Polyploid tubular cells: a shortcut to stress adaptation.

Tubular epithelial cells (TCs) compose the majority of kidney parenchyma and play fundamental roles in maintaining homeostasis. Like other tissues, mostly immature TC with progenitor capabilities are able to replace TC lost during injury via clonal expansion and differentiation. In contrast, differentiated TC lack this capacity. However, as the kidney is frequently exposed to toxic injuries, evolution positively selected a response program that endows differentiated TC to maintain residual kidney function during kidney injury. Recently, we and others have described polyploidization of differentiated TC, a mechanism to augment the function of remnant TC after injury by rapid hypertrophy. Polyploidy is a condition characterized by >2 complete sets of chromosomes. Polyploid cells often display an increased functional capacity and are generally more resilient to stress as evidenced by being conserved across many plants and eukaryote species from flies to mammals. Here, we discuss the occurrence of TC polyploidy in different contexts and conditions and how this integrates into existing concepts of kidney cell responses to injury. Collectively, we aim at stimulating the acquisition of novel knowledge in the kidney field as well as accelerating the translation of this basic response mechanism to the clinical sphere.

Soluble uric acid inhibits ß2 integrin-mediated neutrophil recruitment in innate immunity.

Neutrophils are key players during host defense and sterile inflammation. Neutrophil dysfunction is a characteristic feature of the acquired immunodeficiency during kidney disease. We speculated that the impaired renal clearance of the intrinsic purine metabolite soluble uric acid (sUA) may account for neutrophil dysfunction. Indeed, hyperuricemia (HU, serum UA of 9-12 mg/dL) related or unrelated to kidney dysfunction significantly diminished neutrophil adhesion and extravasation in mice with crystal- and coronavirus-related sterile inflammation using intravital microscopy and an air pouch model. This impaired neutrophil recruitment was partially reversible by depleting UA with rasburicase. We validated these findings in vitro using either neutrophils or serum from patients with kidney dysfunction-related HU with or without UA depletion, which partially normalized the defective migration of neutrophils. Mechanistically, sUA impaired ß2 integrin activity and internalization/recycling by regulating intracellular pH and cytoskeletal dynamics, physiological processes that are known to alter the migratory and phagocytic capability of neutrophils. This effect was fully reversible by blocking intracellular uptake of sUA via urate transporters. In contrast, sUA had no effect on neutrophil extracellular trap formation in neutrophils from healthy subjects or patients with kidney dysfunction. Our results identify an unexpected immunoregulatory role of the intrinsic purine metabolite sUA, which contrasts the well-known immunostimulatory effects of crystalline UA. Specifically targeting UA may help to overcome certain forms of immunodeficiency, for example in kidney dysfunction, but may enhance sterile forms of inflammation.

Renal Progenitors Derived from Urine for Personalized Diagnosis of Kidney Diseases.

BACKGROUND: Chronic kidney disease affects 10% of the world population, and it is associated with progression to end-stage kidney disease and increased morbidity and mortality. The advent of multi-omics technologies has expanded our knowledge on the complexity of kidney diseases, revealing their frequent genetic etiology, particularly in children and young subjects. Genetic heterogeneity and drug screening require patient-derived disease models to establish a correct diagnosis and evaluate new potential treatments and outcomes. SUMMARY: Patient-derived renal progenitors can be isolated from urine to set up proper disease modeling. This strategy allows to make diagnosis of genetic kidney disease in patients carrying unknown significance variants or uncover variants missed from peripheral blood analysis. Furthermore, urinary-derived tubuloids obtained from renal progenitors of patients appear to be potentially valuable for modeling kidney diseases to test ex vivo treatment efficacy or to develop new therapeutic approaches. Finally, renal progenitors derived from urine can provide insights into acute kidney injury and predict kidney function recovery and outcome. KEY MESSAGES: Renal progenitors derived from urine are a promising new noninvasive and easy-to-handle tool, which improves the rate of diagnosis and the therapeutic choice, paving the way toward a personalized healthcare.

Tubular Epithelial Cell HMGB1 Promotes AKI-CKD Transition by Sensitizing Cycling Tubular Cells to Oxidative Stress: A Rationale for Targeting HMGB1 during AKI Recovery.

SIGNIFICANCE STATEMENT: Cells undergoing necrosis release extracellular high mobility group box (HMGB)-1, which triggers sterile inflammation upon AKI in mice. Neither deletion of HMGB1 from tubular epithelial cells, nor HMGB1 antagonism with small molecules, affects initial ischemic tubular necrosis and immediate GFR loss upon unilateral ischemia/reperfusion injury (IRI). On the contrary, tubular cell-specific HMGB1 deficiency, and even late-onset pharmacological HMGB1 inhibition, increased functional and structural recovery from AKI, indicating that intracellular HMGB1 partially counters the effects of extracellular HMGB1. In vitro studies indicate that intracellular HMGB1 decreases resilience of tubular cells from prolonged ischemic stress, as in unilateral IRI. Intracellular HMGB1 is a potential target to enhance kidney regeneration and to improve long-term prognosis in AKI. BACKGROUND: Late diagnosis is a hurdle for treatment of AKI, but targeting AKI-CKD transition may improve outcomes. High mobility group box-1 (HMGB1) is a nuclear regulator of transcription and a driver of necroinflammation in AKI. We hypothesized that HMGB1 would also modulate AKI-CKD transition in other ways. METHODS: We conducted single-cell transcriptome analysis of human and mouse AKI and mouse in vivo and in vitro studies with tubular cell-specific depletion of Hmgb1 and HMGB1 antagonists. RESULTS: HMGB1 was ubiquitously expressed in kidney cells. Preemptive HMGB1 antagonism with glycyrrhizic acid (Gly) and ethyl pyruvate (EP) did not affect postischemic AKI but attenuated AKI-CKD transition in a model of persistent kidney hypoxia. Consistently, tubular Hmgb1 depletion in Pax8 rtTA, TetO Cre, Hmgb1fl/fl mice did not protect from AKI, but from AKI-CKD transition. In vitro studies confirmed that absence of HMGB1 or HMGB1 inhibition with Gly and EP does not affect ischemic necrosis of growth-arrested differentiated tubular cells but increased the resilience of cycling tubular cells that survived the acute injury to oxidative stress. This effect persisted when neutralizing extracellular HMGB1 with 2G7. Consistently, late-onset HMGB1 blockade with EP started after the peak of ischemic AKI in mice prevented AKI-CKD transition, even when 2G7 blocked extracellular HMGB1. CONCLUSION: Treatment of AKI could become feasible when ( 1 ) focusing on long-term outcomes of AKI; ( 2 ) targeting AKI-CKD transition with drugs initiated after the AKI peak; and ( 3 ) targeting with drugs that block HMGB1 in intracellular and extracellular compartments.

Finerenone Added to RAS/SGLT2 Blockade for CKD in Alport Syndrome. Results of a Randomized Controlled Trial with Col4a3-/- Mice.

SIGNIFICANCE STATEMENT: We hypothesized that triple therapy with inhibitors of the renin-angiotensin system (RAS), sodium-glucose transporter (SGLT)-2, and the mineralocorticoid receptor (MR) would be superior to dual RAS/SGLT2 blockade in attenuating CKD progression in Col4a3 -deficient mice, a model of Alport syndrome. Late-onset ramipril monotherapy or dual ramipril/empagliflozin therapy attenuated CKD and prolonged overall survival by 2 weeks. Adding the nonsteroidal MR antagonist finerenone extended survival by 4 weeks. Pathomics and RNA sequencing revealed significant protective effects on the tubulointerstitium when adding finerenone to RAS/SGLT2 inhibition. Thus, triple RAS/SGLT2/MR blockade has synergistic effects and might attenuate CKD progression in patients with Alport syndrome and possibly other progressive chronic kidney disorders. BACKGROUND: Dual inhibition of the renin-angiotensin system (RAS) plus sodium-glucose transporter (SGLT)-2 or the mineralocorticoid receptor (MR) demonstrated additive renoprotective effects in large clinical trials. We hypothesized that triple therapy with RAS/SGLT2/MR inhibitors would be superior to dual RAS/SGLT2 blockade in attenuating CKD progression. METHODS: We performed a preclinical randomized controlled trial (PCTE0000266) in Col4a3 -deficient mice with established Alport nephropathy. Treatment was initiated late (age 6 weeks) in mice with elevated serum creatinine and albuminuria and with glomerulosclerosis, interstitial fibrosis, and tubular atrophy. We block-randomized 40 male and 40 female mice to either nil (vehicle) or late-onset food admixes of ramipril monotherapy (10 mg/kg), ramipril plus empagliflozin (30 mg/kg), or ramipril plus empagliflozin plus finerenone (10 mg/kg). Primary end point was mean survival. RESULTS: Mean survival was 63.7±10.0 days (vehicle), 77.3±5.3 days (ramipril), 80.3±11.0 days (dual), and 103.1±20.3 days (triple). Sex did not affect outcome. Histopathology, pathomics, and RNA sequencing revealed that finerenone mainly suppressed the residual interstitial inflammation and fibrosis despite dual RAS/SGLT2 inhibition. CONCLUSION: Experiments in mice suggest that triple RAS/SGLT2/MR blockade may substantially improve renal outcomes in Alport syndrome and possibly other progressive CKDs because of synergistic effects on the glomerular and tubulointerstitial compartments.

A Clinical Workflow for Cost-Saving High-Rate Diagnosis of Genetic Kidney Diseases.

SIGNIFICANCE STATEMENT: To optimize the diagnosis of genetic kidney disorders in a cost-effective manner, we developed a workflow based on referral criteria for in-person evaluation at a tertiary center, whole-exome sequencing, reverse phenotyping, and multidisciplinary board analysis. This workflow reached a diagnostic rate of 67%, with 48% confirming and 19% modifying the suspected clinical diagnosis. We obtained a genetic diagnosis in 64% of children and 70% of adults. A modeled cost analysis demonstrated that early genetic testing saves 20% of costs per patient. Real cost analysis on a representative sample of 66 patients demonstrated an actual cost reduction of 41%. This workflow demonstrates feasibility, performance, and economic effect for the diagnosis of genetic kidney diseases in a real-world setting. BACKGROUND: Whole-exome sequencing (WES) increases the diagnostic rate of genetic kidney disorders, but accessibility, interpretation of results, and costs limit use in daily practice. METHODS: Univariable analysis of a historical cohort of 392 patients who underwent WES for kidney diseases showed that resistance to treatments, familial history of kidney disease, extrarenal involvement, congenital abnormalities of the kidney and urinary tract and CKD stage ≥G2, two or more cysts per kidney on ultrasound, persistent hyperechoic kidneys or nephrocalcinosis on ultrasound, and persistent metabolic abnormalities were most predictive for genetic diagnosis. We prospectively applied these criteria to select patients in a network of nephrology centers, followed by centralized genetic diagnosis by WES, reverse phenotyping, and multidisciplinary board discussion. RESULTS: We applied this multistep workflow to 476 patients with eight clinical categories (podocytopathies, collagenopathies, CKD of unknown origin, tubulopathies, ciliopathies, congenital anomalies of the kidney and urinary tract, syndromic CKD, metabolic kidney disorders), obtaining genetic diagnosis for 319 of 476 patients (67.0%) (95% in 21 patients with disease onset during the fetal period or at birth, 64% in 298 pediatric patients, and 70% in 156 adult patients). The suspected clinical diagnosis was confirmed in 48% of the 476 patients and modified in 19%. A modeled cost analysis showed that application of this workflow saved 20% of costs per patient when performed at the beginning of the diagnostic process. Real cost analysis of 66 patients randomly selected from all categories showed actual cost reduction of 41%. CONCLUSIONS: A diagnostic workflow for genetic kidney diseases that includes WES is cost-saving, especially if implemented early, and is feasible in a real-world setting.

Polyploid tubular cells initiate a TGF-ß1 controlled loop that sustains polyploidization and fibrosis after acute kidney injury.

Polyploidization of tubular cells (TC) is triggered by acute kidney injury (AKI) to allow survival in the early phase after AKI, but in the long run promotes fibrosis and AKI-chronic kidney disease (CKD) transition. The molecular mechanism governing the link between polyploid TC and kidney fibrosis remains to be clarified. In this study, we demonstrate that immediately after AKI, expression of cell cycle markers mostly identifies a population of DNA-damaged polyploid TC. Using transgenic mouse models and single-cell RNA sequencing we show that, unlike diploid TC, polyploid TC accumulate DNA damage and survive, eventually resting in the G1 phase of the cell cycle. In vivo and in vitro single-cell RNA sequencing along with sorting of polyploid TC shows that these cells acquire a profibrotic phenotype culminating in transforming growth factor (TGF)-ß1 expression and that TGF-ß1 directly promotes polyploidization. This demonstrates that TC polyploidization is a self-sustained mechanism. Interactome analysis by single-cell RNA sequencing revealed that TGF-ß1 signaling fosters a reciprocal activation loop among polyploid TC, macrophages, and fibroblasts to sustain kidney fibrosis and promote CKD progression. Collectively, this study contributes to the ongoing revision of the paradigm of kidney tubule response to AKI, supporting the existence of a tubulointerstitial cross talk mediated by TGF-ß1 signaling produced by polyploid TC following DNA damage.NEW & NOTEWORTHY Polyploidization in tubular epithelial cells has been neglected until recently. Here, we showed that polyploidization is a self-sustained mechanism that plays an important role during chronic kidney disease development, proving the existence of a cross talk between infiltrating cells and polyploid tubular cells. This study contributes to the ongoing revision of kidney adaptation to injury, posing polyploid tubular cells at the center of the process.

Progenitor hierarchy among parietal epithelial cells depicted at the single-cell level.

The role of parietal epithelial cells (PECs) in kidney function and disease was recently revisited. Building on previous studies of human kidney tissue, in the current issue, Liu et al. further characterize PECs using single-cell RNA sequencing data and confirm the crucial pathophysiological role of PECs in murine kidney biology as a reservoir for different types of progenitors.

CKD therapy to improve outcomes of immune-mediated glomerular diseases.

The management of immunoglobulin A nephropathy, membranous nephropathy, lupus nephritis, anti-neutrophil cytoplasmic antibody-associated vasculitis, C3 glomerulonephritis, autoimmune podocytopathies and other immune-mediated glomerular disorders is focused on two major treatment goals, preventing overall mortality and the loss of kidney function. Since minimizing irreversible kidney damage best serves both goals, the management of immune-mediated kidney disorders must focus on the two central pathomechanisms of kidney function decline, i.e., controlling the underlying immune disease process (e.g. with immunotherapies) and controlling the non-immune mechanisms of chronic kidney disease (CKD) progression. Here we review the pathophysiology of these non-immune mechanisms of CKD progression and discuss non-drug and drug interventions to attenuate CKD progression in immune-mediated kidney disorders. Non-pharmacological interventions include reducing salt intake, normalizing body weight, avoiding superimposed kidney injuries, smoking cessation and regular physical activity. Approved drug interventions include inhibitors of the renin-angiotensin-aldosterone system and sodium-glucose cotransporter-2. Numerous additional drugs to improve CKD care are currently being tested in clinical trials. Here we discuss how and when to use these drugs in the different clinical scenarios of immune-mediated kidney diseases.

The five types of glomerulonephritis classified by pathogenesis, activity and chronicity (GN-AC).

Glomerulonephritis (GN) is a diverse group of immune-mediated disorders. Currently, GN is classified largely by histological patterns that are difficult to understand and teach, and most importantly, do not indicate treatment choices. Indeed, altered systemic immunity is the primary pathogenic process and the key therapeutic target in GN. Here, we apply a conceptual framework of immune-mediated disorders to GN guided by immunopathogenesis and hence immunophenotyping: (i) infection-related GN require pathogen identification and control; (ii) autoimmunity-related GN, defined by presence of autoantibodies and (iii) alloimmunity-related GN in transplant recipients both require the suppression of adaptive immunity in lymphoid organs and bone marrow; (iv) autoinflammation-related GN, e.g. inborn errors of immunity diagnosed by genetic testing, requires suppression of single cytokine or complement pathways; and (v) Monoclonal gammopathy-related GN requires B or plasma cell clone-directed therapy. A new GN classification should include disease category, immunological activity to tailor the use of the increasing number of immunomodulatory drugs, and chronicity to trigger standard chronic kidney disease care including the evolving spectrum of cardio-renoprotective drugs. Certain biomarkers allow diagnosis and the assessment of immunological activity and disease chronicity without kidney biopsy. The use of these five GN categories and a therapy-focused GN classification is likely to overcome some of the existing hurdles in GN research, management and teaching by reflecting disease pathogenesis and guiding the therapeutic approach.

Intravenous Glu-plasminogen attenuates cholesterol crystal embolism-induced thrombotic angiopathy, acute kidney injury and kidney infarction.

BACKGROUND: Cholesterol crystal (CC) embolism causes acute kidney injury (AKI) and ischaemic cortical necrosis associated with high mortality. We speculated that sustaining the fibrinolytic system with Glu-plasminogen (Glu-Plg) could be a safe way to attenuate AKI and prevent ischaemic infarction upon CC embolism. METHODS: We induced CC embolism by injecting CC into the left kidney artery of C57BL/6J mice. The primary endpoint was glomerular filtration rate (GFR). RESULTS: Starting as early as 2 h after CC embolism, thrombotic angiopathy progressed gradually in the interlobular, arcuate and interlobar arteries. This was associated with a decrease of GFR reaching a peak at 18 h, i.e. AKI, and progressive ischaemic kidney necrosis developing between 12-48 h after CC injection. Human plasma Glu-Plg extracts injected intravenously 4 h after CC embolism attenuated thrombotic angiopathy, GFR loss as well as ischaemic necrosis in a dose-dependent manner. No bleeding complications occurred after Glu-Plg injection. Injection of an intermediate dose (0.6 mg/kg) had only a transient protective effect on microvascular occlusions lasting for a few hours without a sustained protective effect on AKI at 18-48 h or cortical necrosis, while 1.5 mg/kg were fully protective. Importantly, no bleeding complications occurred. CONCLUSIONS: These results provide the first experimental evidence that Glu-Plg could be an innovative therapeutic strategy to attenuate thrombotic angiopathy, AKI, kidney necrosis and potentially other clinical manifestations of CC embolism syndrome.

Role of Sex Hormones in Prevalent Kidney Diseases.

Chronic kidney disease (CKD) is a constantly growing global health burden, with more than 840 million people affected worldwide. CKD presents sex disparities in the pathophysiology of the disease, as well as in the epidemiology, clinical manifestations, and disease progression. Overall, while CKD is more frequent in females, males have a higher risk to progress to end-stage kidney disease. In recent years, numerous studies have highlighted the role of sex hormones in the health and diseases of several organs, including the kidney. In this review, we present a clinical overview of the sex-differences in CKD and a selection of prominent kidney diseases causing CKD: lupus nephritis, diabetic kidney disease, IgA nephropathy, and autosomal dominant polycystic kidney disease. We report clinical and experimental findings on the role of sex hormones in the development of the disease and its progression to end-stage kidney disease.

Polyploid tubular cells and chronic kidney disease.

Defective DNA repair drives chronic kidney disease (CKD), but mechanisms are unclear. Airik and colleagues use a genetic model of defective DNA repair mimicking karyomegalic nephritis, a form of CKD characterized by tubular epithelial cells (TEC) with large nuclei and tubulointerstitial nephritis. They show that DNA damage in TEC triggers endoreplication leading to polyploid TEC and CKD. Blocking endoreplication preserved kidney function, suggesting that DNA damage triggers CKD via TEC polyploidization, questioning the concept of G2/M-arrest.

Type I interferon-related kidney disorders.

Type I interferon (IFN-I) mediates tissue damage in a wide range of kidney disorders, directly affecting the biology and function of several renal cell types including podocytes, mesangial, endothelial, and parietal epithelial cells. Enhanced IFN-I signaling is observed in the context of viral infections, autoimmunity (e.g., systemic lupus erythematosus), and type 1 interferonopathies, rare monogenic disorders characterized by constitutive activation of the IFN-I pathway. All these IFN-I-related disorders can cause renal dysfunction and share pathogenic and histopathological features. Collapsing glomerulopathy, a histopathological lesion characterized by podocyte loss, collapse of the vascular tuft, and parietal epithelial cell proliferation, is commonly associated with viral infections, has been described in type 1 interferonopathies such as Aicardi-Goutières syndrome and stimulator of IFN genes-associated vasculopathy with onset in infancy, and can also be induced by recombinant IFN therapy. In all these conditions, podocytes and parietal epithelial cells seem to be the primary target of IFN-I-mediated damage. Additionally, immune-mediated glomerular injury is common to viral infections, systemic lupus erythematosus, and type 1 interferonopathies such as coatomer subunit-α syndrome (COPA) and DNASE1L3 deficiency, diseases in which IFN-I apparently promotes immune-mediated kidney injury. Finally, kidney pathology primarily characterized by vascular lesions (e.g., thrombotic microangiopathy and vasculitis) is a hallmark of type 1 interferonopathy adenosine deaminase 2 deficiency as well as of systemic lupus erythematosus, viral infections, and IFN therapy. Defining the nosology, pathogenic mechanisms, and histopathological patterns of IFN-I-related kidney disorders has diagnostic and therapeutic implications, especially considering the likely near-term availability of novel drugs targeting the IFN-I pathway.

DNASE1L3 deficiency, new phenotypes, and evidence for a transient type I IFN signaling.

BACKGROUND: Deoxyribonuclease 1 like 3 (DNASE1L3) is a secreted enzyme that has been shown to digest the extracellular chromatin derived from apoptotic bodies, and DNASE1L3 pathogenic variants have been associated with a lupus phenotype. It is unclear whether interferon signaling is sustained in DNASE1L3 deficiency in humans. OBJECTIVES: To explore interferon signaling in DNASE1L3 deficient patients. To depict the characteristic features of DNASE1L3 deficiencies in human. METHODS: We identified, characterized, and analyzed five new patients carrying biallelic DNASE1L3 variations. Whole or targeted exome and/or Sanger sequencing was performed to detect pathogenic variations in five juvenile systemic erythematosus lupus (jSLE) patients. We measured interferon-stimulated gene (ISG) expression in all patients. We performed a systematic review of all published cases available from its first description in 2011 to March 24th 2022. RESULTS: We identified five new patients carrying biallelic DNASE1L3 pathogenic variations, including three previously unreported mutations. Contrary to canonical type I interferonopathies, we noticed a transient increase of ISGs in blood, which returned to normal with disease remission. Disease in one patient was characterized by lupus nephritis and skin lesions, while four others exhibited hypocomplementemic urticarial vasculitis syndrome. The fourth patient presented also with early-onset inflammatory bowel disease. Reviewing previous reports, we identified 35 additional patients with DNASE1L3 deficiency which was associated with a significant risk of lupus nephritis and a poor outcome together with the presence of anti-neutrophil cytoplasmic antibodies (ANCA). Lung lesions were reported in 6/35 patients. CONCLUSIONS: DNASE1L3 deficiencies are associated with a broad phenotype including frequently lupus nephritis and hypocomplementemic urticarial vasculitis with positive ANCA and rarely, alveolar hemorrhages and inflammatory bowel disease. This report shows that interferon production is transient contrary to anomalies of intracellular DNA sensing and signaling observed in Aicardi-Goutières syndrome or STING-associated vasculitis in infancy (SAVI).

A Report of 2 Cases of Kidney Involvement in ADA2 Deficiency: Different Disease Phenotypes and the Tissue Response to Type I Interferon.

Adenosine deaminase 2 (ADA2) deficiency is a rare autosomal recessive disease that is caused by loss-of-function mutations in the ADA2 gene. It is considered a monogenic form of polyarteritis nodosa and frequently is positive for a type I interferon (IFN) signature. Renal manifestations in ADA2 deficiency are poorly characterized. We herein report 2 cases of ADA2 deficiency with different kidney patterns due, respectively, to a predominantly macroscopic and microscopic vasculopathy, and review the literature on kidney disease in ADA2 deficiency. Patient 1 presented with a spontaneous perirenal hematoma; angiography demonstrated multiple microaneurysms but no further defects of the renal parenchyma; his kidney function remained normal. Patient 2 experienced slowly deteriorating kidney function and proteinuria. No major angiographic abnormalities were detected, while kidney biopsy revealed massive vasculopathy resembling chronic thrombotic microangiopathy (TMA) of the small and medium-sized vessels. Both patients had a positive peripheral type I IFN signature. In immunofluorescence staining of a kidney biopsy sample from patient 2, we observed marked expression of the type I IFN-induced protein MXA within endothelial cells, especially in vessels with TMA, and in infiltrating T cells. Our findings confirm that the kidney phenotype of ADA2 deficiency results from small and medium-sized vessel vasculopathy and suggest that type I IFN may be involved in the pathogenesis of kidney lesions.

Crystal Clots as Therapeutic Target in Cholesterol Crystal Embolism.

RATIONALE: Cholesterol crystal embolism can be a life-threatening complication of advanced atherosclerosis. Pathophysiology and molecular targets for treatment are largely unknown. OBJECTIVE: We aimed to develop a new animal model of cholesterol crystal embolism to dissect the molecular mechanisms of cholesterol crystal (CC)-driven arterial occlusion, tissue infarction, and organ failure. METHODS AND RESULTS: C57BL/6J mice were injected with CC into the left kidney artery. Primary end point was glomerular filtration rate (GFR). CC caused crystal clots occluding intrarenal arteries and a dose-dependent drop in GFR, followed by GFR recovery within 4 weeks, that is, acute kidney disease. In contrast, the extent of kidney infarction was more variable. Blocking necroptosis using mixed lineage kinase domain-like deficient mice or necrostatin-1s treatment protected from kidney infarction but not from GFR loss because arterial obstructions persisted, identifying crystal clots as a primary target to prevent organ failure. CC involved platelets, neutrophils, fibrin, and extracellular DNA. Neutrophil depletion or inhibition of the release of neutrophil extracellular traps had little effects, but platelet P2Y12 receptor antagonism with clopidogrel, fibrinolysis with urokinase, or DNA digestion with recombinant DNase I all prevented arterial occlusions, GFR loss, and kidney infarction. The window-of-opportunity was <3 hours after CC injection. However, combining Nec-1s (necrostatin-1s) prophylaxis given 1 hour before and DNase I 3 hours after CC injection completely prevented kidney failure and infarcts. In vitro, CC did not directly induce plasmatic coagulation but induced neutrophil extracellular trap formation and DNA release mainly from kidney endothelial cells, neutrophils, and few from platelets. CC induced ATP release from aggregating platelets, which increased fibrin formation in a DNase-dependent manner. CONCLUSIONS: CC embolism causes arterial obstructions and organ failure via the formation of crystal clots with fibrin, platelets, and extracellular DNA as critical components. Therefore, our model enables to unravel the pathogenesis of the CC embolism syndrome as a basis for both prophylaxis and targeted therapy.

SGLT2 inhibition requires reconsideration of fundamental paradigms in chronic kidney disease, 'diabetic nephropathy', IgA nephropathy and podocytopathies with FSGS lesions.

In 2020, the Dapagliflozin and Prevention of Adverse Outcomes in Chronic Kidney Disease (DAPA-CKD) trial first demonstrated that inhibition of the sodium-glucose transporter-2 (SGLT2) with dapagliflozin attenuates the progression of chronic kidney disease (CKD) with proteinuria in patients with or without diabetes at an unprecedented effect size. These results have far-reaching implications for a series of traditional concepts in Nephrology. It now became obvious that CKD with and without diabetes involves a predominant SGLT2-driven pathophysiology compared with the other pathogenic pathways currently under consideration. As SGLT2 inhibition is similarly efficacious in diabetic and non-diabetic CKD with proteinuria, treating CKD rather than 'diabetic nephropathy' becomes the central paradigm. Indeed, in older adults with type 2 diabetes, CKD is rather of multifactorial origin. As the DAPA-CKD trial included more patients with immunoglobulin A nephropathy (IgAN) than any of the previous IgAN trials, dual renin-angiotensin/SGLT2 inhibition may become the new standard. The same applies for patients with podocytopathy-related focal segmental glomerulosclerosis lesions. From now on, IgAN and podocytopathy trials without SGLT2 inhibition as background therapy and without glomerular filtration rate decline as primary outcome criterion will be of limited value. These and other potential implications will trigger broad discussions and secondary research activities with conclusions difficult to predict today. However, one is for sure: Nephrology after the DAPA-CKD trial will be not the same as it was before. Finally!

Soluble Uric Acid Is an Intrinsic Negative Regulator of Monocyte Activation in Monosodium Urate Crystal-Induced Tissue Inflammation.

Although monosodium urate (MSU) crystals are known to trigger inflammation, published data on soluble uric acid (sUA) in this context are discrepant. We hypothesized that diverse sUA preparation methods account for this discrepancy and that an animal model with clinically relevant levels of asymptomatic hyperuricemia and gouty arthritis can ultimately clarify this issue. To test this, we cultured human monocytes with different sUA preparation solutions and found that solubilizing uric acid (UA) by prewarming created erroneous results because of UA microcrystal contaminants triggering IL-1ß release. Solubilizing UA with NaOH avoided this artifact, and this microcrystal-free preparation suppressed LPS- or MSU crystal-induced monocyte activation, a process depending on the intracellular uptake of sUA via the urate transporter SLC2A9/GLUT9. CD14+ monocytes isolated from hyperuricemic patients were less responsive to inflammatory stimuli compared with monocytes from healthy individuals. Treatment with plasma from hyperuricemic patients impaired the inflammatory function of CD14+ monocytes, an effect fully reversible by removing sUA from hyperuricemic plasma. Moreover, Alb-creERT2;Glut9 lox/lox mice with hyperuricemia (serum UA of 9-11 mg/dl) showed a suppressed inflammatory response to MSU crystals compared with Glut9 lox/lox controls without hyperuricemia. Taken together, we unravel a technical explanation for discrepancies in the published literature on immune effects of sUA and identify hyperuricemia as an intrinsic suppressor of innate immunity, in which sUA modulates the capacity of monocytes to respond to danger signals. Thus, sUA is not only a substrate for the formation of MSU crystals but also an intrinsic inhibitor of MSU crystal-induced tissue inflammation.

Expectations in children with glomerular diseases from SGLT2 inhibitors.

Chronic kidney disease (CKD) is a global public healthcare concern in the pediatric population, where glomerulopathies represent the second most common cause. Although classification and diagnosis of glomerulopathies still rely mostly on histopathological patterns, patient stratification should complement information supplied by kidney biopsy with clinical data and etiological criteria. Genetic determinants of glomerular injury are particularly relevant in children, with important implications for prognosis and treatment. Targeted therapies addressing the primary cause of the disease are available for a limited number of glomerular diseases. Consequently, in the majority of cases, the treatment of glomerulopathies is actually the treatment of CKD. The efficacy of the currently available strategies is limited, but new prospects evolve. Although the exact mechanisms of action are still under investigation, accumulating data in adults demonstrate the efficacy of sodium-glucose transporter 2 inhibitors (SGLT2i) in slowing the progression of CKD due to diabetic and non-diabetic kidney disease. SGLT2i has proved effective on other comorbidities, such as obesity, glycemic control, and cardiovascular risk that frequently accompany CKD. The use of SGLT2i is not yet approved in children. However, no pathophysiological clues theoretically exclude their application. The hallmark of pediatric CKD is the inevitable imbalance between the metabolic needs of a growing child and the functional capacity of a failing kidney to handle those needs. In this view, developing better strategies to address any modifiable progressor in kidney disease is mandatory, especially considering the long lifespan typical of the pediatric population. By improving the hemodynamic adaptation of the kidney and providing additional beneficial effects on the overall complications of CKD, SGLT2i is a candidate as a potentially innovative drug for the treatment of CKD and glomerular diseases in children.