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1.
Proc Natl Acad Sci U S A ; 114(15): 3921-3926, 2017 04 11.
Artículo en Inglés | MEDLINE | ID: mdl-28348216

RESUMEN

IRBIT [inositol 1,4,5-trisphosphate receptor (IP3R) binding protein released with inositol 1,4,5-trisphosphate (IP3)] is a multifunctional protein that regulates several target molecules such as ion channels, transporters, polyadenylation complex, and kinases. Through its interaction with multiple targets, IRBIT contributes to calcium signaling, electrolyte transport, mRNA processing, cell cycle, and neuronal function. However, the regulatory mechanism of IRBIT binding to particular targets is poorly understood. Long-IRBIT is an IRBIT homolog with high homology to IRBIT, except for a unique N-terminal appendage. Long-IRBIT splice variants have different N-terminal sequences and a common C-terminal region, which is involved in multimerization of IRBIT and Long-IRBIT. In this study, we characterized IRBIT and Long-IRBIT splice variants (IRBIT family). We determined that the IRBIT family exhibits different mRNA expression patterns in various tissues. The IRBIT family formed homo- and heteromultimers. In addition, N-terminal splicing of Long-IRBIT changed the protein stability and selectivity to target molecules. These results suggest that N-terminal diversity of the IRBIT family and various combinations of multimer formation contribute to the functional diversity of the IRBIT family.


Asunto(s)
Adenosilhomocisteinasa/metabolismo , Lectinas Tipo C/metabolismo , Proteínas de la Membrana/metabolismo , Adenosilhomocisteinasa/genética , Animales , Células COS , Señalización del Calcio , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/metabolismo , Chlorocebus aethiops , Femenino , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Lectinas Tipo C/genética , Proteínas de la Membrana/genética , Ratones Endogámicos C57BL , Ratones Endogámicos ICR , Ratones Noqueados , Isoformas de Proteínas , Estabilidad Proteica , Simportadores de Sodio-Bicarbonato/genética , Simportadores de Sodio-Bicarbonato/metabolismo , Intercambiador 3 de Sodio-Hidrógeno/genética , Intercambiador 3 de Sodio-Hidrógeno/metabolismo , Xenopus laevis
2.
Pflugers Arch ; 468(7): 1183-1196, 2016 07.
Artículo en Inglés | MEDLINE | ID: mdl-27044412

RESUMEN

Dent's disease is characterized by defective endocytosis in renal proximal tubules (PTs) and caused by mutations in the 2Cl(-)/H(+) exchanger, CLC-5. However, the pathological role of endosomal acidification in endocytosis has recently come into question. To clarify the mechanism of pathogenesis for Dent's disease, we examined the effects of a novel gating glutamate mutation, E211Q, on CLC-5 functions and endosomal acidification. In Xenopus oocytes, wild-type (WT) CLC-5 showed outward-rectifying currents that were inhibited by extracellular acidosis, but E211Q and an artificial pure Cl(-) channel mutant, E211A, showed linear currents that were insensitive to extracellular acidosis. Moreover, depolarizing pulse trains induced a robust reduction in the surface pH of oocytes expressing WT CLC-5 but not E211Q or E211A, indicating that the E211Q mutant functions as a pure Cl(-) channel similar to E211A. In HEK293 cells, E211A and E211Q stimulated endosomal acidification and hypotonicity-inducible vacuolar-type H(+)-ATPase (V-ATPase) activation at the plasma membrane. However, the stimulatory effects of these mutants were reduced compared with WT CLC-5. Furthermore, gene silencing experiments confirmed the functional coupling between V-ATPase and CLC-5 at the plasma membrane of isolated mouse PTs. These results reveal for the first time that the conversion of CLC-5 from a 2Cl(-)/H(+) exchanger into a Cl(-) channel induces Dent's disease in humans. In addition, defective endosomal acidification as a result of insufficient V-ATPase activation may still be important in the pathogenesis of Dent's disease.


Asunto(s)
Canales de Cloruro/genética , Canales de Cloruro/metabolismo , Enfermedad de Dent/metabolismo , Mutación/genética , ATPasas de Translocación de Protón Vacuolares/metabolismo , Animales , Línea Celular , Membrana Celular/metabolismo , Niño , Enfermedad de Dent/genética , Endocitosis/fisiología , Femenino , Células HEK293 , Homeostasis/fisiología , Humanos , Transporte Iónico/fisiología , Túbulos Renales Proximales/metabolismo , Masculino , Oocitos/metabolismo , Xenopus laevis/metabolismo
3.
Biomed Res Int ; 2014: 504808, 2014.
Artículo en Inglés | MEDLINE | ID: mdl-24982885

RESUMEN

Sodium-coupled bicarbonate absorption from renal proximal tubules (PTs) plays a pivotal role in the maintenance of systemic acid/base balance. Indeed, mutations in the Na(+)-HCO3 (-) cotransporter NBCe1, which mediates a majority of bicarbonate exit from PTs, cause severe proximal renal tubular acidosis associated with ocular and other extrarenal abnormalities. Sodium transport in PTs also plays an important role in the regulation of blood pressure. For example, PT transport stimulation by insulin may be involved in the pathogenesis of hypertension associated with insulin resistance. Type 1 angiotensin (Ang) II receptors in PT are critical for blood pressure homeostasis. Paradoxically, the effects of Ang II on PT transport are known to be biphasic. Unlike in other species, however, Ang II is recently shown to dose-dependently stimulate human PT transport via nitric oxide/cGMP/ERK pathway, which may represent a novel therapeutic target in human hypertension. In this paper, we will review the physiological and pathophysiological roles of PT transport.


Asunto(s)
Equilibrio Ácido-Base , Presión Sanguínea , Túbulos Renales Proximales/metabolismo , Equilibrio Ácido-Base/efectos de los fármacos , Angiotensina II/farmacología , Animales , Transporte Biológico/efectos de los fármacos , Presión Sanguínea/efectos de los fármacos , Humanos , Túbulos Renales Proximales/efectos de los fármacos , Síndrome Metabólico/metabolismo , Síndrome Metabólico/fisiopatología
4.
J Am Soc Nephrol ; 25(7): 1523-32, 2014 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-24511122

RESUMEN

Stimulation of renal proximal tubule (PT) transport by angiotensin II (Ang II) is critical for regulation of BP. Notably, in rats, mice, and rabbits, the regulation of PT sodium transport by Ang II is biphasic: transport is stimulated by picomolar to nanomolar concentrations of Ang II but inhibited by nanomolar to micromolar concentrations of Ang II. However, little is known about the effects of Ang II on human PT transport. By functional analysis with isolated PTs obtained from nephrectomy surgery, we found that Ang II induces a dose-dependent profound stimulation of human PT transport by type 1 Ang II receptor (AT1)-dependent phosphorylation of extracellular signal-regulated kinase (ERK). In PTs of wild-type mice, the nitric oxide (NO) /cGMP/cGMP-dependent kinase II (cGKII) pathway mediated the inhibitory effect of Ang II. In PTs of cGKII-deficient mice, the inhibitory effect of Ang II was lost, but activation of the NO/cGMP pathway failed to phosphorylate ERK. Conversely, in human PTs, the NO/cGMP pathway mediated the stimulatory effect of Ang II by phosphorylating ERK independently of cGKII. These contrasting responses to the NO/cGMP pathway may largely explain the different modes of PT transport regulation by Ang II, and the unopposed marked stimulation of PT transport by high intrarenal concentrations of Ang II may be an important factor in the pathogenesis of human hypertension. Additionally, the previously unrecognized stimulatory effect of the NO/cGMP pathway on PT transport may represent a human-specific therapeutic target in hypertension.


Asunto(s)
Angiotensina II/administración & dosificación , Angiotensina II/fisiología , GMP Cíclico/fisiología , Túbulos Renales Proximales/efectos de los fármacos , Túbulos Renales Proximales/metabolismo , Óxido Nítrico/fisiología , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiología , Relación Dosis-Respuesta a Droga , Humanos , Técnicas In Vitro
5.
Front Physiol ; 4: 270, 2013 Oct 01.
Artículo en Inglés | MEDLINE | ID: mdl-24101904

RESUMEN

The electrogenic Na(+)-HCO3 (-) cotransporter NBCe1 plays an essential role in bicarbonate absorption from renal proximal tubules, but also mediates the other biological processes in extrarenal tissues such as bicarbonate secretion from pancreatic ducts, maintenance of tissue homeostasis in eye, enamel maturation in teeth, or local pH regulation in synapses. Homozygous mutation in NBCe1 cause proximal renal tubular acidosis (pRTA) associated with extrarenal manifestations such as short stature, ocular abnormalities, enamel abnormalities, and migraine. Functional analyses of NBCe1 mutants using different expression systems suggest that at least a 50% reduction of the transport activity may be required to induce severe pRTA. In addition to functional impairments, some NBCe1 mutants show trafficking defects. Some of the pRTA-related NBCe1 mutants showing the cytoplasmic retention have been shown to exert a dominant negative effect through hetero-oligomer complexes with wild-type NBCe1 that may explain the occurrence of extrarenal manifestations in the heterozygous carries of NBCe1 mutations. Both NBCe1 knockout (KO) and W516X knockin (KI) mice showed very severe pRTA and reproduced most of the clinical manifestations observed in human pRTA patients. Functional analysis on isolated renal proximal tubules from W516X KI mice directly confirmed the indispensable role of NBCe1 in bicarbonate absorption from this nephron segment. In this review, we will focus on the molecular mechanisms underling the renal and extrarenal manifestations caused by NBCe1 inactivation.

6.
Curr Hypertens Rev ; 9(2): 148-55, 2013 May.
Artículo en Inglés | MEDLINE | ID: mdl-23971696

RESUMEN

Hypertension is a key factor of cardiovascular disease. Many organs and systems including heart, blood vessel, kidney, sympathetic nerve, and endocrine systems are involved in the regulation of blood pressure. In particular, the kidney plays an essential role in the regulation of blood pressure, but is also quite vulnerable to hypertensive tissue damage. For example, most chronic kidney disease (CKD) patients have hypertension and are revealed to have higher mortality than normal population. Furthermore, hypertensive renal sclerosis is emerging as the third main cause of dialysis patients. This mini review is to summarize the effects of angiotensin II and dopamine on renal proximal tubule transport, which may have important roles in the regulation of blood pressure.


Asunto(s)
Presión Sanguínea/fisiología , Hipertensión/etiología , Túbulos Renales Proximales/fisiología , Riñón/metabolismo , Animales , Transporte Biológico/fisiología , Dopamina/fisiología , Humanos , Hipertensión/metabolismo , Receptor de Angiotensina Tipo 1/fisiología , Sistema Renina-Angiotensina/fisiología
7.
J Am Soc Nephrol ; 24(6): 967-77, 2013 May.
Artículo en Inglés | MEDLINE | ID: mdl-23661805

RESUMEN

Nephronophthisis (NPHP)-related ciliopathies are recessive, single-gene disorders that collectively make up the most common genetic cause of CKD in the first three decades of life. Mutations in 1 of the 15 known NPHP genes explain less than half of all cases with this phenotype, however, and the recently identified genetic causes are exceedingly rare. As a result, a strategy to identify single-gene causes of NPHP-related ciliopathies in single affected families is needed. Although whole-exome resequencing facilitates the identification of disease genes, the large number of detected genetic variants hampers its use. Here, we overcome this limitation by combining homozygosity mapping with whole-exome resequencing in a sibling pair with an NPHP-related ciliopathy. Whole-exome capture revealed a homozygous splice acceptor site mutation (c.698G>T) in the renal Mg(2+) transporter SLC41A1. This mutation resulted in skipping of exon 6 of SLC41A1, resulting in an in-frame deletion of a transmembrane helix. Transfection of cells with wild-type or mutant SLC41A1 revealed that deletion of exon 6 completely blocks the Mg(2+) transport function of SLC41A1. Furthermore, in normal human kidney tissue, endogenous SLC41A1 specifically localized to renal tubules situated at the corticomedullary boundary, consistent with the region of cystogenesis observed in NPHP and related ciliopathies. Last, morpholino-mediated knockdown of slc41a1 expression in zebrafish resulted in ventral body curvature, hydrocephalus, and cystic kidneys, similar to the effects of knocking down other NPHP genes. Taken together, these data suggest that defects in the maintenance of renal Mg(2+) homeostasis may lead to tubular defects that result in a phenotype similar to NPHP.


Asunto(s)
Proteínas de Transporte de Catión/genética , Proteínas de Transporte de Catión/metabolismo , Enfermedades Renales Quísticas/congénito , Magnesio/metabolismo , Animales , Niño , Preescolar , Perros , Exones/genética , Femenino , Genes Recesivos , Células HEK293 , Heterocigoto , Homocigoto , Humanos , Riñón/metabolismo , Riñón/patología , Enfermedades Renales Quísticas/genética , Enfermedades Renales Quísticas/metabolismo , Enfermedades Renales Quísticas/patología , Células de Riñón Canino Madin Darby , Masculino , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Mutación Missense , Linaje , Pez Cebra , Proteínas de Pez Cebra
8.
Pflugers Arch ; 465(9): 1281-91, 2013 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-23559100

RESUMEN

Homozygous mutations in the electrogenic Na(+)-HCO3 (-) cotransporter NBCe1 cause proximal renal tubular acidosis (pRTA) associated with extrarenal manifestations such as ocular abnormalities and migraine. Previously, the NBCe1 cytosolic mutant S982NfsX4 was shown to have a dominant negative effect by forming hetero-oligomer complexes with wild type (WT), which might be responsible for the occurrence of glaucoma and migraine in the heterozygous family members. In this study, we investigated whether the NBCe1 L522P mutant has a similar dominant negative effect. Functional analyses in Xenopus oocytes and HEK293 cells revealed that the L522P mutant had no transport activity due to defective membrane expression. Furthermore, when coexpressed with WT, L522P significantly reduced the transport activity of WT. In HEK293 cells, the cytosolic mutant L522P reduced the membrane expression of NBCe1 by forming hetero-oligomer complexes with WT. Among the artificial Leu(522) mutants, L522I showed proper membrane expression and normal transport activity. However, the other mutants L522R, L522K, L522D, and L522E showed a predominant cytosolic retention. Moreover, L522R had a dominant negative effect, when coexpressed with WT. These results indicate that Leu(522) plays an important role in the structure and trafficking of NBCe1. They also suggest that the NBCe1 mutants retaining in cytoplasm may have the dominant negative effect in common, which may induce some clinical manifestations.


Asunto(s)
Genes Dominantes , Genes Recesivos , Mutación Missense , Simportadores de Sodio-Bicarbonato/metabolismo , Animales , Membrana Celular/metabolismo , Perros , Retículo Endoplásmico/metabolismo , Células HEK293 , Humanos , Células de Riñón Canino Madin Darby , Multimerización de Proteína , Estructura Terciaria de Proteína , Transporte de Proteínas , Simportadores de Sodio-Bicarbonato/química , Simportadores de Sodio-Bicarbonato/genética , Xenopus
9.
Case Rep Neurol Med ; 2012: 704639, 2012.
Artículo en Inglés | MEDLINE | ID: mdl-22937357

RESUMEN

We report a case in whom slow correction of hyponatremia (5 mmol/day for 3 days) induced central pontine myelinolysis (CPM). After the diagnosis was confirmed by imaging, we started to relower serum Na that completely recovered the sign and symptoms of CPM. Rapid correction of serum sodium is known to be associated with CPM. However, it may occur even after slow correction of hyponatremia. Currently, there is no standard therapy for CPM other than supportive therapy. Other therapy includes sterioid, plasmaphresis and IVIG, but these therapies have not been shown to be particularly effective. The pathophysiology of CPM is related to a relative dehydration of the brain during the correction of hyponatremia, resulting in cell death and demyelination, therefore gentle rehydration with lowering serum sodium may not be an unreasonable therapy. The present case provides supportive evidence that reinduction of hyponatremia is effective in treating CPM if started immediately after the diagnosis is suggested. The present case tells us that severe chronic hyponatremia must be managed with extreme care especially in patients with chronic debilitating illness and that relowering serum Na is a treatment of choice when CPM is suggested.

10.
Open Ophthalmol J ; 6: 36-41, 2012.
Artículo en Inglés | MEDLINE | ID: mdl-22798968

RESUMEN

Electrogenic Na(+)-HCO(3) (-) cotransporter NBCe1 is expressed in several tissues such as kidney, eye, and brain, where it may mediate distinct biological processes. In particular, NBCe1 in renal proximal tubules is essential for the regulation of systemic acid/base balance. On the other hand, NBCe1 in eye may be indispensable for the maintenance of tissue homeostasis. Consistent with this view, homozygous mutations in NBCe1 cause severe proximal renal tubular acidosis associated with ocular abnormalities such as band keratopathy, glaucoma, and cataract. The widespread expression of NBCe1 in eye suggests that the inactivation of NBCe1 per se may be responsible for the occurrence of these ocular abnormalities. In this review, we discuss about physiological and pathological roles of NBCe1 in eye.

11.
World J Nephrol ; 1(5): 146-50, 2012 Oct 06.
Artículo en Inglés | MEDLINE | ID: mdl-24175252

RESUMEN

Thiazolidinediones (TZDs), pharmacological activators of peroxisome-proliferator-activated receptors γ (PPARγ), significantly improve insulin resistance and lower plasma glucose concentrations. However, the use of TZDs is associated with plasma volume expansion, the mechanism of which has been a matter of controversy. Originally, PPARγ-mediated enhanced transcription of the epithelial Na channel (ENaC) γ subunit was thought to play a central role in TZD-induced volume expansion. However, later studies suggested that the activation of ENaC alone could not explain TZD-induced volume expansion. We have recently shown that TZDs rapidly stimulate sodium-coupled bicarbonate absorption from renal proximal tubule (PT) in vitro and in vivo. TZD-induced transport stimulation was dependent on PPARγ/Src/EGFR/ERK, and observed in rat, rabbit and human. However, this stimulation was not observed in mouse PTs where Src/EGFR is constitutively activated. Analysis in mouse embryonic fibroblast cells confirmed the existence of PPARγ/Src-dependent non-genomic signaling, which requires the ligand binding ability but not the transcriptional activity of PPARγ. The TZD-induced enhancement of association between PPARγ and Src supports an obligatory role for Src in this signaling. These results support the view that TZD-induced volume expansion is multifactorial. In addition to the PPARγ-dependent enhanced expression of the sodium transport system(s) in distal nephrons, the PPARγ-dependent non-genomic stimulation of renal proximal transport may be also involved in TZD-induced volume expansion.

12.
J Cardiol ; 59(2): 139-46, 2012 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-22154614

RESUMEN

Retroperitoneal fibrosis, inflammatory aortic aneurysm, and pericardial and mediastinal fibrosis are characterized by infiltration of immuno-inflammatory cells and deposition of thickened fibrous tissues. Several recent studies suggested that an immunoglobulin-G4 (IgG4)-related immunological mechanism may play a role in these diseases. By searching the clinical database of patients admitted to our department between 2000 and 2010, we summarized the clinical data of 11 patients who were diagnosed to have these disorders. The diagnoses were idiopathic retroperitoneal fibrosis (8 cases), mediastinal and/or pericardial fibrosis (4 cases), inflammatory abdominal aneurysm (2 cases), and inflammatory coronary periarteritis (1 case). Hypertension, diabetes, and dyslipidemia were found in 45%, 36%, and 55%, respectively, in these patients, and they were all either current or former smokers. Two patients with pericardial involvement showed a rushed clinical course, resulting in in-hospital death. Serum levels of IgG were elevated in 67%, and soluble interleukin-2 receptor was elevated in 75%, when measured. Immunohistochemical analysis showed marked infiltration of IgG4-positive plasma cells in the pericardium in patients who died of constrictive pericarditis. Our data support the notion that immune-inflammatory mechanism, which might be IgG4-related sometimes, may play a role in idiopathic retroperitoneal fibrosis, inflammatory aortic aneurysm, and mediastinal/pericardial fibrosis, although clinical course may differ substantially.


Asunto(s)
Aneurisma de la Aorta/patología , Pericarditis/patología , Fibrosis Retroperitoneal/patología , Adulto , Anciano , Anciano de 80 o más Años , Aneurisma de la Aorta/inmunología , Aneurisma de la Aorta Abdominal/diagnóstico por imagen , Aneurisma de la Aorta Abdominal/patología , Enfermedad Coronaria/patología , Femenino , Humanos , Inmunoglobulina G/análisis , Inmunohistoquímica , Inflamación , Interleucina-2/sangre , Masculino , Mediastinitis/patología , Persona de Mediana Edad , Pericarditis/inmunología , Pericardio , Tomografía de Emisión de Positrones , Fibrosis Retroperitoneal/diagnóstico por imagen , Fibrosis Retroperitoneal/inmunología , Estudios Retrospectivos , Esclerosis/patología , Fumar/efectos adversos , Tomografía Computarizada por Rayos X
13.
Int J Hypertens ; 2011: 391762, 2011.
Artículo en Inglés | MEDLINE | ID: mdl-21629870

RESUMEN

Sodium transport through various nephron segments is quite important in regulating sodium reabsorption and blood pressure. Among several regulators of this process, insulin acts on almost all the nephron segments and is a strong enhancer of sodium reabsorption. Sodium-proton exchanger type 3 (NHE3) is a main regulator of sodium reabsorption in the luminal side of proximal tubule. In the basolateral side of the proximal tubule, sodium-bicarbonate cotransporter (NBCe1) mediates sodium and bicarbonate exit from tubular cells. In the distal nephron and the connecting tubule, epithelial sodium channel (ENaC) is of great importance to sodium reabsorption. NHE3, NBCe1, and ENaC are all regulated by insulin. Recently with-no-lysine (WNK) kinases, responsible for familial hypertension, stimulating sodium reabsorption in the distal nephron, have been found to be also regulated by insulin. We will discuss the regulation of renal sodium transport by insulin and its roles in the pathogenesis of hypertension in insulin resistance.

14.
Cell Metab ; 13(5): 550-61, 2011 May 04.
Artículo en Inglés | MEDLINE | ID: mdl-21531337

RESUMEN

Thiazolidinediones (TZDs) improve insulin resistance by activating a nuclear hormone receptor, peroxisome proliferator-activated receptor γ (PPARγ). However, the use of TZDs is associated with plasma volume expansion through a mechanism that remains to be clarified. Here we showed that TZDs rapidly stimulate sodium-coupled bicarbonate absorption from the renal proximal tubule in vitro and in vivo. TZD-induced transport stimulation is dependent on PPARγ-Src-EGFR-ERK and observed in rat, rabbit and human, but not in mouse proximal tubules where Src-EGFR is constitutively activated. The existence of PPARγ-Src-dependent nongenomic signaling, which requires the ligand-binding ability, but not the transcriptional activity of PPARγ, is confirmed in mouse embryonic fibroblast cells. The enhancement of the association between PPARγ and Src by TZDs supports an indispensable role of Src in this signaling. These results suggest that the PPARγ-dependent nongenomic stimulation of renal proximal transport is also involved in TZD-induced volume expansion.


Asunto(s)
Bicarbonatos/metabolismo , Túbulos Renales Proximales/efectos de los fármacos , Túbulos Renales Proximales/metabolismo , PPAR gamma/metabolismo , Transducción de Señal/efectos de los fármacos , Sodio/metabolismo , Tiazolidinedionas/farmacología , Adipocitos/citología , Adipocitos/efectos de los fármacos , Adipocitos/metabolismo , Animales , Western Blotting , Proteínas de Transporte de Catión/metabolismo , Diferenciación Celular/efectos de los fármacos , Células Cultivadas , Embrión de Mamíferos/citología , Embrión de Mamíferos/efectos de los fármacos , Embrión de Mamíferos/metabolismo , Receptores ErbB/metabolismo , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Fibroblastos/citología , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Humanos , Técnicas para Inmunoenzimas , Inmunoprecipitación , Túbulos Renales Proximales/citología , Ratones , Ratones Endogámicos C57BL , PPAR gamma/genética , Fosforilación/efectos de los fármacos , Conejos , Ratas , Ratas Wistar , Intercambiador 1 de Sodio-Hidrógeno , Intercambiadores de Sodio-Hidrógeno/metabolismo , Especificidad de la Especie , Familia-src Quinasas/metabolismo
16.
Kidney Int ; 79(7): 730-41, 2011 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-21228764

RESUMEN

We have identified a novel homozygous nonsense mutation (W516X) in the kidney-type electrogenic sodium bicarbonate cotransporter 1 (NBC1) in a patient with isolated proximal renal tubular acidosis (pRTA). To specifically address the pathogenesis of this mutation, we created NBC1 W516X knock-in mice to match the patient's abnormalities. The expression of NBC1 mRNA and protein in the kidneys of NBC1(W516X/W516X) mice were virtually absent, indicating that nonsense-mediated mRNA decay (NMD) is involved in the defective transcription and translation of this mutation. These mice not only recapitulated the phenotypes of this patient with growth retardation, pRTA, and ocular abnormalities, but also showed anemia, volume depletion, prerenal azotemia, and several organ abnormalities, culminating in dehydration and renal failure with early lethality before weaning. In isolated renal proximal tubules, both NBC1 activity and the rate of bicarbonate absorption were markedly reduced. Unexpectedly, there was no compensatory increase in mRNA of distal acid/base transporters. Sodium bicarbonate but not saline administration to these mutant mice markedly prolonged their survival, decreased their protein catabolism and attenuated organ abnormalities. The prolonged survival time uncovered the development of corneal opacities due to corneal edema. Thus, NBC1(W516X/W516X) mice with pRTA represent an animal model for metabolic acidosis and may be useful for testing therapeutic inhibition of NMD in vivo.


Asunto(s)
Acidosis Tubular Renal/metabolismo , Acidosis/metabolismo , Túbulos Renales Proximales/metabolismo , Simportadores de Sodio-Bicarbonato/metabolismo , Acidosis/tratamiento farmacológico , Acidosis/genética , Acidosis/patología , Acidosis Tubular Renal/tratamiento farmacológico , Acidosis Tubular Renal/genética , Acidosis Tubular Renal/patología , Factores de Edad , Envejecimiento , Análisis de Varianza , Anemia/genética , Anemia/metabolismo , Animales , Acuaporina 2/metabolismo , Bicarbonatos/metabolismo , Codón sin Sentido , Opacidad de la Córnea/genética , Opacidad de la Córnea/metabolismo , Modelos Animales de Enfermedad , Femenino , Técnicas de Sustitución del Gen , Genotipo , Trastornos del Crecimiento/genética , Trastornos del Crecimiento/metabolismo , Homocigoto , Humanos , Concentración de Iones de Hidrógeno , Túbulos Renales Proximales/efectos de los fármacos , Túbulos Renales Proximales/patología , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Transgénicos , Persona de Mediana Edad , Fenotipo , ARN Mensajero/metabolismo , Índice de Severidad de la Enfermedad , Bicarbonato de Sodio/administración & dosificación , Bicarbonato de Sodio/metabolismo , Simportadores de Sodio-Bicarbonato/genética , Transcripción Genética
17.
Channels (Austin) ; 5(2): 106-9, 2011.
Artículo en Inglés | MEDLINE | ID: mdl-21224720

RESUMEN

The electrogenic Na+ -HCO3⁻ cotransporter NBCe1 plays essential roles in the regulation of systemic and/or local pH. Homozygous inactivating mutations in NBCe1 cause proximal renal tubular acidosis associated with ocular abnormalities. We recently showed that defective membrane expression of NBCe1, caused by several mutations such as Delta65bp (S982NfsX4), is also associated with familial migraine. The Delta65bp mutant is quite unique in that it lacks a putative carbonic anhydrase (CA) II-binding domain but still shows an apparently normal transport activity in Xenopus oocytes. In this addendum, we show that the co-expression of CAII together with the wild-type NBCe1 or the Delta65bp mutant does not enhance the NBCe1 activities in oocytes. Moreover, a carbonic anhydrase inhibitor acetazolamide fails to inhibit the wild-type or the Delta65bp activities co-expressed with CAII. These results indicate that a bicarbonate transport metabolon proposed for the interaction between CAII and NBCe1 does not work at least in Xenopus oocytes.


Asunto(s)
Anhidrasa Carbónica II/química , Oocitos/metabolismo , Simportadores de Sodio-Bicarbonato/metabolismo , Proteínas de Xenopus/metabolismo , Xenopus laevis/metabolismo , Acetazolamida/farmacología , Acidosis Tubular Renal/metabolismo , Secuencia de Aminoácidos , Animales , Aniones , Bicarbonatos/química , Electrofisiología/métodos , Humanos , Datos de Secuencia Molecular , Estructura Terciaria de Proteína , Ratas
18.
Pflugers Arch ; 461(2): 249-59, 2011 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-21234596

RESUMEN

The electrogenic Na(+)-HCO(3)(-) cotransporter NBCe1 encoded by SLC4A4 plays essential roles in the regulation of intracellular/extracellular pH. Homozygous mutations in NBCe1 cause proximal renal tubular acidosis associated with ocular abnormalities. In the present study, we tried to perform functional characterization of the four nonsynonymous single nucleotide polymorphisms (SNPs), E122G, S356Y, K558R, and N640I in NBCe1A. Functional analysis in Xenopus oocytes revealed that while the K558R variant had a significantly reduced transport activity corresponding to 47% of the wild-type activity, the remaining variants E122G, S356Y, and N640I did not change the NBCe1A activity. Apparent Na(+) affinity of K558R was not different from that of wild-type NBCe1A. Immunohistological analyses in HEK293 cells and MDCK cells indicated that none of these SNPs changed the trafficking behaviors of NBCe1A. Functional analysis in HEK293 cells also revealed that only the K558R variant had a reduced transport activity, corresponding to 41-47% of the wild-type activity. From these results, we conclude that among four SNPs, only the K558R variant, which is predicted to lie in transmembrane segment 5, significantly reduces the NBCe1A activity without changing the trafficking behavior or the apparent extracellular Na(+) affinity.


Asunto(s)
Simportadores de Sodio-Bicarbonato/genética , Animales , Células Cultivadas , Perros , Células HEK293 , Humanos , Oocitos/fisiología , Polimorfismo de Nucleótido Simple , Simportadores de Sodio-Bicarbonato/fisiología , Xenopus laevis
19.
J Biol Chem ; 285(45): 35123-32, 2010 Nov 05.
Artículo en Inglés | MEDLINE | ID: mdl-20810651

RESUMEN

The evolutionary loss of hepatic urate oxidase (uricase) has resulted in humans with elevated serum uric acid (urate). Uricase loss may have been beneficial to early primate survival. However, an elevated serum urate has predisposed man to hyperuricemia, a metabolic disturbance leading to gout, hypertension, and various cardiovascular diseases. Human serum urate levels are largely determined by urate reabsorption and secretion in the kidney. Renal urate reabsorption is controlled via two proximal tubular urate transporters: apical URAT1 (SLC22A12) and basolateral URATv1/GLUT9 (SLC2A9). In contrast, the molecular mechanism(s) for renal urate secretion remain unknown. In this report, we demonstrate that an orphan transporter hNPT4 (human sodium phosphate transporter 4; SLC17A3) was a multispecific organic anion efflux transporter expressed in the kidneys and liver. hNPT4 was localized at the apical side of renal tubules and functioned as a voltage-driven urate transporter. Furthermore, loop diuretics, such as furosemide and bumetanide, substantially interacted with hNPT4. Thus, this protein is likely to act as a common secretion route for both drugs and may play an important role in diuretics-induced hyperuricemia. The in vivo role of hNPT4 was suggested by two hyperuricemia patients with missense mutations in SLC17A3. These mutated versions of hNPT4 exhibited reduced urate efflux when they were expressed in Xenopus oocytes. Our findings will complete a model of urate secretion in the renal tubular cell, where intracellular urate taken up via OAT1 and/or OAT3 from the blood exits from the cell into the lumen via hNPT4.


Asunto(s)
Bumetanida/farmacocinética , Diuréticos/farmacocinética , Furosemida/farmacocinética , Túbulos Renales Proximales/metabolismo , Modelos Biológicos , Proteínas Cotransportadoras de Sodio-Fosfato de Tipo I/metabolismo , Ácido Úrico/metabolismo , Animales , Bumetanida/efectos adversos , Bumetanida/farmacología , Diuréticos/efectos adversos , Diuréticos/farmacología , Furosemida/efectos adversos , Furosemida/farmacología , Predisposición Genética a la Enfermedad , Proteínas Facilitadoras del Transporte de la Glucosa/genética , Proteínas Facilitadoras del Transporte de la Glucosa/metabolismo , Gota/genética , Gota/metabolismo , Humanos , Hiperuricemia/inducido químicamente , Hiperuricemia/genética , Hiperuricemia/metabolismo , Transporte Iónico/efectos de los fármacos , Transporte Iónico/genética , Hígado/metabolismo , Mutación Missense , Proteína 1 de Transporte de Anión Orgánico/genética , Proteína 1 de Transporte de Anión Orgánico/metabolismo , Transportadores de Anión Orgánico/genética , Transportadores de Anión Orgánico/metabolismo , Transportadores de Anión Orgánico Sodio-Independiente/genética , Transportadores de Anión Orgánico Sodio-Independiente/metabolismo , Proteínas de Transporte de Catión Orgánico/genética , Proteínas de Transporte de Catión Orgánico/metabolismo , Proteínas Cotransportadoras de Sodio-Fosfato de Tipo I/genética , Urato Oxidasa/genética , Urato Oxidasa/metabolismo , Xenopus laevis
20.
Proc Natl Acad Sci U S A ; 107(36): 15963-8, 2010 Sep 07.
Artículo en Inglés | MEDLINE | ID: mdl-20798035

RESUMEN

Homozygous mutations in SLC4A4, encoding the electrogenic Na(+)-HCO(3)(-) cotransporter NBCe1, have been known to cause proximal renal tubular acidosis (pRTA) and ocular abnormalities. In this study, we report two sisters with pRTA, ocular abnormalities, and hemiplegic migraine. Genetic analysis ruled out pathological mutations in the known genes for familial hemiplegic migraine, but identified a homozygous 65-bp deletion (Delta65bp) in the C terminus of NBCe1, corresponding to the codon change S982NfsX4. Several heterozygous members of this family also presented glaucoma and migraine with or without aura. Despite the normal electrogenic activity in Xenopus oocytes, the Delta65bp mutant showed almost no transport activity due to a predominant cytosolic retention in mammalian cells. Furthermore, coexpression experiments uncovered a dominant negative effect of the mutant through hetero-oligomer formation with wild-type NBCe1. Among other pRTA pedigrees with different NBCe1 mutations, we identified four additional homozygous patients with migraine. The immunohistological and functional analyses of these mutants demonstrate that the near total loss of NBCe1 activity in astrocytes can cause migraine potentially through dysregulation of synaptic pH.


Asunto(s)
Trastornos Migrañosos/genética , Simportadores de Sodio-Bicarbonato/genética , Animales , Línea Celular , Perros , Femenino , Homocigoto , Humanos , Concentración de Iones de Hidrógeno , Masculino , Mutación , Linaje , Xenopus laevis
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