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The volume expansion behavior of low-density polypropylene foams in extrusion is investigated in this paper. Since escape of blowing agent from the foam would cause the foam to contract, and to have low expansion, efforts were made to prevent gas loss during foaming. The basic strategies to the promotion of a large volume expansion ratio are: to use a branched material for preventing cell coalescence; to use a long-chain blowing agent with low diffusivity; to lower the melt temperature for decreasing gas loss during expansion; and to optimize the processing conditions in the die for avoiding too-rapid crystallization. Use of a branched polypropylene resin was required to achieve large volume expansion because prevention of cell coalescence will retard gas loss from the extruded foam to the environment. The foam morphologies of linear and branched polypropylene materials at various processing temperatures were studied using a single-screw tandem foam extrusion system and their volume expansion behaviors were c ompared. Ultra low-density, fine-celled polypropylene foams with very high expansion ratio up to 90-fold were successfully produced from the branched polypropylene resins., INTRODUCTION Thermoplastic foams possess a cellular structure create y the expansion of blowing agent. This internal structure provides unique properties that enable the foamed plastics to be used effectively for [...]

The present study examined the pathogenesis of interstitial inflammation and fibrosis in antihypertensively treated rats with two-kidney, one-clip hypertension. Hypertensive rats were randomized into four groups: no treatment and moderate, intermediate, and intensified lowering of blood pressure with increasing doses of a vasopeptidase inhibitor for 6 wk. The vasopeptidase inhibitor dose dependently lowered blood pressure. The tubulointerstitial damage was accompanied by a diffuse infiltration of mononuclear cells and circumscript mononuclear inflammatory cell cluster formation consisting mainly of T cells and to a lesser degree of macrophages and B cells. Real-time PCR analyses showed a dose-dependent induction of MCP-1 and the Th1-type chemokines IP10 and Mig as well as their receptor CXCR3 and the Th1 cytokine IFN-γ. In situ hybridization and laser microdissection revealed a strong expression of these Th1-associated transcripts in the clusters and, in the case of MCP-1, also diffusely in the interstitium. The inflammation was accompanied by the appearance of myofibroblasts and synthesis of the fibrogenic factor plasminogen activator inhibitor-1 as well as the collagenase matrix metalloproteinase-2, leading to collagen 1 upregulation and interstitial scarring. No inflammation or fibrosis was found in normotensive rats treated with the vasopeptidase inhibitor. The renal injury in the clipped kidney is accompanied by compartment-specific chemokine expression and cell cluster formation of Th1 specificity associated with upregulation of fibrogenic proteins and matrix metalloproteinases. These findings suggest that the Th1 chemokines IP10 and Mig as well as their receptor CXCR3 are potential targets for therapeutic interventions in ischemic nephropathy. [ABSTRACT FROM AUTHOR]

Autoimmunity against the Goodpasture antigen [alpha]3IV-NC1 results in antiglomerular basement membrane glomerulonephritis. Although antibodies are central to the pathogenesis, there is good evidence for the participation of T cells in this disease. To define the contribution of T cells, we used the model of experimental autoimmune glomerulonephritis. Immunization of DBA/1 mice with [alpha]3IV-NC1 resulted in proteinuria, a biphasic course of the disease, and an eventual loss of kidney function. In the initial phase, the mice developed an [alpha]3IV-NC1-specific IgG response, had IgG deposition along the glomerular basement membrane, and steadily increased proteinuria, but only marginal signs of inflammation with limited leukocyte infiltration. After 9-13 weeks, mice proceeded to develop crescentic glomerulonephritis, extensive tubulointerstitial damage, and massive macrophage infiltration. T-cell infiltration was less pronounced, mostly confined to the interstitium, and T cells displayed an activated phenotype with a significant fraction of Th1 or Th17 CD4(+) T cells. Close examination revealed the presence of autoreactive T cells producing IFN[gamma] upon restimulation with [alpha]3IV-NC1. Thus, our results suggest that accumulation of effector T cells, including autoreactive T cells, represents a critical step in the progression from mild glomerulonephritis, with limited glomerular damage, to severe crescentic glomerulonephritis accompanied by tubulointerstitial inflammation and loss of kidney function. [ABSTRACT FROM AUTHOR]

Crescentic glomerulonephritis is mediated by inappropriate humoral and cellular immune responses toward self-antigens that may result from defects in central and peripheral tolerance. Evidence now suggests that regulatory T cells (Tregs) may be of pathophysiological importance in proliferative and crescentic forms of glomerulonephritis. To analyze the role of endogenous Tregs in a T cell-dependent glomerulonephritis model of nephrotoxic nephritis, we used 'depletion of regulatory T cell' (DEREG) mice that express the diphtheria toxin receptor under control of the FoxP3 (forkhead box P3) gene promoter. Toxin injection into these mice efficiently depleted renal and splenic FoxP3(+) Treg cells as determined by fluorescent-activated cell sorting (FACS) and immunohistochemical analyses. Treg depletion exacerbated systemic and renal interferon-γ (IFNγ) expression and increased recruitment of IFNγ-producing Th1 cells into the kidney without an effect on the Th17 immune response. The enhanced Th1 response, following Treg cell depletion, was associated with an aggravated course of glomerulonephritis as measured by glomerular crescent formation. Thus, our results establish the functional importance of endogenous Tregs in the control of a significantly enhanced systemic and renal Th1 immune response in experimental glomerulonephritis. [ABSTRACT FROM AUTHOR]

The evaluation of compound-target interactions (CTIs) is at the heart of drug discovery efforts. Given the substantial time and monetary costs of classical experimental screening, significant efforts have been dedicated to develop deep learning-based models that can accurately predict CTIs. A comprehensive comparison of these models on a large, curated CTI dataset is, however, still lacking. Here, we perform an in-depth comparison of 12 state-of-the-art deep learning architectures that use different protein and compound representations. The models were selected for their reported performance and architectures. To reliably compare model performance, we curated over 300 thousand binding and non-binding CTIs and established several gold-standard datasets of varying size and information. Based on our findings, DeepConv-DTI consistently outperforms other models in CTI prediction performance across the majority of datasets. It achieves an MCC of 0.6 or higher for most of the datasets and is one of the fastest models in training and inference. These results indicate that utilizing convolutional-based windows as in DeepConv-DTI to traverse trainable embeddings is a highly effective approach for capturing informative protein features. We also observed that physicochemical embeddings of targets increased model performance. We therefore modified DeepConv-DTI to include normalized physicochemical properties, which resulted in the overall best performing model Phys-DeepConv-DTI. This work highlights how the systematic evaluation of input features of compounds and targets, as well as their corresponding neural network architectures, can serve as a roadmap for the future development of improved CTI models.Scientific contributionThis work features comprehensive CTI datasets to allow for the objective comparison and benchmarking of CTI prediction algorithms. Based on this dataset, we gained insights into which embeddings of compounds and targets and which deep learning-based algorithms perform best, providing a blueprint for the future development of CTI algorithms. Using the insights gained from this screen, we provide a novel CTI algorithm with state-of-the-art performance., (© 2024. The Author(s).)

Th17 cells play crucial roles in host defense and the pathogenesis of autoimmune diseases in the skin. While their differentiation mechanisms have been extensively studied, the origin of skin Th17 cells remains unclear. In this study, we analyzed single-cell RNA-sequencing data and identify the presence of Th17 cells in the human thymus. Thymic Th17 cells were characterized by high expression levels of Sphingosine-1-Phosphate Receptor 1 (S1PR1), a receptor crucial for T cell egress from lymphoid tissues. In mice, Th17 cell-specific knockout of S1pr1 resulted in the accumulation of Th17 cells in the thymus and a corresponding decrease in their numbers in the skin. Th17 cells that accumulated in the thymus exhibited a lower IL-17A production capacity compared to those in the skin, indicating that the local environment in the skin is important for maintaining the Th17 cell phenotype. Additionally, using a murine psoriasis model, we demonstrated that Th17 cell-specific knockout of S1pr1 reduced their migration to the inflamed skin, thereby ameliorating disease progression. Collectively, our data suggest that S1PR1 mediates Th17 cell migration from the thymus to the skin, thereby modulating their functional engagement in both homeostatic and inflammatory conditions., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Engesser, Wang, Kapffer, Kaffke, Peters, Paust, Geissen, Krebs, Panzer and Asada.)

Antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis is a life-threatening autoimmune disease that often results in kidney failure caused by crescentic glomerulonephritis (GN). To date, treatment of most patients with ANCA-GN relies on non-specific immunosuppressive agents, which may have serious adverse effects and be only partially effective. Here, using spatial and single-cell transcriptome analysis, we characterize inflammatory niches in kidney samples from 34 patients with ANCA-GN and identify proinflammatory, cytokine-producing CD4 + and CD8 + T cells as a pathogenic signature. We then utilize these transcriptomic profiles for digital pharmacology and identify ustekinumab, a monoclonal antibody targeting IL-12 and IL-23, as the strongest therapeutic drug to use. Moreover, four patients with relapsing ANCA-GN are treated with ustekinumab in combination with low-dose cyclophosphamide and steroids, with ustekinumab given subcutaneously (90 mg) at weeks 0, 4, 12, and 24. Patients are followed up for 26 weeks to find this treatment well-tolerated and inducing clinical responses, including improved kidney function and Birmingham Vasculitis Activity Score, in all ANCA-GN patients. Our findings thus suggest that targeting of pathogenic T cells in ANCA-GN patients with ustekinumab might represent a potential approach and warrants further investigation in clinical trials., (© 2024. The Author(s).)

Autoantibody-mediated glomerulonephritis (AGN) arises from dysregulated renal inflammation, with urgent need for improved treatments. IL-17 is implicated in AGN and drives pathology in a kidney-intrinsic manner via renal tubular epithelial cells (RTECs). Nonetheless, downstream signaling mechanisms provoking kidney pathology are poorly understood. A noncanonical RNA binding protein (RBP), Arid5a, was upregulated in human and mouse AGN. Arid5a-/- mice were refractory to AGN, with attenuated myeloid infiltration and impaired expression of IL-17-dependent cytokines and transcription factors (C/EBPβ, C/EBPδ). Transcriptome-wide RIP-Seq revealed that Arid5a inducibly interacts with conventional IL-17 target mRNAs, including CEBPB and CEBPD. Unexpectedly, many Arid5a RNA targets corresponded to translational regulation and RNA processing pathways, including rRNAs. Indeed, global protein synthesis was repressed in Arid5a-deficient cells, and C/EBPs were controlled at the level of protein rather than RNA accumulation. IL-17 prompted Arid5a nuclear export and association with 18S rRNA, a 40S ribosome constituent. Accordingly, IL-17-dependent renal autoimmunity is driven by Arid5a at the level of ribosome interactions and translation., (© 2024 Li et al.)

Adaptation of immune cells to tissue-specific microenvironments is a crucial process in homeostasis and inflammation. Here, we show that murine effector type 2 innate lymphoid cells (ILC2s) from various organs are equally effective in repopulating ILC2 niches in other anatomical locations where they adapt tissue-specific phenotypes of target organs. Single-cell transcriptomics of ILC2 populations revealed upregulation of retinoic acid (RA) signaling in ILC2s during adaptation to the small intestinal microenvironment, and RA signaling mediated reprogramming of kidney effector ILC2s toward the small intestinal phenotype in vitro and in vivo. Inhibition of intestinal ILC2 adaptation by blocking RA signaling impaired worm expulsion during Strongyloides ratti infection, indicating functional importance of ILC2 tissue imprinting. In conclusion, this study highlights that effector ILC2s retain the ability to adapt to changing tissue-specific microenvironments, enabling them to exert tissue-specific functions, such as promoting control of intestinal helminth infections., (© 2023 Shaikh et al.)

Mucosal-associated invariant T (MAIT) cells have been implicated in various inflammatory diseases of barrier organs, but so far, their role in kidney disease is unclear. Here we report that MAIT cells that recognize their prototypical ligand, the vitamin B2 intermediate 5-OP-RU presented by MR1, reside in human and mouse kidneys. Single cell RNAseq analysis reveals several intrarenal MAIT subsets, and one, carrying the genetic fingerprint of tissue-resident MAIT17 cells, is activated and expanded in a murine model of crescentic glomerulonephritis (cGN). An equivalent subset is also present in kidney biopsies of patients with anti-neutrophil cytoplasmatic antibody (ANCA)-associated cGN. MAIT cell-deficient MR1 mice show aggravated disease, whereas B6-MAIT CAST mice, harboring higher MAIT cell numbers, are protected from cGN. The expanded MAIT17 cells express anti-inflammatory mediators known to suppress cGN, such as CTLA-4, PD-1, and TGF-β. Interactome analysis predicts CXCR6 - CXCL16-mediated cross-talk with renal mononuclear phagocytes, known to drive cGN progression. In line, we find that cGN is aggravated upon CXCL16 blockade. Finally, we present an optimized 5-OP-RU synthesis method which we apply to attenuating cGN in mice. In summary, we propose that CXCR6 + MAIT cells might play a protective role in cGN, implicating them as a potential target for anti-inflammatory therapies., (© 2023. The Author(s).)

Significance Statement: T-cell infiltration is a hallmark of crescentic GN (cGN), often caused by ANCA-associated vasculitis. Pathogenic T-cell subsets, their clonality, and downstream effector mechanisms leading to kidney injury remain to be fully elucidated. Single-cell RNA sequencing and T-cell receptor sequencing revealed activated, clonally expanded cytotoxic CD4 + and CD8 + T cells in kidneys from patients with ANCA-associated cGN. In experimental cGN, kidney-infiltrating CD8 + T cells expressed the cytotoxic molecule, granzyme B (GzmB), which induced apoptosis in renal tissue cells by activation of procaspase-3, and aggravated disease pathology. These findings describe a pathogenic function of (clonally expanded) cytotoxic T cells in cGN and identify GzmB as a mediator and potential therapeutic target in immune-mediated kidney disease., Background: Crescentic GN (cGN) is an aggressive form of immune-mediated kidney disease that is an important cause of end stage renal failure. Antineutrophilic cytoplasmic antibody (ANCA)-associated vasculitis is a common cause. T cells infiltrate the kidney in cGN, but their precise role in autoimmunity is not known., Methods: Combined single-cell RNA sequencing and single-cell T-cell receptor sequencing were conducted on CD3 + T cells isolated from renal biopsies and blood of patients with ANCA-associated cGN and from kidneys of mice with experimental cGN. Functional and histopathological analyses were performed with Cd8a-/- and GzmB-/- mice., Results: Single-cell analyses identified activated, clonally expanded CD8 + and CD4 + T cells with a cytotoxic gene expression profile in the kidneys of patients with ANCA-associated cGN. Clonally expanded CD8 + T cells expressed the cytotoxic molecule, granzyme B (GzmB), in the mouse model of cGN. Deficiency of CD8 + T cells or GzmB ameliorated the course of cGN. CD8 + T cells promoted macrophage infiltration and GzmB activated procaspase-3 in renal tissue cells, thereby increasing kidney injury., Conclusions: Clonally expanded cytotoxic T cells have a pathogenic function in immune-mediated kidney disease., (Copyright © 2023 by the American Society of Nephrology.)

Ischemia-reperfusion injury (IRI) is one of the major causes of acute kidney injury (AKI), and experimental work has revealed detailed insight into the inflammatory response in the kidney. T cells and NFκB pathway play an important role in IRI. Therefore, we examined the regulatory role and mechanisms of IkappaB kinase 1 (IKK1) in CD4 + T lymphocytes in an experimental model of IRI. IRI was induced in CD4cre and CD4IKK1Δ mice. Compared to control mice, conditional deficiency of IKK1 in CD4 + T lymphocyte significantly decreased serum creatinine, blood urea nitrogen (BUN) level, and renal tubular injury score. Mechanistically, lack in IKK1 in CD4 + T lymphocytes reduced the ability of CD4 lymphocytes to differentiate into Th1/Th17 cells. Similar to IKK1 gene ablation, pharmacological inhibition of IKK also protected mice from IRI. Together, lymphocyte IKK1 plays a pivotal role in IRI by promoting T cells differentiation into Th1/Th17 and targeting lymphocyte IKK1 may be a novel therapeutic strategy for IRI., (© 2023. The Author(s).)

Membranous nephropathy (MN) is an antibody-mediated autoimmune disease characterized by glomerular immune complexes containing complement components. However, both the initiation pathways and the pathogenic significance of complement activation in MN are poorly understood. Here, we show that components from all three complement pathways (alternative, classical and lectin) are found in renal biopsies from patients with MN. Proximity ligation assays to directly visualize complement assembly in the tissue reveal dominant activation via the classical pathway, with a close correlation to the degree of glomerular C1q-binding IgG subclasses. In an antigen-specific autoimmune mouse model of MN, glomerular damage and proteinuria are reduced in complement-deficient mice compared with wild-type littermates. Severe disease with progressive ascites, accompanied by extensive loss of the integral podocyte slit diaphragm proteins, nephrin and neph1, only occur in wild-type animals. Finally, targeted silencing of C3 using RNA interference after the onset of proteinuria significantly attenuates disease. Our study shows that, in MN, complement is primarily activated via the classical pathway and targeting complement components such as C3 may represent a promising therapeutic strategy., (© 2023. The Author(s).)

The discovery of tissue-resident memory T cells (T RM cells) reinterpreted the potential of human tissue-specific immunity. Following T cell receptor (TCR) activation and clonal expansion, effector T cells migrate to peripheral tissues where they remain long-term and differentiate to T RM cells after antigen clearance. This allows for prompt immunological responses upon antigen re-encounter. In addition to their protective properties in acute infections, recent studies have revealed that T RM cells might lead to aggravation of autoimmune diseases, such as lupus nephritis (LN) and anti-neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis (GN). These diseases present as proliferative and crescentic glomerulonephritis (cGN), which is a life-threatening condition leading to end-stage renal disease (ESRD) if left untreated. A better understanding of renal T RM cells might lead to identifying new therapeutic targets for relapsing autoimmune diseases of the kidney. In this review, we summarize the current knowledge of renal T RM cells and discuss their potential pathophysiological roles in renal autoimmune diseases., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Ginsberg, Panzer and Asada.)

Glucocorticoids remain a cornerstone of therapeutic regimes for autoimmune and chronic inflammatory diseases - for example, in different forms of crescentic glomerulonephritis - because of their rapid antiinflammatory effects, low cost, and wide availability. Despite their routine use for decades, the underlying cellular mechanisms by which steroids exert their therapeutic effects need to be fully elucidated. Here, we demonstrate that high-dose steroid treatment rapidly reduced the number of proinflammatory CXCR3+CD4+ T cells in the kidney by combining high-dimensional single-cell and morphological analyses of kidney biopsies from patients with antineutrophil cytoplasmic antibody-associated (ANCA-associated) crescentic glomerulonephritis. Using an experimental model of crescentic glomerulonephritis, we show that the steroid-induced decrease in renal CD4+ T cells is a consequence of reduced T cell recruitment, which is associated with an ameliorated disease course. Mechanistic in vivo and in vitro studies revealed that steroids act directly on renal tissue cells, such as tubular epithelial cells, but not on T cells, which resulted in an abolished renal expression of CXCL9 and CXCL10 as well as in the prevention of CXCR3+CD4+ T cell recruitment to the inflamed kidneys. Thus, we identified the CXCL9/CXCL10-CXCR3 axis as a previously unrecognized cellular and molecular target of glucocorticoids providing protection from immune-mediated pathology.

γδ T cells are involved in the control of Staphylococcus aureus infection, but their importance in protection compared to other T cells is unclear. We used a mouse model of systemic S. aureus infection associated with high bacterial load and persistence in the kidney. Infection caused fulminant accumulation of γδ T cells in the kidney. Renal γδ T cells acquired tissue residency and were maintained in high numbers during chronic infection. At day 7, up to 50% of renal γδ T cells produced IL-17A in situ and a large fraction of renal γδ T cells remained IL-17A + during chronic infection. Controlled depletion revealed that γδ T cells restricted renal S. aureus replication in the acute infection and provided protection during chronic renal infection and upon reinfection. Our results demonstrate that kidney-resident γδ T cells are nonredundant in limiting local S. aureus growth during chronic infection and provide enhanced protection against reinfection.

Staphylococcus aureus is frequently detected in patients with sepsis and thus represents a major health burden worldwide. CD4+ T helper cells are involved in the immune response to S. aureus by supporting antibody production and phagocytosis. In particular, Th1 and Th17 cells secreting IFN-γ and IL-17A, are involved in the control of systemic S. aureus infections in humans and mice. To investigate the role of T cells in severe S. aureus infections, we established a mouse sepsis model in which the kidney was identified to be the organ with the highest bacterial load and abundance of Th17 cells. In this model, IL-17A but not IFN-γ was required for bacterial control. Using Il17aCre × R26YFP mice we could show that Th17 fate cells produce Th17 and Th1 cytokines, indicating a high degree of Th17 cell plasticity. Single cell RNA-sequencing of renal Th17 fate cells uncovered their heterogeneity and identified a cluster with a Th1 expression profile within the Th17 cell population, which was absent in mice with T-bet/Tbx21-deficiency in Th17 cells (Il17aCre x R26eYFP x Tbx21-flox). Blocking Th17 to Th1 transdifferentiation in Th17 fate cells in these mice resulted in increased S. aureus tissue loads. In summary, we highlight the impact of Th17 cells in controlling systemic S. aureus infections and show that T-bet expression by Th17 cells is required for bacterial clearance. While targeting the Th17 cell immune response is an important therapeutic option in autoimmunity, silencing Th17 cells might have detrimental effects in bacterial infections., Competing Interests: The authors have declared that no competing interests exist.

Background: IL-17A-producing CD4 + T helper (T H 17) cells play a critical role in autoimmune and chronic inflammatory diseases, such as crescentic GN. The proinflammatory effects of IL-17 are mediated by the activation of the IL-17RA/IL-17RC complex. Although the expression of these receptors on epithelial and endothelial cells is well characterized, the IL-17 receptor expression pattern and function on hematopoietic cells, e.g. , CD4 + T cell subsets, remains to be elucidated., Methods: Crescentic GN (nephrotoxic nephritis) was induced in IL-17A, IFN γ , and Foxp3 triple-reporter mice for sorting of renal CD4 + T cell subsets and subsequent single-cell RNA sequencing. Moreover, we generated T H 17 cell-specific IL-17RA and IL-17RC gene-deficient mice and studied the functional role of IL-17 signaling in T H 17 cells in crescentic GN, imiquimod-induced psoriasis, and in the CD4 + CD45RB high T cell transfer colitis model., Results: We identified a specific expression of the IL-17 receptor A/C complex on CD4 + T H 17 cells. Single-cell RNA sequencing of T H 17 cells revealed the activation of the IL-17 receptor signaling pathway in experimental crescentic GN. Disruption of the IL-17RC signaling pathway in CD4 + T cells and, most importantly, specifically in CD4 + T H 17 cells, potentiates the IL-17 cytokine response and results in an accelerated course of experimental crescentic GN. Comparable results were observed in experimental models of psoriasis and colitis., Conclusions: Our findings indicate that IL-17 receptor C signaling has a previously unrecognized function in the regulation of CD4 + T H 17 cells and in the control of organ-specific autoimmunity and might provide new insights into the development of more efficient anti-T H 17 treatment strategies., (Copyright © 2021 by the American Society of Nephrology.)

Immune-mediated kidney diseases are a leading cause of end-stage renal disease. Despite recent discoveries, the immunopathogenesis of this heterogeneous disease group remains incompletely understood, which is a major reason for the lack of specific therapies and targeted interventions. Accumulating evidence suggests that cytokines related to the T cell response play an important role in renal autoimmunity. In this issue of the JCI, Li et al. demonstrate that IL-23 directly regulates the metabolism of parenchymal kidney cells, thereby generating a proinflammatory microenvironment that exacerbates T cell-driven renal tissue damage. These findings identify the IL-23/IL-17 axis as a key mediator of renal tissue injury and open new avenues for the development of pathogenesis-based treatment strategies in renal inflammatory diseases.

BK polyomavirus-associated nephropathy is a common complication after kidney transplantation leading to reduced graft function or loss. The molecular pathogenesis of BK polyomavirus-induced nephropathy is not well understood. A recent study had described a protective effect of the activating natural killer cell receptor KIR3DS1 in BK polyomavirus-associated nephropathy, suggesting a role of NK cells in modulating disease progression. Using an in vitro cell culture model of human BK polyomavirus infection and kidney biopsy samples from patients with BK polyomavirus-associated nephropathy, we observed significantly increased surface expression of the ligand for KIR3DS1, HLA-F, on BK polyomavirus-infected kidney tubular cells. Upregulation of HLA-F expression resulted in significantly increased binding of KIR3DS1 to BK polyomavirus-infected cells and activation of primary KIR3DS-positive natural killer cells. Thus, our data provide a mechanism by which KIR3DS-positive natural killer cells can control BK polyomavirus infection of the kidney, and rationale for exploring HLA-F/KIR3DS1 interactions for immunotherapeutic approaches in BK polyomavirus-associated nephropathy., (Copyright © 2020 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)

Morphologic examination of tissue biopsies is essential for histopathological diagnosis. However, accurate and scalable cellular quantification in human samples remains challenging. Here, we present a deep learning-based approach for antigen-specific cellular morphometrics in human kidney biopsies, which combines indirect immunofluorescence imaging with U-Net-based architectures for image-to-image translation and dual segmentation tasks, achieving human-level accuracy. In the kidney, podocyte loss represents a hallmark of glomerular injury and can be estimated in diagnostic biopsies. Thus, we profiled over 27,000 podocytes from 110 human samples, including patients with antineutrophil cytoplasmic antibody-associated glomerulonephritis (ANCA-GN), an immune-mediated disease with aggressive glomerular damage and irreversible loss of kidney function. We identified previously unknown morphometric signatures of podocyte depletion in patients with ANCA-GN, which allowed patient classification and, in combination with routine clinical tools, showed potential for risk stratification. Our approach enables robust and scalable molecular morphometric analysis of human tissues, yielding deeper biological insights into the human kidney pathophysiology.

The randomized, controlled STOP-IgAN trial in patients with IgA nephropathy (IgAN) and substantial proteinuria showed no benefit of immunosuppression added on top of supportive care on renal function over three years. As a follow-up we evaluated renal outcomes in patients over a follow-up of up to ten years in terms of serum creatinine, proteinuria, end-stage kidney disease (ESKD), and death. The adapted primary endpoint was the time to first occurrence of a composite of death, ESKD, or a decline of over 40% in the estimated glomerular filtration rate (eGFR) compared to baseline at randomization into STOP-IgAN. Data were analyzed by Cox-regression models. Follow-up data were available for 149 participants, representing 92% of the patients originally randomized. Median follow-up was 7.4 years (inter quartile range 5.7 to 8.3 years). The primary endpoint was reached in 36 of 72 patients randomized to supportive care and 35 of 77 patients randomized to additional immunosuppression (hazard ratio 1.20; 95% confidence interval 0.75 to 1.92). ESKD occurred in 17 of the patients with supportive care and in 20 of the patients with additional immunosuppression. Additionally, the rates of eGFR loss over 40% and annual eGFR loss did not differ between groups. Two patients died with supportive care and three with additional immunosuppression. Thus, within the limitations of a retrospective study, over a follow-up of up to ten years, and using an adapted primary endpoint, we failed to detect differences in key clinical outcomes in IgAN patients randomized to receive added immunosuppression on top of supportive care versus supportive care alone., (Copyright © 2020 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)

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., (Copyright © 2020 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)

Background: Recent studies have identified the EGF receptor (EGFR) ligand amphiregulin (AREG) as an important mediator of inflammatory diseases. Both pro- and anti-inflammatory functions have been described, but the role of AREG in GN remains unknown., Methods: The nephrotoxic nephritis model of GN was studied in AREG -/- mice after bone marrow transplantation, and in mice with myeloid cell-specific EGFR deficiency. Therapeutic utility of AREG neutralization was assessed. Furthermore, AREG's effects on renal cells and monocytes/macrophages (M/M) were analyzed. Finally, we evaluated AREG expression in human renal biopsies., Results: Renal AREG mRNA was strongly upregulated in murine GN. Renal resident cells were the most functionally relevant source of AREG. Importantly, the observation that knockout mice showed significant amelioration of disease indicates that AREG is pathogenic in GN. AREG enhanced myeloid cell responses via inducing chemokine and colony stimulating factor 2 (CSF2) expression in kidney resident cells. Furthermore, AREG directly skewed M/M to a proinflammatory M1 phenotype and protected them from apoptosis. Consequently, anti-AREG antibody treatment dose-dependently ameliorated GN. Notably, selective abrogation of EGFR signaling in myeloid cells was sufficient to protect against nephritis. Finally, strong upregulation of AREG expression was also detected in kidneys of patients with two forms of crescentic GN., Conclusions: AREG is a proinflammatory mediator of GN via ( 1 ) enhancing renal pathogenic myeloid cell infiltration and ( 2 ) direct effects on M/M polarization, proliferation, and cytokine secretion. The AREG/EGFR axis is a potential therapeutic target for acute GN., (Copyright © 2020 by the American Society of Nephrology.)

Background: Neutrophil gelatinase-associated lipocalin (NGAL) is a diagnostic marker of intrinsic kidney injury produced by damaged renal cells and by neutrophils. ANCA-associated vasculitis features necrotizing crescentic GN (NCGN), and ANCA-activated neutrophils contribute to NCGN. Whether NGAL plays a mechanistic role in ANCA-associated vasculitis is unknown., Methods: We measured NGAL in patients with ANCA-associated vasculitis and mice with anti-myeloperoxidase (anti-MPO) antibody-induced NCGN. We compared kidney histology, neutrophil functions, T cell proliferation and polarization, renal infiltrating cells, and cytokines in wild-type and NGAL-deficient chimeric mice with anti-MPO antibody-induced NCGN. To assess the role of T H 17 immunity, we transplanted irradiated MPO-immunized MPO-deficient mice with bone marrow from either wild-type or NGAL-deficient mice; we also transplanted irradiated MPO-immunized MPO/IL-17A double-deficient mice with bone marrow from either IL-17A-deficient or NGAL/IL-17A double-deficient mice., Results: Mice and patients with active ANCA-associated vasculitis demonstrated strongly increased serum and urinary NGAL levels. ANCA-stimulated neutrophils released NGAL. Mice with NGAL-deficient bone marrow developed worsened MPO-ANCA-induced NCGN. Intrinsic neutrophil functions were similar in NGAL-deficient and wild-type neutrophils, whereas T cell immunity was increased in chimeric mice with NGAL-deficient neutrophils with more renal infiltrating T H 17 cells. NGAL-expressing neutrophils and CD3 + T cells were in close proximity in kidney and spleen. CD4 + T cells showed no intrinsic difference in proliferation and polarization in vitro , whereas iron siderophore-loaded NGAL suppressed T H 17 polarization. We found significantly attenuated NCGN in IL-17A-deficient chimeras compared with MPO-deficient mice receiving wild-type bone marrow, as well as in NGAL/IL-17A-deficient chimeras compared with NGAL-deficient chimeras., Conclusions: Our findings support that bone marrow-derived, presumably neutrophil, NGAL protects from ANCA-induced NCGN by downregulating T H 17 immunity., (Copyright © 2020 by the American Society of Nephrology.)

Therapeutic targeting of IL-17A and its receptor IL-17RA with antibodies has turned out to be a tremendous success in the treatment of several autoimmune conditions. As the IL-17 cytokine family consists of six members (IL-17A to F), it is intriguing to elucidate the biological function of these five other molecules to identify more potential targets. In the past decade, IL-17C has emerged as quite a unique member of this pro-inflammatory cytokine group. In contrast to the well-described IL-17A and IL-17F, IL-17C is upregulated at very early timepoints of several disease settings. Also, the cellular source of the homodimeric cytokine differs from the other members of the family: Epithelial rather than hematopoietic cells were identified as the producers of IL-17C, while its receptor IL-17RE is expressed on T H 17 cells as well as the epithelial cells themselves. Numerous investigations led to the current understanding that IL-17C (a) maintains an autocrine loop in the epithelium reinforcing innate immune barriers and (b) stimulates highly inflammatory T H 17 cells. Functionally, the IL-17C/RE axis has been described to be involved in the pathogenesis of several diseases ranging from infectious and autoimmune conditions to cancer development and progression. This body of evidence has paved the way for the first clinical trials attempting to neutralize IL-17C in patients. Here, we review the latest knowledge about identification, regulation, and function of the IL-17C/IL-17receptor E pathway in inflammation and immunity, with a focus on the mechanisms underlying tissue injury. We also discuss the rationale for the translation of these findings into new therapeutic approaches in patients with immune-mediated disease., (Copyright © 2020 Nies and Panzer.)

Background: GATA3 is a dual-zinc finger transcription factor that regulates gene expression in many developing tissues. In the kidney, GATA3 is essential for ureteric bud branching, and mice without it fail to develop kidneys. In humans, autosomal dominant GATA3 mutations can cause renal aplasia as part of the hypoparathyroidism, renal dysplasia, deafness (HDR) syndrome that includes mesangioproliferative GN. This suggests that GATA3 may have a previously unrecognized role in glomerular development or injury., Methods: To determine GATA3's role in glomerular development or injury, we assessed GATA3 expression in developing and mature kidneys from Gata3 heterozygous ( + /-) knockout mice, as well as injured human and rodent kidneys., Results: We show that GATA3 is expressed by FOXD1 lineage stromal progenitor cells, and a subset of these cells mature into mesangial cells (MCs) that continue to express GATA3 in adult kidneys. In mice, we uncover that GATA3 is essential for normal glomerular development, and mice with haploinsufficiency of Gata3 have too few MC precursors and glomerular abnormalities. Expression of GATA3 is maintained in MCs of adult kidneys and is markedly increased in rodent models of mesangioproliferative GN and in IgA nephropathy, suggesting that GATA3 plays a critical role in the maintenance of glomerular homeostasis., Conclusions: These results provide new insights on the role GATA3 plays in MC development and response to injury. It also shows that GATA3 may be a novel and robust nuclear marker for identifying MCs in tissue sections., (Copyright © 2019 by the American Society of Nephrology.)

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.

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 T H 17 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.

Anti-glomerular basement membrane (anti-GBM) disease is characterized by antibodies and T cells directed against the Goodpasture antigen, the noncollagenous domain of the α3-chain of type IV collagen [α3(IV)NC1] of the GBM. Consequences are the deposition of autoantibodies along the GBM and the development of crescentic glomerulonephritis (GN) with rapid loss of renal function. Forkhead box protein P3 (Foxp3) + regulatory T (Treg) cells are crucial for the maintenance of peripheral tolerance to self-antigens and the prevention of immunopathology. Here, we use the mouse model of experimental autoimmune GN to characterize the role of Treg cells in anti-GBM disease. Immunization of DBA/1 mice with α3(IV)NC1 induced the formation of α3(IV)NC1-specific T cells and antibodies and, after 8-10 wk, the development of crescentic GN. Immunization resulted in increased frequencies of peripheral Treg cells and renal accumulation of these cells in the stage of acute GN. Depletion of Treg cells during immunization led to enhanced generation of α3(IV)NC1-specific antibodies and T cells and to aggravated GN. In contrast, depletion or expansion of the Treg cell population in mice with established autoimmunity had only minor consequences for renal inflammation and did not alter the severity of GN. In conclusion, our results indicate that in anti-GBM disease, Treg cells restrict the induction of autoimmunity against α3(IV)NC1. However, Treg cells are inefficient in preventing crescentic GN after autoimmunity has been established.

Immune-mediated glomerular diseases like crescentic glomerulonephritis (cGN) are driven by inappropriately regulated cellular and humoral immune responses subsequently leading to renal tissue injury. Recent studies demonstrated the crucial role for regulatory T cells (Tregs) in suppressing pathogenic T-cell responses during nephrotoxic nephritis (NTN), a murine model of cGN. However, mechanisms of immune regulation in cGN are less clear. Here, we aim at investigating the role of the co-inhibitory PD-1/PD-L1 pathway in Treg-mediated suppression of renal inflammation. We demonstrated that Foxp3 + Tregs expressing PD-L1 infiltrate the kidney during NTN. Inhibition of PD-L1 signalling by using PD-L1 -/- mice or by blockage of PD-L1 in wildtype mice resulted in an increased Treg frequency in the inflamed kidney. However, mice lacking PD-L1 developed more severe NTN associated with an elevated pathogenic renal Th1 immune response, which was reversed by blockage of IFNγ in these mice. Interestingly, lack of PD-L1 altered the gene expression profile of Tregs in homeostasis and kidney inflammation. Functionally, Tregs from nephritic PD-L1 -/- mice had impaired suppressive capacity in vitro and failed to protect from NTN in vivo. Thus, PD-L1 displays a protective role in NTN, which is related to Treg-mediated suppression of the Th1 immune response.

Predicting renal outcome in antineutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis (GN) remains a major challenge. We aimed to identify reliable predictors of end-stage renal disease (ESRD) and to develop and validate a clinicopathologic score to predict renal outcome in ANCA-associated GN. In a prospective training cohort of 115 patients, the percentage of normal glomeruli (without scarring, crescents, or necrosis within the tuft) was the strongest independent predictor of death-censored ESRD. Regression tree analysis identified predictive cutoff values for three parameters: percentage normal glomeruli (N0 >25%, N1 10 to 25%, N2 <10%), percentage tubular atrophy and interstitial fibrosis (T0 ≤25%, T1 >25%), and estimated glomerular filtration rate at the time of diagnosis (G0 >15 ml/min/1.73 m 2 , G1 ≤15 ml/min/1.73 m 2 ). Cox regression analysis was used to assign points to each parameter (N1 = 4, N2 = 6, T1 = 2, G1 = 3 points), and the resulting risk score was used to classify predicted ESRD risk as low (0), intermediate (2 to 7), or high (8 to 11 points). The risk score accurately predicted ESRD at 36 months in the training cohort (0%, 26%, and 68%, respectively) and in an independent validation cohort of 90 patients (0%, 27%, and 78%, respectively). Here, we propose a clinically applicable renal risk score for ANCA-associated GN that highlights the importance of unaffected glomeruli as a predictor of renal outcome and allows early risk prediction of ESRD., (Copyright © 2018 International Society of Nephrology. All rights reserved.)