSK6 in PCSK6-associated but not in PLA2R-positive MN. Thus, PCSK6 may be a likely novel antigenic target in MN in patients with prolonged NSAID use.

EXT1 and NCAM1-associated membranous lupus nephritis in a cohort of patients undergoing repeat kidney biopsies.

BACKGROUND: Exostosin 1/2 (EXT1/2) and neural cell adhesion molecule 1 (NCAM1) associated membranous lupus nephritis (MLN) may represent distinct disease phenotypes with prognostic significance. METHODS: We searched our archives for patients with systemic lupus erythematous (SLE) and at least two kidney biopsies demonstrating MLN. Each biopsy was stained for EXT1 and NCAM1 and scored as positive or negative. Histopathologic and clinical data were reviewed. RESULTS: We identified 31 patients with a clinical diagnosis of SLE and at least two kidney biopsies with MLN. A total of 28 patients (90%) showed concordant staining for EXT1 and NCAM1 in both biopsies; 8 patients (26%) were EXT1 positive and NCAM1 negative, 18 patients (58%) were EXT1 negative and NCAM1 negative and 2 patients (7%) were EXT1 negative and NCAM1 positive. A total of three patients (10%) had discordant EXT1 staining between their first and last biopsies; two patients (7%) were EXT1 positive in their first biopsy and EXT1 negative in the last biopsy and one patient (3%) was EXT1 negative in his first biopsy and EXT1 positive in the last biopsy. Compared with the EXT1-negative cohort at the time of the first biopsy, the EXT1-positive cohort had a higher average estimated glomerular filtration rate (eGFR; 141 versus 108 mL/min/1.73 m2; P = 0.04), lower average percent global glomerulosclerosis (0.5 versus 12%; P = 0.05), lower average interstitial fibrosis and tubular atrophy (2.5 versus 11.7%; P = 0.06) and lower average total National Institutes of Health (NH) chronicity scores (0.75 versus 2.33; P = 0.05). On long-term follow-up, the rate of change in eGFR did not significantly differ between the two groups (P = 0.24). One EXT1-positive patient (12.5%) developed stage 4 chronic kidney disease (CKD) or end-stage kidney disease (ESKD) compared with four patients (20%) in the EXT-negative group and two of the three EXT1-discordant patients (P = 0.38). CONCLUSIONS: We performed the largest retrospective repeat-biopsy study to evaluate EXT1 and NCAM1 autoantigens in MLN. Our data demonstrate that EXT1 positivity is associated with better kidney function at the time of diagnosis and raises the possibility that EXT1 status may change throughout the disease course of MLN.

Nephronophthisis: a pathological and genetic perspective.

Nephronophthisis (NPHP) is an autosomal recessive cystic kidney disease and is one of the most frequent genetic causes for kidney failure (KF) in children and adolescents. Over 20 genes cause NPHP and over 90 genes contribute to renal ciliopathies often involving multiple organs. About 15-20% of NPHP patients have additional extrarenal symptoms affecting other organs than the kidneys. The involvement of additional organ systems in syndromic forms of NPHP is explained by shared expression of most NPHP gene products in centrosomes and primary cilia, a sensory organelle present in most mammalian cells. This finding resulted in the classification of NPHP as a ciliopathy. If extrarenal symptoms are present in addition to NPHP, these disorders are defined as NPHP-related ciliopathies (NPHP-RC) and can involve the retina (e.g., with Senior-Løken syndrome), CNS (central nervous system) (e.g., with Joubert syndrome), liver (e.g., Boichis and Arima syndromes), or bone (e.g., Mainzer-Saldino and Sensenbrenner syndromes). This review focuses on the pathological findings and the recent genetic advances in NPHP and NPHP-RC. Different mechanisms and signaling pathways are involved in NPHP ranging from planar cell polarity, sonic hedgehog signaling (Shh), DNA damage response pathway, Hippo, mTOR, and cAMP signaling. A number of therapeutic interventions appear to be promising, ranging from vasopressin receptor 2 antagonists such as tolvaptan, cyclin-dependent kinase inhibitors such as roscovitine, Hh agonists such as purmorphamine, and mTOR inhibitors such as rapamycin.

The characteristics of patients with kidney light chain deposition disease concurrent with light chain amyloidosis.

The type of monoclonal light chain nephropathy is thought to be largely a function of the structural and physiochemical properties of light chains; hence most affected patients have only one light chain kidney disease type. Here, we report the first series of kidney light chain deposition disease (LCDD) concomitant with light chain amyloidosis (LCDD+AL), with or without light chain cast nephropathy (LCCN). Our LCDD+AL cohort consisted of 37 patients (54% females, median age 70 years (range 40-86)). All cases showed Congo red-positive amyloid deposits staining for one light chain isotype on immunofluorescence (62% lambda), and LCDD with diffuse linear staining of glomerular and tubular basement membranes for one light chain isotype (97% same isotype as the amyloidogenic light chain) and ultrastructural non-fibrillar punctate deposits. Twelve of 37 cases (about 1/3 of patients) had concomitant LCCN of same light chain isotype. Proteomic analysis of amyloid and/or LCDD deposits in eight revealed a single light chain variable domain mutable subgroup in all cases (including three with separate microdissections of LCDD and amyloid light chain deposits). Clinical data on 21 patients showed proteinuria (100%), hematuria (75%), kidney insufficiency and nephrotic syndrome (55%). Extra-kidney involvement was present in 43% of the patients. Multiple myeloma occurred in 68% (about 2/3) of these patients; none had lymphoma. On follow up (median 16 months), 63% developed kidney failure and 56% died. The median kidney and patient survivals were 12 and 32 months, respectively. LCDD+AL mainly affected patients 60 years of age or older. Thus, LCDD+AL could be caused by two pathological light chains produced by subclones stemming from one immunoglobulin light chain lambda or kappa rearrangement, with a distinct mutated complementary determining region.

Membranous Nephropathy With Extensive Tubular Basement Membrane Deposits Following Allogeneic Hematopoietic Cell Transplant: A Report of 5 Cases.

Tubular basement membrane (TBM) deposits are very uncommon in non-lupus membranous nephropathy. We report 5 patients with membranous nephropathy and extensive TBM deposits following allogeneic hematopoietic cell transplant. Patients presented with nephrotic syndrome (3 also had acute kidney injury) late post-transplant in association with chronic graft-versus-host disease (cGVHD). Kidney biopsies revealed global subepithelial and extensive TBM immune complex deposits, accompanied by acute tubular injury (n = 4) and tubulointerstitial inflammation (n = 4). Proteomic analysis of glomeruli in 4 cases identified PLA2R in 1, with no significant protein spectra for PLA2R, THSD7A, EX1/2, NELL-1, PCDH7, NCAM1, or SEMA3B detected in the remaining 3. On follow-up (for a mean 42 months), 4 patients had complete and 1 partial remission following prednisone and/or rituximab therapy. We propose that membranous nephropathy with extensive TBM deposits is a distinctive clinicopathologic lesion associated with allogeneic hematopoietic cell transplant. Pathogenesis likely involves cGVHD-driven antibodies against glomerular and TBM components, the identity of which remains to be elucidated.

NELL1 is a target antigen in malignancy-associated membranous nephropathy.

Patients with membranous nephropathy have an increased risk of malignancy compared to the general population, but the target antigen for malignancy-associated membranous nephropathy is unknown. To explore this, we utilized mass spectrometry for antigen discovery in malignancy-associated membranous nephropathy examining immune complexes eluted from frozen kidney biopsy tissue using protein G bead immunoglobulin capture. Antigen discovery was performed comparing cases of membranous nephropathy of unknown and known type. Mass spectrophotometric analysis revealed that nerve epidermal growth factor-like 1 (NELL1) immune complexes were uniquely present within the biopsy tissue in membranous nephropathy. Additional NELL1-positive cases were subsequently identified by immunofluorescence. In a consecutive series, 3.8% of PLA2R- and THSD7A-negative cases were NELL1-positive. These NELL1-positive cases had segmental to incomplete IgG capillary loop staining (93.4%) and dominant or co-dominant IgG1-subclass staining (95.5%). The mean age of patients with NELL1-positive membranous nephropathy was 66.8 years, with a slight male predominance (58.2%) and 33% had concurrent malignancy. Compared with PLA2R- and THSD7A-positive cases of membranous nephropathy, there was a greater proportion of cases with malignancies in the NELL1-associated group. Thus, NELL1-associated membranous nephropathy has a unique histopathology characterized by incomplete capillary loop staining, IgG1-predominance, and is more often associated with malignancy than other known types of membranous nephropathy.

Neural cell adhesion molecule 1 is a novel autoantigen in membranous lupus nephritis.

Membranous lupus nephritis is a frequent cause of nephrotic syndrome in patients with systemic lupus erythematosus. It has been shown in phospholipase A2 receptor positive membranous nephropathy that known antibodies can be detected within sera, determination of the target autoantigen can have diagnostic significance, inform prognosis, and enable non-invasive monitoring of disease activity. Here we utilized mass spectrometry for antigen discovery in laser captured microdissected glomeruli from formalin-fixed paraffin embedded tissue and tissue protein G immunoprecipitation studies to interrogate immune complexes from frozen kidney biopsy tissue. We identified neural cell adhesion molecule 1 (NCAM1) to be a target antigen in some cases of membranous lupus nephritis and within rare cases of primary membranous nephropathy. The prevalence of NCAM1 association was 6.6% of cases of membranous lupus nephritis and in 2.0% of primary membranous nephropathy cases. NCAM1 was found to colocalize with IgG within glomerular immune deposits by confocal microscopy. Additionally, serum from patients with NCAM1-associated membranous nephropathy showed reactivity to NCAM1 recombinant protein on Western blotting and by indirect immunofluorescence assay, demonstrating the presence of circulating antibodies. Thus, we propose that NCAM1 is a target autoantigen in a subset of patients with membranous lupus nephritis. Future studies are needed to determine whether anti-NCAM1 antibody levels correlate with disease activity or response to therapy.

A multi-center retrospective cohort study defines the spectrum of kidney pathology in Coronavirus 2019 Disease (COVID-19).

Kidney failure is common in patients with Coronavirus Disease-19 (COVID-19), resulting in increased morbidity and mortality. In an international collaboration, 284 kidney biopsies were evaluated to improve understanding of kidney disease in COVID-19. Diagnoses were compared to five years of 63,575 native biopsies prior to the pandemic and 13,955 allograft biopsies to identify diseases that have increased in patients with COVID-19. Genotyping for APOL1 G1 and G2 alleles was performed in 107 African American and Hispanic patients. Immunohistochemistry for SARS-CoV-2 was utilized to assess direct viral infection in 273 cases along with clinical information at the time of biopsy. The leading indication for native biopsy was acute kidney injury (45.4%), followed by proteinuria with or without concurrent acute kidney injury (42.6%). There were more African American patients (44.6%) than patients of other ethnicities. The most common diagnosis in native biopsies was collapsing glomerulopathy (25.8%), which was associated with high-risk APOL1 genotypes in 91.7% of cases. Compared to the five-year biopsy database, the frequency of myoglobin cast nephropathy and proliferative glomerulonephritis with monoclonal IgG deposits was also increased in patients with COVID-19 (3.3% and 1.7%, respectively), while there was a reduced frequency of chronic conditions (including diabetes mellitus, IgA nephropathy, and arterionephrosclerosis) as the primary diagnosis. In transplants, the leading indication was acute kidney injury (86.4%), for which rejection was the predominant diagnosis (61.4%). Direct SARS-CoV-2 viral infection was not identified. Thus, our multi-center large case series identified kidney diseases that disproportionately affect patients with COVID-19 and demonstrated a high frequency of APOL1 high-risk genotypes within this group, with no evidence of direct viral infection within the kidney.

Immunoglobulin-Negative DNAJB9-Associated Fibrillary Glomerulonephritis: A Report of 9 Cases.

Fibrillary glomerulonephritis (FGN) was previously defined by glomerular deposition of haphazardly oriented fibrils that stain with antisera to immunoglobulins but do not stain with Congo red. We report what is to our knowledge the first series of immunoglobulin-negative FGN, consisting of 9 adults (7 women and 2 men) with a mean age at diagnosis of 66 years. Patients presented with proteinuria (100%; mean protein excretion, 3g/d), hematuria (100%), and elevated serum creatinine level (100%). Comorbid conditions included carcinoma in 3 and hepatitis C virus infection in 2; no patient had hypocomplementemia or monoclonal gammopathy. Histologically, glomeruli were positive for DNAJB9, showed mostly mild mesangial hypercellularity and/or sclerosis, and were negative for immunoglobulins by immunofluorescence on frozen and paraffin tissue. Ultrastructurally, randomly oriented fibrils measuring 13 to 20nm in diameter were seen intermingling with mesangial matrix in all and infiltrating glomerular basement membranes in 5. On follow-up (mean duration, 21 months), 2 had disease remission, 4 had persistently elevated serum creatinine levels and proteinuria, and 3 required kidney replacement therapy. Thus, rare cases of FGN are not associated with glomerular immunoglobulin deposition, and the diagnosis of FGN in these cases can be confirmed by DNAJB9 immunostaining. Pathogenesis remains to be elucidated.

COVID-19-Associated Kidney Injury: A Case Series of Kidney Biopsy Findings.

BACKGROUND: Reports show that AKI is a common complication of severe coronavirus disease 2019 (COVID-19) in hospitalized patients. Studies have also observed proteinuria and microscopic hematuria in such patients. Although a recent autopsy series of patients who died with severe COVID-19 in China found acute tubular necrosis in the kidney, a few patient reports have also described collapsing glomerulopathy in COVID-19. METHODS: We evaluated biopsied kidney samples from ten patients at our institution who had COVID-19 and clinical features of AKI, including proteinuria with or without hematuria. We documented clinical features, pathologic findings, and outcomes. RESULTS: Our analysis included ten patients who underwent kidney biopsy (mean age: 65 years); five patients were black, three were Hispanic, and two were white. All patients had proteinuria. Eight patients had severe AKI, necessitating RRT. All biopsy samples showed varying degrees of acute tubular necrosis, and one patient had associated widespread myoglobin casts. In addition, two patients had findings of thrombotic microangiopathy, one had pauci-immune crescentic GN, and another had global as well as segmental glomerulosclerosis with features of healed collapsing glomerulopathy. Interestingly, although the patients had confirmed severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection by RT-PCR, immunohistochemical staining of kidney biopsy samples for SARS-CoV-2 was negative in all ten patients. Also, ultrastructural examination by electron microscopy showed no evidence of viral particles in the biopsy samples. CONCLUSIONS: The most common finding in our kidney biopsy samples from ten hospitalized patients with AKI and COVID-19 was acute tubular necrosis. There was no evidence of SARS-CoV-2 in the biopsied kidney tissue.

AKI and Collapsing Glomerulopathy Associated with COVID-19 and APOL 1 High-Risk Genotype.

BACKGROUND: Kidney involvement is a feature of COVID-19 and it can be severe in Black patients. Previous research linked increased susceptibility to collapsing glomerulopathy, including in patients with HIV-associated nephropathy, to apo L1 (APOL1) variants that are more common in those of African descent. METHODS: To investigate genetic, histopathologic, and molecular features in six Black patients with COVID-19 presenting with AKI and de novo nephrotic-range proteinuria, we obtained biopsied kidney tissue, which was examined by in situ hybridization for viral detection and by NanoString for COVID-19 and acute tubular injury-associated genes. We also collected peripheral blood for APOL1 genotyping. RESULTS: This case series included six Black patients with COVID-19 (four men, two women), mean age 55 years. At biopsy day, mean serum creatinine was 6.5 mg/dl and mean urine protein-creatinine ratio was 11.5 g. Kidney biopsy specimens showed collapsing glomerulopathy, extensive foot process effacement, and focal/diffuse acute tubular injury. Three patients had endothelial reticular aggregates. We found no evidence of viral particles or SARS-CoV-2 RNA. NanoString showed elevated chemokine gene expression and changes in expression of genes associated with acute tubular injury compared with controls. All six patients had an APOL1 high-risk genotype. Five patients needed dialysis (two of whom died); one partially recovered without dialysis. CONCLUSIONS: Collapsing glomerulopathy in Black patients with COVID-19 was associated with high-risk APOL1 variants. We found no direct viral infection in the kidneys, suggesting a possible alternative mechanism: a "two-hit" combination of genetic predisposition and cytokine-mediated host response to SARS-CoV-2 infection. Given this entity's resemblance with HIV-associated nephropathy, we propose the term COVID-19-associated nephropathy to describe it.

Proteomic Analysis Identifies Distinct Glomerular Extracellular Matrix in Collapsing Focal Segmental Glomerulosclerosis.

BACKGROUND: The mechanisms leading to extracellular matrix (ECM) replacement of areas of glomerular capillaries in histologic variants of FSGS are unknown. This study used proteomics to test the hypothesis that glomerular ECM composition in collapsing FSGS (cFSGS) differs from that of other variants. METHODS: ECM proteins in glomeruli from biopsy specimens of patients with FSGS not otherwise specified (FSGS-NOS) or cFSGS and from normal controls were distinguished and quantified using mass spectrometry, verified and localized using immunohistochemistry (IHC) and confocal microscopy, and assessed for gene expression. The analysis also quantified urinary excretion of ECM proteins and peptides. RESULTS: Of 58 ECM proteins that differed in abundance between cFSGS and FSGS-NOS, 41 were more abundant in cFSGS and 17 in FSGS-NOS. IHC showed that glomerular tuft staining for cathepsin B, cathepsin C, and annexin A3 in cFSGS was significantly greater than in other FSGS variants, in minimal change disease, or in membranous nephropathy. Annexin A3 colocalized with cathepsin B and C, claudin-1, phosphorylated ERK1/2, and CD44, but not with synaptopodin, in parietal epithelial cells (PECs) infiltrating cFSGS glomeruli. Transcripts for cathepsins B and C were increased in FSGS glomeruli compared with normal controls, and urinary excretion of both cathepsins was significantly greater in cFSGS compared with FSGS-NOS. Urinary excretion of ECM-derived peptides was enhanced in cFSGS, although in silico analysis did not identify enhanced excretion of peptides derived from cathepsin B or C. CONCLUSIONS: ECM differences suggest that glomerular sclerosis in cFSGS differs from that in other FSGS variants. Infiltration of activated PECs may disrupt ECM remodeling in cFSGS. These cells and their cathepsins may be therapeutic targets.

Detection of SARS-CoV-2 in formalin-fixed paraffin-embedded tissue sections using commercially available reagents.

Coronavirus Disease-19 (COVID-19), caused by the coronavirus SARS-CoV-2, was initially recognized in Wuhan, China and subsequently spread to all continents. The disease primarily affects the lower respiratory system, but may involve other organs and systems. Histopathologic evaluation of tissue from affected patients is crucial for diagnostic purposes, but also for advancing our understanding of the disease. For that reason, we developed immunohistochemical (IHC) and in situ hybridization (ISH) assays for detection of the. virus. A total of eight autopsy lungs, one placenta, and ten kidney biopsies from COVID-19 patients were stained with a panel of commercially available antibodies for IHC and commercially available RNA probes for ISH. Similarly, autopsy lungs, placentas and renal biopsies from non-COVID-19 patients were stained with the same antibodies and probes. All eight lungs and the placenta from COVID-19 patients stained positive by IHC and ISH, while the kidney biopsies stained negative by both methodologies. As expected, all specimens from non-COVID-19 patients were IHC and ISH negative. These two assays represent a sensitive and specific method for detecting the virus in tissue samples. We provide the protocols and the list of commercially available antibodies and probes for these assays, so they can be readily implemented in pathology laboratories and medical examiner offices for diagnostic and research purposes.

Serum amyloid P deposition is a sensitive and specific feature of membranous-like glomerulopathy with masked IgG kappa deposits.

Membranous-like glomerulopathy with masked IgG kappa deposits (MGMID) is a recently described pattern of glomerulonephritis with a unique histopathology. The pattern is characterized by subepithelial and/or mesangial immune deposits that are "masked", to immunoglobulin staining by routine immunofluorescence but strongly stain for IgG and kappa light chain after protease digestion. Patients with this pattern of glomerulonephritis are most commonly young females presenting with proteinuria and a vague history of autoimmune disease such as low titer antinuclear antibodies. Here we compared the mass spectrometry profile of laser capture microdissected glomeruli from nine MGMID renal biopsies with eight biopsies showing other patterns of membranous glomerulopathy. The protein most significantly increased in MGMID was serum amyloid P. Immunostaining showed serum amyloid P colocalized with IgG in the glomeruli of MGMID but not with PLA2R-associated membranous glomerulopathy. Serum amyloid P was positive in the glomeruli of all 32 MGMID biopsies but negative in biopsies of other types of membranous glomerulopathies such as those associated with PLA2R and THSD7A. There were four biopsies with glomerular serum amyloid P staining among the 173 biopsies that did not fulfill criteria for MGMID or amyloidosis. All four of these biopsies with positive serum amyloid P staining had a membranous pattern of glomerulopathy with IgG kappa deposits that only differed from MGMID by the lack of "masking". Thus, positive staining within glomerular deposits for serum amyloid P identifies a unique form of glomerulonephritis likely sharing a common pathophysiologic mechanism of disease.

Light chain only variant of proliferative glomerulonephritis with monoclonal immunoglobulin deposits is associated with a high detection rate of the pathogenic plasma cell clone.

IgG (mainly IgG3) is the most commonly involved isotype in proliferative glomerulonephritis with monoclonal immunoglobulin deposits (PGNMID). Here we describe the first series of PGNMID with deposition of monoclonal immunoglobulin light chain only (PGNMID-light chain). This multicenter cohort of 17 patients presented with nephritic or nephrotic syndrome with underlying hematologic conditions of monoclonal gammopathy of renal significance (71%) or multiple myeloma (29%). Monoclonal immunoglobulin was identified by serum and urine immunofixation in 65% and 73%, respectively, with abnormal serum free light chain in 83%, and a detectable bone marrow plasma cell clone in 88% of patients. Renal biopsy showed a membranoproliferative pattern in most patients. By immunofluorescence, deposits were restricted to glomeruli and composed of restricted light chain (kappa in 71%) and C3, with granular appearance and subendothelial, mesangial and subepithelial distribution by electron microscopy. Proteomic analysis in four cases of kappa PGNMID-light chain revealed spectra for kappa constant and variable domains, without evidence of Ig heavy chains; spectra for proteins of the alternative pathway of complement and terminal complex were detected in three. The classical pathway was not detected in three cases. After median follow up of 70 months, the renal response was dependent on a hematologic response and occurred in six of ten patients treated with plasma cell-directed chemotherapy but none of five patients receiving other therapies. Thus, PGNMID-light chain differs from PGNMID-IgG by higher frequency of a detectable pathogenic plasma cell clone. Hence, proper recognition is crucial as anti-myeloma agents may improve renal prognosis. Activation of an alternative pathway of complement by monoclonal immunoglobulin light chain likely plays a role in its pathogenesis.

APOL1-Associated Collapsing Focal Segmental Glomerulosclerosis in a Patient With Stimulator of Interferon Genes (STING)-Associated Vasculopathy With Onset in Infancy (SAVI).

Apolipoprotein L1 (APOL1) risk variants G1 and G2 are known to result in risk for kidney disease in patients of African ancestry. APOL1-associated nephropathy typically occurs in association with certain environmental factors or systemic diseases. As such, there has been increasing evidence of the role of interferon (IFN) pathways in the pathogenesis of APOL1-associated collapsing glomerulopathy in patients with human immunodeficiency virus (HIV) infection and systemic lupus erythematosus, 2 conditions that are associated with high IFN levels. Collapsing glomerulopathy has also been described in patients receiving exogenous IFN therapy administered for various medical conditions. We describe a patient with a genetic condition that results in an increased IFN state, stimulator of IFN genes (STING)-associated vasculopathy with onset in infancy (SAVI), who developed collapsing glomerulopathy during a flare of his disease. The patient was found to have APOL1 G1 and G2 risk variants. This case supports the role of IFN in inducing APOL1-associated collapsing glomerulopathy.

A diagnostician's field guide to crystalline nephropathies.

The kidney's role in filtration of blood and production of urine occurs via a combination of size and charge filtration at the glomerular basement membrane and resorption and excretion of molecules through a complex tubular system embedded within an ion gradient. This delicate system provides the kidney with a unique propensity for substrate saturation and crystal nucleation within the nephron. While crystalline nephropathies may seem exotic to the uninitiated, they are comprised of easily recognizable morphologies and generally lack complicated classification schemas. Additionally, unlike many intrinsic kidney diseases, crystalline nephropathies are often associated with systemic conditions that, upon further investigation, may elucidate critically important information. This review focuses on practical, diagnostically relevant and high yield information that can be utilized by diagnosticians. Our hope is to equip the reader who reviews renal tissue with a practical toolkit that they feel empowered to use when faced with crystal formation in a kidney biopsy, pre-implantation biopsy, or nephrectomy specimen. Short Abstract The kidney's role in filtration of blood and production of urine provides a unique propensity for substrate saturation and crystal nucleation within the nephron. While crystalline nephropathies may seem exotic to the uninitiated, they are comprised of easily recognizable morphologies and generally lack complicated classification. Additionally, crystalline nephropathies are often associated with systemic conditions that, upon further investigation, may elucidate critically important information.

A practical approach to the pathology of renal intratubular casts.

The identification and proper characterization of pathologic renal intratubular casts can be an arduous task, especially since they often admixed with non-pathologic casts, obfuscating debris and inflammation. The list of pathologic intratubular casts is long, and they can be easily missed or misdiagnosed without a thorough understanding of their pathophysiology and morphologic variety. Correct characterization of tubular casts is important since each cast type has a unique pathogenic mechanism, with specific treatment and prognostic implications. This review discusses the clinicopathologic characteristics of the six most common pathologic casts: light chain, hemoglobin, myoglobin, red cell, neutrophilic and bile casts. We also discuss hyaline and uromodulin casts, the commonly encountered "benign" cast types that share certain histologic features with pathologic casts. We limit the discussion to proteinaceous and cellular intratubular casts, with crystalline casts discussed in a separate review within the same journal issue. While not exhaustive, this review covers pathogenesis, clinical and prognostic significance, and a practical discussion of the histomorphologic spectrum of each cast type, along with commonly encountered pitfalls.