Introduction The maintenance of a healthy epithelial-endothelial juxtaposition requires crosstalk within glomerular cellular niches. Here, we sought to understand the spatially anchored regulation and transition of endothelial and mesangial cells from health to injury in DKD. Methods From 132 human kidney samples, an integrated multiomics approach was leveraged to identify cellular niches, cell-cell communication, cell injury trajectories, and regulatory transcription factor networks in glomerular capillary endothelial (EC-GC) and mesangial cells. Data were culled from single nucleus RNA, ATAC-sequencing and four orthogonal spatial transcriptomic technologies for correlation with histopathological and clinical trial data. Results We identified a cellular niche in diabetic glomeruli enriched in a proliferative endothelial cell subtype (prEC) and altered vascular smooth muscle cells (VSMCs). Cellular communication within this niche maintained pro-angiogenic signaling with loss of anti-angiogenic factors. We identified a transcription factor network of MEF2C, MEF2A, and TRPS1 which regulated SEMA6A and PLXNA2, a receptor-ligand pair opposing angiogenesis. In silico knockout of the transcription factor network accelerated the transition from healthy EC-GCs toward a degenerative injured endothelial phenotype, with concomitant disruption of EC-GC and prEC expression patterns. Glomeruli enriched in the prEC niche had histologic evidence of neovascularization. MEF2C activity was increased in diabetic glomeruli with nodular mesangial sclerosis. The gene regulatory network (GRN) of MEF2C was dysregulated in EC-GCs of patients with DKD, but sodium glucose transporter-2 inhibitor (SGLT2i) treatment reversed the MEF2C GRN effects of DKD. Conclusions The MEF2C, MEF2A, and TRPS1 transcription factor network carefully balances the fate of the EC-GC in DKD. When the transcription factor network is “on” or over-expressed in DKD, EC-GCs may progress to a prEC state, while transcription factor suppression leads to cell death. SGLT2i therapy may restore the balance of MEF2C activity.

The study by Caza et al. is the largest study to date to examine the prevalence of nondiabetic kidney disease in clinically indicated kidney biopsies obtained from patients with diabetes. Nondiabetic kidney disease was identified in patients both with and without concurrent diabetic nephropathy and encompassed a wide spectrum of kidney diseases. Clinical data available at biopsy had limitations for predicting nondiabetic kidney disease and distinguishing affected patients from those with diabetic nephropathy only.

Background: The Kidney Precision Medicine Project (KPMP) consortium aims to redefine chronic kidney disease (CKD) by integrating clinical, pathological, and molecular tissue data from kidney biopsies. Here, we demonstrate how biopsy data in CKD can clarify disease etiology and contribute to understandings of disease pathophysiology and clinical prognosis. Methods: The KPMP is obtaining research kidney biopsies from individuals with CKD (defined as an estimated glomerular filtration rate [eGFR] < 60 mL/min/1.73m2 and/or albuminuria >30 mg/g creatinine) and diabetes (enrolled as diabetes and CKD or DKD) or hypertension (enrolled as hypertension and CKD or HCKD). A team of kidney pathologists and nephrologists adjudicated the primary clinico-pathological diagnosis for 258 participants with CKD. We compared pathological features and kidney transcriptional signatures between participants with a primary adjudicated diagnosis of diabetic nephropathy and those with other causes of CKD. We developed a model using clinical and biomarker data that predicted the probability of diabetic nephropathy and tested associations of the signature with CKD progression among Chronic Renal Insufficiency Cohort (CRIC) participants with diabetes (n=229). Results: Among 183 participants enrolled as DKD, 102 (56%) had a primary adjudicated clinico-pathologic diagnosis of diabetic nephropathy. Among 75 participants enrolled as HCKD, 42 (56%) had a primary diagnosis of hypertension-associated kidney disease. Those with diabetic nephropathy, compared with other diagnoses, had more severe interstitial fibrosis, tubular atrophy, tubular injury, segmental sclerosis, and severe arteriolar hyalinosis, and single-nucleus and single-cell transcriptional analyses revealed upregulation of immune and inflammatory pathways and downregulation of oxidative phosphorylation. A combination of age, hemoglobin A1c, urine albumin-creatinine ratio, and serum KIM-1 and sTNFR1 predicted a clinico-pathologic diagnosis of diabetic nephropathy in the KPMP (AUC 0.82, 95% CI 0.75-0.89) and was associated with an increased risk of CKD progression among patients with diabetes enrolled in CRIC (HR 1.48 [95% CI 1.27-1.73] per 10% higher predicted probability of diabetic nephropathy). Conclusion: In common presentations of CKD, kidney biopsies may alter a priori impressions, reveal a diversity of diagnosis, structure, and function that is associated with clinical outcomes and can impact therapeutic decisions. ### Competing Interest Statement The authors have declared no competing interest. ### Clinical Protocols ### Funding Statement The Kidney Precision Medicine Project (KPMP) is supported by the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) through the following grants: U01DK133081, U01DK133091, U01DK133092, U01DK133093, U01DK133095, U01DK133097, U01DK114866, U01DK114908, U01DK133090, U01DK133113, U01DK133766, U01DK133768, U01DK114907, U01DK114920, U01DK114923, U01DK114933, U24DK114886, UH3DK114926, UH3DK114861, UH3DK114915, and UH3DK114937. Funding for the CRIC Study was obtained under a cooperative agreement from National Institute of Diabetes and Digestive and Kidney Diseases (U01DK060990, U01DK060984, U01DK061022, U01DK061021, U01DK061028,U01DK060980, U01DK060963, U01DK060902 and U24DK060990). In addition, this work was supported in part by: the Perelman School of Medicine at the University of Pennsylvania Clinical and Translational Science Award NIH/NCATS UL1TR000003, Johns Hopkins University UL1 TR-000424, University of Maryland GCRC M01 RR-16500, Clinical and Translational Science Collaborative of Cleveland, UL1TR000439 from the National Center for Advancing Translational Sciences (NCATS) component of the National Institutes of Health and NIH roadmap for Medical Research, Michigan Institute for Clinical and Health Research (MICHR) UL1TR000433, University of Illinois at Chicago CTSA UL1RR029879, Tulane COBRE for Clinical and Translational Research in Cardiometabolic Diseases P20 GM109036, Kaiser Permanente NIH/NCRR UCSF-CTSI UL1 RR-024131, Department of Internal Medicine, University of New Mexico School of Medicine Albuquerque, NM R01DK119199. We gratefully acknowledge the essential contributions of our patient participants and the support of the American public through their tax dollars. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: The study was approved by the University of Washington Institutional Review Board (IRB 20190213). Written informed consent was received from all participants prior to study participation. I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes Clinical and histopathological data used in the study are available upon request to the KPMP via kpmp.org. Biomarker and molecular data used in the study are available online at kpmp.org.

The endothelin receptor antagonist atrasentan improved kidney outcomes in the SONAR trial for type 2 diabetes and chronic kidney disease (NCT01858532), though individual responses varied. To identify molecular biomarkers of atrasentan response and outcome, we conducted a nested case-control proteomics study (N = 180) within the SONAR trial population and identified urinary clusterin (uCLU) as the top candidate. Transcriptomic analyses of human kidney biopsies at tissue and single cell level from independent cohorts revealed higher CLU mRNA levels associated with worse kidney function and outcomes. An endothelin signaling activation score derived from pathway genes was reduced by atrasentan in mice with diabetic kidney disease. In the SONAR trial (N = 3,060) population, higher uCLU predicted worse outcomes, while atrasentan reduced uCLU by 42.6% over six weeks. Early uCLU changes independently predict improved kidney outcomes. In summary, uCLU is associated with kidney disease progression and response to atrasentan treatment, supporting its potential as a pharmacodynamic biomarker to target therapy.

BACKGROUND Acute interstitial nephritis (AIN) is a common cause of acute kidney injury (AKI), but the diagnosis may be missed as kidney biopsies are rarely obtained when acute tubular injury (ATI) is suspected.METHODS The Kidney Precision Medicine Project is a cohort study that obtains kidney biopsies from individuals with AKI, which undergo pathologic and molecular interrogation. We compared ATI and AIN cases among the first 60 AKI participants.RESULTS On clinicopathologic adjudication, 30 patients (50%) had a primary adjudicated diagnosis of ATI, 13 (22%) patients had AIN, 9 (15%) had diabetic nephropathy, and 3 (5%) had other conditions. There were increased interstitial white blood cells and tubulitis (P < 0.05 for both) in AIN compared with ATI. Prior to biopsy, the treating clinician suspected ATI in 83% of the cases with adjudicated ATI, while the treating clinician suspected AIN in 54% of the cases with AIN. Tissue transcriptomic signatures showed enrichment of proinflammatory signaling and increased expression of CXCL9, a chemokine induced by IFN-γ, in myeloid cells of participants with AIN. CXCL9 localized to inflammatory infiltration in spatial transcriptomic data.CONCLUSION Adjudication of kidney biopsies revealed distinct pathologic and molecular profiles between ATI and AIN. Kidney biopsy should be considered more frequently in AKI, as AIN is clinically underrecognized.TRIAL REGISTRATION ClinicalTrials.gov NCT04334707.FUNDING National Institute of Diabetes and Digestive and Kidney Diseases grants U01DK133081, U01DK133091, U01DK133092, U01DK133093, U01DK133095, U01DK133097, U01DK114866, U01DK114908, U01DK133090, U01DK133113, U01DK133766, U01DK133768, U01DK114907, U01DK114920, U01DK114923, U01DK114933, U24DK114886, UH3DK114926, UH3DK114861, UH3DK114915, and UH3DK114937.

Acute kidney injury (AKI) and chronic kidney disease (CKD) are two interconnected clinical conditions, both defined by degree of functional impairment, but with heterogeneous clinical trajectories. Using new transcriptomic technologies, recent studies have described the cellular diversity in the healthy and injured kidney at the single cell level. Here, we used single nucleus transcriptomics to investigate the molecular diversity and commonalities in kidney biopsies from over 150 participants with AKI and CKD enrolled within the Kidney Precision Medicine Project (KPMP) and did so at the patient participant level. Using an unsupervised approach, we identified two multi-cellular programs associated with clinical and histopathological features of acute injury and chronic damage, respectively. We found that these programs are expressed across patients with AKI and CKD, supporting shared, rather than distinct, underlying molecular mechanisms. These programs capture tissue-level compositional changes towards adaptive and failed-repair states in tubular epithelial cells, as well as intra-cellular molecular changes characteristic of stress in all cell types. We identified subunits of the NFkB and AP-1 complexes, as well as members of the STAT family, as putative upstream regulators of the acute and chronic programs. We were able to map these continuous molecular measures of acute injury and chronic damage to urine and plasma protein profiles obtained at time of biopsy. These non-invasive protein signatures were predictive of renal outcomes in an independent cohort of 44 thousand participants from the UK biobank. In summary, unbiased identification of cellular programs in kidney disease biopsies defined molecular programs of injury cutting across conventional disease categorization and established a non-invasive molecular link to long term patient outcomes.

Building upon a foundational Human Kidney resource, we present a comprehensive multi-modal atlas that defines spatially resolved versus unresolved repair states and mechanisms in human kidney disease. Homeostatic interactions between injured kidney epithelium and its surrounding milieu determine successful repair outcomes, while pathogenic signaling promotes unresolved inflammation and fibrosis leading to chronic disease. We integrated multiple single-cell and spatial modalities across ~700 samples from >350 patients (~250 research biopsies), analyzing ~1.7 million cells alongside complementary mouse multi-omic profiles spanning acute-to-chronic injury and aging (>300,000 cells) and spatial transcriptomic analysis of >150 human biopsies. This cross-species atlas delineates functional pathways and druggable targets across the nephron and defines gene regulatory networks and chromatin landscapes governing tubular, fibroblast, and immune cell transitions from injury to either recovery or failed repair states. We identified distinct cellular states associated with specific pathological features that show dynamic distributions between acute kidney injury (AKI) and chronic kidney disease (CKD), organized within unique spatial niches that reveal progression mechanisms from early injury to unresolved disease. Gene regulatory analyses prioritized key transcription factor activities (SOX4, SOX9, NFKB1, REL, KLFs) and their target networks establishing disease states and tissue microenvironments. These regulatory programs were directly linked to clinical outcomes, identifying molecular signatures of recovery and secreted biomarkers predictive of AKI-to-CKD progression, providing a key resource for therapeutic development and precision medicine approaches in kidney disease.

Rationale & Objective:The Kidney Precision Medicine Project is obtaining kidney biopsies from people with chronic kidney disease (CKD) and acute kidney injury for comprehensive clinical, histopathological, and molecular characterization. Here, we describe histopathological findings from a subset of kidney biopsies from adults with CKD. Study Design:Descriptive case series of histopathology findings from adjudicated CKD biopsies. Setting & Participants:Kidney Precision Medicine Project enrolled adults with CKD and diabetes (DKD) and/or hypertension (HCKD) with persistent eGFR 30-59 mL/min/1.73 m2, urine albumin-creatinine ratio ≥30 mg/g, or urine protein-creatinine ratio ≥150 mg/g. Clinicopathological adjudication by study nephrologists and kidney pathologists was completed as part of a pilot program for 39 participants enrolled 2019-2022. Clinicians completed surveys to assess impacts of biopsies on diagnosis, prognosis, and management. Exposures:This is a descriptive study without defined exposures. Outcomes:Characterization of glomerular, tubulointerstitial, and vascular histopathological features across CKD biopsies. Analytical Approach:Continuous variables were summarized as mean (standard deviation) or median (interquartile range); categorial variables were summarized as count (percentage). Results:Participants' mean age was 59 years, 59% were female. Mean eGFR was 55 mL/min/1.73 m2; median urine albumin-creatinine ratio and urine protein-creatinine ratio were 81 mg/g and 211 mg/g, respectively. Among DKD-enrolled participants (N = 28), 15 (54%) had a primary diagnosis of diabetic nephropathy, 3 (11%) had hypertension-associated nephropathy, 2 (7%) had other glomerular diseases, and 8 (29%) had nonspecific findings. Among HCKD-enrolled participants (N = 11), 5 (46%) had hypertension-associated nephropathy and 6 (55%) had nonspecific findings. A range of glomerular, tubulointerstitial, and vascular findings was observed. 26% of clinicians stated results were different than expected; 77% stated results affected prognostic discussions. Limitations:Small sample size and lack of longitudinal data limit generalizability. Conclusions:Kidney biopsies in people with common causes of CKD demonstrate a broad range of histopathology and may have clinical utility. Unsuspected disease processes and unexpected and nonspecific findings precluding a definitive diagnosis are often present.

The phase 3 DUPLEX trial is evaluating sparsentan, a novel, nonimmunosuppressive, single-molecule dual endothelin angiotensin receptor antagonist, in patients with focal segmental glomerulosclerosis (FSGS).

Diabetes increases the risk of both cardiovascular disease and kidney disease. Notably, most of the excess cardiovascular risk in people with diabetes is in those with kidney disease. Apolipoprotein C3 (APOC3) is a key regulator of plasma triglycerides, and it has recently been suggested to play a role in both type 1 diabetes-accelerated atherosclerosis and kidney disease progression. To investigate if APOC3 plays a role in kidney disease in people with type 2 diabetes, we analyzed plasma levels of APOC3 from the Veterans Affairs Diabetes Trial. Elevated baseline APOC3 levels predicted a greater loss of renal function. To mechanistically test if APOC3 plays a role in diabetic kidney disease and associated atherosclerosis, we treated black and tan, brachyury, WT and leptin-deficient (OB; diabetic) mice, a model of type 2 diabetes, with an antisense oligonucleotide (ASO) to APOC3 or a control ASO, all in the setting of human-like dyslipidemia. Silencing APOC3 prevented diabetes-augmented albuminuria, renal glomerular hypertrophy, monocyte recruitment, and macrophage accumulation, partly driven by reduced ICAM1 expression. Furthermore, reduced levels of APOC3 suppressed atherosclerosis associated with diabetes. This suggests that targeting APOC3 might benefit both diabetes-accelerated atherosclerosis and kidney disease.

In 2003, the title of a paper by Rollino et al asked the following question: "Is it possible to diagnose primary anti-phospholipid syndrome (PAPS) on the basis of renal thrombotic microangiopathy (PAPS nephropathy) in the absence of other thrombotic process?"1 The authors concluded, in essence, based on their experience with 3 patients and a literature review, that thrombotic microangiopathy (TMA) in the kidney in a patient with antiphospholipid antibodies (aPL) constituted a distinct clinicopathologic entity falling within the spectrum of PAPS.

Little is known about what constitutes the dense deposits of dense deposit disease (DDD), apart from components of the complement pathway. This study presents the novel finding that large accumulations of apolipoprotein E are present in the deposits of DDD, as revealed by mass spectroscopy and confirmed by both confocal microscopy and immunohistochemistry. The findings suggest a new modality for diagnosis of DDD and introduce potential new mechanisms for understanding DDD pathophysiology.

Immunoglobulin A (IgA)-dominant infection-related glomerulonephritis is a morphologic variant of acute infection-related glomerulonephritis that typically occurs in association with staphylococcal infection. It is most common in older patients and those with diabetes, and rare in children. Patients typically present with acute kidney injury, proteinuria, and hematuria. Hypocomplementemia is present in the vast majority of the patients. The prognosis is poor, with less than 20% of patients fully recovering kidney function, most likely owing to age-related underlying kidney changes and comorbidities, including diabetic kidney disease. The most common site of infection is skin, including cellulitis, surgical wound infection, skin abscesses, and intravenous line infection. The most common infectious agents are Staphylococcus, including methicillin-resistant Staphylococcus aureus (MRSA). Light microscopy: Glomeruli show segmental to global endocapillary hypercellularity with frequent neutrophils. Crescents may be present. Underlying changes of diabetic kidney disease may be present, including nodular glomerulosclerosis and arteriolar hyalinosis. Immunofluorescence microscopy: Glomeruli show chunky, irregular mesangial and capillary wall polyclonal IgA with lesser staining for IgG and prominent C3. The deposits often show a garland or starry-sky appearance as in typical poststreptococcal-related glomerulonephritis. Unlike IgA nephropathy, λ light chain is not typically dominant. Electron microscopy: Glomeruli show endocapillary hypercellularity, frequent intracapillary neutrophils, occasional mesangial and small subendothelial deposits, and hump-type large subepithelial deposits without surrounding glomerular basement membrane reaction. In more chronic phases, hump-type deposits may be sparse, located only in the hinge region. Although the pathogenesis of IgA nephropathy is linked to presence of galactose-deficient IgA1, the pathogenic mechanism of the selective IgA deposition in IgA-dominant infection-related glomerulonephritis is not well understood but likely involves specific host-pathogen interaction. Immune complexes may be circulating or the bacterial-derived antigen can bind within the glomerulus, with antibody and complement activation occurring in situ. Increased serum IgA levels have been reported in some, suggesting activation of selective IgA immune response. Polyclonal increases of serum IgA and IgG, high levels of circulating IgA and IgG immune complexes, and massive T-cell activation are hypothesized to play a role in this entity. Staphylococcal enterotoxin has been proposed as a causative agent in this entity following MRSA infection. Superantigens (enterotoxin) stimulate resting T cells to proliferate, causing a massive activation of T cells and a subsequent release of T cell–derived lymphokines (eg, interleukin 1, 2, or 6) and cytokines (eg, tumor necrosis factor α or interferon γ). More than 50% of patients with IgA-dominant infection-related glomerulonephritis have diabetes. Therefore, it is possible that the diabetic milieu plays a role in the mounting of an IgA-dominant immune response to the infection. IgA-dominant infection-related glomerulonephritis should be distinguished from IgA nephropathy and IgA vasculitis (Henoch-Schönlein purpura nephritis). Clinical and histopathologic features that suggest IgA-dominant infection-related glomerulonephritis include initial presentation in elderly patients, acute kidney injury at presentation, staphylococcal infection, low serum complement, diffuse endocapillary hypercellularity with prominent neutrophil infiltration, stronger staining for C3 than IgA and lack of λ light chain predominance on immunofluorescence, and the presence of hump-type subepithelial deposits on electron microscopy. Some studies suggest that weak or negative staining for galactose-deficient IgA1 may favor IgA-dominant infection-related glomerulonephritis over IgA nephropathy, but this staining is not currently used in clinical practice and additional studies are needed to confirm this notion. •Global endocapillary hypercellularity, with frequent neutrophils•Dominant irregular capillary wall and mesangial C3, with IgA-dominant staining, comparing to other immunoglobulins, and thus lesser staining for IgG•Subepithelial hump-like depositsFigure 2Crescents and fibrinoid necrosis may occur in IgA-dominant infection-related glomerulonephritis (Jones silver stain).View Large Image Figure ViewerDownload Hi-res image Download (PPT)Figure 3IgA-dominant infection-related glomerulonephritis with irregular chunky capillary loop and mesangial staining for immunoglobulin A (immunofluorescence microscopy).View Large Image Figure ViewerDownload Hi-res image Download (PPT)Figure 4Scattered, irregular capillary wall and mesangial course granular staining (starry-sky pattern) for C3 in IgA-dominant infection-related glomerulonephritis (immunofluorescence microscopy).View Large Image Figure ViewerDownload Hi-res image Download (PPT)Figure 5Subepithelial hump-like deposits in IgA-dominant infection-related glomerulonephritis. The deposits sit on the top of glomerular basement membrane, with no adjacent basement membrane reaction. The glomerular basement membrane is thickened, consistent with underlying diabetic nephropathy (electron microscopy).View Large Image Figure ViewerDownload Hi-res image Download (PPT) Paisit Paueksakon, MD, Behzad Najafian, MD, Charles E. Alpers, MD, and Agnes B. Fogo, MD. None. The authors declare that they have no relevant financial interests.

Introduction:The phase 3 DUPLEX trial is evaluating sparsentan, a novel, nonimmunosuppressive, single-molecule dual endothelin angiotensin receptor antagonist, in patients with focal segmental glomerulosclerosis (FSGS). Methods:DUPLEX (NCT03493685) is a global, multicenter, randomized, double-blind, parallel-group, active-controlled study evaluating the efficacy and safety of sparsentan 800 mg once daily versus irbesartan 300 mg once daily in patients aged 8 to 75 years (USA/UK) and 18 to 75 years (ex-USA/UK) weighing ≥20 kg with biopsy-proven FSGS or documented genetic mutation in a podocyte protein associated with FSGS, and urine protein-to-creatinine ratio (UP/C) ≥1.5 g/g. Baseline characteristics blinded to treatment allocation are reported descriptively. Results:The primary analysis population includes 371 patients (336 adult, 35 pediatric [<18 years]) who were randomized and received study drug (median age, 42 years). Patients were White (73.0%), Asian (13.2%), Black/African American (6.7%), or Other race (7.0%); and from North America (38.8%), Europe (36.1%), South America (12.7%), or Asia Pacific (12.4%). Baseline median UP/C was 3.0 g/g; 42.6% in nephrotic-range (UP/C >3.5 g/g [adults]; >2.0 g/g [pediatrics]). Patients were evenly distributed across estimated glomerular filtration rate (eGFR) categories corresponding to chronic kidney disease (CKD) stages 1 to 3b. Thirty-three patients (9.4% of 352 evaluable samples) had pathogenic or likely pathogenic (P/LP) variants of genes essential to podocyte structural integrity and function, 27 (7.7%) had P/LP collagen gene (COL4A3/4/5) variants, and 14 (4.0%) had high-risk APOL1 genotypes. Conclusions:Patient enrollment in DUPLEX, the largest interventional study in FSGS to date, will enable important characterization of the treatment effect of sparsentan in a geographically broad and clinically diverse FSGS population.