Accumulation of α-synuclein mediates podocyte injury in Fabry nephropathy.

Current therapies for Fabry disease are based on reversing intracellular accumulation of globotriaosylceramide (Gb3) by enzyme replacement therapy (ERT) or chaperone-mediated stabilization of the defective enzyme, thereby alleviating lysosomal dysfunction. However, their effect in the reversal of end-organ damage, like kidney injury and chronic kidney disease, remains unclear. In this study, ultrastructural analysis of serial human kidney biopsies showed that long-term use of ERT reduced Gb3 accumulation in podocytes but did not reverse podocyte injury. Then, a CRISPR/Cas9-mediated α-galactosidase knockout podocyte cell line confirmed ERT-mediated reversal of Gb3 accumulation without resolution of lysosomal dysfunction. Transcriptome-based connectivity mapping and SILAC-based quantitative proteomics identified α-synuclein (SNCA) accumulation as a key event mediating podocyte injury. Genetic and pharmacological inhibition of SNCA improved lysosomal structure and function in Fabry podocytes, exceeding the benefits of ERT. Together, this work reconceptualizes Fabry-associated cell injury beyond Gb3 accumulation, and introduces SNCA modulation as a potential intervention, especially for patients with Fabry nephropathy.

CD4+ T cells produce GM-CSF and drive immune-mediated glomerular disease by licensing monocyte-derived cells to produce MMP12.

GM-CSF in glomerulonephritisDespite glomerulonephritis being an immune-mediated disease, the contributions of individual immune cell types are not clear. To address this gap in knowledge, Paust et al. characterized pathological immune cells in samples from patients with glomerulonephritis and in samples from mice with the disease. The authors found that CD4+ T cells producing granulocyte-macrophage colony-stimulating factor (GM-CSF) licensed monocytes to promote disease by producing matrix metalloproteinase 12 and disrupting the glomerular basement membrane. Targeting GM-CSF to inhibit this axis reduced disease severity in mice, implicating this cytokine as a potential therapeutic target for patients with glomerulonephritis. -CM.

Antigen Cross-Presentation by Murine Proximal Tubular Epithelial Cells Induces Cytotoxic and Inflammatory CD8+ T Cells.

Immune-mediated glomerular diseases are characterized by infiltration of T cells, which accumulate in the periglomerular space and tubulointerstitium in close contact to proximal and distal tubuli. Recent studies described proximal tubular epithelial cells (PTECs) as renal non-professional antigen-presenting cells that stimulate CD4+ T-cell activation. Whether PTECs have the potential to induce activation of CD8+ T cells is less clear. In this study, we aimed to investigate the capacity of PTECs for antigen cross-presentation thereby modulating CD8+ T-cell responses. We showed that PTECs expressed proteins associated with cross-presentation, internalized soluble antigen via mannose receptor-mediated endocytosis, and generated antigenic peptides by proteasomal degradation. PTECs induced an antigen-dependent CD8+ T-cell activation in the presence of soluble antigen in vitro. PTEC-activated CD8+ T cells expressed granzyme B, and exerted a cytotoxic function by killing target cells. In murine lupus nephritis, CD8+ T cells localized in close contact to proximal tubuli. We determined enhanced apoptosis in tubular cells and particularly PTECs up-regulated expression of cleaved caspase-3. Interestingly, induction of apoptosis in the inflamed kidney was reduced in the absence of CD8+ T cells. Thus, PTECs have the capacity for antigen cross-presentation thereby inducing cytotoxic CD8+ T cells in vitro, which may contribute to the pathology of immune-mediated glomerulonephritis.

Ectodomain shedding by ADAM proteases as a central regulator in kidney physiology and disease.

Besides its involvement in blood and bone physiology, the kidney's main function is to filter substances and thereby regulate the electrolyte composition of body fluids, acid-base balance and toxin removal. Depending on underlying conditions, the nephron must undergo remodeling and cellular adaptations. The proteolytic removal of cell surface proteins via ectodomain shedding by A Disintegrin and Metalloproteases (ADAMs) is of importance for the regulation of cell-cell and cell-matrix adhesion of renal cells. ADAM10 controls glomerular and tubule development in a Notch1 signaling-dependent manner and regulates brush border composition. ADAM17 regulates the renin angiotensin system and is together with ADAM10 involved in calcium phosphate homeostasis. In kidney disease ADAMs, especially ADAM17 contribute to inflammation through their involvement in IL-6 trans-signaling, Notch-, epithelial growth factor receptor-, and tumor necrosis factor α signaling. ADAMs are interesting drug targets to reduce the inflammatory burden, defective cell adhesion and impaired signaling pathways in kidney diseases.

CD73-mediated adenosine production by CD8 T cell-derived extracellular vesicles constitutes an intrinsic mechanism of immune suppression.

Immune cells at sites of inflammation are continuously activated by local antigens and cytokines, and regulatory mechanisms must be enacted to control inflammation. The stepwise hydrolysis of extracellular ATP by ectonucleotidases CD39 and CD73 generates adenosine, a potent immune suppressor. Here we report that human effector CD8 T cells contribute to adenosine production by releasing CD73-containing extracellular vesicles upon activation. These extracellular vesicles have AMPase activity, and the resulting adenosine mediates immune suppression independently of regulatory T cells. In addition, we show that extracellular vesicles isolated from the synovial fluid of patients with juvenile idiopathic arthritis contribute to T cell suppression in a CD73-dependent manner. Our results suggest that the generation of adenosine upon T cell activation is an intrinsic mechanism of human effector T cells that complements regulatory T cell-mediated suppression in the inflamed tissue. Finally, our data underscore the role of immune cell-derived extracellular vesicles in the control of immune responses.

The ubiquitin-proteasome system in kidney physiology and disease.

Intracellular proteins continuously turn over by degradation and synthesis in all organ tissues. Owing to its irreversible nature, protein degradation is a highly selective process to avoid irreparable breakdown of cellular constituents, thereby disrupting cellular stability, integrity and signalling. The majority of intracellular proteins are degraded by the ubiquitin-proteasome system (UPS), a multi-enzyme process that involves the covalent conjugation of ubiquitin to a substrate protein and its recognition and degradation by the core multicomponent proteolytic complex of the UPS, the proteasome. In addition to labelling misfolded, damaged, aggregation-prone and intact but unneeded proteins for proteasomal degradation, ubiquitylation regulates a multitude of cellular processes, such as transcription, translation, endocytosis, and receptor activity and subcellular localization. In addition, the proteasome generates peptides for antigen presentation in the immune system and for further degradation by peptidases to provide amino acids for protein biosynthesis and gluconeogenesis. Alterations of the UPS or of protein substrates that render them more or less susceptible to degradation are responsible for disorders associated with renal cell dysfunction. In this Review, we provide insight into the elegant and complex nature of UPS-mediated proteostasis and focus on its established and potential roles in renal cell physiology and pathophysiology.

Renal proximal tubular epithelial cells exert immunomodulatory function by driving inflammatory CD4+ T cell responses.

In immune-mediated glomerular diseases like crescentic glomerulonephritis (cGN), inflammatory CD4+ T cells accumulate within the tubulointerstitial compartment in close contact to proximal and distal tubular epithelial cells and drive renal inflammation and tissue damage. However, whether renal epithelial cell populations play a role in the pathogenesis of cGN by modulating CD4+ T cell responses is less clear. In the present study, we aimed to investigate the potential of renal epithelial cells to function as antigen-presenting cells, thereby stimulating CD4+ T cell responses. Using a FACS-based protocol that allowed comparative analysis of cortical epithelial cell populations, we showed that particularly proximal tubular epithelial cells (PTECs) express molecules linked with antigen-presenting cell function, including major histocompatibility complex class II (MHCII), CD74, CD80, and CD86 in homeostasis and nephrotoxic nephritis, a murine model of cGN. Protein expression was visualized at the PTEC single cell level by imaging flow cytometry. Interestingly, we found inflammation-dependent regulation of epithelium-expressed CD74, CD80, and CD86, whereas MHCII expression was not altered. Antigen-specific stimulation of CD4+ T cells by PTECs in vitro supported CD4+ T cell survival and induced CD4+ T cell activation, proliferation, and inflammatory cytokine production. In patients with antineutrophil cytoplasmic antibody-associated glomerulonephritis, MHCII and CD74 were expressed by both proximal and distal tubules, whereas CD86 was predominantly expressed by proximal tubules. Thus, particularly PTECs have the potential to induce an inflammatory phenotype in CD4+ T cells in vitro, which might also play a role in the pathology of immune-mediated kidney disease.

The chemokine receptor CX3CR1 reduces renal injury in mice with angiotensin II-induced hypertension.

The role of CX3CR1, also known as fractalkine receptor, in hypertension is unknown. The present study determined the role of the fractalkine receptor CX3CR1 in hypertensive renal and cardiac injury. Expression of CX3CR1 was determined using CX3CR1GFP/+ mice that express a green fluorescent protein (GFP) reporter in CX3CR1+ cells. FACS analysis of leukocytes isolated from the kidney showed that 34% of CD45+ cells expressed CX3CR1. Dendritic cells were the majority of positive cells (67%) followed by macrophages (10%), NK cells (6%), and T cells (10%). With the use of confocal microscopy, the receptor was detected in the kidney only on infiltrating cells but not on resident renal cells. To evaluate the role of CX3CR1 in hypertensive end-organ injury, an aggravated model of hypertension was used. Unilateral nephrectomy was performed followed by infusion of angiotensin II (ANG II, 1.5 ng·g-1·min-1) and a high-salt diet in wild-type ( n = 15) and CX3CR1-deficient mice ( n = 18). CX3CR1 deficiency reduced the number of renal dendritic cells and increased the numbers of renal CD11b/F4/80+ macrophages and CD11b/Ly6G+ neutrophils in ANG II-infused mice. Surprisingly, CX3CR1-deficient mice exhibited increased albuminuria, glomerular injury, and reduced podocyte density in spite of similar levels of arterial hypertension. In contrast, cardiac damage as assessed by increased heart weight, cardiac fibrosis, and expression of fetal genes, and matrix components were not different between both genotypes. Our findings suggest that CX3CR1 exerts protective properties by modulating the invasion of inflammatory cells in hypertensive renal injury. CX3CR1 inhibition should be avoided in hypertension because it may promote hypertensive renal injury.

Role of Apolipoprotein L1 in Human Parietal Epithelial Cell Transition.

Human parietal epithelial cells (PECs) are progenitor cells that sustain podocyte homeostasis. We hypothesized that the lack of apolipoprotein (APO) L1 ensures the PEC phenotype, but its induction initiates PEC transition (expression of podocyte markers). APOL1 expression and down-regulation of miR193a coincided with the expression of podocyte markers during the transition. The induction of APOL1 also stimulated transition markers in human embryonic kidney cells (cells with undetectable APOL1 protein expression). APOL1 silencing in PECs up-regulated miR193a expression, suggesting the possibility of a reciprocal feedback relationship between APOL1 and miR193a. HIV, interferon-γ, and vitamin D receptor agonist down-regulated miR193a expression and induced APOL1 expression along with transition markers in PECs. Luciferase assay suggested a putative interaction between miR193a and APOL1. Since silencing of APOL1 attenuated HIV-, vitamin D receptor agonist-, miR193a inhibitor-, and interferon-γ-induced expression of transition markers, APOL1 appears to be a critical functional constituent of the miR193a- APOL1 axis in PECs. This notion was confirmed by further enhanced expression of PEC markers in APOL1 mRNA-silenced PECs. In vivo studies, glomeruli in patients with HIV, and HIV/APOL1 transgenic mice had foci of PECs expressing synaptopodin, a transition marker. APOL1 likely regulates PEC molecular phenotype through modulation of miR193a expression, and APOL1 and miR193a share a reciprocal feedback relationship.

An Indirect Immunofluorescence Method Facilitates Detection of Thrombospondin Type 1 Domain-Containing 7A-Specific Antibodies in Membranous Nephropathy.

Thrombospondin type 1 domain-containing 7A (THSD7A) is a target antigen identified in adult membranous nephropathy (MN) along with the major antigen phospholipase A2 receptor 1 (PLA2R1). The prevalence of THSD7A-Ab-positive patients is unknown, and it is unclear whether the clinical presentation differs between patients positive for PLA2R1-Ab or THSD7A-Ab. We screened serum samples of 1276 patients with MN from three different cohorts for the presence of THSD7A-Ab by Western blot analysis and a newly developed indirect immunofluorescence test (IFT). Compared with Western blot analysis, the IFT had a 92% sensitivity and a 100% specificity. The prevalence of THSD7A-associated MN in a prospective cohort of 345 patients with MN was 2.6%, and most were women. In this cohort, the percentage of patients with THSD7A-associated MN and malignant disease significantly exceeded that of patients with PLA2R1-associated MN and malignant disease. In all cohorts, we identified 40 patients with THSD7A-associated MN, eight of whom developed a malignancy within a median time of 3 months from diagnosis of MN. In one patient with THSD7A-associated MN and metastases of an endometrial carcinoma, immunohistochemistry showed THSD7A expression on the metastatic cells and within follicular dendritic cells of the metastasis-infiltrated lymph node. We conclude that the IFT allows sensitive and specific measurement of circulating THSD7A-Ab in patients with MN. Patients with THSD7A-associated MN differ in their clinical characteristics from patients with PLA2R1-associated MN, and more intensive screening for the presence of malignancies may be warranted in those with THSD7A-associated MN.

Nanobodies that block gating of the P2X7 ion channel ameliorate inflammation.

Ion channels are desirable therapeutic targets, yet ion channel-directed drugs with high selectivity and few side effects are still needed. Unlike small-molecule inhibitors, antibodies are highly selective for target antigens but mostly fail to antagonize ion channel functions. Nanobodies-small, single-domain antibody fragments-may overcome these problems. P2X7 is a ligand-gated ion channel that, upon sensing adenosine 5'-triphosphate released by damaged cells, initiates a proinflammatory signaling cascade, including release of cytokines, such as interleukin-1ß (IL-1ß). To further explore its function, we generated and characterized nanobodies against mouse P2X7 that effectively blocked (13A7) or potentiated (14D5) gating of the channel. Systemic injection of nanobody 13A7 in mice blocked P2X7 on T cells and macrophages in vivo and ameliorated experimental glomerulonephritis and allergic contact dermatitis. We also generated nanobody Dano1, which specifically inhibited human P2X7. In endotoxin-treated human blood, Dano1 was 1000 times more potent in preventing IL-1ß release than small-molecule P2X7 antagonists currently in clinical development. Our results show that nanobody technology can generate potent, specific therapeutics against ion channels, confirm P2X7 as a therapeutic target for inflammatory disorders, and characterize a potent new drug candidate that targets P2X7.

Autoantibodies against thrombospondin type 1 domain-containing 7A induce membranous nephropathy.

Membranous nephropathy (MN) is the most common cause of nephrotic syndrome in adults, and one-third of patients develop end-stage renal disease (ESRD). Circulating autoantibodies against the podocyte surface antigens phospholipase A2 receptor 1 (PLA2R1) and the recently identified thrombospondin type 1 domain-containing 7A (THSD7A) are assumed to cause the disease in the majority of patients. The pathogenicity of these antibodies, however, has not been directly proven. Here, we have reported the analysis and characterization of a male patient with THSD7A-associated MN who progressed to ESRD and subsequently underwent renal transplantation. MN rapidly recurred after transplantation. Enhanced staining for THSD7A was observed in the kidney allograft, and detectable anti-THSD7A antibodies were present in the serum before and after transplantation, suggesting that these antibodies induced a recurrence of MN in the renal transplant. In contrast to PLA2R1, THSD7A was expressed on both human and murine podocytes, enabling the evaluation of whether anti-THSD7A antibodies cause MN in mice. We demonstrated that human anti-THSD7A antibodies specifically bind to murine THSD7A on podocyte foot processes, induce proteinuria, and initiate a histopathological pattern that is typical of MN. Furthermore, anti-THSD7A antibodies induced marked cytoskeletal rearrangement in primary murine glomerular epithelial cells as well as in human embryonic kidney 293 cells. Our findings support a causative role of anti-THSD7A antibodies in the development of MN.

Immune Complex-Type Deposits in the Fischer-344 to Lewis Rat Model of Renal Transplantation and a Subset of Human Transplant Glomerulopathy.

BACKGROUND: Antibody-mediated rejection is a leading cause for renal transplant loss. Rodent models are useful to dissect pathomechanisms and to develop treatment strategies. Although used for decades as a model, glomerular histopathological findings of Fischer-344 kidneys transplanted into Lewis rats have never been comprehensively described. METHODS: Kidneys from Fischer-344 rats were transplanted into Lewis rats as life-sustaining allografts without immunosuppression. Lewis isografts and normal Fischer-344 kidneys served as controls. Grafts were harvested at 9 days, 6 and 26 weeks. Histopathological examination included light microscopy, immunohistochemistry, and morphometry. Findings were compared with 51 human biopsies with transplant glomerulopathy. RESULTS: Most glomerular findings in rat allografts resembled human acute and chronic antibody-mediated rejection with glomerulitis, microthrombosis, microaneurysms, glomerular hypertrophy, podocyte loss, glomerular basement membrane splitting, and secondary focal and segmental glomerulosclerosis. In line with previous reports on nonendothelial antigens, glomerular immunoglobulin and C4d deposition was mostly nonendothelial. Only in 26-week allografts, we found mesangial and subendothelial immune complex-type electron-dense deposits. Similar deposits were found in 8 of 51 human biopsies with transplant glomerulopathy after rigorous exclusion of immune complexes of other cause, particularly recurrent glomerulonephritis and hepatitis C. CONCLUSIONS: Thus, our model closely reflects the glomerular changes of acute antibody-mediated rejection in humans and of a special subset of human transplant glomerulopathy. The significance of alloimmune immune complex-type deposits in human transplants deserves further investigation.

CD38 is expressed on inflammatory cells of the intestine and promotes intestinal inflammation.

The enzyme CD38 is expressed on a variety of hematopoietic and non-hematopoietic cells and is involved in diverse processes such as generation of calcium-mobilizing metabolites, cell activation, and chemotaxis. Here, we show that under homeostatic conditions CD38 is highly expressed on immune cells of the colon mucosa of C57BL/6 mice. Myeloid cells recruited to this tissue upon inflammation also express enhanced levels of CD38. To determine the role of CD38 in intestinal inflammation, we applied the dextran sulfate sodium (DSS) colitis model. Whereas wild-type mice developed severe colitis, CD38-/- mice had only mild disease following DSS-treatment. Histologic examination of the colon mucosa revealed pronounced inflammatory damage with dense infiltrates containing numerous granulocytes and macrophages in wild-type animals, while these findings were significantly attenuated in CD38-/- mice. Despite attenuated histological findings, the mRNA expression of inflammatory cytokines and chemokines was only marginally lower in the colons of CD38-/- mice as compared to wild-type mice. In conclusion, our results identify a function for CD38 in the control of inflammatory processes in the colon.

Glomerulopathy induced by immunization with a peptide derived from the goodpasture antigen α3IV-NC1.

Mouse experimental autoimmune glomerulonephritis, a model of human antiglomerular basement membrane disease, depends on both Ab and T cell responses to the Goodpasture Ag noncollagenous domain 1 of the α3-chain of type IV collagen (α3IV-NC1). The aim of our study was to further characterize the T cell-mediated immune response. Repeated immunization with mouse α3IV-NC1 caused fatal glomerulonephritis in DBA/1 mice. Although two immunizations were sufficient to generate high α3IV-NC1-specific IgG titers, Ab and complement deposition along the glomerular basement membranes, and a nephrotic syndrome, two additional immunizations were needed to induce a necrotizing/crescentic glomerulonephritis. Ten days after the first immunization, α3IV-NC1-specific CD4(+) cells producing TNF-α, IFN-γ, or IL-17A were detected in the spleen. With the emergence of necrotizing/crescentic glomerulonephritis, ∼0.15% of renal CD4(+) cells were specific for α3IV-NC1. Using peptides spanning the whole α3IV-NC1 domain, three immunodominant T cell epitopes were identified. Immunization with these peptides did not lead to clinical signs of experimental autoimmune glomerulonephritis or necrotizing/crescentic glomerulonephritis. However, mice immunized with one of the peptides (STVKAGDLEKIISRC) developed circulating Abs against mouse α3IV-NC1 first detected at 8 wk, and 50% of the mice showed mild proteinuria at 18-24 wk due to membranous glomerulopathy. Taken together, our results suggest that autoreactive T cells are able to induce the formation of pathologic autoantibodies. The quality and quantity of α3IV-NC1-specific Ab and T cell responses are critical for the phenotype of the glomerulonephritis.

MicroRNA-193a Regulates the Transdifferentiation of Human Parietal Epithelial Cells toward a Podocyte Phenotype.

Parietal epithelial cells have been identified as potential progenitor cells in glomerular regeneration, but the molecular mechanisms underlying this process are not fully defined. Here, we established an immortalized polyclonal human parietal epithelial cell (hPEC) line from naive human Bowman's capsule cells isolated by mechanical microdissection. These hPECs expressed high levels of PEC-specific proteins and microRNA-193a (miR-193a), a suppressor of podocyte differentiation through downregulation of Wilms' tumor 1 in mice. We then investigated the function of miR-193a in the establishment of podocyte and PEC identity and determined whether inhibition of miR-193a influences the behavior of PECs in glomerular disease. After stable knockdown of miR-193a, hPECs adopted a podocyte-like morphology and marker expression, with decreased expression levels of PEC markers. In mice, inhibition of miR-193a by complementary locked nucleic acids resulted in an upregulation of the podocyte proteins synaptopodin and Wilms' tumor 1. Conversely, overexpression of miR-193a in vivo resulted in the upregulation of PEC markers and the loss of podocyte markers in isolated glomeruli. Inhibition of miR-193a in a mouse model of nephrotoxic nephritis resulted in reduced crescent formation and decreased proteinuria. Together, these results show the establishment of a human PEC line and suggest that miR-193a functions as a master switch, such that glomerular epithelial cells with high levels of miR-193a adopt a PEC phenotype and cells with low levels of miR-193a adopt a podocyte phenotype. miR-193a-mediated maintenance of PECs in an undifferentiated reactive state might be a prerequisite for PEC proliferation and migration in crescent formation.

CXCL5 drives neutrophil recruitment in TH17-mediated GN.

Neutrophil trafficking to sites of inflammation is essential for the defense against bacterial and fungal infections, but also contributes to tissue damage in TH17-mediated autoimmunity. This process is regulated by chemokines, which often show an overlapping expression pattern and function in pathogen- and autoimmune-induced inflammatory reactions. Using a murine model of crescentic GN, we show that the pathogenic TH17/IL-17 immune response induces chemokine (C-X-C motif) ligand 5 (CXCL5) expression in kidney tubular cells, which recruits destructive neutrophils that contribute to renal tissue injury. By contrast, CXCL5 was dispensable for neutrophil recruitment and effective bacterial clearance in a murine model of acute bacterial pyelonephritis. In line with these findings, CXCL5 expression was highly upregulated in the kidneys of patients with ANCA-associated crescentic GN as opposed to patients with acute bacterial pyelonephritis. Our data therefore identify CXCL5 as a potential therapeutic target for the restriction of pathogenic neutrophil infiltration in TH17-mediated autoimmune diseases while leaving intact the neutrophil function in protective immunity against invading pathogens.

Thrombospondin type-1 domain-containing 7A in idiopathic membranous nephropathy.

BACKGROUND: Idiopathic membranous nephropathy is an autoimmune disease. In approximately 70% of patients, it is associated with autoantibodies against the phospholipase A2 receptor 1 (PLA2R1). Antigenic targets in the remaining patients are unknown. METHODS: Using Western blotting, we screened serum samples from patients with idiopathic membranous nephropathy, patients with other glomerular diseases, and healthy controls for antibodies against human native glomerular proteins. We partially purified a putative new antigen, identified this protein by means of mass spectrometry of digested peptides, and validated the results by analysis of recombinant protein expression, immunoprecipitation, and immunohistochemical analysis. RESULTS: Serum samples from 6 of 44 patients in a European cohort and 9 of 110 patients in a Boston cohort with anti-PLA2R1-negative idiopathic membranous nephropathy recognized a glomerular protein that was 250 kD in size. None of the serum samples from the 74 patients with idiopathic membranous nephropathy who were seropositive for anti-PLA2R1 antibodies, from the 76 patients with other glomerular diseases, and from the 44 healthy controls reacted against this antigen. Although this newly identified antigen is clearly different from PLA2R1, it shares some biochemical features, such as N-glycosylation, membranous location, and reactivity with serum only under nonreducing conditions. Mass spectrometry identified this antigen as thrombospondin type-1 domain-containing 7A (THSD7A). All reactive serum samples recognized recombinant THSD7A and immunoprecipitated THSD7A from glomerular lysates. Moreover, immunohistochemical analyses of biopsy samples from patients revealed localization of THSD7A to podocytes, and IgG eluted from one of these samples was specific for THSD7A. CONCLUSIONS: In our cohort, 15 of 154 patients with idiopathic membranous nephropathy had circulating autoantibodies to THSD7A but not to PLA2R1, a finding that suggests a distinct subgroup of patients with this condition. (Funded by the French National Center for Scientific Research and others.).

UCH-L1 induces podocyte hypertrophy in membranous nephropathy by protein accumulation.

Podocytes are terminally differentiated cells of the glomerular filtration barrier that react with hypertrophy in the course of injury such as in membranous nephropathy (MGN). The neuronal deubiquitinase ubiquitin C-terminal hydrolase L1 (UCH-L1) is expressed and activated in podocytes of human and rodent MGN. UCH-L1 regulates the mono-ubiquitin pool and induces accumulation of poly-ubiquitinated proteins in affected podocytes. Here, we investigated the role of UCH-L1 in podocyte hypertrophy and in the homeostasis of the hypertrophy associated "model protein" p27(Kip1). A better understanding of the basic mechanisms leading to podocyte hypertrophy is crucial for the development of specific therapies in MGN. In human and rat MGN, hypertrophic podocytes exhibited a simultaneous up-regulation of UCH-L1 and of cytoplasmic p27(Kip1) content. Functionally, inhibition of UCH-L1 activity and knockdown or inhibition of UCH-L1 attenuated podocyte hypertrophy by decreasing the total protein content in isolated glomeruli and in cultured podocytes. In contrast, UCH-L1 levels and activity increased podocyte hypertrophy and total protein content in culture, specifically of cytoplasmic p27(Kip1). UCH-L1 enhanced cytoplasmic p27(Kip1) levels by nuclear export and decreased poly-ubiquitination and proteasomal degradation of p27(Kip1). In parallel, UCH-L1 increased podocyte turnover, migration and cytoskeletal rearrangement, which are associated with known oncogenic functions of cytoplasmic p27(Kip1) in cancer. We propose that UCH-L1 induces podocyte hypertrophy in MGN by increasing the total protein content through altered degradation and accumulation of proteins such as p27(Kip1) in the cytoplasm of podocytes. Modification of both UCH-L1 activity and levels could be a new therapeutic avenue to podocyte hypertrophy in MGN.