RESUMEN
BACKGROUND: Acute kidney injury (AKI) is common in lupus nephritis (LN) and a risk factor for development of chronic kidney disease. In adults with LN, AKI severity correlates with the incidence of kidney failure and patient survival. Data on AKI outcomes in children with LN, particularly those requiring kidney replacement therapy (KRT), are limited. METHODS: A multicenter, retrospective cohort study was performed in children diagnosed between 2010 and 2019 with LN and AKI stage 3 treated with dialysis (AKI stage 3D). Descriptive statistics were used to characterize demographics, clinical data, and kidney biopsy findings; treatment data for LN were not included. Logistic regression was used to examine the association of these variables with kidney failure. RESULTS: Fifty-nine patients (mean age 14.3 years, 84.7% female) were identified. The most common KRT indications were fluid overload (86.4%) and elevated blood urea nitrogen/creatinine (74.6%). Mean follow-up duration was 3.9 ± 2.9 years. AKI recovery without progression to kidney failure occurred in 37.3% of patients. AKI recovery with later progression to kidney failure occurred in 25.4% of patients, and there was no kidney recovery from AKI in 35.6% of patients. Older age, severe (> 50%) tubular atrophy and interstitial fibrosis, and National Institutes of Health (NIH) chronicity index score > 4 on kidney biopsy were associated with kidney failure. CONCLUSIONS: Children with LN and AKI stage 3D have a high long-term risk of kidney failure. Severe tubular atrophy and interstitial fibrosis at the time of AKI, but not AKI duration, are predictive of kidney disease progression. A higher resolution version of the Graphical abstract is available as Supplementary information.
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Lesión Renal Aguda , Artritis Juvenil , Nefritis Lúpica , Nefrología , Reumatología , Adulto , Niño , Humanos , Femenino , Adolescente , Masculino , Nefritis Lúpica/complicaciones , Nefritis Lúpica/terapia , Nefritis Lúpica/diagnóstico , Estudios de Cohortes , Estudios Retrospectivos , Artritis Juvenil/complicaciones , Diálisis Renal , Lesión Renal Aguda/epidemiología , Lesión Renal Aguda/etiología , Lesión Renal Aguda/terapia , Fibrosis , Atrofia/complicacionesRESUMEN
The urinary tract is usually culture negative despite its close proximity to microbial flora. The precise mechanism by which the kidneys and urinary tract defends against infection is not well understood. The initial kidney cells to encounter ascending pathogens are the collecting tubule cells that consist of principal cells (PCs) that express aquaporin 2 (AQP2) and intercalated cells (ICs) that express vacuolar H+-ATPase (V-ATPase, B1 subunit). We have previously shown that ICs are involved with the human renal innate immune defense. Here we generated two reporter mice, VATPase B1-cre+tdT+ mice to fluorescently label ICs and AQP2-cre+tdT+ mice to fluorescently label PCs, and then performed flow sorting to enrich PCs and ICs for analysis. Isolated ICs and PCs along with proximal tubular cells were used to measure antimicrobial peptide (AMP) mRNA expression. ICs and PCs were significantly enriched for AMPs. Isolated ICs responded to uropathogenic Escherichia coli (UPEC) challenge in vitro and had higher RNase4 gene expression than control while both ICs and PCs responded to UPEC challenge in vivo by upregulating Defb1 mRNA expression. To our knowledge, this is the first report of isolating murine collecting tubule cells and performing targeted analysis for multiple classes of AMPs.
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Acuaporina 2/inmunología , Células Epiteliales/metabolismo , Túbulos Renales Colectores/inmunología , Reacción en Cadena de la Polimerasa , Animales , Acuaporina 2/genética , Inmunidad Innata/inmunología , Riñón/inmunología , Riñón/metabolismo , Ratones Transgénicos , Reacción en Cadena de la Polimerasa/métodos , Regulación hacia Arriba/inmunología , ATPasas de Translocación de Protón Vacuolares/inmunología , ATPasas de Translocación de Protón Vacuolares/metabolismoRESUMEN
Hypertension affects one billion people and is a principal reversible risk factor for cardiovascular disease. Pseudohypoaldosteronism type II (PHAII), a rare Mendelian syndrome featuring hypertension, hyperkalaemia and metabolic acidosis, has revealed previously unrecognized physiology orchestrating the balance between renal salt reabsorption and K(+) and H(+) excretion. Here we used exome sequencing to identify mutations in kelch-like 3 (KLHL3) or cullin 3 (CUL3) in PHAII patients from 41 unrelated families. KLHL3 mutations are either recessive or dominant, whereas CUL3 mutations are dominant and predominantly de novo. CUL3 and BTB-domain-containing kelch proteins such as KLHL3 are components of cullin-RING E3 ligase complexes that ubiquitinate substrates bound to kelch propeller domains. Dominant KLHL3 mutations are clustered in short segments within the kelch propeller and BTB domains implicated in substrate and cullin binding, respectively. Diverse CUL3 mutations all result in skipping of exon 9, producing an in-frame deletion. Because dominant KLHL3 and CUL3 mutations both phenocopy recessive loss-of-function KLHL3 mutations, they may abrogate ubiquitination of KLHL3 substrates. Disease features are reversed by thiazide diuretics, which inhibit the Na-Cl cotransporter in the distal nephron of the kidney; KLHL3 and CUL3 are expressed in this location, suggesting a mechanistic link between KLHL3 and CUL3 mutations, increased Na-Cl reabsorption, and disease pathogenesis. These findings demonstrate the utility of exome sequencing in disease gene identification despite the combined complexities of locus heterogeneity, mixed models of transmission and frequent de novo mutation, and establish a fundamental role for KLHL3 and CUL3 in blood pressure, K(+) and pH homeostasis.
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Proteínas Portadoras/genética , Proteínas Cullin/genética , Hipertensión/genética , Mutación/genética , Seudohipoaldosteronismo/genética , Desequilibrio Hidroelectrolítico/genética , Proteínas Adaptadoras Transductoras de Señales , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Presión Sanguínea/genética , Proteínas Portadoras/química , Estudios de Cohortes , Proteínas Cullin/química , Electrólitos , Exones/genética , Femenino , Perfilación de la Expresión Génica , Genes Dominantes/genética , Genes Recesivos/genética , Genotipo , Homeostasis/genética , Humanos , Concentración de Iones de Hidrógeno , Hipertensión/complicaciones , Hipertensión/fisiopatología , Masculino , Ratones , Proteínas de Microfilamentos , Modelos Moleculares , Datos de Secuencia Molecular , Fenotipo , Potasio/metabolismo , Seudohipoaldosteronismo/complicaciones , Seudohipoaldosteronismo/fisiopatología , Cloruro de Sodio/metabolismo , Desequilibrio Hidroelectrolítico/complicaciones , Desequilibrio Hidroelectrolítico/fisiopatologíaRESUMEN
Genetic inactivation of the epithelial Na(+) channel α-subunit (αENaC) in the renal collecting duct (CD) does not interfere with Na(+) and K(+) homeostasis in mice. However, inactivation in the CD and a part of the connecting tubule (CNT) induces autosomal recessive pseudohypoaldosteronism type 1 (PHA-1) symptoms in subjects already on a standard diet. In the present study, we further examined the importance of αENaC in the CNT. Knockout mice with αENaC deleted primarily in a part of the CNT (CNT-KO) were generated using Scnn1a(lox/lox) mice and Atp6v1b1::Cre mice. With a standard diet, plasma Na(+) concentration ([Na(+)]) and [K(+)], and urine Na(+) and K(+) output were unaffected. Seven days of Na(+) restriction (0.01% Na(+)) led to a higher urine Na(+) output only on days 3-5, and after 7 days plasma [Na(+)] and [K(+)] were unaffected. In contrast, the CNT-KO mice were highly susceptible to a 2-day 5% K(+) diet and showed lower food intake and relative body weight, lower plasma [Na(+)], higher fractional excretion (FE) of Na(+), higher plasma [K(+)], and lower FE of K(+). The higher FE of Na(+) coincided with lower abundance and phosphorylation of the Na(+)-Cl(-) cotransporter. In conclusion, reducing ENaC expression in the CNT induces clear PHA-1 symptoms during high dietary K(+) loading.
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Canales Epiteliales de Sodio/biosíntesis , Túbulos Renales Colectores/metabolismo , Potasio/metabolismo , Seudohipoaldosteronismo/genética , Seudohipoaldosteronismo/metabolismo , Aldosterona/metabolismo , Animales , Peso Corporal , Colon/metabolismo , Dieta , Ingestión de Alimentos , Canales Epiteliales de Sodio/genética , Femenino , Túbulos Renales Colectores/patología , Masculino , Ratones , Ratones Noqueados , Fosforilación , Potasio/sangre , Seudohipoaldosteronismo/patología , Sodio/sangre , Sodio/metabolismo , Miembro 1 de la Familia de Transportadores de Soluto 12/biosíntesis , Miembro 1 de la Familia de Transportadores de Soluto 12/genética , Miembro 3 de la Familia de Transportadores de Soluto 12/biosíntesis , Miembro 3 de la Familia de Transportadores de Soluto 12/genéticaRESUMEN
BACKGROUND: Cross-sectional studies of children with prevalent nephrotic syndrome (NS) have shown 25-vitamin D (25(OH)D) deficiency rates of 20-100 %. Information on 25(OH)D status in incident patients or following remission is limited. This study aimed to assess 25(OH)D status of incident idiopathic NS children at presentation and longitudinally with short-term observation. METHODS: Multicenter longitudinal study of children (2-18 years old) from 14 centers across the Midwest Pediatric Nephrology Consortium with incident idiopathic NS. 25(OH)D levels were assessed at diagnosis and 3 months later. RESULTS: Sixty-one children, median age 5 (3, 11) years, completed baseline visit and 51 completed second visit labs. All 61 (100 %) had 25(OH)D < 20 ng/ml at diagnosis. Twenty-seven (53 %) had 25(OH)D < 20 ng/ml at follow-up. Fourteen (28 %) children were steroid resistant. Univariate analysis showed that children prescribed vitamin D supplements were less likely to have 25(OH)D deficiency at follow-up (OR 0.2, 95 % CI 0.04, 0.6). Steroid response, age, and season did not predict 25(OH)D deficiency. Multivariable linear regression modeling showed higher 25(OH)D levels at follow-up by 13.2 ng/ml (SE 4.6, p < 0.01) in children supplemented with vitamin D. CONCLUSIONS: In this incident idiopathic NS cohort, all children at diagnosis had 25(OH)D deficiency and the majority continued to have a deficiency at 2-4 months. Supplemental vitamin D decreased the odds of 25(OH)D deficiency at follow-up, supporting a role for supplementation in incident NS.
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Síndrome Nefrótico/epidemiología , Deficiencia de Vitamina D/epidemiología , Vitamina D/análogos & derivados , Adolescente , Biomarcadores/sangre , Niño , Preescolar , Suplementos Dietéticos , Femenino , Humanos , Incidencia , Modelos Lineales , Modelos Logísticos , Estudios Longitudinales , Masculino , Medio Oeste de Estados Unidos/epidemiología , Análisis Multivariante , Síndrome Nefrótico/diagnóstico , Oportunidad Relativa , Hormona Paratiroidea/sangre , Estudios Prospectivos , Factores de Tiempo , Resultado del Tratamiento , Vitamina D/sangre , Vitamina D/uso terapéutico , Deficiencia de Vitamina D/sangre , Deficiencia de Vitamina D/diagnóstico , Deficiencia de Vitamina D/tratamiento farmacológicoRESUMEN
Inherited and acquired disorders that enhance the activity of transporters mediating renal tubular Na(+) reabsorption are well established causes of hypertension. It is unclear, however, whether primary activation of an Na(+)-independent chloride transporter in the kidney can also play a pathogenic role in this disease. Here, mice overexpressing the chloride transporter pendrin in intercalated cells of the distal nephron (Tg(B1-hPDS) mice) displayed increased renal absorption of chloride. Compared with normal mice, these transgenic mice exhibited a delayed increase in urinary NaCl and ultimately, developed hypertension when exposed to a high-salt diet. Administering the same sodium intake as NaHCO3 instead of NaCl did not significantly alter BP, indicating that the hypertension in the transgenic mice was chloride-sensitive. Moreover, excessive chloride absorption by pendrin drove parallel absorption of sodium through the epithelial sodium channel ENaC and the sodium-driven chloride/bicarbonate exchanger (Ndcbe), despite an appropriate downregulation of these sodium transporters in response to the expanded vascular volume and hypertension. In summary, chloride transport in the distal nephron can play a primary role in driving NaCl transport in this part of the kidney, and a primary abnormality in renal chloride transport can provoke arterial hypertension. Thus, we conclude that the chloride/bicarbonate exchanger pendrin plays a major role in controlling net NaCl absorption, thereby influencing BP under conditions of high salt intake.
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Presión Sanguínea/fisiología , Antiportadores de Cloruro-Bicarbonato/metabolismo , Cloruros/metabolismo , Hipertensión/metabolismo , Riñón/metabolismo , Proteínas de Transporte de Membrana/metabolismo , Nefronas/metabolismo , Cloruro de Sodio/metabolismo , Animales , Humanos , Inmunohistoquímica , Transporte Iónico , Ratones , Ratones Transgénicos , Transportadores de SulfatoRESUMEN
BACKGROUND: Nephritis is a common manifestation of IgA vasculitis and is morphologically indistinguishable from IgA nephropathy. While MEST-C scores are predictive of kidney outcomes in IgA nephropathy, their value in IgA vasculitis nephritis has not been investigated in large multiethnic cohorts. METHODS: Biopsies from 262 children and 99 adults with IgA vasculitis nephritis ( N =361) from 23 centers in North America, Europe, and Asia were independently scored by three pathologists. MEST-C scores were assessed for correlation with eGFR/proteinuria at biopsy. Because most patients ( N =309, 86%) received immunosuppression, risk factors for outcomes were evaluated in this group using latent class mixed models to identify classes of eGFR trajectories over a median follow-up of 2.7 years (interquartile range, 1.2-5.1). Clinical and histologic parameters associated with each class were determined using logistic regression. RESULTS: M, E, T, and C scores were correlated with either eGFR or proteinuria at biopsy. Two classes were identified by latent class mixed model, one with initial improvement in eGFR followed by a late decline (class 1, N =91) and another with stable eGFR (class 2, N =218). Class 1 was associated with a higher risk of an established kidney outcome (time to ≥30% decline in eGFR or kidney failure; hazard ratio, 5.84; 95% confidence interval, 2.37 to 14.4). Among MEST-C scores, only E1 was associated with class 1 by multivariable analysis. Other factors associated with class 1 were age 18 years and younger, male sex, lower eGFR at biopsy, and extrarenal noncutaneous disease. Fibrous crescents without active changes were associated with class 2. CONCLUSIONS: Kidney outcome in patients with biopsied IgA vasculitis nephritis treated with immunosuppression was determined by clinical risk factors and endocapillary hypercellularity (E1) and fibrous crescents, which are features that are not part of the International Study of Diseases of Children classification.
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Glomerulonefritis por IGA , Vasculitis por IgA , Nefritis , Adulto , Niño , Humanos , Masculino , Adolescente , Glomerulonefritis por IGA/complicaciones , Glomerulonefritis por IGA/tratamiento farmacológico , Glomerulonefritis por IGA/patología , Vasculitis por IgA/complicaciones , Vasculitis por IgA/tratamiento farmacológico , Vasculitis por IgA/patología , Tasa de Filtración Glomerular , Riñón/patología , Nefritis/complicaciones , Proteinuria/etiología , Biopsia , Estudios RetrospectivosRESUMEN
The kidney collecting system develops from branching morphogenesis of the ureteric bud (UB). This process requires signaling by growth factors such as glial cell line derived neurotrophic factor (GDNF) and fibroblast growth factors (FGFs) as well as cell extracellular matrix interactions mediated by integrins. The importance of integrin signaling in UB development was investigated by deleting integrin beta1 at initiation (E10.5) and late (E18.5) stages of development. Deletion at E10.5 resulted in a severe branching morphogenesis phenotype. Deletion at E18.5 did not alter renal development but predisposed the collecting system to severe injury following ureteric obstruction. beta1 integrin was required for renal tubular epithelial cells to mediate GDNF- and FGF-dependent signaling despite normal receptor localization and activation in vitro. Aberrations in the same signaling molecules were present in the beta1-null UBs in vivo. Thus beta1 integrins can regulate organ branching morphogenesis during development by mediating growth-factor-dependent signaling in addition to their well-defined role as adhesion receptors.
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Integrina beta1/metabolismo , Túbulos Renales Colectores/embriología , Túbulos Renales Colectores/metabolismo , Uréter/embriología , Uréter/metabolismo , Animales , Adhesión Celular , Línea Celular , Movimiento Celular , Proliferación Celular , Eliminación de Gen , Regulación del Desarrollo de la Expresión Génica , Edad Gestacional , Sustancias de Crecimiento/metabolismo , Integrina beta1/genética , Túbulos Renales Colectores/citología , Ratones , Ratones Noqueados , Ratones Transgénicos , Morfogénesis , Técnicas de Cultivo de Órganos , Transducción de SeñalRESUMEN
We used a systems biology-based approach to investigate the basis of cell-specific expression of the water channel aquaporin-2 (AQP2) in the renal collecting duct. Computational analysis of the 5'-flanking region of the AQP2 gene (Genomatix) revealed 2 conserved clusters of putative transcriptional regulator (TR) binding elements (BEs) centered at -513 bp (corresponding to the SF1, NFAT, and FKHD TR families) and -224 bp (corresponding to the AP2, SRF, CREB, GATA, and HOX TR families). Three other conserved motifs corresponded to the ETS, EBOX, and RXR TR families. To identify TRs that potentially bind to these BEs, we carried out mRNA profiling (Affymetrix) in mouse mpkCCDc14 collecting duct cells, revealing expression of 25 TRs that are also expressed in native inner medullary collecting duct. One showed a significant positive correlation with AQP2 mRNA abundance among mpkCCD subclones (Ets1), and 2 showed a significant negative correlation (Elf1 and an orphan nuclear receptor Nr1h2). Transcriptomic profiling in native proximal tubules (PT), medullary thick ascending limbs (MTAL), and IMCDs from kidney identified 14 TRs (including Ets1 and HoxD3) expressed in the IMCD but not PT or MTAL (candidate AQP2 enhancer roles), and 5 TRs (including HoxA5, HoxA9 and HoxA10) expressed in PT and MTAL but not in IMCD (candidate AQP2 repressor roles). In luciferase reporter assays, overexpression of 3 ETS family TRs transactivated the mouse proximal AQP2 promoter. The results implicate ETS family TRs in cell-specific expression of AQP2 and point to HOX, RXR, CREB and GATA family TRs as playing likely additional roles.
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Acuaporina 2/biosíntesis , Túbulos Renales Colectores/metabolismo , Animales , Acuaporina 2/fisiología , Clonación Molecular , Biología Computacional , Genes Reporteros , Ratones , Microscopía Confocal , Microscopía Fluorescente , Modelos Biológicos , Modelos Genéticos , Análisis de Secuencia por Matrices de Oligonucleótidos , Sondas ARN , ARN Mensajero/metabolismo , Biología de SistemasRESUMEN
AVP resistance of the medullary collecting duct (mCD) in postobstructive uropathy (POU) has been attributed to increased production of PGE2. P2Y2 receptor activation causes production of PGE2 by the mCD. We hypothesize that increased P2Y2 receptor expression and/or activity may contribute to the diuresis of POU. Sprague-Dawley rats were subjected to bilateral ureteral obstruction for 24 h followed by release (BUO/R, n = 17) or sham operation (SHM/O, n = 15) and euthanized after 1 wk or 12 days. BUO/R rats developed significant polydipsia, polyuria, urinary concentration defect, and increased urinary PGE2 and decreased aquaporin-2 protein abundance in the inner medulla compared with SHM/O rats. After BUO/R, the relative mRNA expression of P2Y2 and P2Y6 receptors was increased by 2.7- and 4.9-fold, respectively, without significant changes in mRNA expression of P2Y1 or P2Y4 receptor. This was associated with a significant 3.5-fold higher protein abundance of the P2Y2 receptor in BUO/R than SHM/O rats. When freshly isolated mCD fractions were challenged with different types of nucleotides (ATPgammaS, ADP, UTP, or UDP), BUO/R and SHM/O rats responded to only ATPgammaS and UTP and released PGE2, consistent with involvement of the P2Y2, but not P2Y6, receptor. ATPgammaS- or UTP-stimulated increases in PGE2 were much higher in BUO/R (3.20- and 2.28-fold, respectively, vs. vehicle controls) than SHM/O (1.68- and 1.30-fold, respectively, vs. vehicle controls) rats. In addition, there were significant 2.4- and 2.1-fold increases in relative mRNA expression of prostanoid EP1 and EP3 receptors, respectively, in the inner medulla of BUO/R vs. SHM/O rats. Taken together, these data suggest that increased production of PGE2 by the mCD in POU may be due to increased expression and activity of the P2Y2 receptor. Increased mRNA expression of EP1 and EP3 receptors in POU may also help accentuate PGE2-induced signaling in the mCD.
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Diuresis , Túbulos Renales Colectores/metabolismo , Receptores Purinérgicos P2/metabolismo , Obstrucción Ureteral/metabolismo , Adenosina Trifosfato/análogos & derivados , Adenosina Trifosfato/farmacología , Animales , Dinoprostona/orina , Modelos Animales de Enfermedad , Diuresis/efectos de los fármacos , Diuresis/genética , Regulación de la Expresión Génica , Capacidad de Concentración Renal , Túbulos Renales Colectores/efectos de los fármacos , Túbulos Renales Colectores/fisiopatología , Masculino , Poliuria/etiología , Poliuria/metabolismo , Poliuria/fisiopatología , Agonistas del Receptor Purinérgico P2 , ARN Mensajero/metabolismo , Ratas , Ratas Sprague-Dawley , Receptores de Prostaglandina E/metabolismo , Subtipo EP1 de Receptores de Prostaglandina E , Subtipo EP3 de Receptores de Prostaglandina E , Receptores Purinérgicos P2/genética , Receptores Purinérgicos P2Y2 , Transducción de Señal , Factores de Tiempo , Regulación hacia Arriba , Obstrucción Ureteral/complicaciones , Obstrucción Ureteral/genética , Obstrucción Ureteral/fisiopatología , Uridina Trifosfato/metabolismoRESUMEN
Rh C glycoprotein (Rhcg) is an NH(3)-specific transporter expressed in both intercalated cells (IC) and principal cells (PC) in the renal collecting duct. Recent studies show that deletion of Rhcg from both intercalated and principal cells inhibits both basal and acidosis-stimulated renal ammonia excretion. The purpose of the current studies was to better understand the specific role of Rhcg expression in intercalated cells in basal and metabolic acidosis-stimulated renal ammonia excretion. We generated mice with intercalated cell-specific Rhcg deletion (IC-Rhcg-KO) using Cre-loxP techniques; control (C) mice were floxed Rhcg but Cre negative. Under basal conditions, IC-Rhcg-KO and C mice excreted urine with similar ammonia content and pH. Mice were then acid loaded by adding HCl to their diet. Ammonia excretion after acid loading increased similarly in IC-Rhcg-KO and C mice during the first 2 days of acid loading but on day 3 was significantly less in IC-Rhcg-KO than in C mice. During the first 2 days of acid loading, urine was significantly more acidic in IC-Rhcg-KO mice than in C mice; there was no difference on day 3. In IC-Rhcg-KO mice, acid loading increased principal cell Rhcg expression in both the cortex and outer medulla as well as expression of another ammonia transporter, Rh glycoprotein B (Rhbg), in principal cells in the outer medulla. We conclude that 1) Rhcg expression in intercalated cells is necessary for the normal renal response to metabolic acidosis; 2) principal cell Rhcg contributes to both basal and acidosis-stimulated ammonia excretion; and 3) adaptations in Rhbg expression occur in response to acid-loading.
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Acidosis/metabolismo , Amoníaco/orina , Proteínas de Transporte de Catión/deficiencia , Túbulos Renales Colectores/metabolismo , Glicoproteínas de Membrana/deficiencia , Acidosis/genética , Acidosis/fisiopatología , Adaptación Fisiológica , Animales , Bicarbonatos/sangre , Proteínas de Transporte de Catión/genética , Modelos Animales de Enfermedad , Glicoproteínas/metabolismo , Concentración de Iones de Hidrógeno , Integrasas/genética , Túbulos Renales Colectores/fisiopatología , Glicoproteínas de Membrana/genética , Proteínas de Transporte de Membrana/metabolismo , Ratones , Ratones Noqueados , Potasio/sangre , Regiones Promotoras Genéticas , Sodio/sangre , Factores de Tiempo , ATPasas de Translocación de Protón Vacuolares/genéticaRESUMEN
Rh B glycoprotein (Rhbg) is a member of the Rh glycoprotein family of ammonia transporters. In the current study, we examine Rhbg's role in basal and acidosis-stimulated acid-base homeostasis. Metabolic acidosis induced by HCl administration increased Rhbg expression in both the cortex and outer medulla. To test the functional significance of increased Rhbg expression, we used a Cre-loxP approach to generate mice with intercalated cell-specific Rhbg knockout (IC-Rhbg-KO). On normal diet, intercalated cell-specific Rhbg deletion did not alter urine ammonia excretion, pH, or titratable acid excretion significantly, but it did decrease glutamine synthetase expression in the outer medulla significantly. After metabolic acidosis was induced, urinary ammonia excretion was significantly less in IC-Rhbg-KO than in control (C) mice on days 2-4 of acid loading, but not on day 5. Urine pH and titratable acid excretion and dietary acid intake did not differ significantly between acid-loaded IC-Rhcg-KO and C mice. In IC-Rhbg-KO mice, acid loading increased connecting segment (CNT) cell and outer medullary collecting duct principal cell Rhbg expression. In both C and IC-Rhbg-KO mice, acid loading decreased glutamine synthetase in both the cortex and outer medulla; the decrease on day 3 was similar in IC-Rhbg-KO and C mice, but on day 5 it was significantly greater in IC-Rhbg-KO than in C mice. We conclude 1) intercalated cell Rhbg contributes to acidosis-stimulated renal ammonia excretion, 2) Rhbg in CNT and principal cells may contribute to renal ammonia excretion, and 3) decreased glutamine synthetase expression may enable normal rates of ammonia excretion under both basal conditions and on day 5 of acid loading in IC-Rhbg-KO mice.
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Amoníaco/orina , Glicoproteínas/metabolismo , Riñón/metabolismo , Proteínas de Transporte de Membrana/metabolismo , Acidosis/genética , Acidosis/metabolismo , Ácidos/farmacología , Alelos , Animales , Bicarbonatos/sangre , Western Blotting , Dieta , Electrólitos/metabolismo , Eliminación de Gen , Regulación Enzimológica de la Expresión Génica/fisiología , Glutamato-Amoníaco Ligasa/biosíntesis , Glicoproteínas/genética , Inmunohistoquímica , Proteínas de Transporte de Membrana/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Potasio/sangreRESUMEN
Background: Kidney transplant (KT) recipients have higher incidence of malignancies, including Human Papillomavirus (HPV)-associated cancers. Thus, HPV vaccines may have an important role in preventing HPV-related disease in this population; however, immunogenicity and safety data are lacking. Objective: To examine the immunological response and tolerability to HPV vaccination in pediatric KT recipients compared to future KT candidates. Methods: The quadrivalent HPV vaccine was administered to girls and boys age 9-18 recruited from seven centers part of the Pediatric Nephrology Research Consortium. Subjects were recruited for three groups: (1) CKD: chronic kidney disease stages 3, 4, and 5 not on dialysis; (2) Dialysis; (3) KT recipients. The outcome consisted of antibody concentrations against HPV 6, 11, 16, and 18. Geometric mean titers (GMTs) and seroconversion rates were compared. Vaccine tolerability was assessed. Results: Sixty-five participants were recruited: 18 in the CKD, 18 in the dialysis, and 29 into the KT groups. KT patients had significantly lower GMTs after vaccination for all serotypes. The percentages of subjects who reached seroconversion were overall lower for the KT group, reaching statistical significance for HPV 6, 11, and 18. Comparing immunosuppressed subjects (anyone taking immunosuppression medications, whether KT recipient or not) with the non-immunosuppressed participants, the former had significantly lower GMTs for all the HPV serotypes and lower seroconversion rates for HPV 6, 11, and 18. KT females had higher GMTs and seroconversion rates for certain serotypes. There were no adverse events in either group. Conclusions: HPV vaccine was well-tolerated in this population. Pediatric KT recipients had in general lower GMTs and seroconversion rates compared to their peers with CKD or on dialysis. Immunosuppression played a role in the lack of seroconversion. Our results emphasize the importance of advocating for HPV vaccination prior to KT and acknowledge its safety post transplantation. Future studies are needed to investigate the effect of a supplemental dose of HPV vaccine in KT recipients who do not seroconvert and to evaluate the long-term persistence of antibodies post-KT.
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An amendment to this paper has been published and can be accessed via a link at the top of the paper.
RESUMEN
The collecting duct of the kidney is composed of two morphologically and physiologically distinct cell types, principal and intercalated cells. To better understand intercalated cell function we generated a transgenic mouse expressing Cre recombinase under the control of a cell type- specific promoter. We used 7 kb of the ATP6V1B1 5' untranslated region (B1 promoter), a gene found in the intercalated cells of the kidney and the male reproductive tract. We first crossed these B1-Cre transgenic mice with the ROSA26-loxP-stop-loxP-yellow fluorescent protein reporter mice to assess the specificity of Cre expression. Immunohistochemistry and confocal fluorescence microscopy showed that Cre is selectively active in all intercalated cells (type A, type B, and non-A/B cells) within the collecting duct and most cells of the connecting segment. About half of the principal cells of the connecting segment also expressed Cre, a pattern also seen in B1-driven enhanced green fluorescent protein transgenic mice. Cre was found to be active in the male reproductive tract and at a low level in limited non-ATP6V1B1 expressing tissues. The B1-Cre transgenic mice are healthy, breed normally, produce regular sized litters, and transmit the transgene in Mendelian fashion. This new cell-specific Cre expressing mouse should prove useful for the study of intercalated cell physiology and development.
Asunto(s)
Integrasas/biosíntesis , Túbulos Renales Colectores/enzimología , Riñón/citología , Regiones Promotoras Genéticas/fisiología , ATPasas de Translocación de Protón Vacuolares/genética , Animales , Femenino , Genitales Masculinos , Humanos , Integrasas/análisis , Riñón/enzimología , Proteínas Luminiscentes , Masculino , Ratones , Ratones Transgénicos , ATPasas de Translocación de Protón Vacuolares/análisisRESUMEN
Renal cysts in autosomal dominant polycystic kidney disease arise from cells throughout the nephron, but there is an uncertainty as to whether both the intercalated cells (ICs) and principal cells (PCs) within the collecting duct give rise to cysts. To determine this, we crossed mice containing loxP sites within introns 1 and 4 of the Pkd1 gene with transgenic mice expressing Cre recombinase under control of the aquaporin-2 promoter or the B1 subunit of the proton ATPase promoter, thereby generating PC- or IC-specific knockout of Pkd1, respectively. Mice, that had Pkd1 deleted in the PCs, developed progressive cystic kidney disease evident during the first postnatal week and had an average lifespan of 8.2 weeks. There was no change in the cellular cAMP content or membrane aquaporin-2 expression in their kidneys. Cysts were present in the cortex and outer medulla but were absent in the papilla. Mice in which PKd1 was knocked out in the ICs had a very mild cystic phenotype as late as 13 weeks of age, limited to 1-2 cysts and confined to the outer rim of the kidney cortex. These mice lived to at least 1.5 years of age without evidence of early mortality. Our findings suggest that PCs are more important than ICs for cyst formation in polycystic kidney disease.
Asunto(s)
Enfermedades Renales Quísticas/genética , Enfermedades Renales Quísticas/patología , Canales Catiónicos TRPP/genética , Animales , Corteza Renal/patología , Médula Renal/patología , Túbulos Renales Colectores/patología , Ratones , Ratones Noqueados , Ratones Transgénicos , FenotipoRESUMEN
The kidneys play a critical role in the maintenance of water homeostasis. This is achieved by the inherent architecture of the nephron along with the expression of various membrane transporters and channels that are responsible for the vectorial transport of salt and water. The collecting duct has become a focus of attention by virtue of its ability to transport water independent of solutes (free-water transport), and its apparent involvement in various water balance disorders. It was originally believed that the water transport capability of the collecting duct was solely under the influence of the circulating hormone, arginine vasopressin (AVP). However, during the past decade, locally produced autocrine and/or paracrine factors have emerged as potent modulators of transport of water by the collecting duct. Recently, much attention has been focused on the purinergic regulation of renal water transport. This review focuses on the role of the P2Y(2) receptor, the predominant purinergic receptor expressed in the collecting duct, in the modulation of water transport in physiological and pathophysiological conditions, and its therapeutic potential as a drug target to treat water balance disorders in the clinic. Studies carried out by us and other investigators are unravelling potent interactions among AVP, prostanoid and purinergic systems in the medullary collecting duct, and the perturbations of these interactions in water balance disorders such as acquired nephrogenic diabetes insipidus. Future studies should address the potential therapeutic benefits of modulators of P2Y(2) receptor signalling in water balance disorders, which are extremely prevalent in hospitalised patients irrespective of the underlying pathology.
RESUMEN
The renal collecting duct consists of intercalated cells (ICs) and principal cells (PCs). We have previously demonstrated that collecting ducts have a role in the innate immune defense of the kidney. Transcriptomics is an important tool used to enhance systems-level understanding of cell biology. However, transcriptomics performed on whole kidneys provides limited insight of collecting duct cell gene expression, because these cells comprise a small fraction of total kidney cells. Recently we generated reporter mouse models to enrich collecting duct specific PC and ICs and reported targeted gene expression of anti-microbial peptide genes. Here we report transcriptomics on enriched ICs and PCs and performed a pilot study sequencing four single ICs. We identified 3,645 genes with increased relative expression in ICs compared to non-ICs. In comparison to non-PCs, 2,088 genes had higher relative expression in PCs. IC associated genes included the innate interleukin 1 receptor, type 1 and the antimicrobial peptide(AMP) adrenomedullin. The top predicted canonical pathway for enriched ICs was lipopolysaccharide/Interleukin 1 mediated inhibition of Retinoid X Receptor alpha function and decreased Retinoid X Receptor expression was confirmed to occur 1-hour post experimental murine UTI in ICs but not in non-ICs.
Asunto(s)
Células Epiteliales/metabolismo , Perfilación de la Expresión Génica/métodos , Túbulos Renales Colectores/citología , Túbulos Renales Colectores/inmunología , Lipopolisacáridos/metabolismo , Receptor alfa X Retinoide/antagonistas & inhibidores , Receptor alfa X Retinoide/metabolismo , Animales , Acuaporina 2/genética , Acuaporina 2/metabolismo , Femenino , Inmunidad Innata/genética , Interleucina-1/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Proyectos Piloto , Transducción de Señal/genética , Transcriptoma/inmunología , ATPasas de Translocación de Protón Vacuolares/genética , ATPasas de Translocación de Protón Vacuolares/metabolismoRESUMEN
In vitro studies suggest that collecting duct-derived (CD-derived) endothelin-1 (ET-1) can regulate renal Na reabsorption; however, the physiologic role of CD-derived ET-1 is unknown. Consequently, the physiologic effect of selective disruption of the ET-1 gene in the CD of mice was determined. Mice heterozygous for aquaporin2 promoter Cre recombinase and homozygous for loxP-flanked exon 2 of the ET-1 gene (called CD-specific KO of ET-1 [CD ET-1 KO] mice) were generated. These animals had no CD ET-1 mRNA and had reduced urinary ET-1 excretion. CD ET-1 KO mice on a normal Na diet were hypertensive, while body weight, Na excretion, urinary aldosterone excretion, and plasma renin activity were unchanged. CD ET-1 KO mice on a high-Na diet had worsened hypertension, reduced urinary Na excretion, and excessive weight gain, but showed no differences between aldosterone excretion and plasma renin activity. Amiloride or furosemide reduced BP in CD ET-1 KO mice on a normal or high-Na diet and prevented excessive Na retention in salt-loaded CD ET-1 KO mice. These studies indicate that CD-derived ET-1 is an important physiologic regulator of renal Na excretion and systemic BP.
Asunto(s)
Endotelina-1/genética , Endotelina-1/fisiología , Hipernatremia/etiología , Hipertensión/etiología , Túbulos Renales Colectores/fisiología , Aldosterona/orina , Amilorida/farmacología , Animales , Presión Sanguínea/efectos de los fármacos , Diuréticos/farmacología , Furosemida/farmacología , Heterocigoto , Homocigoto , Hibridación in Situ , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , ARN Mensajero/genética , Cloruro de Sodio Dietético/farmacología , Cloruro de Sodio Dietético/orina , Aumento de Peso/efectos de los fármacosRESUMEN
Diffuse leiomyomatosis is associated with the inherited kidney disease Alport syndrome, and characterized by visceral smooth muscle overgrowth within the respiratory, gastrointestinal and female reproductive tracts. Although partial deletions of the type IV collagen genes COL4A5 and COL4A6, paired head-to-head on chromosome Xq22, are known to cause diffuse leiomyomatosis, loss of function for type IV collagen does not explain smooth muscle overgrowth. To further clarify pathogenic mechanisms, we have characterized novel deletions in patients with Alport syndrome-diffuse leiomyomatosis or Alport syndrome alone. A 27.6-kb deletion, in a female with Alport syndrome-diffuse leiomyomatosis, is marked by the most proximal, i.e. most 5', COL4A5 breakpoint described to date. By comparing this deletion to others described here and previously, we have defined a minimal overlap region, only 4.2 kb in length and containing the COL4A5-COL4A6 proximal promoters, loss of which contributes to smooth muscle overgrowth. A novel deletion in a male with Alport syndrome alone is>1.4 Mb in length, encompassing COL4A5 and COL4A6 entirely, as well as neighboring genes. We postulate that loss of the 4.2-kb region in diffuse leiomyomatosis causes misregulation of neighboring genes, contributing to smooth muscle overgrowth. Deletion of the neighboring genes themselves may afford protection from this condition.