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Introduction: Environmental contributors to kidney disease progression remain elusive. We explored how residential air pollution affects disease progression in patients with primary glomerulopathies. Methods: Nephrotic Syndrome Study Network (NEPTUNE) and CureGlomerulonephropathy (CureGN) participants with residential census tract data and ≥2 years of follow-up were included. Using Cox proportional hazards models, the associations per doubling in annual average baseline concentrations of total particulate matter with diameter ≤2.5 µm (PM2.5) and its components, black carbon (BC), and sulfate, with time to ≥40% decline in estimated glomerular filtration rate (eGFR) or kidney failure were estimated. Serum tumour necrosis factor levels and kidney tissue transcriptomic inflammatory pathway activation scores were used as molecular markers of disease progression. Results: PM2.5, BC, and sulfate exposures were comparable in NEPTUNE (n = 228) and CureGN (n = 697). In both cohorts, participants from areas with higher levels of pollutants had lower eGFR, were older and more likely self-reported racial and ethnic minorities. In a fully adjusted model combining both cohorts, kidney disease progression was associated with PM2.5 (adjusted hazard ratio 1.55 [95% confidence interval: 1.00-2.38], P = 0.0489) and BC (adjusted hazard ratio 1.43 [95% confidence interval: 0.98-2.07], P = 0.0608) exposure. Sulfate and PM2.5 exposure were positively correlated with serum tumour necrosis factor (TNF) (P = 0.003) and interleukin-1ß levels (P = 0.03), respectively. Sulfate exposure was also directly associated with transcriptional activation of the TNF and JAK-STAT signaling pathways in kidneys (r = 0.55-0.67, P-value <0.01). Conclusion: Elevated exposure to select air pollutants is associated with increased risk of disease progression and systemic inflammation in patients with primary.
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R-loop-triggered collateral single-stranded DNA (ssDNA) nuclease activity within Class 1 Type I CRISPR-Cas systems holds immense potential for nucleic acid detection. However, the hyperactive ssDNase activity of Cas3 introduces unwanted noise and false-positive results. In this study, we identified a novel Type I-A Cas3 variant derived from Thermococcus siculi, which remains in an auto-inhibited state until it is triggered by Cascade complex and R-loop formation. This Type I-A CRISPR-Cas3 system not only exhibits an expanded protospacer adjacent motif (PAM) recognition capability but also demonstrates remarkable intolerance towards mismatched sequences. Furthermore, it exhibits dual activation modes-responding to both DNA and RNA targets. The culmination of our research efforts has led to the development of the Hyper-Active-Verification Establishment (HAVE, ). This innovation enables swift and precise human papillomavirus (HPV) diagnosis in clinical samples, providing a robust molecular diagnostic tool based on the Type I-A CRISPR-Cas3 system. Our findings contribute to understanding type I-A CRISPR-Cas3 system regulation and facilitate the creation of advanced diagnostic solutions with broad clinical applicability.
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Sistemas CRISPR-Cas , Infecções por Papillomavirus , Humanos , Infecções por Papillomavirus/diagnóstico , Infecções por Papillomavirus/virologia , Infecções por Papillomavirus/genética , Papillomaviridae/genética , Proteínas Associadas a CRISPR/metabolismo , Proteínas Associadas a CRISPR/genética , Papillomavirus HumanoRESUMO
Type I CRISPR-Cas systems employ multi-subunit effector Cascade and helicase-nuclease Cas3 to target and degrade foreign nucleic acids, representing the most abundant RNA-guided adaptive immune systems in prokaryotes. Their ability to cause long fragment deletions have led to increasing interests in eukaryotic genome editing. While the Cascade structures of all other six type I systems have been determined, the structure of the most evolutionarily conserved type I-B Cascade is still missing. Here, we present two cryo-EM structures of the Synechocystis sp. PCC 6714 (Syn) type I-B Cascade, revealing the molecular mechanisms that underlie RNA-directed Cascade assembly, target DNA recognition, and local conformational changes of the effector complex upon R-loop formation. Remarkably, a loop of Cas5 directly intercalated into the major groove of the PAM and facilitated PAM recognition. We further characterized the genome editing profiles of this I-B Cascade-Cas3 in human CD3+ T cells using mRNA-mediated delivery, which led to unidirectional 4.5 kb deletion in TRAC locus and achieved an editing efficiency up to 41.2%. Our study provides the structural basis for understanding target DNA recognition by type I-B Cascade and lays foundation for harnessing this system for long range genome editing in human T cells.
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Sistemas CRISPR-Cas , Microscopia Crioeletrônica , Edição de Genes , Synechocystis , Edição de Genes/métodos , Humanos , Synechocystis/genética , Proteínas Associadas a CRISPR/metabolismo , Proteínas Associadas a CRISPR/genética , Proteínas Associadas a CRISPR/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/química , Linfócitos T/metabolismo , Estruturas R-Loop/genéticaRESUMO
Tissue regeneration is limited in several organs, including the kidney, contributing to the high prevalence of kidney disease globally. However, evolutionary and physiological adaptive responses and the presence of renal progenitor cells suggest an existing remodeling capacity. This study uncovered endogenous tissue remodeling mechanisms in the kidney that were activated by the loss of body fluid and salt and regulated by a unique niche of a minority renal cell type called the macula densa (MD). Here, we identified neuronal differentiation features of MD cells that sense the local and systemic environment and secrete angiogenic, growth, and extracellular matrix remodeling factors, cytokines and chemokines, and control resident progenitor cells. Serial intravital imaging, MD nerve growth factor receptor and Wnt mouse models, and transcriptome analysis revealed cellular and molecular mechanisms of these MD functions. Human and therapeutic translation studies illustrated the clinical potential of MD factors, including CCN1, as a urinary biomarker and therapeutic target in chronic kidney disease. The concept that a neuronally differentiated key sensory and regulatory cell type responding to organ-specific physiological inputs controls local progenitors to remodel or repair tissues may be applicable to other organs and diverse tissue-regenerative therapeutic strategies.
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Diferenciação Celular , Regeneração , Animais , Camundongos , Humanos , Rim/metabolismo , Neurônios/metabolismo , Neurônios/patologia , Insuficiência Renal Crônica/patologia , Insuficiência Renal Crônica/metabolismo , Insuficiência Renal Crônica/fisiopatologia , Insuficiência Renal Crônica/genética , MasculinoRESUMO
Current classification of chronic kidney disease (CKD) into stages using indirect systemic measures (estimated glomerular filtration rate (eGFR) and albuminuria) is agnostic to the heterogeneity of underlying molecular processes in the kidney thereby limiting precision medicine approaches. To generate a novel CKD categorization that directly reflects within kidney disease drivers we analyzed publicly available transcriptomic data from kidney biopsy tissue. A Self-Organizing Maps unsupervised artificial neural network machine-learning algorithm was used to stratify a total of 369 patients with CKD and 46 living kidney donors as healthy controls. Unbiased stratification of the discovery cohort resulted in identification of four novel molecular categories of disease termed CKD-Blue, CKD-Gold, CKD-Olive, CKD-Plum that were replicated in independent CKD and diabetic kidney disease datasets and can be further tested on any external data at kidneyclass.org. Each molecular category spanned across CKD stages and histopathological diagnoses and represented transcriptional activation of distinct biological pathways. Disease progression rates were highly significantly different between the molecular categories. CKD-Gold displayed rapid progression, with significant eGFR-adjusted Cox regression hazard ratio of 5.6 [1.01-31.3] for kidney failure and hazard ratio of 4.7 [1.3-16.5] for composite of kidney failure or a 40% or more eGFR decline. Urine proteomics revealed distinct patterns between the molecular categories, and a 25-protein signature was identified to distinguish CKD-Gold from other molecular categories. Thus, patient stratification based on kidney tissue omics offers a gateway to non-invasive biomarker-driven categorization and the potential for future clinical implementation, as a key step towards precision medicine in CKD.
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Progressão da Doença , Taxa de Filtração Glomerular , Rim , Medicina de Precisão , Insuficiência Renal Crônica , Transcriptoma , Humanos , Medicina de Precisão/métodos , Insuficiência Renal Crônica/patologia , Insuficiência Renal Crônica/urina , Insuficiência Renal Crônica/diagnóstico , Insuficiência Renal Crônica/fisiopatologia , Pessoa de Meia-Idade , Feminino , Masculino , Rim/patologia , Rim/fisiopatologia , Idoso , Biópsia , Adulto , Redes Neurais de Computação , Estudos de Casos e Controles , Perfilação da Expressão Gênica , Aprendizado de Máquina não SupervisionadoRESUMO
Chronic kidney disease (CKD) is a public health problem driven by myofibroblast accumulation, leading to interstitial fibrosis. Heterogeneity is a recently recognized characteristic in kidney fibroblasts in CKD, but the role of different populations is still unclear. Here, we characterize a proinflammatory fibroblast population (named CXCL-iFibro), which corresponds to an early state of myofibroblast differentiation in CKD. We demonstrate that CXCL-iFibro co-localize with macrophages in the kidney and participate in their attraction, accumulation, and switch into FOLR2+ macrophages from early CKD stages on. In vitro, macrophages promote the switch of CXCL-iFibro into ECM-secreting myofibroblasts through a WNT/ß-catenin-dependent pathway, thereby suggesting a reciprocal crosstalk between these populations of fibroblasts and macrophages. Finally, the detection of CXCL-iFibro at early stages of CKD is predictive of poor patient prognosis, which shows that the CXCL-iFibro population is an early player in CKD progression and demonstrates the clinical relevance of our findings.
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Receptor 2 de Folato , Insuficiência Renal Crônica , Humanos , Rim/patologia , Insuficiência Renal Crônica/patologia , Fibroblastos/metabolismo , Miofibroblastos/metabolismo , Fibrose , Macrófagos/metabolismo , Receptor 2 de Folato/metabolismoRESUMO
Glomerular diseases are classified using a descriptive taxonomy that is not reflective of the heterogeneous underlying molecular drivers. This limits not only diagnostic and therapeutic patient management, but also impacts clinical trials evaluating targeted interventions. The Nephrotic Syndrome Study Network (NEPTUNE) is poised to address these challenges. The study has enrolled >850 pediatric and adult patients with proteinuric glomerular diseases who have contributed to deep clinical, histologic, genetic, and molecular profiles linked to long-term outcomes. The NEPTUNE Knowledge Network, comprising combined, multiscalar data sets, captures each participant's molecular disease processes at the time of kidney biopsy. In this editorial, we describe the design and implementation of NEPTUNE Match, which bridges a basic science discovery pipeline with targeted clinical trials. Noninvasive biomarkers have been developed for real-time pathway analyses. A Molecular Nephrology Board reviews the pathway maps together with clinical, laboratory, and histopathologic data assembled for each patient to compile a Match report that estimates the fit between the specific molecular disease pathway(s) identified in an individual patient and proposed clinical trials. The NEPTUNE Match report is communicated using established protocols to the patient and the attending nephrologist for use in their selection of available clinical trials. NEPTUNE Match represents the first application of precision medicine in nephrology with the aim of developing targeted therapies and providing the right medication for each patient with primary glomerular disease.
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Nefropatias , Síndrome Nefrótica , Adulto , Criança , Humanos , Biomarcadores , Ensaios Clínicos como Assunto , Glomérulos Renais/patologia , Síndrome Nefrótica/diagnóstico , Síndrome Nefrótica/genética , Síndrome Nefrótica/terapiaRESUMO
BACKGROUND: Insulin resistance (IR) is a pathophysiologic hallmark of type 2 diabetes and associated with the presence of chronic kidney disease (CKD). Experimental studies suggest that endothelin-1 increases IR. We assessed the association between IR and cardio-renal outcomes and the effect of the selective endothelin receptor antagonist atrasentan on IR in patients with type 2 diabetes and CKD. METHODS: We used data from the RADAR and SONAR trials that recruited participants with type 2 diabetes and CKD [eGFR 25-75 mL/min/1.73 m², urine albumin-to-creatinine ratio of 300-5000 mg/g]. IR was calculated using the homeostatic model assessment (HOMA-IR). The association between HOMA-IR and the pre-specified cardio-renal outcomes was assessed using multivariable Cox proportional hazards regression, and effects of atrasentan on HOMA-IR by a linear mixed effect model. RESULTS: In the SONAR trial, each log-unit increase in HOMA-IR was associated with an increased risk of the composite cardio-renal outcome [hazard ratio 1.32 (95%CI 1.09,1.60; p = 0.004)], kidney outcome [hazard ratio 1.30 (95%CI 1.00,1.68; p-value = 0.048)], and the kidney or all-cause mortality outcome [hazard ratio 1.25 (95%CI 1.01,1.55; p-value = 0.037)]. After 12 weeks treatment in the RADAR trial (N = 123), atrasentan 0.75 mg/day and 1.25 mg/day compared to placebo reduced HOMA-IR by 19.1 (95%CI -17.4, 44.3) and 26.7% (95%CI -6.4, 49.5), respectively. In the SONAR trial (N = 1914), atrasentan 0.75 mg/day compared to placebo reduced HOMA-IR by 9.6% (95%CI 0.6, 17.9). CONCLUSIONS: More severe IR is associated with increased risk of cardio-renal outcomes. The endothelin receptor antagonist atrasentan reduced IR. TRIAL REGISTRATION: RADAR trial (Reducing Residual Albuminuria in Subjects With Diabetes and Nephropathy With AtRasentan): NCT01356849. SONAR trial (The Study Of Diabetic Nephropathy With AtRasentan) NCT01858532.
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Diabetes Mellitus Tipo 2 , Resistência à Insulina , Insuficiência Renal Crônica , Humanos , Atrasentana/efeitos adversos , Diabetes Mellitus Tipo 2/diagnóstico , Diabetes Mellitus Tipo 2/tratamento farmacológico , Diabetes Mellitus Tipo 2/epidemiologia , Rim , Insuficiência Renal Crônica/diagnóstico , Insuficiência Renal Crônica/tratamento farmacológico , Insuficiência Renal Crônica/epidemiologia , Antagonistas dos Receptores de Endotelina/efeitos adversosRESUMO
Kidney organoids are a promising model to study kidney disease, but their use is constrained by limited knowledge of their functional protein expression profile. Here, we define the organoid proteome and transcriptome trajectories over culture duration and upon exposure to TNFα, a cytokine stressor. Older organoids increase deposition of extracellular matrix but decrease expression of glomerular proteins. Single cell transcriptome integration reveals that most proteome changes localize to podocytes, tubular and stromal cells. TNFα treatment of organoids results in 322 differentially expressed proteins, including cytokines and complement components. Transcript expression of these 322 proteins is significantly higher in individuals with poorer clinical outcomes in proteinuric kidney disease. Key TNFα-associated protein (C3 and VCAM1) expression is increased in both human tubular and organoid kidney cell populations, highlighting the potential for organoids to advance biomarker development. By integrating kidney organoid omic layers, incorporating a disease-relevant cytokine stressor and comparing with human data, we provide crucial evidence for the functional relevance of the kidney organoid model to human kidney disease.
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Nefropatias , Fator de Necrose Tumoral alfa , Humanos , Fator de Necrose Tumoral alfa/metabolismo , Proteoma/metabolismo , Rim , Nefropatias/genética , Nefropatias/metabolismo , Organoides/metabolismoRESUMO
Underlying molecular mechanisms of the kidney protective effects of sodium glucose co-transporter 2 (SGLT2) inhibitors are not fully elucidated. Therefore, we studied the association between urinary epidermal growth factor (uEGF), a mitogenic factor involved in kidney repair, and kidney outcomes in patients with type 2 diabetes (T2D). The underlying molecular mechanisms of the SGLT2 inhibitor canagliflozin on EGF using single-cell RNA sequencing from kidney tissue were examined. Urinary EGF-to-creatinine ratio (uEGF/Cr) was measured in 3521 CANagliflozin cardioVascular Assessment Study (CANVAS) participants at baseline and week 52. Associations of uEGF/Cr with kidney outcome were assessed using multivariable-adjusted Cox regression models. Single-cell RNA sequencing was performed using protocol kidney biopsy tissue from ten young patients with T2D on SGLT2i, six patients with T2D on standard care only, and six healthy controls (HCs). In CANVAS, each doubling in baseline uEGF/Cr was associated with a 12% (95% confidence interval 1-22) decreased risk of kidney outcome. uEGF/Cr decreased after 52 weeks with placebo and remained stable with canagliflozin (between-group difference +7.3% (2.0-12.8). In young persons with T2D, EGF mRNA was primarily expressed in the thick ascending loop of Henle. Expression in biopsies from T2D without SGLT2i was significantly lower compared to HCs, whereas treatment with SGLT2i increased EGF levels closer to the healthy state. In young persons with T2D without SGLT2i, endothelin-1 emerged as a key regulator of the EGF co-expression network. SGLT2i treatment was associated with a shift towards normal EGF expression. Thus, decreased uEGF represents increased risk of kidney disease progression in patients with T2D. Canagliflozin increased kidney tissue expression of EGF and was associated with a downstream signaling cascade linked to tubular repair and reversal of tubular injury.
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Doenças Cardiovasculares , Diabetes Mellitus Tipo 2 , Inibidores do Transportador 2 de Sódio-Glicose , Humanos , Canagliflozina/farmacologia , Canagliflozina/uso terapêutico , Doenças Cardiovasculares/tratamento farmacológico , Diabetes Mellitus Tipo 2/complicações , Fator de Crescimento Epidérmico/genética , Glucose , Sódio/metabolismo , Transportador 2 de Glucose-Sódio/genética , Transportador 2 de Glucose-Sódio/metabolismo , Inibidores do Transportador 2 de Sódio-Glicose/farmacologia , Inibidores do Transportador 2 de Sódio-Glicose/uso terapêuticoRESUMO
The relationship between urinary endothelial growth factor (uEGF) and cardiovascular autonomic neuropathy (CAN) in adults with type 1 diabetes was evaluated. uEGF levels at baseline and standardized CAN measures were collected at baseline and annually for 3 years for type 1 diabetes adults. Linear regression analysis and linear mixed effects model were used for analysis. In this cohort (n = 44, 59% women, mean ± standard deviation age 34 ± 13 years and diabetes duration 14 ± 6 years), lower baseline uEGF levels correlated with lower baseline expiration : inspiration ratios (P = 0.03) and greater annual declines in Valsalva ratios (P = 0.02) in the unadjusted model, and correlated with lower low-frequency power : high-frequency power ratios (P = 0.01) and greater annual changes in low-frequency power : high-frequency power ratios (P = 0.01) after adjustment for age, sex, body mass index, and hemoglobin A1C. In conclusion, baseline uEGF levels correlate to baseline and longitudinal changes in CAN indices. A large-scale, long-term study is needed to validate uEGF as a reliable CAN biomarker.
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Doenças do Sistema Nervoso Autônomo , Diabetes Mellitus Tipo 1 , Neuropatias Diabéticas , Humanos , Adulto , Feminino , Adulto Jovem , Pessoa de Meia-Idade , Masculino , Fator de Crescimento Epidérmico/urina , Diabetes Mellitus Tipo 1/complicações , Sistema Nervoso Autônomo , Biomarcadores/urina , Doenças do Sistema Nervoso Autônomo/complicações , Doenças do Sistema Nervoso Autônomo/diagnóstico , Neuropatias Diabéticas/etiologiaRESUMO
This study aimed to assess the intraindividual variations of urinary biomarkers in hospitalized children with glomerular diseases. Hospitalized children with glomerular diseases participated in the study. For each patient, an overnight (9:00 p.m.-7:00 a.m.) urine was collected, followed by a 24-h urine (classified into four distinct periods: morning 7:00 a.m.-12:00 p.m., afternoon 12:00 p.m.-4:00 p.m., evening 4:00 p.m.-9:00 p.m., and overnight 9:00 p.m.-7:00 a.m.). The concentrations of protein, albumin, N-acetyl-beta-D-glucosaminidase, and epidermal growth factor (EGF) were measured and normalized by three correction factors (creatinine, osmolality, or specific gravity, respectively). Additionally, the 2nd overnight urine sample was grouped into different aliquots according to centrifugation, additives, storage temperature, or delayed processing. Twenty (14 boys, 6 girls) children were enrolled, with an average age of 11.3 years. Among the three correction factors, creatinine-normalized biomarkers provided the best agreements among different periods over 24 h. There were significant diurnal variations during 24 h in the concentrations of urinary protein, albumin, N-acetyl-beta-D-glucosaminidase, and EGF (p = 0.001, p = 0.003, p = 0.003, and p = 0.003, respectively). Evening urine overestimated 24-h urinary protein and albumin, while overnight urine underestimated 24-h urinary albumin. Urinary EGF showed low variability within a day or between the 2 days (coefficients of variation 10.2% and 10.6%, respectively) and excellent agreements (intraclass correlation coefficients > 0.9) with 24-h urinary concentration. Furthermore, urinary EGF was not affected by centrifugation, additives, storage temperature, or delayed processing of urine samples (all p > 0.05). Conclusion: Given the diurnal variations of urinary biomarkers, urine samples should be collected during the same time period in clinical practice if possible. The results also extend the evidence for urinary EGF as a relatively stable biomarker applied in the future clinical practice. What is Known: ⢠Urinary biomarkers have been widely used or discussed in making diagnoses and therapy regimens and estimating the prognosis of pediatric glomerular diseases. It remains unclear whether their levels would be affected by the time of sample collection, processing methods, and storage conditions in hospitalized children with glomerular diseases. What is New: ⢠The levels of both commonly used biomarkers and novel biomarkers exhibited diurnal variations in hospitalized children with glomerular diseases. ⢠Our results extend the evidence for urinary EGF as a relatively stable biomarker applied in the future clinical practice.
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Acetilglucosaminidase , Criança Hospitalizada , Masculino , Feminino , Humanos , Criança , Creatinina/urina , Acetilglucosaminidase/urina , Biomarcadores/urina , AlbuminasRESUMO
BACKGROUND: This study investigated the association between urinary epidermal growth factor (EGF) and complete remission (CR) of proteinuria in children with IgA nephropathy (IgAN). METHODS: We included 108 patients from the Registry of IgA Nephropathy in Chinese Children. The urinary EGF at the baseline and follow-up were measured and normalized by urine creatinine (expressed as uEGF/Cr). The person-specific uEGF/Cr slopes were estimated using linear mixed-effects models for the subset of patients with longitudinal data of uEGF/Cr. Cox models were used to analyze the associations of baseline uEGF/Cr and uEGF/Cr slope with CR of proteinuria. RESULTS: Patients with high baseline uEGF/Cr were more likely to achieve CR of proteinuria (adjusted HR 2.24, 95% CI: 1.05-4.79). The addition of high baseline uEGF/Cr on the traditional parameters significantly improved the model fit for predicting CR of proteinuria. In the subset of patients with longitudinal data of uEGF/Cr, high uEGF/Cr slope was associated with a higher likelihood of CR of proteinuria (adjusted HR 4.03, 95% CI: 1.02-15.88). CONCLUSIONS: Urinary EGF may be a useful noninvasive biomarker for predicting and monitoring CR of proteinuria in children with IgAN. IMPACT: High levels of baseline uEGF/Cr (>21.45 ng/mg) could serve as an independent predictor for CR of proteinuria. The addition of baseline uEGF/Cr on the traditional clinical pathological parameters significantly improved the fitting ability for the prediction of CR of proteinuria. Longitudinal data of uEGF/Cr were also independently associated with CR of proteinuria. Our study provides evidence that urinary EGF may be a useful noninvasive biomarker in the prediction of CR of proteinuria as well as monitoring therapeutic response, thus guiding treatment strategies in clinical practice for children with IgAN.
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Fator de Crescimento Epidérmico , Glomerulonefrite por IGA , Humanos , Criança , Glomerulonefrite por IGA/complicações , População do Leste Asiático , Taxa de Filtração Glomerular , Proteinúria , Creatinina , BiomarcadoresRESUMO
The diagnosis of nephrotic syndrome relies on clinical presentation and descriptive patterns of injury on kidney biopsies, but not specific to underlying pathobiology. Consequently, there are variable rates of progression and response to therapy within diagnoses. Here, an unbiased transcriptomic-driven approach was used to identify molecular pathways which are shared by subgroups of patients with either minimal change disease (MCD) or focal segmental glomerulosclerosis (FSGS). Kidney tissue transcriptomic profile-based clustering identified three patient subgroups with shared molecular signatures across independent, North American, European, and African cohorts. One subgroup had significantly greater disease progression (Hazard Ratio 5.2) which persisted after adjusting for diagnosis and clinical measures (Hazard Ratio 3.8). Inclusion in this subgroup was retained even when clustering was limited to those with less than 25% interstitial fibrosis. The molecular profile of this subgroup was largely consistent with tumor necrosis factor (TNF) pathway activation. Two TNF pathway urine markers were identified, tissue inhibitor of metalloproteinases-1 (TIMP-1) and monocyte chemoattractant protein-1 (MCP-1), that could be used to predict an individual's TNF pathway activation score. Kidney organoids and single-nucleus RNA-sequencing of participant kidney biopsies, validated TNF-dependent increases in pathway activation score, transcript and protein levels of TIMP-1 and MCP-1, in resident kidney cells. Thus, molecular profiling identified a subgroup of patients with either MCD or FSGS who shared kidney TNF pathway activation and poor outcomes. A clinical trial testing targeted therapies in patients selected using urinary markers of TNF pathway activation is ongoing.
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Glomerulosclerose Segmentar e Focal , Nefrologia , Nefrose Lipoide , Síndrome Nefrótica , Humanos , Glomerulosclerose Segmentar e Focal/patologia , Nefrose Lipoide/diagnóstico , Inibidor Tecidual de Metaloproteinase-1 , Síndrome Nefrótica/diagnóstico , Fatores de Necrose Tumoral/uso terapêuticoRESUMO
Diabetic kidney disease (DKD) is the leading cause of end-stage kidney disease (ESKD). Prognostic biomarkers reflective of underlying molecular mechanisms are critically needed for effective management of DKD. A three-marker panel was derived from a proteomics analysis of plasma samples by an unbiased machine learning approach from participants (N = 58) in the Clinical Phenotyping and Resource Biobank study. In combination with standard clinical parameters, this panel improved prediction of the composite outcome of ESKD or a 40% decline in glomerular filtration rate. The panel was validated in an independent group (N = 68), who also had kidney transcriptomic profiles. One marker, plasma angiopoietin 2 (ANGPT2), was significantly associated with outcomes in cohorts from the Cardiovascular Health Study (N = 3,183) and the Chinese Cohort Study of Chronic Kidney Disease (N = 210). Glomerular transcriptional angiopoietin/Tie (ANG-TIE) pathway scores, derived from the expression of 154 ANG-TIE signaling mediators, correlated positively with plasma ANGPT2 levels and kidney outcomes. Higher receptor expression in glomeruli and higher ANG-TIE pathway scores in endothelial cells corroborated potential functional effects in the kidney from elevated plasma ANGPT2 levels. Our work suggests that ANGPT2 is a promising prognostic endothelial biomarker with likely functional impact on glomerular pathogenesis in DKD.
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Diabetes Mellitus , Nefropatias Diabéticas , Falência Renal Crônica , Humanos , Angiopoietina-1/genética , Receptor TIE-2/genética , Nefropatias Diabéticas/genética , Estudos de Coortes , Células Endoteliais , Angiopoietina-2/genética , Angiopoietinas , Transdução de Sinais , Biomarcadores , Progressão da DoençaRESUMO
Phospholipase Cε (PLCε) is a phospholipase C isoform with a wide range of physiological functions. It has been implicated in aortic valve disorders, but its role in frequently associated aortic disease remains unclear. To determine the role of PLCε in thoracic aortic aneurysm and dissection (TAAD) we used PLCε-deficient mice, which develop aortic valve insufficiency and exhibit aortic dilation of the ascending thoracic aorta and arch without histopathological evidence of injury. Fourteen days of infusion of Plce1+/+ and Plce1-/- mice with angiotensin II (ANG II), which induces aortic dilation and dissection, led to sudden death secondary to ascending aortic dissection in 43% of Plce1-/- versus 5% of Plce1+/+ mice (P < 0.05). Medial degeneration and TAAD were detected in 80% of Plce1-/- compared with 10% of Plce1+/+ mice (P < 0.05) after 4 days of ANG II. Treatment with ANG II markedly increased PLCε expression within the ascending aortic adventitia. Total RNA sequencing demonstrated marked upregulation of inflammatory and fibrotic pathways mediated by interleukin-1ß, interleukin-6, and tumor necrosis factor-α. In silico analysis of whole exome sequences of 258 patients with type A dissection identified 5 patients with nonsynonymous PLCE1 variants. Our data suggest that PLCε deficiency plays a role in the development of TAAD and aortic insufficiency.NEW & NOTEWORTHY We describe a novel phenotype by which PLCε deficiency predisposes to aortic valve insufficiency and ascending aortic aneurysm, dissection, and sudden death in the setting of ANG II-mediated hypertension. We demonstrate PLCE1 variants in patients with type A aortic dissection and aortic insufficiency, suggesting that PLCE1 may also play a role in human aortic disease. This finding is of very high significance because it has not been previously demonstrated that PLCε directly mediates aortic dissection.
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Aneurisma da Aorta Ascendente , Aneurisma da Aorta Torácica , Aneurisma Aórtico , Dissecção Aórtica , Insuficiência da Valva Aórtica , Hipertensão , Humanos , Camundongos , Animais , Insuficiência da Valva Aórtica/genética , Camundongos Endogâmicos C57BL , Aneurisma Aórtico/genética , Aneurisma Aórtico/patologia , Dissecção Aórtica/genética , Angiotensina II , Morte Súbita , Aneurisma da Aorta Torácica/genética , Aneurisma da Aorta Torácica/metabolismoRESUMO
Research on kidney diseases is being transformed by the rapid expansion and innovations in omics technologies. The analysis, integration, and interpretation of big data, however, have been an impediment to the growing interest in applying these technologies to understand kidney function and failure. Targeting this urgent need, the University of Michigan O'Brien Kidney Translational Core Center (MKTC) and its Administrative Core established the Applied Systems Biology Core. The Core provides need-based support for the global kidney community centered on enabling incorporation of systems biology approaches by creating web-based, user-friendly analytic and visualization tools, like Nephroseq and Nephrocell, guiding with experimental design, and processing, analysis, and integration of large data sets. The enrichment core supports systems biology education and dissemination through workshops, seminars, and individualized training sessions. Meanwhile, the Pilot and Feasibility Program of the MKTC provides pilot funding to both early-career and established investigators new to the field, to integrate a systems biology approach into their research projects. The relevance and value of the portfolio of training and services offered by MKTC are reflected in the expanding community of young investigators, collaborators, and users accessing resources and engaging in systems biology-based kidney research, thereby motivating MKTC to persevere in its mission to serve the kidney research community by enabling access to state-of-the-art data sets, tools, technologies, expertise, and learning opportunities for transformative basic, translational, and clinical studies that will usher in solutions to improve the lives of people impacted by kidney disease.
Assuntos
Nefropatias , Biologia de Sistemas , Humanos , Rim , Michigan , Pesquisa Translacional BiomédicaRESUMO
OBJECTIVE: Understanding mechanisms underlying rapid estimated glomerular filtration rate (eGFR) decline is important to predict and treat kidney disease in type 1 diabetes (T1D). RESEARCH DESIGN AND METHODS: We performed a case-control study nested within four T1D cohorts to identify urinary proteins associated with rapid eGFR decline. Case and control subjects were categorized based on eGFR decline ≥3 and <1 mL/min/1.73 m2/year, respectively. We used targeted liquid chromatography-tandem mass spectrometry to measure 38 peptides from 20 proteins implicated in diabetic kidney disease. Significant proteins were investigated in complementary human cohorts and in mouse proximal tubular epithelial cell cultures. RESULTS: The cohort study included 1,270 participants followed a median 8 years. In the discovery set, only cathepsin D peptide and protein were significant on full adjustment for clinical and laboratory variables. In the validation set, associations of cathepsin D with eGFR decline were replicated in minimally adjusted models but lost significance with adjustment for albuminuria. In a meta-analysis with combination of discovery and validation sets, the odds ratio for the association of cathepsin D with rapid eGFR decline was 1.29 per SD (95% CI 1.07-1.55). In complementary human cohorts, urine cathepsin D was associated with tubulointerstitial injury and tubulointerstitial cathepsin D expression was associated with increased cortical interstitial fractional volume. In mouse proximal tubular epithelial cell cultures, advanced glycation end product-BSA increased cathepsin D activity and inflammatory and tubular injury markers, which were further increased with cathepsin D siRNA. CONCLUSIONS: Urine cathepsin D is associated with rapid eGFR decline in T1D and reflects kidney tubulointerstitial injury.
Assuntos
Diabetes Mellitus Tipo 1 , Nefropatias Diabéticas , Albuminúria , Animais , Biomarcadores/metabolismo , Estudos de Casos e Controles , Catepsina D , Estudos de Coortes , Progressão da Doença , Taxa de Filtração Glomerular , Humanos , Camundongos , Proteômica/métodosRESUMO
Fibrinogen-like 2 (FGL2) was recently found to be associated with fibrosis in a mouse model of kidney damage and was proposed as a potential therapeutic target in chronic kidney disease (CKD). We assessed the association of renal FGL2 mRNA expression with the disease outcome in two independent CKD cohorts (NEPTUNE and Innsbruck CKD cohort) using Kaplan Meier survival analysis. The regulation of FGL2 in kidney biopsies of CKD patients as compared to healthy controls was further assessed in 13 human CKD transcriptomics datasets. The FGL2 protein expression in human renal tissue sections was determined via immunohistochemistry. The regulators of FGL2 mRNA expression in renal tissue were identified in the co-expression and upstream regulator analysis of FGL2-positive renal cells via the use of single-cell RNA sequencing data from the kidney precision medicine project (KPMP). Higher renal FGL2 mRNA expression was positively associated with kidney fibrosis and negatively associated with eGFR. Renal FGL2 mRNA expression was upregulated in CKD as compared with healthy controls and associated with CKD progression in the Innsbruck CKD cohort (p-value = 0.0036) and NEPTUNE cohort (p-value = 0.0048). The highest abundance of FGL2 protein in renal tissue was detected in the thick ascending limb of the loop of Henle and macula densa, proximal tubular cells, as well as in glomerular endothelial cells. The upstream regulator analysis identified TNF, IL1B, IFNG, NFKB1, and SP1 as factors potentially inducing FGL2-co-expressed genes, whereas factors counterbalancing FGL2-co-expressed genes included GLI1, HNF1B, or PPARGC1A. In conclusion, renal FGL2 mRNA expression is elevated in human CKD, and higher FGL2 levels are associated with fibrosis and worse outcomes.