Oxysterol-binding protein like 7 deficiency leads to ER stress mediated apoptosis in podocytes and proteinuria.

Chronic kidney disease (CKD) is associated with renal lipid dysmetabolism among a variety of other pathways. We recently demonstrated that oxysterol-binding protein like 7 (OSBPL7) modulates the expression and function of ATP Binding Cassette Subfamily A Member 1 (ABCA1) in podocytes, a specialized type of cell essential for kidney filtration. Drugs that target OSBPL7 lead to improved renal outcomes in several experimental models of CKD. However, the role of OSBPL7 in podocyte injury remains unclear. Employing mouse models and cellular assays, we investigated the influence of OSBPL7 deficiency on podocytes. We demonstrated that reduced renal OSBPL7 levels as observed in two different models of experimental CKD are linked to increased podocyte apoptosis, primarily mediated by heightened endoplasmic reticulum (ER) stress. While as expected the absence of OSBPL7 also resulted in lipid dysregulation (increased lipid droplets and triglycerides content), OSBPL7-deficiency related lipid dysmetabolism did not contribute to podocyte injury. Similarly, we demonstrated that the decreased autophagic flux we observed in OSBPL7-deficient podocytes was not the mechanistic link between OSBPL7-deficiency and apoptosis. In a complementary zebrafish model, osbpl7 knockdown was sufficient to induce proteinuria and morphological damage to the glomerulus, underscoring its physiological relevance. Our study shed new light on the mechanistic link between OSBPL7 deficiency and podocyte injury in glomerular diseases associated with CKD, and it strengthen the role of OSBPL7 as a novel therapeutic target.

APOL1 risk variants affect podocyte lipid homeostasis and energy production in focal segmental glomerulosclerosis.

Lipotoxicity was recently reported in several forms of kidney disease, including focal segmental glomerulosclerosis (FSGS). Susceptibility to FSGS in African Americans is associated with the presence of genetic variants of the Apolipoprotein L1 gene (APOL1) named G1 and G2. If and how endogenous APOL1 may alter mitochondrial function by the modifying cellular lipid metabolism is unknown. Using transgenic mice expressing the APOL1 variants (G0, G1 or G2) under endogenous promoter, we show that APOL1 risk variant expression in transgenic mice does not impair kidney function at baseline. However, APOL1 G1 expression worsens proteinuria and kidney function in mice characterized by the podocyte inducible expression of nuclear factor of activated T-cells (NFAT), which we have found to cause FSGS. APOL1 G1 expression in this FSGS-model also results in increased triglyceride and cholesterol ester contents in kidney cortices, where lipid accumulation correlated with loss of renal function. In vitro, we show that the expression of endogenous APOL1 G1/G2 in human urinary podocytes is associated with increased cellular triglyceride content and is accompanied by mitochondrial dysfunction in the presence of compensatory oxidative phosphorylation (OXPHOS) complexes elevation. Our findings indicate that APOL1 risk variant expression increases the susceptibility to lipid-dependent podocyte injury, ultimately leading to mitochondrial dysfunction.

Benefit of B7-1 staining and abatacept for treatment-resistant post-transplant focal segmental glomerulosclerosis in a predominantly pediatric cohort: time for a reappraisal.

BACKGROUND: Primary FSGS manifests with nephrotic syndrome and may recur following KT. Failure to respond to conventional therapy after recurrence results in poor outcomes. Evaluation of podocyte B7-1 expression and treatment with abatacept (a B7-1 antagonist) has shown promise but remains controversial. METHODS: From 2012 to 2020, twelve patients developed post-KT FSGS with nephrotic range proteinuria, failed conventional therapy, and were treated with abatacept. Nine/twelve (< 21 years old) experienced recurrent FSGS; three adults developed de novo FSGS, occurring from immediately, up to 8 years after KT. KT biopsies were stained for B7-1. RESULTS: Nine KTRs (75%) responded to abatacept. Seven of nine KTRs were B7-1 positive and responded with improvement/resolution of proteinuria. Two patients with rFSGS without biopsies resolved proteinuria after abatacept. Pre-treatment UPCR was 27.0 ± 20.4 (median 13, range 8-56); follow-up UPCR was 0.8 ± 1.3 (median 0.2, range 0.07-3.9, p < 0.004). Two patients who were B7-1 negative on multiple KT biopsies did not respond to abatacept and lost graft function. One patient developed proteinuria while receiving belatacept, stained B7-1 positive, but did not respond to abatacept. CONCLUSIONS: Podocyte B7-1 staining in biopsies of KTRs with post-transplant FSGS identifies a subset of patients who may benefit from abatacept. A higher resolution version of the Graphical abstract is available as Supplementary information.

Activation of Stimulator of IFN Genes (STING) Causes Proteinuria and Contributes to Glomerular Diseases.

BACKGROUND: The signaling molecule stimulator of IFN genes (STING) was identified as a crucial regulator of the DNA-sensing cyclic GMP-AMP synthase (cGAS)-STING pathway, and this signaling pathway regulates inflammation and energy homeostasis under conditions of obesity, kidney fibrosis, and AKI. However, the role of STING in causing CKD, including diabetic kidney disease (DKD) and Alport syndrome, is unknown. METHODS: To investigate whether STING activation contributes to the development and progression of glomerular diseases such as DKD and Alport syndrome, immortalized human and murine podocytes were differentiated for 14 days and treated with a STING-specific agonist. We used diabetic db/db mice, mice with experimental Alport syndrome, C57BL/6 mice, and STING knockout mice to assess the role of the STING signaling pathway in kidney failure. RESULTS: In vitro, murine and human podocytes express all of the components of the cGAS-STING pathway. In vivo, activation of STING renders C57BL/6 mice susceptible to albuminuria and podocyte loss. STING is activated at baseline in mice with experimental DKD and Alport syndrome. STING activation occurs in the glomerular but not the tubulointerstitial compartment in association with autophagic podocyte death in Alport syndrome mice and with apoptotic podocyte death in DKD mouse models. Genetic or pharmacologic inhibition of STING protects from progression of kidney disease in mice with DKD and Alport syndrome and increases lifespan in Alport syndrome mice. CONCLUSION: The activation of the STING pathway acts as a mediator of disease progression in DKD and Alport syndrome. Targeting STING may offer a therapeutic option to treat glomerular diseases of metabolic and nonmetabolic origin or prevent their development, progression, or both.

Sphingosine-1-Phosphate Metabolism and Signaling in Kidney Diseases.

In the past few decades, sphingolipids and sphingolipid metabolites have gained attention because of their essential role in the pathogenesis and progression of kidney diseases. Studies in models of experimental and clinical nephropathies have described accumulation of sphingolipids and sphingolipid metabolites, and it has become clear that the intracellular sphingolipid composition of renal cells is an important determinant of renal function. Proper function of the glomerular filtration barrier depends heavily on the integrity of lipid rafts, which include sphingolipids as key components. In addition to contributing to the structural integrity of membranes, sphingolipid metabolites, such as sphingosine-1-phosphate (S1P), play important roles as second messengers regulating biologic processes, such as cell growth, differentiation, migration, and apoptosis. This review will focus on the role of S1P in renal cells and how aberrant extracellular and intracellular S1P signaling contributes to the pathogenesis and progression of kidney diseases.

Sterol-O-acyltransferase-1 has a role in kidney disease associated with diabetes and Alport syndrome.

Defective cholesterol metabolism primarily linked to reduced ATP-binding cassette transporter A1 (ABCA1) expression is closely associated with the pathogenesis and progression of kidney diseases, including diabetic kidney disease and Alport Syndrome. However, whether the accumulation of free or esterified cholesterol contributes to progression in kidney disease remains unclear. Here, we demonstrate that inhibition of sterol-O-acyltransferase-1 (SOAT1), the enzyme at the endoplasmic reticulum that converts free cholesterol to cholesterol esters, which are then stored in lipid droplets, effectively reduced cholesterol ester and lipid droplet formation in human podocytes. Furthermore, we found that inhibition of SOAT1 in podocytes reduced lipotoxicity-mediated podocyte injury in diabetic kidney disease and Alport Syndrome in association with increased ABCA1 expression and ABCA1-mediated cholesterol efflux. In vivo, Soat1 deficient mice did not develop albuminuria or mesangial expansion at 10-12 months of age. However, Soat1 deficiency/inhibition in experimental models of diabetic kidney disease and Alport Syndrome reduced cholesterol ester content in kidney cortices and protected from disease progression. Thus, targeting SOAT1-mediated cholesterol metabolism may represent a new therapeutic strategy to treat kidney disease in patients with diabetic kidney disease and Alport Syndrome, like that suggested for Alzheimer's disease and cancer treatments.

Lipid mediators of insulin signaling in diabetic kidney disease.

Diabetic kidney disease (DKD) affects ∼40% of patients with diabetes and is associated with high mortality rates. Among different cellular targets in DKD, podocytes, highly specialized epithelial cells of the glomerular filtration barrier, are injured in the early stages of DKD. Both clinical and experimental data support the role of preserved insulin signaling as a major contributor to podocyte function and survival. However, little is known about the key modulators of podocyte insulin signaling. This review summarizes the novel knowledge that intracellular lipids such as cholesterol and sphingolipids are major determinants of podocyte insulin signaling. In particular, the implications of these lipids on DKD development, progression, and treatment will be addressed.

Crosstalk Between Lipids and Mitochondria in Diabetic Kidney Disease.

PURPOSE OF REVIEW: The goal of this review is to review the role that renal parenchymal lipid accumulation plays in contributing to diabetic kidney disease (DKD), specifically contributing to the mitochondrial dysfunction observed in glomerular renal cells in the context of DKD development and progression. RECENT FINDINGS: Mitochondrial dysfunction has been observed in experimental and clinical DKD. Recently, Ayanga et al. demonstrate that podocyte-specific deletion of a protein involved in mitochondrial dynamics protects from DKD progression. Furthermore, our group has recently shown that ATP-binding cassette A1 (a protein involved in cholesterol and phospholipid efflux) is significantly reduced in clinical and experimental DKD and that genetic or pharmacological induction of ABCA1 is sufficient to protect from DKD. ABCA1 deficiency in podocytes leads to mitochondrial dysfunction observed with alterations of mitochondrial lipids, in particular, cardiolipin (a mitochondrial-specific phospholipid). However, through pharmacological reduction of cardiolipin peroxidation DKD progression is reverted. Lipid metabolism is significantly altered in the diabetic kidney and renders cellular components, such as the podocyte, susceptible to injury leading to worsened DKD progression. Dysfunction of the lipid metabolism pathway can also lead to mitochondrial dysfunction and mitochondrial lipid alteration. Future research aimed at targeting mitochondrial lipids content and function could prove to be beneficial for the treatment of DKD.

Hydroxypropyl-ß-cyclodextrin protects from kidney disease in experimental Alport syndrome and focal segmental glomerulosclerosis.

Studies suggest that altered renal lipid metabolism plays a role in the pathogenesis of diabetic kidney disease and that genetic or pharmacological induction of cholesterol efflux protects from the development of diabetic kidney disease and focal segmental glomerulosclerosis (FSGS). Here we tested whether altered lipid metabolism contributes to renal failure in the Col4a3 knockout mouse model for Alport Syndrome. There was an eight-fold increase in the cholesterol content in renal cortexes of mice with Alport Syndrome. This was associated with increased glomerular lipid droplets and cholesterol crystals. Treatment of mice with Alport Syndrome with hydroxypropyl-ß-cyclodextrin (HPßCD) reduced cholesterol content in the kidneys of mice with Alport Syndrome and protected from the development of albuminuria, renal failure, inflammation and tubulointerstitial fibrosis. Cholesterol efflux and trafficking-related genes were primarily affected in mice with Alport Syndrome and were differentially regulated in the kidney cortex and isolated glomeruli. HPßCD also protected from proteinuria and mesangial expansion in a second model of non-metabolic kidney disease, adriamycin-induced nephropathy. Consistent with our experimental findings, microarray analysis confirmed dysregulation of several lipid-related genes in glomeruli isolated from kidney biopsies of patients with primary FSGS enrolled in the NEPTUNE study. Thus, lipid dysmetabolism occurs in non-metabolic glomerular disorders such as Alport Syndrome and FSGS, and HPßCD improves renal function in experimental Alport Syndrome and FSGS.

Sphingomyelinase-like phosphodiesterase 3b mediates radiation-induced damage of renal podocytes.

The molecular mechanisms responsible for the development of proteinuria and glomerulosclerosis in radiation nephropathy remain largely unknown. Podocytes are increasingly recognized as key players in the pathogenesis of proteinuria in primary and secondary glomerular disorders. The lipid-modulating enzyme sphingomyelin phosphodiesterase acid-like 3B (SMPDL3b) is a key determinant of podocyte injury and a known off target of the anti-CD20 antibody rituximab (RTX). The current study investigates the role of sphingolipids in radiation-induced podocytopathy. After a single dose of radiation (8 Gy), several ceramide species were significantly elevated. In particular, C16:00, C24:00, and C24:1 ceramides were the most abundant ceramide species detected. These changes were paralleled by a time-dependent drop in SMPDL3b protein, sphingosine, and sphingosine-1-phosphate levels. Interestingly, SMPDL3b-overexpressing podocytes had higher basal levels of sphingosine-1-phosphate and maintained basal ceramide levels after irradiation. Morphologically, irradiated podocytes demonstrated loss of filopodia and remodeling of cortical actin. Furthermore, the actin binding protein ezrin relocated from the plasma membrane to the cytosol as early as 2 h after radiation. In contrast, SMPDL3b overexpressing podocytes were protected from radiation-induced cytoskeletal remodeling. Treatment with RTX before radiation exposure partially protected podocytes from SMPDL3b loss, cytoskeletal remodeling, and caspase 3 cleavage. Our results demonstrate that radiation injury induces early cytoskeletal remodeling, down-regulation of SMPDL3b, and elevation of cellular ceramide levels. Overexpression of SMPDL3b and pretreatment with RTX confer a radioprotective effect in cultured podocytes. These findings indicate a potential role for SMPDL3b and RTX in radiation-induced podocytopathy.-Ahmad, A., Mitrofanova, A., Bielawski, J., Yang, Y., Marples, B., Fornoni, A., Zeidan, Y. H. Sphingomyelinase-like phosphodiesterase 3b mediates radiation-induced damage of renal podocytes.

Genetic determination of the vascular reactions in humans in response to the diving reflex.

The purpose of this study was to investigate the genetic mechanisms of the defense vascular reactions in response to the diving reflex in humans with polymorphisms in the genes ADBR2, ACE, AGTR1, BDKRB2, and REN We hypothesized that protective vascular reactions, in response to the diving reflex, are genetically determined and are distinguished in humans with gene polymorphisms of the renin-angiotensin and kinin-bradykinin system. A total of 80 subjects (19 ± 1.4 yr) participated in the study. The intensity of the vascular response was estimated using photoplethysmogram. The I/D polymorphism (rs4340) of ACE was analyzed by PCR. REN (G/A, rs2368564), AGTR1 (A/C, rs5186), BDKRB2 (T/C, rs1799722), and ADBR2 (A/G, rs1042713) polymorphisms were examined using the two-step multiplex PCR followed by carrying allele hybridization on the biochip. Subjects with the BDKRB2 (C/C), ACE (D/D), and ADBR2 (G/G, G/A) genotypes exhibited the strongest peripheral vasoconstriction in response to diving. In subjects with a combination of the BDKRB2 (C/C) plus ACE (D/D) genotypes, we observed the lowest pulse wave amplitude and pulse transit time values and the highest arterial blood pressure during face immersion compared with the heterozygous individuals, suggesting that these subjects are more susceptible to diving hypoxia. This study observed that humans with gene polymorphisms of the renin-angiotensin and kinin-bradykinin systems demonstrate various expressions of protective vascular reactions in response to the diving reflex. The obtained results might be used in estimation of resistance to hypoxia of any origin in human beings or in a medical practice.NEW & NOTEWORTHY Our study demonstrates that the vascular reactions in response to the diving reflex are genetically determined and depend on gene polymorphisms of the kinin-bradykinin and the renin-angiotensin systems.

Nephrin Contributes to Insulin Secretion and Affects Mammalian Target of Rapamycin Signaling Independently of Insulin Receptor.

Nephrin belongs to a family of highly conserved proteins with a well characterized function as modulators of cell adhesion and guidance, and nephrin may have a role in metabolic pathways linked to podocyte and pancreatic ß-cell survival. However, this role is incompletely characterized. In this study, we developed floxed nephrin mice for pancreatic ß-cell-specific deletion of nephrin, which had no effect on islet size and glycemia. Nephrin deficiency, however, resulted in glucose intolerance in vivo and impaired glucose-stimulated insulin release ex vivo Glucose intolerance was also observed in eight patients with nephrin mutations compared with three patients with other genetic forms of nephrotic syndrome or nine healthy controls.In vitro experiments were conducted to investigate if nephrin affects autocrine signaling through insulin receptor A (IRA) and B (IRB), which are both expressed in human podocytes and pancreatic islets. Coimmunoprecipitation of nephrin and IRB but not IRA was observed and required IR phosphorylation. Nephrin per se was sufficient to induce phosphorylation of p70S6K in an phosphatidylinositol 3-kinase-dependent but IR/Src-independent manner, which was not augmented by exogenous insulin. These results suggest a role for nephrin as an independent modulator of podocyte and pancreatic ß-cell nutrient sensing in the fasting state and the potential of nephrin as a drug target in diabetes.

Sphingomyelinase-like phosphodiesterase 3b expression levels determine podocyte injury phenotypes in glomerular disease.

Diabetic kidney disease (DKD) is the most common cause of ESRD in the United States. Podocyte injury is an important feature of DKD that is likely to be caused by circulating factors other than glucose. Soluble urokinase plasminogen activator receptor (suPAR) is a circulating factor found to be elevated in the serum of patients with FSGS and causes podocyte αVß3 integrin-dependent migration in vitro. Furthermore, αVß3 integrin activation occurs in association with decreased podocyte-specific expression of acid sphingomyelinase-like phosphodiesterase 3b (SMPDL3b) in kidney biopsy specimens from patients with FSGS. However, whether suPAR-dependent αVß3 integrin activation occurs in diseases other than FSGS and whether there is a direct link between circulating suPAR levels and SMPDL3b expression in podocytes remain to be established. Our data indicate that serum suPAR levels are also elevated in patients with DKD. However, unlike in FSGS, SMPDL3b expression was increased in glomeruli from patients with DKD and DKD sera-treated human podocytes, where it prevented αVß3 integrin activation by its interaction with suPAR and led to increased RhoA activity, rendering podocytes more susceptible to apoptosis. In vivo, inhibition of acid sphingomyelinase reduced proteinuria in experimental DKD but not FSGS, indicating that SMPDL3b expression levels determined the podocyte injury phenotype. These observations suggest that SMPDL3b may be an important modulator of podocyte function by shifting suPAR-mediated podocyte injury from a migratory phenotype to an apoptotic phenotype and that it represents a novel therapeutic glomerular disease target.

Structural determinants of the insulin receptor-related receptor activation by alkali.

IRR is a member of the insulin receptor (IR) family that does not have any known agonist of a peptide nature but can be activated by mildly alkaline medium and was thus proposed to function as an extracellular pH sensor. IRR activation by alkali is defined by its N-terminal extracellular region. To reveal key structural elements involved in alkali sensing, we developed an in vitro method to quantify activity of IRR and its mutants. Replacing the IRR L1C domains (residues 1-333) or L2 domain (residues 334-462) or both with the homologous fragments of IR reduced the receptor activity to 35, 64, and 7% percent, respectively. Within L1C domains, five amino acid residues (Leu-135, Gly-188, Arg-244, and vicinal His-318 and Lys-319) were identified as IRR-specific by species conservation analysis of the IR family. These residues are exposed and located in junctions between secondary structure folds. The quintuple mutation of these residues to alanine had the same negative effect as the entire L1C domain replacement, whereas none of the single mutations was as effective. Separate mutations of these five residues and of L2 produced partial negative effects that were additive. The pH dependence of cell-expressed mutants (L1C and L2 swap, L2 plus triple LGR mutation, and L2 plus quintuple LGRHK mutation) was shifted toward alkalinity and, in contrast with IRR, did not show significant positive cooperativity. Our data suggest that IRR activation is not based on a single residue deprotonation in the IRR ectodomain but rather involves synergistic conformational changes at multiple points.

Role of the Innate Immune Response in Glomerular Disease Pathogenesis: Focus on Podocytes.

Accumulating evidence indicates that inflammatory and immunologic processes play a significant role in the development and progression of glomerular diseases. Podocytes, the terminally differentiated epithelial cells, are crucial for maintaining the integrity of the glomerular filtration barrier. Once injured, podocytes cannot regenerate, leading to progressive proteinuric glomerular diseases. However, emerging evidence suggests that podocytes not only maintain the glomerular filtration barrier and are important targets of immune responses but also exhibit many features of immune-like cells, where they are involved in the modulation of the activity of innate and adaptive immunity. This dual role of podocytes may lead to the discovery and development of new therapeutic targets for treating glomerular diseases. This review aims to provide an overview of the innate immunity mechanisms involved in podocyte injury and the progression of proteinuric glomerular diseases.

Transplantation: platform to study recurrence of disease.

Beyond the direct benefit that a transplanted organ provides to an individual recipient, the study of the transplant process has the potential to create a better understanding of the pathogenesis, etiology, progression and possible therapy for recurrence of disease after transplantation while at the same time providing insight into the original disease. Specific examples of this include: 1) recurrence of focal segmental glomerulosclerosis (FSGS) after kidney transplantation, 2) recurrent autoimmunity after pancreas transplantation, and 3) recurrence of disease after orthotopic liver transplantation (OLT) for cirrhosis related to progressive steatosis secondary to jejuno-ileal bypass (JIB) surgery. Our team has been studying these phenomena and their immunologic underpinnings, and we suggest that expanding the concept to other pathologic processes and/or transplanted organs that harbor the risk for recurrent disease may provide novel insight into the pathogenesis of a host of other disease processes that lead to organ failure.

Kidney lipid dysmetabolism and lipid droplet accumulation in chronic kidney disease.

Chronic kidney disease (CKD) is a global health problem with rising incidence and prevalence. Among several pathogenetic mechanisms responsible for disease progression, lipid accumulation in the kidney parenchyma might drive inflammation and fibrosis, as has been described in fatty liver diseases. Lipids and their metabolites have several important structural and functional roles, as they are constituents of cell and organelle membranes, serve as signalling molecules and are used for energy production. However, although lipids can be stored in lipid droplets to maintain lipid homeostasis, lipid accumulation can become pathogenic. Understanding the mechanisms linking kidney parenchymal lipid accumulation to CKD of metabolic or non-metabolic origin is challenging, owing to the tremendous variety of lipid species and their functional diversity across different parenchymal cells. Nonetheless, multiple research reports have begun to emphasize the effect of dysregulated kidney lipid metabolism in CKD progression. For example, altered cholesterol and fatty acid metabolism contribute to glomerular and tubular cell injury. Newly developed lipid-targeting agents are being tested in clinical trials in CKD, raising expectations for further therapeutic development in this field.

Management of Lupus Nephritis: New Treatments and Updated Guidelines.

Management of lupus nephritis has evolved considerably over the past years. Here, we provide a comprehensive review of clinical trials that form the basis for the Kidney Disease: Improving Global Outcomes and EULAR/ERA-EDTA updated guidelines and present day trials that will change the landscape of lupus nephritis therapy in years to come. In addition, we highlight the issues related to cost of therapy, resistant disease, and downstream adverse effects of specific therapies.

The podocyte: glomerular sentinel at the crossroads of innate and adaptive immunity.

Focal segmental glomerulosclerosis (FSGS) is a common glomerular disorder that manifests clinically with the nephrotic syndrome and has a propensity to recur following kidney transplantation. The pathophysiology and therapies available to treat FSGS currently remain elusive. Since the podocyte appears to be the target of apparent circulating factor(s) that lead to recurrence of proteinuria following kidney transplantation, this article is focused on the podocyte. In the context of kidney transplantation, the performance of pre- and post-reperfusion biopsies, and the establishment of in vitro podocyte liquid biopsies/assays allow for the development of clinically relevant studies of podocyte biology. This has given insight into new pathways, involving novel targets in innate and adaptive immunity, such as SMPDL3b, cGAS-STING, and B7-1. Elegant experimental studies suggest that the successful clinical use of rituximab and abatacept, two immunomodulating agents, in our case series, may be due to direct effects on the podocyte, in addition to, or perhaps distinct from their immunosuppressive functions. Thus, tissue biomarker-directed therapy may provide a rational approach to validate the mechanism of disease and allow for the development of new therapeutics for FSGS. This report highlights recent progress in the field and emphasizes the importance of kidney transplantation and recurrent FSGS (rFSGS) as a platform for the study of primary FSGS.

ABCA1 deficiency contributes to podocyte pyroptosis priming via the APE1/IRF1 axis in diabetic kidney disease.

Decreased ATP Binding Cassette Transporter A1 (ABCA1) expression and caspase-4-mediated noncanonical inflammasome contribution have been described in podocytes in diabetic kidney disease (DKD). To investigate a link between these pathways, we evaluated pyroptosis-related mediators in human podocytes with stable knockdown of ABCA1 (siABCA1) and found that mRNA levels of IRF1, caspase-4, GSDMD, caspase-1 and IL1ß were significantly increased in siABCA1 compared to control podocytes and that protein levels of caspase-4, GSDMD and IL1ß were equally increased. IRF1 knockdown in siABCA1 podocytes prevented increases in caspase-4, GSDMD and IL1ß. Whereas TLR4 inhibition did not decrease mRNA levels of IRF1 and caspase-4, APE1 protein expression increased in siABCA1 podocytes and an APE1 redox inhibitor abrogated siABCA1-induced expression of IRF1 and caspase-4. RELA knockdown also offset the pyroptosis priming, but ChIP did not demonstrate increased binding of NFκB to IRF1 promoter in siABCA1 podocytes. Finally, the APE1/IRF1/Casp1 axis was investigated in vivo. APE1 IF staining and mRNA levels of IRF1 and caspase 11 were increased in glomeruli of BTBR ob/ob compared to wildtype. In conclusion, ABCA1 deficiency in podocytes caused APE1 accumulation, which reduces transcription factors to increase the expression of IRF1 and IRF1 target inflammasome-related genes, leading to pyroptosispriming.