Prognostic models can strongly support individualized care provision and well-informed shared decision making. There has been an upsurge of prognostic research in the field of nephrology, but the... Show morePrognostic models can strongly support individualized care provision and well-informed shared decision making. There has been an upsurge of prognostic research in the field of nephrology, but the uptake of prognostic models in clinical practice remains limited. Therefore, we map out the research field of prognostic models for kidney patients and provide directions on how to proceed from here. We performed a scoping review of studies developing, validating, or updating a prognostic model for patients with CKD. We searched all published models in PubMed and Embase and report predicted outcomes, methodological quality, and validation and/or updating efforts. We found 602 studies, of which 30.1% concerned CKD populations, 31.6% dialysis populations, and 38.4% kidney transplantation populations. The most frequently predicted outcomes were mortality (n=129), kidney disease progression (n=75), and kidney graft survival (n=54). Most studies provided discrimination measures (80.4%), but much less showed calibration results (43.4%). Of the 415 development studies, 28.0% did not perform any validation and 57.6% performed only internal validation. Moreover, only 111 models (26.7%) were externally validated either in the development study itself or in an independent external validation study. Finally, in 45.8% of development studies no useable version of the model was reported. To conclude, many prognostic models have been developed for patients with CKD, mainly for outcomes related to kidney disease progression and patient/graft survival. To bridge the gap between prediction research and kidney patient care, patient-reported outcomes, methodological rigor, complete reporting of prognostic models, external validation, updating, and impact assessment urgently need more attention. Show less
Background Cystatin C is recommended as a confirmatory test to eGFR when more precise estimates are needed for clinical decision making. Although eGFR on the basis of both creatinine and cystatin ... Show moreBackground Cystatin C is recommended as a confirmatory test to eGFR when more precise estimates are needed for clinical decision making. Although eGFR on the basis of both creatinine and cystatin (eGFR(cr-cys)) is the most accurate estimate in research studies, it is uncertain whether this is true in real-world settings, particularly when there are large discordances between eGFR based on creatinine (eGFR(cr)) and that based on cystatin C (eGFR(cys))Methods We included 6185 adults referred for measured GFR (mGFR) using plasma clearance of iohexol in Stockholm, Sweden, who had 9404 concurrent measurements of creatinine, cystatin C, and iohexol clearance. The performance of eGFR(cr), eGFR(cys), and eGFR(cr-cys) was assessed against mGFR with median bias, P-30, and correct classification of GFR categories. We stratified analyses within three categories: eGFR(cys) at least 20% lower than eGFR(cr) (eGFR(cys)eGFR(cr)).Results eGFR(cr) and eGFR(cys) were similar in 4226 (45%) samples, and among these samples all three estimating equations performed similarly. By contrast, eGFR(cr-cys) was much more accurate in cases of discordance. For example, when eGFR(cys)eGFR(cr) (8% of samples), the median biases were -4.5, 8.4, and 1.4 ml/min per 1.73m(2). The findings were consistent among individuals with cardiovascular disease, heart failure, diabetes mellitus, liver disease, and cancer.Conclusions When eGFR(cr) and eGFR(cys) are highly discordant in clinical practice, eGFR(cr-cys) is more accurate than either eGFR(cr) or eGFR(cys). Show less
Background:The prognosis for kidney survival is poor in patients presenting with circulating anti-glomerular basement membrane (GBM) antibodies and severe kidney injury. It is unknown if treat-ment... Show moreBackground:The prognosis for kidney survival is poor in patients presenting with circulating anti-glomerular basement membrane (GBM) antibodies and severe kidney injury. It is unknown if treat-ment with an endopeptidase that cleaves circulating and kidney bound IgG can alter the prognosis. Methods: An investigator-driven phase 2a one-arm study (EudraCT 2016-004082-39) was performed in 17 hospitals in five European countries. A single dose of 0.25 mg/kg of imlifidase was given to 15 adults with circulating anti-GBM antibodies and an eGFR < 15 ml/min per 1.73m(2). All patients received standard treatment with cyclophosphamide and corticosteroids, but plasma exchange only if autoantibodies rebounded. The primary outcomes were safety and dialysis independency at 6 months.Results: At inclusion, ten patients were dialysis dependent and the other five had eGFR levels between 7 and 14 ml/min per 1.73m(2). The median age was 61 years (range 19-77), six were women, and six were also positive for anti-neutrophil cytoplasmic antibodies. Then 6 hours after imlifidase infusion, all patients had anti-GBM antibodies levels below the reference range of a prespecified assay. At 6 months 67% (ten out of 15) were dialysis independent. This is significantly higher compared with 18% (nine out of 50) in a historical control cohort (P < 0.001, Fisher's exact test). Eight serious adverse events (including one death) were reported, none assessed as probably or possibly related to the study drug. Conclusions: In this pilot study, the use of imlifidase was associated with a better outcome compared with earlier publications, without major safety issues, but the findings need to be confirmed in a randomized controlled trial. Show less
Significance Statement This randomized trial demonstrates the safety and efficacy of conversion from calcineurin inhibitor (CNI)? to belatacept-based maintenance immunosuppression in renal... Show moreSignificance Statement This randomized trial demonstrates the safety and efficacy of conversion from calcineurin inhibitor (CNI)? to belatacept-based maintenance immunosuppression in renal transplant recipients 6?60 months post-transplant. Patients converted to belatacept showed sustained improvement in renal function associated with an acceptable safety profile consistent with prior experience and a smaller treatment difference in acute rejection postconversion compared with that observed in earlier studies in de novo renal allograft recipients. These results favor the use of belatacept as an alternative to continued long-term CNI-based maintenance immunosuppression, which is particularly relevant for CNI-intolerant patients, including those who experience nephrotoxicity. These data help inform clinical practice guidelines regarding the conversion of such patients to an alternative immunosuppressive drug regimen.Background Calcineurin inhibitors (CNIs) are standard of care after kidney transplantation, but they are associated with nephrotoxicity and reduced long-term graft survival. Belatacept, a selective T cell costimulation blocker, is approved for the prophylaxis of kidney transplant rejection. This phase 3 trial evaluated the efficacy and safety of conversion from CNI-based to belatacept-based maintenance immunosuppression in kidney transplant recipients.Methods Stable adult kidney transplant recipients 6?60 months post-transplantation under CNI-based immunosuppression were randomized (1:1) to switch to belatacept or continue treatment with their established CNI. The primary end point was the percentage of patients surviving with a functioning graft at 24 months.Results Overall, 446 renal transplant recipients were randomized to belatacept conversion (n=223) or CNI continuation (n=223). The 24-month rates of survival with graft function were 98% and 97% in the belatacept and CNI groups, respectively (adjusted difference, 0.8; 95.1% CI, ?2.1 to 3.7). In the belatacept conversion versus CNI continuation groups, 8% versus 4% of patients experienced biopsy-proven acute rejection (BPAR), respectively, and 1% versus 7% developed de novo donor-specific antibodies (dnDSAs), respectively. The 24-month eGFR was higher with belatacept (55.5 versus 48.5 ml/min per 1.73 m(2) with CNI). Both groups had similar rates of serious adverse events, infections, and discontinuations, with no unexpected adverse events. One patient in the belatacept group had post-transplant lymphoproliferative disorder.Conclusions Switching stable renal transplant recipients from CNI-based to belatacept-based immunosuppression was associated with a similar rate of death or graft loss, improved renal function, and a numerically higher BPAR rate but a lower incidence of dnDSA. Clinical Trial registry name and registration number: A Study in Maintenance Kidney Transplant Recipients Following Conversion to Nulojix? (Belatacept)-Based, NCT01820572 Show less
Background IgAnephropathy (IgAN) is the most common primary glomerular disease worldwide and is a leading cause of renal failure. The disease mechanisms are not completely understood, but a higher... Show moreBackground IgAnephropathy (IgAN) is the most common primary glomerular disease worldwide and is a leading cause of renal failure. The disease mechanisms are not completely understood, but a higher abundance of galactose-deficient IgA is recognized to play a crucial role in IgAN pathogenesis. Although both types of human IgA (IgA1 and IgA2) have several N-glycans as post-translational modification, only IgA1 features extensive hinge-region O-glycosylation. IgA1 galactose deficiency on the O-glycans is commonly detected by a lectin-based method. To date, limited detail is known about IgA O- and N-glycosylation in IgAN.Methods To gain insights into the complex O- and N-glycosylation of serum IgA1 and IgA2 in IgAN, we used liquid chromatography-mass spectrometry (LC-MS) for the analysis of tryptic glycopeptides of serum IgA from 83 patients with IgAN and 244 age- and sex-matched healthy controls.Results Multiple structural features of N-glycosylation of IgA1 and IgA2 were associated with IgAN and glomerular function in our cross-sectional study. These features included differences in galactosylation, sialylation, bisection, fucosylation, and N-glycan complexity. Moreover, IgA1 O-glycan sialylation was associated with both the disease and glomerular function. Finally, glycopeptides were a better predictor of IgAN and glomerular function than galactose-deficient IgA1 levels measured by lectin-based ELISA.Conclusions Our high-resolution data suggest that IgA O- and N-glycopeptides are promising targets for future investigations on the pathophysiology of IgAN and as potential noninvasive biomarkers for disease prediction and deteriorating kidney function. Show less
Khan, A.; Shang, N.; Petukhova, L.; Zhang, J.; Shen, Y.F.; Hebbring, S.J.; ... ; Kiryluk, K. 2021
Background Genetic variants in complement genes have been associated with a wide range of human disease states, but well-powered genetic association studies of complement activation have not been... Show moreBackground Genetic variants in complement genes have been associated with a wide range of human disease states, but well-powered genetic association studies of complement activation have not been performed in large multiethnic cohorts. Methods We performed medical records?based genome-wide and phenome-wide association studies for plasma C3 and C4 levels among participants of the Electronic Medical Records and Genomics (eMERGE) network. Results In a GWAS for C3 levels in 3949 individuals, we detected two genome-wide significant loci: chr.1q31.3 (CFH locus; rs3753396-A; ?=0.20; 95% CI, 0.14 to 0.25; P=1.52x10(-11)) and chr.19p13.3 (C3 locus; rs11569470-G; ?=0.19; 95% CI, 0.13 to 0.24; P=1.29x10(-8)). These two loci explained approximately 2% of variance in C3 levels. GWAS for C4 levels involved 3998 individuals and revealed a genome-wide significant locus at chr.6p21.32 (C4 locus; rs3135353-C; ?=0.40; 95% CI, 0.34 to 0.45; P=4.58x10(-35)). This locus explained approximately 13% of variance in C4 levels. The multiallelic copy number variant analysis defined two structural genomic C4 variants with large effect on blood C4 levels: C4-BS (?=?0.36; 95% CI, ?0.42 to ?0.30; P=2.98x10(-22)) and C4-AL-BS (?=0.25; 95% CI, 0.21 to 0.29; P=8.11x10(-23)). Overall, C4 levels were strongly correlated with copy numbers of C4A and C4B genes. In comprehensive phenome-wide association studies involving 102,138 eMERGE participants, we cataloged a full spectrum of autoimmune, cardiometabolic, and kidney diseases genetically related to systemic complement activation. Conclusions We discovered genetic determinants of plasma C3 and C4 levels using eMERGE genomic data linked to electronic medical records. Genetic variants regulating C3 and C4 levels have large effects and multiple clinical correlations across the spectrum of complement-related diseases in humans.Significance Statement The complement pathway represents one of the critical arms of the innate immune system. We combined genome-wide and phenome-wide association studies using medical records data for C3 and C4 levels to discover common genetic variants controlling systemic complement activation. Three genome-wide significant loci had large effects on complement levels. These loci encode three critical complement genes: CFH, C3, and C4. We performed detailed functional annotations of the significant loci, including multiallelic copy number variant analysis of the C4 locus to define two structural genomic variants with large effects on C4 levels. Blood C4 levels were strongly correlated with the copy number of C4A and C4B genes. Lastly, using genome-wide genetic correlations and electronic health records?based phenome-wide association studies in 102,138 participants, we catalogued a spectrum of human diseases genetically related to systemic complement activation, including inflammatory, autoimmune, cardiometabolic, and kidney diseases. Show less
Background Crumbs2 is expressed at embryonic stages as well as in the retina, brain, and glomerular podocytes. Recent studies identified CRB2 mutations as a novel cause of steroid-resistant... Show moreBackground Crumbs2 is expressed at embryonic stages as well as in the retina, brain, and glomerular podocytes. Recent studies identified CRB2 mutations as a novel cause of steroid-resistant nephrotic syndrome (SRNS).Methods To study the function of Crb2 at the renal filtration barrier, mice lacking Crb2 exclusively in podocytes were generated. Gene expression and histologic studies as well as transmission and scanning electron microscopy were used to analyze these Crb2(podKO) knockout mice and their littermate controls. Furthermore, high-resolution expansion microscopy was used to investigate Crb2 distribution in murine glomeruli. For pull-down experiments, live cell imaging, and transcriptome analyses, cell lines were applied that inducibly express fluorescent protein-tagged CRB2 wild type and mutants.Results Crb2(podKO) mice developed proteinuria directly after birth that preceded a prominent development of disordered and effaced foot processes, upregulation of renal injury and inflammatory markers, and glomerulosclerosis. Pull-down assays revealed an interaction of CRB2 with Nephrin, mediated by their extracellular domains. Expansion microscopy showed that in mice glomeruli, Crb2 and Nephrin are organized in adjacent clusters. SRNS-associated CRB2 protein variants and a mutant that lacks a putative conserved O-glycosylation site were not transported to the cell surface. Instead, mutants accumulated in the ER, showed altered glycosylation pattern, and triggered an ER stress response.Conclusions Crb2 is an essential component of the podocyte's slit diaphragm, interacting with Nephrin. Loss of slit diaphragm targeting and increasing ER stress are pivotal factors for onset and progression of CRB2-related SRNS. Show less
Ramspek, C.L.; Evans, M.; Wanner, C.; Drechsler, C.; Chesnaye, N.C.; Szymczak, M.; ... ; EQUAL Study Investigators 2021
Background: Various prediction models have been developed to predict the risk of kidney failure in patients with CKD. However, guideline-recommended models have yet to be compared head to head,... Show moreBackground: Various prediction models have been developed to predict the risk of kidney failure in patients with CKD. However, guideline-recommended models have yet to be compared head to head, their validation in patients with advanced CKD is lacking, and most do not account for competing risks.Methods: To externally validate 11 existing models of kidney failure, taking the competing risk of death into account, we included patients with advanced CKD from two large cohorts: the European Quality Study (EQUAL), an ongoing European prospective, multicenter cohort study of older patients with advanced CKD, and the Swedish Renal Registry (SRR), an ongoing registry of nephrology-referred patients with CKD in Sweden. The outcome of the models was kidney failure (defined as RRT-treated ESKD). We assessed model performance with discrimination and calibration.Results: The study included 1580 patients from EQUAL and 13,489 patients from SRR. The average c statistic over the 11 validated models was 0.74 in EQUAL and 0.80 in SRR, compared with 0.89 in previous validations. Most models with longer prediction horizons overestimated the risk of kidney failure considerably. The 5-year Kidney Failure Risk Equation (KFRE) overpredicted risk by 10%-18%. The four- and eight-variable 2-year KFRE and the 4-year Grams model showed excellent calibration and good discrimination in both cohorts.Conclusions: Some existing models can accurately predict kidney failure in patients with advanced CKD. KFRE performed well for a shorter time frame (2 years), despite not accounting for competing events. Models predicting over a longer time frame (5 years) overestimated risk because of the competing risk of death. The Grams model, which accounts for the latter, is suitable for longer-term predictions (4 years). Show less
Fu, E.L.; Evans, M.; Clase, C.M.; Tomlinson, L.A.; Diepen, M. van; Dekker, F.W.; Carrero, J.J. 2021
Background It is unknown whether stopping renin-angiotensin system (RAS) inhibitor therapy in patients with advanced CKD affects outcomes.Methods We studied patients referred to nephrologist care,... Show moreBackground It is unknown whether stopping renin-angiotensin system (RAS) inhibitor therapy in patients with advanced CKD affects outcomes.Methods We studied patients referred to nephrologist care, listed on the Swedish Renal Registry during 2007-2017, who developed advanced CKD (eGFR <30 ml/min per 1.73m(2)) while on RAS inhibitor therapy. Using target trial emulation techniques on the basis of cloning, censoring, and weighting, we compared the risks of stopping within 6 months and remaining off treatment versus continuing RAS inhibitor therapy. These included risks of subsequent 5-year all-cause mortality, major adverse cardiovascular events, and initiation of kidney replacement therapy (KRT).Results Of 10,254 prevalent RAS inhibitor users (median age 72 years, 36% female) with new-onset eGFR >30 ml/min per 1.73 m(2), 1553 (15%) stopped RAS inhibitor therapy within 6 months. Median eGFR was 23 ml/min per 1.73 m(2). Compared with continuing RAS inhibition, stopping this therapy was associated with a higher absolute 5-year risk of death (40.9% versus 54.5%) and major adverse cardiovascular events (47.6% versus 59.5%), but with a lower risk of KRT (36.1% versus 27.9%); these corresponded to absolute risk differences of 13.6 events per 100 patients, 11.9 events per 100 patients, and -8.3 events per 100 patients, respectively. Results were consistent whether patients stopped RAS inhibition at higher or lower eGFR, across prespecified subgroups, after adjustment and stratification for albuminuria and potassium, and when modeling RAS inhibition as a time-dependent exposure using a marginal structural model.Conclusions In this nationwide observational study of people with advanced CKD, stopping RAS inhibition was associated with higher absolute risks of mortality and major adverse cardiovascular events, but also with a lower absolute risk of initiating KRT. Show less
Background During mammalian kidney development, nephron progenitors undergo a mesenchymal-to-epithelial transition and eventually differentiate into the various tubular segments of the nephron.... Show moreBackground During mammalian kidney development, nephron progenitors undergo a mesenchymal-to-epithelial transition and eventually differentiate into the various tubular segments of the nephron. Recently, Drop-seq single-cell RNA sequencing technology for measuring gene expression from thousands of individual cells identified the different cell types in the developing kidney. However, that analysis did not include the additional layer of heterogeneity that alternative mRNA splicing creates.Methods Full transcript length single-cell RNA sequencing characterized the transcriptomes of 544 individual cells from mouse embryonic kidneys.Results Gene expression levels measured with full transcript length single-cell RNA sequencing identified each cell type. Further analysis comprehensively characterized splice isoform switching during the transition between mesenchymal and epithelial cellular states, which is a key transitional process in kidney development. The study also identified several putative splicing regulators, including the genes Esrp1/2 and Rbfox1/2.Conclusions Discovery of the sets of genes that are alternatively spliced as the fetal kidney mesenchyme differentiates into tubular epithelium will improve our understanding of the molecular mechanisms that drive kidney development. Show less
Berg, C.W. van den; Koudijs, A.; Ritsma, L.; Rabelink, T.J. 2020
Significance StatementThe ability to differentiate human induced pluripotent stem cells to kidney organoids in vitro holds promise for disease modeling, drug discovery, and clinical application.... Show moreSignificance StatementThe ability to differentiate human induced pluripotent stem cells to kidney organoids in vitro holds promise for disease modeling, drug discovery, and clinical application. The authors differentiated such cells to kidney tissue comprising glomerular, proximal, and distal tubular structures. Earlier research demonstrated that these structures become vascularized upon transplantation in mice and show advanced maturation. To investigate whether human induced pluripotent stem cell?derived kidney organoids can also become functional in vivo, they applied high-resolution intravital multiphoton imaging through a titanium imaging window. They demonstrated in vivo glomerular filtration and size-selective glomerular barrier function in the transplanted organoids. This technique can be instrumental for further developing stem cell?derived organoids toward clinical applications. BackgroundThe utility of kidney organoids in regenerative medicine will rely on the functionality of the glomerular and tubular structures in these tissues. Recent studies have demonstrated the vascularization and subsequent maturation of human pluripotent stem cell?derived kidney organoids after renal subcapsular transplantation. This raises the question of whether the glomeruli also become functional upon transplantation.MethodsWe transplanted kidney organoids under the renal capsule of the left kidney in immunodeficient mice followed by the implantation of a titanium imaging window on top of the kidney organoid. To assess glomerular function in the transplanted human pluripotent stem cell?derived kidney tissue 1, 2, and 3 weeks after transplantation, we applied high-resolution intravital multiphoton imaging through the imaging window during intravenous infusion of fluorescently labeled low and high molecular mass dextran molecules or albumin.ResultsAfter vascularization, glomerular structures in the organoid displayed dextran and albumin size selectivity across their glomerular filtration barrier. We also observed evidence of proximal tubular dextran reuptake.ConclusionsOur results demonstrate that human pluripotent stem cell?derived glomeruli can develop an appropriate barrier function and discriminate between molecules of varying size. These characteristics together with tubular presence of low molecular mass dextran provide clear evidence of functional filtration. This approach to visualizing glomerular filtration function will be instrumental for translation of organoid technology for clinical applications as well as for disease modeling. Show less
Dumas, S.J.; Meta, E.; Borri, M.; Goveia, J.; Rohlenova, K.; Conchinha, N.V.; ... ; Carmeliet, P. 2020
Background Renal endothelial cells from glomerular, cortical, and medullary kidney compartments are exposed to different microenvironmental conditions and support specific kidney processes. However... Show moreBackground Renal endothelial cells from glomerular, cortical, and medullary kidney compartments are exposed to different microenvironmental conditions and support specific kidney processes. However, the heterogeneous phenotypes of these cells remain incompletely inventoried. Osmotic homeostasis is vitally important for regulating cell volume and function, and in mammals, osmotic equilibrium is regulated through the countercurrent system in the renal medulla, where water exchange through endothelium occurs against an osmotic pressure gradient. Dehydration exposes medullary renal endothelial cells to extreme hyperosmolarity, and how these cells adapt to and survive in this hypertonic milieu is unknown.Methods We inventoried renal endothelial cell heterogeneity by single-cell RNA sequencing >40,000 mouse renal endothelial cells, and studied transcriptome changes during osmotic adaptation upon water deprivation. We validated our findings by immunostaining and functionally by targeting oxidative phosphorylation in a hyperosmolarity model in vitro and in dehydrated mice in vivo.Results We identified 24 renal endothelial cell phenotypes (of which eight were novel), highlighting extensive heterogeneity of these cells between and within the cortex, glomeruli, and medulla. In response to dehydration and hypertonicity, medullary renal endothelial cells upregulated the expression of genes involved in the hypoxia response, glycolysis, and-surprisingly-oxidative phosphorylation. Endothelial cells increased oxygen consumption when exposed to hyperosmolarity, whereas blocking oxidative phosphorylation compromised endothelial cell viability during hyperosmotic stress and impaired urine concentration during dehydration.Conclusions This study provides a high-resolution atlas of the renal endothelium and highlights extensive renal endothelial cell phenotypic heterogeneity, as well as a previously unrecognized role of oxidative phosphorylation in the metabolic adaptation of medullary renal endothelial cells to water deprivation. Show less
Berg, C.W. van den; Koudijs, A.; Ritsma, L.; Rabelink, T.J. 2020
Background The utility of kidney organoids in regenerative medicine will rely on the functionality of the glomerular and tubular structures in these tissues. Recent studies have demonstrated the... Show moreBackground The utility of kidney organoids in regenerative medicine will rely on the functionality of the glomerular and tubular structures in these tissues. Recent studies have demonstrated the vascularization and subsequent maturation of human pluripotent stem cell–derived kidney organoids after renal subcapsular transplantation. This raises the question of whether the glomeruli also become functional upon transplantation.Methods We transplanted kidney organoids under the renal capsule of the left kidney in immunodeficient mice followed by the implantation of a titanium imaging window on top of the kidney organoid. To assess glomerular function in the transplanted human pluripotent stem cell–derived kidney tissue 1, 2, and 3 weeks after transplantation, we applied high-resolution intravital multiphoton imaging through the imaging window during intravenous infusion of fluorescently labeled low and high molecular mass dextran molecules or albumin.Results After vascularization, glomerular structures in the organoid displayed dextran and albumin size selectivity across their glomerular filtration barrier. We also observed evidence of proximal tubular dextran reuptake.Conclusions Our results demonstrate that human pluripotent stem cell–derived glomeruli can develop an appropriate barrier function and discriminate between molecules of varying size. These characteristics together with tubular presence of low molecular mass dextran provide clear evidence of functional filtration. This approach to visualizing glomerular filtration function will be instrumental for translation of organoid technology for clinical applications as well as for disease modeling. Show less
Ravani, P.; Fiocco, M.; Liu, P.; Quinn, R.R.; Hemmelgarn, B.; James, M.; ... ; Tonelli, M. 2019
Background A glycocalyx envelope consisting of proteoglycans and adhering proteins covers endothelial cells, both the luminal and abluminal surface. We previously demonstrated that short-term loss... Show moreBackground A glycocalyx envelope consisting of proteoglycans and adhering proteins covers endothelial cells, both the luminal and abluminal surface. We previously demonstrated that short-term loss of integrity of the luminal glycocalyx layer resulted in perturbed glomerular filtration barrier function.Methods To explore the role of the glycocalyx layer of the endothelial extracellular matrix in renal function, we generated mice with an endothelium-specific and inducible deletion of hyaluronan synthase 2 (Has2), the enzyme that produces hyaluronan, the main structural component of the endothelial glycocalyx layer. We also investigated the presence of endothelial hyaluronan in human kidney tissue from patients with varying degrees of diabetic nephropathy.Results Endothelial deletion of Has2 in adult mice led to substantial loss of the glycocalyx structure, and analysis of their kidneys and kidney function showed vascular destabilization, characterized by mesangiolysis, capillary ballooning, and albuminuria. This process develops over time into glomerular capillary rarefaction and glomerulosclerosis, recapitulating the phenotype of progressive human diabetic nephropathy. Using a hyaluronan-specific probe, we found loss of glomerular endothelial hyaluronan in association with lesion formation in tissue from patients with diabetic nephropathy. We also demonstrated that loss of hyaluronan, which harbors a specific binding site for angiopoietin and a key regulator of endothelial quiescence and maintenance of EC barrier function results in disturbed angiopoietin 1 Tie2.Conclusions Endothelial loss of hyaluronan results in disturbed glomerular endothelial stabilization. Glomerular endothelial hyaluronan is a previously unrecognized key component of the extracelluar matrixthat is required for glomerular structure and function and lost in diabetic nephropathy. Show less
Hermsen, M.; Bel, T. de; Boer, M. den; Steenbergen, E.J.; Kers, J.; Florquin, S.; ... ; Laak, J.A.W.M. van der 2019
Background The development of deep neural networks is facilitating more advanced digital analysis of histopathologic images. We trained a convolutional neural network for multiclass segmentation of... Show moreBackground The development of deep neural networks is facilitating more advanced digital analysis of histopathologic images. We trained a convolutional neural network for multiclass segmentation of digitized kidney tissue sections stained with periodic acid-Schiff (PAS).Methods We trained the network using multiclass annotations from 40 whole-slide images of stained kidney transplant biopsies and applied it to four independent data sets. We assessed multiclass segmentation performance by calculating Dice coefficients for ten tissue classes on ten transplant biopsies from the Radboud University Medical Center in Nijmegen, The Netherlands, and on ten transplant biopsies from an external center for validation. We also fully segmented 15 nephrectomy samples and calculated the network's glomerular detection rates and compared network-based measures with visually scored histologic components (Banff classification) in 82 kidney transplant biopsies.Results The weighted mean Dice coefficients of all classes were 0.80 and 0.84 in ten kidney transplant biopsies from the Radboud center and the external center, respectively. The best segmented class was "glomeruli" in both data sets (Dice coefficients, 0.95 and 0.94, respectively), followed by "tubuli combined" and "interstitium." The network detected 92.7% of all glomeruli in nephrectomy samples, with 10.4% false positives. In whole transplant biopsies, the mean intraclass correlation coefficient for glomerular counting performed by pathologists versus the network was 0.94. We found significant correlations between visually scored histologic components and network-based measures.Conclusions This study presents the first convolutional neural network for multiclass segmentation of PAS-stained nephrectomy samples and transplant biopsies. Our network may have utility for quantitative studies involving kidney histopathology across centers and provide opportunities for deep learning applications in routine diagnostics. Show less
Kamburova, E.G.; Wisse, B.W.; Joosten, I.; Allebes, W.A.; Meer, A. van der; Hilbrands, L.B.; ... ; Otten, H.G. 2018
