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Antibody-mediated autoimmune pathologies like membranous nephropathy are difficult to model, particularly in the absence of local target antigen expression in model organisms such as mice and rats; as is the case for phospholipase A2 receptor 1 (PLA2R1), the major autoantigen in membranous nephropathy. Here, we generated a transgenic mouse line expressing the full-length human PLA2R1 in podocytes, which has no kidney impairment after birth. Beginning from the age of three weeks, these mice spontaneously developed anti-human PLA2R1 antibodies, a nephrotic syndrome with progressive albuminuria and hyperlipidemia, and the typical morphological signs of membranous nephropathy with granular glomerular deposition of murine IgG in immunofluorescence and subepithelial electron-dense deposits by electron microscopy. Importantly, human PLA2R1-expressing Rag2

BACKGROUND: Primary membranous nephropathy (MN) is an autoimmune kidney disease in which immune complexes are deposited beneath the epithelium in the glomeruli. The condition introduces a high risk for end-stage kidney disease. Seventy to 80 percent of patients with MN have circulating antibodies against phospholipase A2 receptor 1 (PLA2R1), and levels correlate with treatment response and prognosis. However, experimental evidence that human anti-PLA2R1 antibodies induce MN has been elusive.METHODS: In passive transfer experiments, minipigs received plasma or purified IgG from patients with PLA2R1-associated MN or from healthy controls. Anti-PLA2R1 antibodies and proteinuria were monitored using Western blot, ELISA, and Coomassie staining. Kidney tissues were analyzed using immunohistochemistry, immunofluorescence, electron microscopy, and proteomic analyses.RESULTS: Minipigs, like humans, express PLA2R1 on podocytes. Human anti-PLA2R1 antibodies bound to minipig PLA2R1 in vitro and in vivo. Passive transfer of human anti-PLA2R1 antibodies from patients with PLA2R1-associated MN to minipigs led to histological characteristics of human early-stage MN, activation of components of the complement cascade, and low levels of proteinuria. We observed development of an autologous, later phase of disease.CONCLUSIONS: A translational approach from humans to minipigs showed that human anti-PLA2R1 antibodies are pathogenic in MN, although in the heterologous phase of disease only low-level proteinuria developed.

Changes in habitat availability and prey abundance are predicted to adversely influence survival and reproduction of wildlife in the Southern Ocean. Some populations of southern right whale (SRW; Eubalaena australis) are showing dramatic changes in habitat use. Surveys were undertaken in the austral winters of 2020 and 2021 at the key nursery and socialising ground for New Zealand SRWs: Port Ross, Auckland Islands, with 548 encounters and 599 skin biopsy samples collected. Data from these two surveys spanned peak periods of use and were used to test the hypothesis there have been shifts in the phenology, demographic composition and behaviour of SRWs using the Auckland Islands over the past three decades. The behavioural phenology and demographic composition of SRW resembles that observed in the 1990s. In contrast, the proportion of groups containing cow-calf pairs increased from 20% in the 1998 survey to 50% in 2020/21. These changes are consistent with a growing population undergoing strong recruitment, not limited by food resources. Continued use of Port Ross by all SRW demographic classes confirms this as key habitat for SRW in New Zealand waters, and we support increased enforcement of existing management measures to reduce whale-vessel interactions in this remote subantarctic archipelago.

Calcimimetic agents allosterically increase the calcium ion sensitivity of the calcium-sensing receptor (CaSR), which is expressed in the tubular system and to a lesser extent in podocytes. Activation of this receptor can reduce glomerular proteinuria and structural damage in proteinuric animal models. However, the precise role of the podocyte CaSR remains unclear. Here, a CaSR knockdown in cultured murine podocytes and a podocyte-specific CaSR knockout in BALB/c mice were generated to study its role in proteinuria and kidney function. Podocyte CaSR knockdown abolished the calcimimetic R-568 mediated calcium ion-influx, disrupted the actin cytoskeleton, and reduced cellular attachment and migration velocity. Adriamycin-induced proteinuria enhanced glomerular CaSR expression in wild type mice. Albuminuria, podocyte foot process effacement, podocyte loss and glomerular sclerosis were significantly more pronounced in adriamycin-treated podocyte-specific CaSR knockout mice compared to wild type littermates. Co-treatment of wild type mice with adriamycin and the calcimimetic cinacalcet reduced proteinuria in wild type, but not in podocyte-specific CaSR knockout mice. Additionally, four children with nephrotic syndrome, whose parents objected to glucocorticoid therapy, were treated with cinacalcet for one to 33 days. Proteinuria declined transiently by up to 96%, serum albumin increased, and edema resolved. Thus, activation of podocyte CaSR regulates key podocyte functions in vitro and reduced toxin-induced proteinuria and glomerular damage in mice. Hence, our findings suggest a potential novel role of CaSR signaling in control of glomerular disease.

Purpose:Prostate-specific membrane antigen (PSMA) targeting radioligands deliver radiation to PSMA-expressing cells. However, the relationship between PSMA levels and intralesion heterogeneity of PSMA expression, and cytotoxic radiation by radioligand therapy (RLT) is unknown. Here we investigate RLT efficacy as function of PSMA levels/cell, and the fraction of PSMA+ cells in a tumor.Experimental Design:RM1 cells expressing different levels of PSMA (PSMA−, PSMA+, PSMA++, PSMA+++; study 1) or a mix of PSMA+ and PSMA− RM1 (study 2, 4) or PC-3/PC-3-PIP (study 3) cells at various ratios were injected into mice. Mice received 177Lu- (studies 1–3) or 225Ac- (study 4) PSMA617. Tumor growth was monitored. Two days post-RLT, tumors were resected in a subset of mice. Radioligand uptake and DNA damage were quantified.Results:177Lu-PSMA617 efficacy increased with increasing PSMA levels (study 1) and fractions of PSMA positive cells (studies 2, 3) in both, the RM1 and PC-3-PIP models. In tumors resected 2 days post-RLT, PSMA expression correlated with 177Lu-PSMA617 uptake and the degree of DNA damage. Compared with 177Lu-PSMA617, 225Ac-PSMA617 improved overall antitumor effectiveness and tended to enhance the differences in therapeutic efficacy between experimental groups.Conclusions:In the current models, both the degree of PSMA expression and the fraction of PSMA+ cells correlate with 177Lu-/225Ac-PSMA617 tumor uptake and DNA damage, and thus, RLT efficacy. Low or heterogeneous PSMA expression represents a resistance mechanism to RLT.See related commentary by Ravi Kumar and Hofman, p. 2774

Absolute tumor volumes and bodyweights

Relative tumor volumes, 177Lu-/225Ac-PSMA617 uptake and 53BP1 IRS

GM-CSF in glomerulonephritis Despite 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

Abstract Chronic hyperglycemia, as in diabetes mellitus, may cause glomerular damage with microalbuminuria as an early sign. Noteworthy, even acute hyperglycemia can increase glomerular permeability before structural damage of the glomerular filter can be detected. Despite intensive research, specific antiproteinuric therapy is not available so far. Thus, a deeper understanding of the molecular mechanisms of albuminuria is desirable. P38 MAPK signaling is involved in the development of hyperglycemia-induced albuminuria. However, the mechanism of increased p38 MAPK activity leading to increased permeability and albuminuria remained unclear. Recently, we demonstrated that acute hyperglycemia triggers endocytosis of nephrin, the key molecule of the slit diaphragm, and induces albuminuria. Here, we identify p38 MAPK as a pivotal regulator of hyperglycemia-induced nephrin endocytosis. Activated p38 MAPK phosphorylates the nephrin c-terminus at serine 1146, facilitating the interaction of PKCα with nephrin. PKCα phosphorylates nephrin at threonine residues 1120 and 1125, mediating the binding of β-arrestin2 to nephrin. β-arrestin2 triggers endocytosis of nephrin by coupling it to the endocytic machinery, leading to increased glomerular permeability. Pharmacological inhibition of p38 MAPK preserves nephrin surface expression and significantly attenuates albuminuria. Key messages Acute hyperglycemia triggers endocytosis of nephrin. Activated p38 MAPK phosphorylates the nephrin c-terminus at serine 1146, facilitating the interaction of PKCα with nephrin. PKCα phosphorylates nephrin at threonine residues 1120 and 1125, mediating the binding of β-arrestin2 to nephrin. β-arrestin2 triggers endocytosis of nephrin by coupling it to the endocytic machinery, leading to a leaky glomerular filter. Pharmacological inhibition of p38 MAPK preserves nephrin surface expression and significantly attenuates albuminuria under hyperglycemic conditions.

γδ T cells are involved in the control of

Background Renal denervation (RDN) is an invasive intervention to treat drug-resistant arterial hypertension. Its therapeutic value is contentious. Here we examined the effects of RDN on inflammatory and infectious kidney disease models in mice. Methods Mice were unilaterally or bilaterally denervated, or sham operated, then three disease models were induced: nephrotoxic nephritis (NTN, a model for crescentic GN), pyelonephritis, and acute endotoxemic kidney injury (as a model for septic kidney injury). Analytical methods included measurement of renal glomerular filtration, proteinuria, flow cytometry of renal immune cells, immunofluorescence microscopy, and three-dimensional imaging of optically cleared kidney tissue by light-sheet fluorescence microscopy followed by algorithmic analysis. Results Unilateral RDN increased glomerular filtration in denervated kidneys, but decreased it in the contralateral kidneys. In the NTN model, more nephritogenic antibodies were deposited in glomeruli of denervated kidneys, resulting in stronger inflammation and injury in denervated compared with contralateral nondenervated kidneys. Also, intravenously injected LPS increased neutrophil influx and inflammation in the denervated kidneys, both after unilateral and bilateral RDN. When we induced pyelonephritis in bilaterally denervated mice, both kidneys contained less bacteria and neutrophils. In unilaterally denervated mice, pyelonephritis was attenuated and intrarenal neutrophil numbers were lower in the denervated kidneys. The nondenervated contralateral kidneys harbored more bacteria, even compared with sham-operated mice, and showed the strongest influx of neutrophils. Conclusions Our data suggest that the increased perfusion and filtration in denervated kidneys can profoundly influence concomitant inflammatory diseases. Renal deposition of circulating nephritic material is higher, and hence antibody- and endotoxin-induced kidney injury was aggravated in mice. Pyelonephritis was attenuated in denervated murine kidneys, because the higher glomerular filtration facilitated better flushing of bacteria with the urine, at the expense of contralateral, nondenervated kidneys after unilateral denervation.

Crescentic glomerulonephritis (cGN), most often caused by anti-neutrophil cytoplasmic autoantibody (ANCA)-associated vasculitis, is an aggressive form of immune-mediated kidney disease and represents an important cause of end-stage renal failure. Although it is known that T cells infiltrate the kidney in cGN, their precise role in autoimmune kidney disease remains to be fully elucidated. By performing single-cell analysis, we identified activated, clonally expanded CD8+ T cells with a cytotoxic gene expression profile in the kidneys of patients with ANCA-associated cGN. Using an experimental model of cGN, we demonstrated that clonally expanded murine CD8+ T cells highly expressed the cytotoxic molecule granzyme B. Moreover, lack of CD8+ T cells or granzyme B resulted in an ameliorated course of cGN. This was associated with reduced cleaved caspase-3 induction in renal tissue cells. Our data indicate that clonally expanded cytotoxic CD8+ T cells have a previously unrecognized pathogenic function in aggravating immune-mediated kidney disease.

Recovery-oriented perspectives have become accepted worldwide as an alternative to the biomedical approach to conceptualizing and managing severe mental health problems. It has been proposed that one advantage of this is to support self-efficacy amongst people with a lived experience of psychosis, especially when recovery messages are presented by lived experience peers. The aim of the present study was to investigate the proposed psychological benefits of the recovery paradigm, by testing for possible differential impacts of recovery versus biomedical messages on self-efficacy beliefs and positive emotional state amongst people with experience of psychosis. It was hypothesized that (1) recovery-oriented messages, when presented by lived experience peers, would generate improvements in self-efficacy and positive emotions relative to biomedical messages presented by a professional and (2) recovery-oriented messages delivered by a professional would generate improvements in self-efficacy and positive emotions relative to biomedical messages delivered by a professional. We also explored whether recovery-oriented messages were more impactful when delivered by a lived experience peer versus a professional.Experimental design with three within-subject video-based conditions.Fifty-three participants with lived experience of psychosis viewed three videos: (i) people with lived experience sharing their experiences of recovery; (ii) mental health professionals presenting traditional biomedical conceptualizations of psychosis; and (iii) mental health professionals presenting recovery perspectives. Participants provided baseline clinical and demographic information, and post-viewing ratings of experienced changes in self-efficacy and emotional state.Hypothesis 1 was supported: both self-efficacy and positive emotions were significantly increased by a video of peers sharing personal recovery stories relative to professionals presenting biomedical messages. Hypothesis 2 was partially supported: when comparing videos of recovery versus biomedical messages delivered by professionals, significant relative benefits were found for positive emotions, but not self-efficacy.This experimental investigation generated a pattern of findings broadly supportive of the assumed psychological benefits of the recovery paradigm for people with lived experiences of psychosis. Findings must be interpreted with caution given the limitations of the present design, but encourage further experimental research to directly test the interpersonal impacts of the recovery paradigm.

Introduction: Tc-99m-labeled Mas3-y-nal-k(Sub-KuE) (Tc-99m-PSMA I&S) is a prostate specific membrane antigen (PSMA) tracer that can be used for planar and SPECT/CT gamma imaging and radio-guided surgery (RGS). The primary aim of this study was to estimate the dosimetry of Tc-99m-PSMA I&S using a hybrid method (sequential gamma planar imaging and one single SPECT/CT) in healthy volunteers. The secondary aim was to depict the tracer biodistribution and tumor-to-background ratios (TBR) in patients with prostate cancer (PCa). Methods: Dosimetry of Tc-99m-PSMA I&S was investigated in four healthy volunteers. Whole-body planar imaging was acquired at 1, 2, 3, 6 and 24 hours, and SPECT/CT at 6h after tracer injection. Contours of organs were drawn on all acquisitions to determine organ activity at each timepoint. Absorbed dose was estimated using two Methods: 1) independent curve-fitting manual method (Levenberg-Marquardt-based algorithm using dose factors from Radiation Dose Assessment Resource (RADAR) web site) and 2) OLINDA/EXM® v 2.0 software (HERMES Medical Solutions). Biodistribution of Tc-99m-PSMA I&S was assessed in ten patients with PCa on SPECT/CT images at 6 h. Tumor uptake (SUVmax), and TBR (tumor SUVmax/ background organ SUVmean) using muscle (T/M), bladder (T/B) and intestine (T/I) as background organs were determined. Results: The mean injected activity of Tc-99m-PSMA I&S was 717 MBq (range: 562-828). No adverse events related to the injection of Tc-99m-PSMA-I&S were reported. The average radiation effective dose was 0.0055 mSv/MBq with the RADAR manual method and 0.0052 mSv/MBq with OLINDA/EXM®. Total body effective dose ranged between 3.33-4.42 and 3.11-4.23 mSv, respectively. All PCa patients showed high tracer uptake in primary and metastatic lesions with T/M, T/B, T/I ranging from 5.29 - 110, 0.11 – 7.02 and 0.96 – 16.30, respectively. Conclusion: Effective doses of Tc-99m-PSMA I&S were comparable to those known for most of the Tc-99m tracers and was lower than 68Ga-labelled and 18F-labelled agents. Tc-99m-PSMA I&S SPECT/CT showed high TBR in PCa patients. This study can provide required data for translation and approval of Tc-99m-PSMA I&S by regulatory agencies.

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.

A wide spectrum of immunological functions has been attributed to Interleukin 9 (IL-9), including effects on the survival and proliferation of immune and parenchymal cells. In recent years, emerging evidence suggests that IL-9 expression can promote tissue repair in inflammatory conditions. However, data about the involvement of IL-9 in kidney tissue protection is very limited. Here, we investigated the role of IL-9 in Adriamycin-induced nephropathy (AN), a mouse model for proteinuric chronic kidney disease. Compared to wild type mice, IL-9 knockout (Il9−/−) mice with AN displayed accelerated development of proteinuria, aggravated glomerulosclerosis and deterioration of kidney function. At an early stage of disease, the Il9−/− mice already displayed a higher extent of glomerular podocyte injury and loss of podocyte number compared to wild type mice. In the kidney, T cells and innate lymphoid cells produced IL-9. However, selective deficiency of IL-9 in the innate immune system in Il9−/−Rag2−/− mice that lack T and B cells did not alter the outcome of AN, indicating that IL-9 derived from the adaptive immune system was the major driver of tissue protection in this model. Mechanistically, we could show that podocytes expressed the IL-9 receptor in vivo and that IL-9 signaling protects podocytes from Adriamycin-induced apoptosis in vitro. Finally, in vivo treatment with IL-9 effectively protected wild type mice from glomerulosclerosis and kidney failure in the AN model. The detection of increased serum IL-9 levels in patients with primary focal and segmental glomerulosclerosis further suggests that IL-9 production is induced by glomerular injury in humans. Thus, IL-9 confers protection against experimental glomerulosclerosis, identifying the IL-9 pathway as a potential therapeutic target in proteinuric chronic kidney disease.

The phospholipase A2 receptor 1 (PLA2R1) is the major autoantigen in patients suffering from membranous nephropathy. To date, the lack of endogenous glomerular expression of PLA2R1 in mice and rats has impeded the establishment of PLA2R1-dependent animal models of this disease. Here, we generated a transgenic mouse line expressing murine full-length PLA2R1 in podocytes. Furthermore, expression of murine PLA2R1 did not result in any morphological disturbance as high-resolution confocal microscopy demonstrated an intact nephrin distribution with normal foot processes. Transfer of rabbit anti-mPLA2R1 antibodies to these mice induced nephrotic range proteinuria, hypercholesterolemia, and histomorphological signs of membranous nephropathy. Immunohistochemical and immunofluorescence analyses revealed enhanced staining for murine PLA2R1 in the presence of unaffected staining for murine thrombospondin type-1 domain-containing 7A in the diseased mice, resembling what is classically found in patients with PLA2R1-associated membranous nephropathy Thus, our mouse model of membranous nephropathy will allow investigation of PLA2R1-specific pathomechanisms and may help to develop and assess antigen-specific treatments in vivo.

Anti-Thy1.1 transgenic mice develop glomerular lesions that mimic collapsing focal segmental glomerulosclerosis (FSGS) in humans with collapse of the glomerular tuft and marked hyperplasia of the parietal epithelial cells (PECs). Immunostaining of phosphor-S6 ribosomal protein (pS6RP) revealed high mTOR activity in PECs of the FSGS lesions of these mice. In this study we questioned whether the mTOR inhibitor rapamycin (sirolimus) could attenuate the development and progression of glomerulosclerotic lesions in the anti-Thy1.1 transgenic mice. We observed reduced mTOR signalling and proliferation in human parietal epithelial cells after rapamycin treatment. Experiments with anti-Thy1.1. mice showed that early treatment with sirolimus reduced the development of glomerular lesions and glomerular cell proliferation at day 4. Levels of albuminuria, podocyte injury and podocyte number were similar in the sirolimus and vehicle treated groups. The initial beneficial effects of sirolimus treatment were not observed at day 7. Late sirolimus treatment did not reduce albuminuria or the progression of glomerulosclerosis. Taken together, rapamycin attenuated PEC proliferation and the formation of early FSGS lesions in experimental FSGS and reduced human PEC proliferation in vitro. However, the initial inhibition of PEC proliferation did not translate into a decline of albuminuria nor in a sustained reduction in sclerotic lesions.

Mucopolysaccharidosis type VI (MPS-VI), caused by mutational inactivation of the glycosaminoglycan-degrading enzyme arylsulfatase B (Arsb), is a lysosomal storage disorder primarily affecting the skeleton. We have previously reported that Arsb-deficient mice display high trabecular bone mass and impaired skeletal growth. In the present study, we treated them by weekly injection of recombinant human ARSB (rhARSB) to analyze the impact of enzyme replacement therapy (ERT) on skeletal growth and bone remodeling. We found that all bone-remodeling abnormalities of Arsb-deficient mice were prevented by ERT, whereas chondrocyte defects were not. Likewise, histologic analysis of the surgically removed femoral head from an ERT-treated MPS-VI patient revealed that only chondrocytes were pathologically affected. Remarkably, a side-by-side comparison with other cell types demonstrated that chondrocytes have substantially reduced capacity to endocytose rhARSB, together with low expression of the mannose receptor. We finally took advantage of Arsb-deficient mice to establish quantification of chondroitin sulfation for treatment monitoring. Our data demonstrate that bone-remodeling cell types are accessible to systemically delivered rhARSB, whereas the uptake into chondrocytes is inefficient. ispartof: HUMAN MOLECULAR GENETICS vol:29 issue:5 pages:803-816 ispartof: location:England status: published

Background The glomerulus comprises podocytes, mesangial cells, and endothelial cells, which jointly determine glomerular filtration. Understanding this intricate functional unit beyond the transcriptome requires bulk isolation of these cell types for biochemical investigations. We developed a globally applicable tripartite isolation method for murine mesangial and endothelial cells and podocytes (timMEP). Methods We separated glomerular cell types from wild-type or mT/mG mice via a novel FACS approach, and validated their purity. Cell type proteomes were compared between strains, ages, and sex. We applied timMEP to the podocyte-targeting, immunologic, THSD7A-associated, model of membranous nephropathy. Results timMEP enabled protein-biochemical analyses of podocytes, mesangial cells, and endothelial cells derived from reporter-free mice, and allowed for the characterization of podocyte, endothelial, and mesangial proteomes of individual mice. We identified marker proteins for mesangial and endothelial proteins, and outlined protein-based, potential communication networks and phosphorylation patterns. The analysis detected cell type-specific proteome differences between mouse strains and alterations depending on sex, age, and transgene. After exposure to anti-THSD7A antibodies, timMEP resolved a fine-tuned initial stress response, chiefly in podocytes, that could not be detected by bulk glomerular analyses. The combination of proteomics with super-resolution imaging revealed a specific loss of slit diaphragm, but not of other foot process proteins, unraveling a protein-based mechanism of podocyte injury in this animal model. Conclusion timMEP enables glomerular cell type-resolved investigations at the transcriptional and protein-biochemical level in health and disease, while avoiding reporter-based artifacts, paving the way toward the comprehensive and systematic characterization of glomerular cell biology.

The deubiquitinating enzyme ubiquitin C-terminal hydrolase-L1 (UCH-L1) is required for the maintenance of axonal integrity in neurons and is thought to regulate the intracellular pool of ubiquitin in the brain. In this study, we show that UCH-L1 has an immunological function in dendritic cell (DC) Ag cross-presentation. UCH-L1 is expressed in mouse kidney, spleen, and bone marrow–derived DCs, and its expression and activity are regulated by the immune stimuli LPS and IFN-γ. UCH-L1–deficient mice have significantly reduced ability to cross-prime CD8 T cells in vivo and in vitro because of a reduced ability of DCs to generate MHC class I (MHC I) peptide complexes for cross-presented Ags. Mechanistically, Ag uptake by phagocytosis and receptor-mediated endocytosis as well as phagosome maturation are unaffected by loss of UCH-L1 in DCs. Rather, MHC I recycling is reduced by loss of UCH-L1, which affects the colocalization of intracellular MHC I with late endosomal/lysosomal compartments necessary for cross-presentation of Ag. These results demonstrate a hitherto unrecognized role of the deubiquitinating enzyme UCH-L1 in DC Ag processing.

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.

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.

Background About 3%–5% of adults with membranous nephropathy have autoantibodies directed against thrombospondin type 1 domain–containing 7A (THSD7A), a podocyte-expressed transmembrane protein. However, the temporal and spatial expression of THSD7A and its biologic function for podocytes are unknown, information that is needed to understand the effects of THSD7A autoantibodies in this disease. Methods Using a variety of microscopic techniques, we analyzed THSD7A localization in postnatal, adult, and autoantibody-injected mice as well as in human podocytes. We also analyzed THSD7A function in human podocytes using confocal microscopy; Western blotting; and adhesion and migration assays. Results We found that THSD7A expression begins on glomerular vascularization with slit diaphragm formation in development. THSD7A localizes to the basal aspect of foot processes, closely following the meanders of the slit diaphragm in human and mice. Autoantibodies binding to THSD7A localize to the slit diaphragm. In human podocytes, THSD7A expression is accentuated at filopodia and thin arborized protrusions, an expression pattern associated with decreased membrane activity of cytoskeletal regulators. We also found that, phenotypically, THSD7A expression in human podocytes is associated not only with increases in cell size, enhanced adhesion, and reduced detachment from collagen type IV–coated plates but also, with decreased ability to migrate. Conclusions Our findings suggest that THSD7A functions as a foot process protein involved in the stabilization of the slit diaphragm of mature podocytes and that autoantibodies to THSD7A, on the basis of their localization, might structurally and functionally alter the slit diaphragm’s permeability to protein.

In recent years, the cytokine interleukin (IL)-22 attracted considerable attention due to its important immunoregulatory function in barrier tissues, such as the gut, lung, and skin. Although a regenerative role of IL-22 in renal tubular damage has been demonstrated, the role of IL-22 in the immunopathogenesis of glomerular injury is still unknown. Here, we demonstrate that the IL-22 receptor is expressed in the glomerular compartment of the kidney and that IL-22 expression increases in the renal cortex after induction of glomerular injury in a mouse model for crescentic glomerulonephritis (cGN, nephrotoxic nephritis). We identified γδ T cells and TH17 cells as major sources for IL-22 in the nephritic kidney. However, neither genetic or antibody-mediated deletion of IL-22 nor genetic deficiency in its endogenous inhibitor IL-22Rα2 (IL-22 binding protein) resulted in substantial phenotypic differences in mice with cGN with respect to crescent formation, tubulointerstitial damage, and kidney function impairment. Similarly, we did not observe significant differences between wild-type or IL-22-deficient mice in a mouse model of secondary focal and segmental glomerulosclerosis (adriamycin-induced nephropathy). As shown previously, we detected concomitant upregulation of IL-17A and IFN-γ production by T cells during the course of cGN, providing alternative cytokine pathways that mediate glomerular injury in this model. In conclusion, we show here that endogenous IL-22 expression is redundant in different forms of glomerular injury, indicating that the IL-22-directed therapies that are being tested in various human diseases might not affect the kidney in patients with glomerular disease.