TACI haploinsufficiency protects against BAFF-driven humoral autoimmunity in mice.

Polymorphisms in TACI, a BAFF family cytokine receptor, are linked to diverse human immune disorders including common variable immunodeficiency (CVID) and systemic lupus erythematosus (SLE). Functional studies of individual variants show modest impacts on surface TACI expression and/or downstream signal transduction, indicating that relatively subtle variation in TACI activity can impact human B-cell biology. However, significant complexity underlies TACI biology, including both positive and negative regulation of physiologic and pathogenic B-cell responses. To model these contradictory events, we compared the functional impact of TACI deletion on separate models of murine SLE driven by T cell-independent and -dependent breaks in B-cell tolerance. First, we studied whether reduced surface TACI expression was sufficient to protect against progressive BAFF-mediated systemic autoimmunity. Strikingly, despite a relatively modest impact on surface TACI levels, TACI haploinsufficiency markedly reduced pathogenic RNA-associated autoantibody titers and conferred long-term protection from BAFF-driven lupus nephritis. In contrast, B cell-intrinsic TACI deletion exerted a limited impact of autoantibody generation in murine lupus characterized by spontaneous germinal center formation and T cell-dependent humoral autoimmunity. Together, these combined data provide new insights into TACI biology and highlight how TACI signals must be tightly regulated during protective and pathogenic B-cell responses.

Regression of diabetic nephropathy by treatment with empagliflozin in BTBR ob/ob mice.

BACKGROUND: The sodium-glucose cotransporter-2 (SGLT2) inhibitor empagliflozin lowers blood glucose via reduced tubular reabsorption of filtered glucose and is an important new therapy for diabetic nephropathy (DN). This study tested whether treatment with empagliflozin would ameliorate proteinuria and the pathologic alterations of DN including podocyte number and integrity in the leptin-deficient BTBR ob/ob mouse model of DN. METHODS: Study cohorts included wild-type (WT) BTBR mice, untreated diabetic BTBR ob/ob mice and mice treated with empagliflozin for 6 weeks after development of established DN at 18 weeks of age. RESULTS: Hyperglycemia, proteinuria, serum creatinine, accumulation of mesangial matrix and the extent of mesangiolysis were reversed with empagliflozin treatment. Treatment with empagliflozin resulted in an increased podocyte number and podocyte density, improvement in the degree of podocyte foot process effacement and parietal epithelial cell activation. SGLT2 inhibition reduced renal oxidative stress, measured by urinary excretion of markers of RNA/DNA damage and in situ demonstration of decreased carbonyl oxidation. There was no discernable difference in accumulations of advanced glycation end-products by immunohistochemistry. CONCLUSION: The structural improvements seen in BTBR ob/ob mice treated with empagliflozin provide insights into potential long-term benefits for humans with DN, for whom there is no comparable biopsy information to identify structural changes effected by SGLT2 inhibition. The findings suggest SGLT2 inhibition may ameliorate DN through glucose lowering-dependent and -independent mechanisms that lead to podocyte restoration and delay or reversal of disease progress.

Complement Deficiencies Result in Surrogate Pathways of Complement Activation in Novel Polygenic Lupus-like Models of Kidney Injury.

Lupus nephritis (LN) is a major contributor to morbidity and mortality in lupus patients, but the mechanisms of kidney damage remain unclear. In this study, we introduce, to our knowledge, novel models of LN designed to resemble the polygenic nature of human lupus by embodying three key genetic alterations: the Sle1 interval leading to anti-chromatin autoantibodies; Mfge8-/- , leading to defective clearance of apoptotic cells; and either C1q-/- or C3-/- , leading to low complement levels. We report that proliferative glomerulonephritis arose only in the presence of all three abnormalities (i.e., in Sle1.Mfge8 -/- C1q -/- and Sle1.Mfge8 -/- C3 -/- triple-mutant [TM] strains [C1q -/-TM and C3-/- TM, respectively]), with structural kidney changes resembling those in LN patients. Unexpectedly, both TM strains had significant increases in autoantibody titers, Ag spread, and IgG deposition in the kidneys. Despite the early complement component deficiencies, we observed assembly of the pathogenic terminal complement membrane attack complex in both TM strains. In C1q-/- TM mice, colocalization of MASP-2 and C3 in both the glomeruli and tubules indicated that the lectin pathway likely contributed to complement activation and tissue injury in this strain. Interestingly, enhanced thrombin activation in C3-/- TM mice and reduction of kidney injury following attenuation of thrombin generation by argatroban in a serum-transfer nephrotoxic model identified thrombin as a surrogate pathway for complement activation in C3-deficient mice. These novel mouse models of human lupus inform the requirements for nephritis and provide targets for intervention.

Reversal of hypertriglyceridemia in diabetic BTBR ob/ob mice does not prevent nephropathy.

The etiology of diabetic nephropathy in type 2 diabetes is multifactorial. Sustained hyperglycemia is a major contributor, but additional contributions come from the hypertension, obesity, and hyperlipidemia that are also commonly present in patients with type 2 diabetes and nephropathy. The leptin deficient BTBR ob/ob mouse is a model of type 2 diabetic nephropathy in which hyperglycemia, obesity, and hyperlipidemia, but not hypertension, are present. We have shown that reversal of the constellation of these metabolic abnormalities with leptin replacement can reverse the morphologic and functional manifestations of diabetic nephropathy. Here we tested the hypothesis that reversal specifically of the hypertriglyceridemia, using an antisense oligonucleotide directed against ApoC-III, an apolipoprotein that regulates the interactions of VLDL (very low density lipoproteins) with the LDL receptor, is sufficient to ameliorate the nephropathy of Type 2 diabetes. Antisense treatment resulted in reduction of circulating ApoC-III protein levels and resulted in substantial lowering of triglycerides to near-normal levels in diabetic mice versus controls. Antisense treatment did not ameliorate proteinuria or pathologic manifestations of diabetic nephropathy, including podocyte loss. These studies indicate that pathologic manifestations of diabetic nephropathy are unlikely to be reduced by lipid-lowering therapeutics alone, but does not preclude a role for such interventions to be used in conjunction with other therapeutics commonly employed in the treatment of diabetes and its complications.

Non-invasive metabolic biomarkers for early diagnosis of diabetic nephropathy: Meta-analysis of profiling metabolomics studies.

AIM: Diabetic nephropathy (DN) is one of the worst complications of diabetes. Despite a growing number of DN metabolite profiling studies, most studies are suffering from inconsistency in their findings. The main goal of this meta-analysis was to reach to a consensus panel of significantly dysregulated metabolites as potential biomarkers in DN. DATA SYNTHESIS: To identify the significant dysregulated metabolites, meta-analysis was performed by "vote-counting rank" and "robust rank aggregation" strategies. Bioinformatics analyses were performed to identify the most affected genes and pathways. Among 44 selected studies consisting of 98 metabolite profiles, 17 metabolites (9 up-regulated and 8 down-regulated metabolites), were identified as significant ones by both the meta-analysis strategies (p-value<0.05 and OR>2 or <0.5) and selected as DN metabolite meta-signature. Furthermore, enrichment analyses confirmed the involvement of various effective biological pathways in DN pathogenesis, such as urea cycle, TCA cycle, glycolysis, and amino acid metabolisms. Finally, by performing a meta-analysis over existing time-course studies in DN, the results indicated that lactic acid, hippuric acid, allantoin (in urine), and glutamine (in blood), are the topmost non-invasive early diagnostic biomarkers. CONCLUSION: The identified metabolites are potentially involved in diabetic nephropathy pathogenesis and could be considered as biomarkers or drug targets in the disease. PROSPERO REGISTRATION NUMBER: CRD42020197697.

Beneficial effect on podocyte number in experimental diabetic nephropathy resulting from combined atrasentan and RAAS inhibition therapy.

Podocyte loss and proteinuria are both key features of human diabetic nephropathy (DN). The leptin-deficient BTBR mouse strain with the ob/ob mutation develops progressive weight gain, type 2 diabetes, and diabetic nephropathy that has many features of advanced human DN, including increased mesangial matrix, mesangiolysis, podocyte loss, and proteinuria. Selective antagonism of the endothelin-1 type A receptor (ETAR) by atrasentan treatment in combination with renin-angiotensin-aldosterone system inhibition with losartan has been shown to have the therapeutic benefit of lowering proteinuria in patients with DN, but the underlying mechanism for this benefit is not well understood. Using a similar therapeutic approach in diabetic BTBR ob/ob mice, this treatment regimen significantly increased glomerular podocyte number compared with diabetic BTBR ob/ob controls and suggested that parietal epithelial cells were a source for podocyte restoration. Atrasentan treatment alone also increased podocyte number but to a lesser degree. Mice treated with atrasentan demonstrated a reduction in proteinuria, matching the functional improvement reported in humans. This is a first demonstration that treatment with the highly selective ETAR antagonist atrasentan can lead to restoration of the diminished podocyte number characteristic of DN in humans and thereby underlies the reduction in proteinuria in patients with diabetes undergoing similar treatment. The benefit of ETAR antagonism in DN extended to a decrease in mesangial matrix as measured by a reduction in accumulations of collagen type IV in both the atrasentan and atrasentan + losartan-treated groups compared with untreated controls.

High-protein diet accelerates diabetes and kidney disease in the BTBRob/ob mouse.

There is a need for improved animal models that better translate to human kidney disease to predict outcome of pharmacological effects in the patient. The diabetic BTBRob/ob mouse model mimics key features of early diabetic nephropathy in humans, but with chronic injury limited to glomeruli. To explore if we could induce an accelerated and more advanced disease phenotype that closer translates to human disease, we challenged BTBRob/ob mice with a high-protein diet (HPD; 30%) and followed the progression of metabolic and renal changes up to 20 wk of age. Animals on the HPD showed enhanced metabolic derangements, evidenced by further increased levels of glucose, HbA1C, cholesterol, and alanine aminotransferase. The urinary albumin-to-creatinine ratio was markedly increased with a 53-fold change compared with lean controls, whereas BTBRob/ob mice on the standard diet only presented an 8-fold change. HPD resulted in more advanced mesangial expansion already at 14 wk of age compared with BTBRob/ob mice on the standard diet and also aggravated glomerular pathology as well as interstitial fibrosis. Gene expression analysis revealed that HPD triggered expression of markers of fibrosis and inflammation in the kidney and increased oxidative stress markers in urine. This study showed that HPD significantly aggravated renal injury in BTBRob/ob mice by further advancing albuminuria, glomerular, and tubulointerstitial pathology by 20 wk of age. This mouse model offers closer translation to humans and enables exploration of new end points for pharmacological efficacy studies that also holds promise to shorten study length.

Integrated Functional Genomic Analysis Enables Annotation of Kidney Genome-Wide Association Study Loci.

BACKGROUND: Linking genetic risk loci identified by genome-wide association studies (GWAS) to their causal genes remains a major challenge. Disease-associated genetic variants are concentrated in regions containing regulatory DNA elements, such as promoters and enhancers. Although researchers have previously published DNA maps of these regulatory regions for kidney tubule cells and glomerular endothelial cells, maps for podocytes and mesangial cells have not been available. METHODS: We generated regulatory DNA maps (DNase-seq) and paired gene expression profiles (RNA-seq) from primary outgrowth cultures of human glomeruli that were composed mainly of podocytes and mesangial cells. We generated similar datasets from renal cortex cultures, to compare with those of the glomerular cultures. Because regulatory DNA elements can act on target genes across large genomic distances, we also generated a chromatin conformation map from freshly isolated human glomeruli. RESULTS: We identified thousands of unique regulatory DNA elements, many located close to transcription factor genes, which the glomerular and cortex samples expressed at different levels. We found that genetic variants associated with kidney diseases (GWAS) and kidney expression quantitative trait loci were enriched in regulatory DNA regions. By combining GWAS, epigenomic, and chromatin conformation data, we functionally annotated 46 kidney disease genes. CONCLUSIONS: We demonstrate a powerful approach to functionally connect kidney disease-/trait-associated loci to their target genes by leveraging unique regulatory DNA maps and integrated epigenomic and genetic analysis. This process can be applied to other kidney cell types and will enhance our understanding of genome regulation and its effects on gene expression in kidney disease.

Gene deletion of the Na+-glucose cotransporter SGLT1 ameliorates kidney recovery in a murine model of acute kidney injury induced by ischemia-reperfusion.

Renal Na+-glucose cotransporter SGLT1 mediates glucose reabsorption in the late proximal tubule, a hypoxia-sensitive tubular segment that enters the outer medulla. Gene deletion in mice (Sglt1-/-) was used to determine the role of the cotransporter in acute kidney injury induced by ischemia-reperfusion (IR), including the initial injury and subsequent recovery phase. On days 1 and 16 after IR, absolute and fractional urinary glucose excretion remained greater in Sglt1-/- mice versus wild-type (WT) littermates, consistent with a sustained contribution of SGLT1 to tubular glucose reabsorption in WT mice. Absence of SGLT1 did not affect the initial kidney impairment versus WT mice, as indicated by similar increases on day 1 in plasma concentrations of creatinine and urinary excretion of the tubular injury marker kidney injury molecule-1 as well as a similar rise in plasma osmolality and fall in urine osmolality as indicators of impaired urine concentration. Recovery of kidney function on days 14/16, however, was improved in Sglt1-/- versus WT mice, as indicated by lower plasma creatinine, higher glomerula filtration rate (by FITC-sinistrin in awake mice), and more completely restored urine and plasma osmolality. This was associated with a reduced tubular injury score in the cortex and outer medulla, better preserved renal mRNA expression of tubular transporters (Sglt2 and Na+-K+-2Cl- cotransporter Nkcc2), and a lesser rise in renal mRNA expression of markers of injury, inflammation, and fibrosis [kidney injury molecule-1, chemokine (C-C motif) ligand 2, fibronectin 1, and collagen type I-α1] in Sglt1-/- versus WT mice. These results suggest that SGLT1 activity in the late proximal tubule may have deleterious effects during recovery of IR-induced acute kidney injury and identify SGLT1 as a potential therapeutic target.

Differential expression of parietal epithelial cell and podocyte extracellular matrix proteins in focal segmental glomerulosclerosis and diabetic nephropathy.

In healthy glomeruli, parietal epithelial cell (PEC)-derived extracellular matrix (ECM) proteins include laminin-ß1, perlecan, and collagen type IV-α2 and podocyte-specific ECM proteins include laminin-ß2, agrin, and collagen type IV-α4. This study aimed to define individual ECM protein isoform expression by PECs in both experimental and human focal segmental glomerulosclerosis (FSGS) and diabetic nephropathy (DN) and to determine if changes were CD44 dependent. In experimental FSGS induced with a cytotoxic podocyte antibody and in the BTBR ob/ob mouse model of DN, PEC-derived protein staining was significantly increased in PECs. Dual staining also showed de novo expression of the podocyte-specific ECM proteins laminin-ß2 and agrin in PECs. Similar findings were observed in biopsies from patients with FSGS and DN. Increases in individual ECM proteins colocalized with CD44 in PECs in disease. To determine the role of CD44, FSGS was induced in CD44-/- and CD44+/+ mice. PEC staining for perlecan, collagen type IV-α2, laminin-ß2, and agrin were significantly lower in diseased CD44-/- mice compared with diseased CD44+/+ mice. These results show that in experimental and human FSGS and DN, PECs typically in an activated state, produce both PEC-derived and podocyte-specific ECM protein isoforms, and that the majority of these changes were dependent on CD44.

A Novel Type 2 Diabetes Mouse Model of Combined Diabetic Kidney Disease and Atherosclerosis.

Diabetic kidney disease and atherosclerotic disease are major causes of morbidity and mortality associated with type 2 diabetes (T2D), and diabetic kidney disease is a major cardiovascular risk factor. The black and tan, brachyury (BTBR) mouse strain with leptin deficiency (Lepob) has emerged as one of the best models of human diabetic kidney disease. However, no T2D mouse model of combined diabetic kidney disease and atherosclerosis exists. Our goal was to generate such a model. To this end, the low-density lipoprotein (LDL) receptor was targeted for degradation via inducible degrader of the LDL receptor (IDOL) overexpression, using liver-targeted adenoassociated virus serotype DJ/8 (AAV-DJ/8) in BTBR wild-type and BTBR Lepob mice. Liver-targeted IDOL-AAV-DJ/8 increased plasma LDL cholesterol compared with the control enhanced green fluorescent protein AAV-DJ/8. IDOL-induced dyslipidemia caused formation of atherosclerotic lesions of an intermediate stage, which contained both macrophages and smooth muscle cells. BTBR Lepob mice exhibited diabetic kidney disease. IDOL-induced dyslipidemia worsened albuminuria and glomerular macrophage accumulation but had no effect on mesangial expansion or podocyte numbers. Thus, by inducing hepatic degradation of the LDL receptor, we generated a T2D model of combined kidney disease and atherosclerosis. This model provides a new tool to study mechanisms, interactions, and treatment strategies of kidney disease and atherosclerosis in T2D.

TACI deletion protects against progressive murine lupus nephritis induced by BAFF overexpression.

B cells are known to promote the pathogenesis of systemic lupus erythematosus (SLE) via the production of pathogenic anti-nuclear antibodies. However, the signals required for autoreactive B cell activation and the immune mechanisms whereby B cells impact lupus nephritis pathology remain poorly understood. The B cell survival cytokine B cell activating factor of the TNF Family (BAFF) has been implicated in the pathogenesis of SLE and lupus nephritis in both animal models and human clinical studies. Although the BAFF receptor has been predicted to be the primary BAFF family receptor responsible for BAFF-driven humoral autoimmunity, in the current study we identify a critical role for signals downstream of Transmembrane Activator and CAML Interactor (TACI) in BAFF-dependent lupus nephritis. Whereas transgenic mice overexpressing BAFF develop progressive membranoproliferative glomerulonephritis, albuminuria and renal dysfunction, TACI deletion in BAFF-transgenic mice provided long-term (about 1 year) protection from renal disease. Surprisingly, disease protection in this context was not explained by complete loss of glomerular immune complex deposits. Rather, TACI deletion specifically reduced endocapillary, but not mesangial, immune deposits. Notably, although excess BAFF promoted widespread breaks in B cell tolerance, BAFF-transgenic antibodies were enriched for RNA- relative to DNA-associated autoantigen reactivity. These RNA-associated autoantibody specificities were specifically reduced by TACI or Toll-like receptor 7 deletion. Thus, our study provides important insights into the autoantibody specificities driving proliferative lupus nephritis, and suggests that TACI inhibition may be novel and effective treatment strategy in lupus nephritis.

Histopathology of Veins Obtained at Hemodialysis Arteriovenous Fistula Creation Surgery.

Stenosis from venous neointimal hyperplasia is common in native arteriovenous fistulas (AVFs). However, the preexisting histologic characteristics of veins at fistula creation, and associations thereof with baseline patient factors, have not been well characterized. In this study, we conducted histologic analysis of a segment of the vein used for anastomosis creation, obtained during AVF creation from 554 of the 602 participants in the multicenter Hemodialysis Fistula Maturation Cohort Study. We quantified intimal and medial areas and lengths of the internal and external elastic lamina by morphometry and assessed venous wall cells by immunohistochemistry, extracellular matrix with Movat stain, and calcium deposition by alizarin red stain. We also studied a representative subset of veins for markers of monocyte/macrophage content, cell proliferation, apoptosis, and neoangiogenesis. Neointima occupied >20% of the lumen in 57% of fully circumferential vein samples, and neointimal hyperplasia associated positively with age and inversely with black race. The neointima was usually irregularly thickened, sometimes concentric, and contained α-smooth muscle actin-expressing cells of smooth muscle or myofibroblast origin. Proteoglycans admixed with lesser amounts of collagen constituted the predominant matrix in the neointima. In 82% of vein samples, the media of vessel walls contained large aggregates of collagen. A minority of veins expressed markers of inflammation, cell proliferation, cell death, calcification, or neoangiogenesis. In conclusion, we observed preexisting abnormalities, including neointimal hyperplasia and prominent accumulation of extracellular matrix, in veins used for AVF creation from a substantial proportion of this cohort.

The mitochondrial-targeted peptide, SS-31, improves glomerular architecture in mice of advanced age.

Although age-associated changes in kidney glomerular architecture have been described in mice and man, the mechanisms are unknown. It is unclear if these changes can be prevented or even reversed by systemic therapies administered at advanced age. Using light microscopy and transmission electron microscopy, our results showed glomerulosclerosis with injury to mitochondria in glomerular epithelial cells in mice aged 26 months (equivalent to a 79-year-old human). To test the hypothesis that reducing mitochondrial damage in late age would result in lowered glomerulosclerosis, we administered the mitochondrial targeted peptide, SS-31, to aged mice. Baseline (24-month-old) mice were randomized to receive 8 weeks of SS-31, or saline, and killed at 26 months of age. SS-31 treatment improved age-related mitochondrial morphology and glomerulosclerosis. Assessment of glomeruli revealed that SS-31 reduced senescence (p16, senescence-associated-ß-Gal) and increased the density of parietal epithelial cells. However, SS-31 treatment reduced markers of parietal epithelial cell activation (Collagen IV, pERK1/2, and α-smooth muscle actin). SS-31 did not impact podocyte density, but it reduced markers of podocyte injury (desmin) and improved cytoskeletal integrity (synaptopodin). This was accompanied by higher glomerular endothelial cell density (CD31). Thus, despite initiating therapy in late-age mice, a short course of SS-31 has protective benefits on glomerular mitochondria, accompanied by temporal changes to the glomerular architecture. This systemic pharmacological intervention in old-aged animals limits glomerulosclerosis and senescence, reduces parietal epithelial cell activation, and improves podocyte and endothelial cell integrity.

The role of PDGF-D in healthy and fibrotic kidneys.

Platelet-derived growth factor (PDGF)-D, a specific PDGF receptor ß (PDGFR-ß) ligand, mediates mesangial proliferation in vitro and in vivo. However, its role in renal development, physiology, and fibrosis is relatively unknown. In healthy murine kidneys, PDGF-D was found to be expressed on renal mesenchymal cells (mesangial cells, fibroblasts, and vascular smooth muscle cells). During renal fibrosis, PDGF-D and its receptor PDGFR-ß were markedly and similarly upregulated in both human and murine kidneys on activated mesenchymal cells, but PDGF-D was also expressed de novo in injured renal tubular cells. The functional role of PDGF-D was studied in Pdgfd-/- mice, which showed no obvious spontaneous renal phenotype at a young age or during aging. Compared with wild-type littermates, Pdgfd-/- mice had significantly reduced renal interstitial fibrosis in two models of renal scarring: unilateral ureteral obstruction and unilateral ischemia/reperfusion injury. This was associated with reduced phosphorylation of PDGFR-ß and its downstream mediator p38. Systemic adenoviral overexpression of PDGF-D in healthy mice resulted in increased collagen deposition in the kidney interstitium. Thus, PDGF-D is upregulated in murine and human kidney fibrosis, may mediate renal scarring, and is dispensable for normal kidney development and physiological functions. PDGF-D may be a suitable therapeutic target to combat kidney fibrosis.

Opposing impact of B cell-intrinsic TLR7 and TLR9 signals on autoantibody repertoire and systemic inflammation.

Systemic lupus erythematosus is a multisystem autoimmune disease characterized by autoantibodies targeting nucleic acid-associated Ags. The endosomal TLRs TLR7 and TLR9 are critical for generation of Abs targeting RNA- or DNA-associated Ags, respectively. In murine lupus models, deletion of TLR7 limits autoimmune inflammation, whereas deletion of TLR9 exacerbates disease. Whether B cell or myeloid TLR7/TLR9 signaling is responsible for these effects has not been fully addressed. In this study, we use a chimeric strategy to evaluate the effect of B cell-intrinsic deletion of TLR7 versus TLR9 in parallel lupus models. We demonstrate that B cell-intrinsic TLR7 deletion prevents RNA-associated Ab formation, decreases production of class-switched Abs targeting nonnuclear Ags, and limits systemic autoimmunity. In contrast, B cell-intrinsic TLR9 deletion results in decreased DNA-reactive Ab, but increased Abs targeting a broad range of systemic autoantigens. Further, we demonstrate that B cell-intrinsic TLR9 deletion results in increased systemic inflammation and immune complex glomerulonephritis, despite intact TLR signaling within the myeloid compartment. These data stress the critical importance of dysregulated B cell-intrinsic TLR signaling in the pathogenesis of systemic lupus erythematosus.

Deficient Autophagy Results in Mitochondrial Dysfunction and FSGS.

FSGS is a heterogeneous fibrosing disease of the kidney, the cause of which remains poorly understood. In most cases, there is no effective treatment to halt or retard progression to renal failure. Increasing evidence points to mitochondrial dysfunction and the generation of reactive oxygen species in the pathogenesis of CKD. Autophagy, a major intracellular lysosomal degradation system, performs homeostatic functions linked to metabolism and organelle turnover. We prevented normal autophagic pathways in nephrons of mice by mutating critical autophagy genes ATG5 or ATG7 during nephrogenesis. Mutant mice developed mild podocyte and tubular dysfunction within 2 months, profound glomerular and tubular changes bearing close similarity to human disease by 4 months, and organ failure by 6 months. Ultrastructurally, podocytes and tubular cells showed vacuolization, abnormal mitochondria, and evidence of endoplasmic reticulum stress, features that precede the appearance of histologic or clinical disease. Similar changes were observed in human idiopathic FSGS kidney biopsy specimens. Biochemical analysis of podocytes and tubules of 2-month-old mutant mice revealed elevated production of reactive oxygen species, activation of endoplasmic reticulum stress pathways, phosphorylation of p38, and mitochondrial dysfunction. Furthermore, cultured proximal tubule cells isolated from mutant mice showed marked mitochondrial dysfunction and elevated mitochondrial reactive oxygen species generation that was suppressed by a mitochondrial superoxide scavenger. We conclude that mitochondrial dysfunction and endoplasmic reticulum stress due to impaired autophagic organelle turnover in podocytes and tubular epithelium are sufficient to cause many of the manifestations of FSGS in mice.

Serum amyloid A and inflammation in diabetic kidney disease and podocytes.

Inflammatory pathways are central mechanisms in diabetic kidney disease (DKD). Serum amyloid A (SAA) is increased by chronic inflammation, but SAA has not been previously evaluated as a potential DKD mediator. The aims of this study were to determine whether SAA is increased in human DKD and corresponding mouse models and to assess effects of SAA on podocyte inflammatory responses. SAA was increased in the plasma of people with DKD characterized by overt proteinuria and inversely correlated with estimated glomerular filtration rate (creatinine-based CKD-EPI). SAA was also elevated in plasma of diabetic mouse models including type 1 diabetes (streptozotocin/C57BL/6) and type 2 diabetes (BTBR-ob/ob). SAA mRNA (Nephromine) was increased in human DKD compared with non-diabetic and/or glomerular disease controls (glomerular fold change 1.5, P=0.017; tubulointerstitium fold change 1.4, P=0.021). The kidneys of both diabetic mouse models also demonstrated increased SAA mRNA (quantitative real-time PCR) expression compared with non-diabetic controls (type 1 diabetes fold change 2.9; type 2 diabetes fold change 42.5, P=0.009; interaction by model P=0.57). Humans with DKD and the diabetic mouse models exhibited extensive SAA protein deposition in the glomeruli and tubulointerstitium in similar patterns by immunohistochemistry. SAA localized within podocytes of diabetic mice. Podocytes exposed to advanced glycation end products, metabolic mediators of inflammation in diabetes, increased expression of SAA mRNA (fold change 15.3, P=0.004) and protein (fold change 38.4, P=0.014). Podocytes exposed to exogenous SAA increased NF-κB activity, and pathway array analysis revealed upregulation of mRNA for NF-κB-dependent targets comprising numerous inflammatory mediators, including SAA itself (fold change 17.0, P=0.006). Inhibition of NF-κB reduced these pro-inflammatory responses. In conclusion, SAA is increased in the blood and produced in the kidneys of people with DKD and corresponding diabetic mouse models. Podocytes are likely to be key responder cells to SAA-induced inflammation in the diabetic kidney. SAA is a compelling candidate for DKD therapeutic and biomarker discovery.

The phenotypes of podocytes and parietal epithelial cells may overlap in diabetic nephropathy.

Reversal of diabetic nephropathy (DN) has been achieved in humans and mice, but only rarely and under special circumstances. As progression of DN is related to podocyte loss, reversal of DN requires restoration of podocytes. Here, we identified and quantified potential glomerular progenitor cells that could be a source for restored podocytes. DN was identified in 31 human renal biopsy cases and separated into morphologically early or advanced lesions. Markers of podocytes (WT-1, p57), parietal epithelial cells (PECs) (claudin-1), and cell proliferation (Ki-67) were identified by immunohistochemistry. Podocyte density was progressively reduced with DN. Cells marking as podocytes (p57) were present infrequently on Bowman's capsule in controls, but significantly increased in histologically early DN. Ki-67-expressing cells were identified on the glomerular tuft and Bowman's capsule in DN, but rarely in controls. Cells marking as PECs were present on the glomerular tuft, particularly in morphologically advanced DN. These findings show evidence of phenotypic plasticity in podocyte and PEC populations and are consistent with studies in the BTBR ob/ob murine model in which reversibility of DN occurs with podocytes potentially regenerating from PEC precursors. Thus, our findings support, but do not prove, that podocytes may regenerate from PEC progenitors in human DN. If so, progression of DN may represent a modifiable net balance between podocyte loss and regeneration.

Foam cells and the pathogenesis of kidney disease.

PURPOSE OF REVIEW: Foam cells in human glomeruli can be encountered in various renal diseases including focal segmental glomerulosclerosis and diabetic nephropathy. Although foam cells are key participants in atherosclerosis, surprisingly little is known about their pathogenicity in the kidney. We review our understanding (or lack thereof) of foam cells in the kidney, as well as insights gained in studies of foam cells and macrophages involved in atherosclerosis to suggest areas of investigation that will allow better characterization of the role of these cells in renal disease. RECENT FINDINGS: There is a general dearth of animal models of disease with renal foam cell accumulation, limiting progress in our understanding of the pathobiology of these cells. Recent genetic modifications of hyperlipidemic mice have resulted in some new disease models with renal foam cell accumulation. Recent studies have challenged older paradigms by findings that indicate that many tissue macrophages are derived from cells permanently residing in the tissue from birth rather than circulating monocytes. SUMMARY: Renal foam cells remain an enigma. Extrapolating from studies of atherosclerosis suggests that therapeutics targeting mitochondrial reactive oxygen species production, or modulating cholesterol and lipoprotein uptake or egress from these cells, may prove beneficial for kidney diseases in which foam cells are present.