RESUMEN
Nephrotic syndrome, characterized by proteinuria and hypoalbuminemia, results from the dysregulation of glomerular podocytes and is a significant cause of end-stage kidney disease. Patients with idiopathic nephrotic syndrome are generally treated with immunosuppressive agents; however, these agents produce various adverse effects. Previously, we reported the renoprotective effects of a stimulator of the mitochondrial ATP-dependent K+ channel (MitKATP), nicorandil, in a remnant kidney model. Nonetheless, the cellular targets of these effects remain unknown. Here, we examined the effect of nicorandil on puromycin aminonucleoside-induced nephrosis (PAN) rats, a well-established model of podocyte injury and human nephrotic syndrome. PAN was induced using a single intraperitoneal injection. Nicorandil was administered orally at 30 mg/kg/day. We found that proteinuria and hypoalbuminemia in PAN rats were significantly ameliorated following nicorandil treatment. Immunostaining and ultrastructural analysis under electron microscopy demonstrated that podocyte injury in PAN rats showed a significant partial attenuation following nicorandil treatment. Nicorandil ameliorated the increase in the oxidative stress markers nitrotyrosine and 8-hydroxy-2-deoxyguanosine in glomeruli. Conversely, nicorandil prevented the decrease in levels of the antioxidant enzyme manganese superoxide dismutase in PAN rats. We found that mitochondrial Ca2+ uniporter levels in glomeruli were higher in PAN rats than in control rats, and this increase was significantly attenuated by nicorandil. We conclude that stimulation of MitKATP by nicorandil reduces proteinuria by attenuating podocyte injury in PAN nephrosis, which restores mitochondrial antioxidative capacity, possibly through mitochondrial Ca2+ uniporter modulation. These data indicate that MitKATP may represent a novel target for podocyte injury and nephrotic syndrome.NEW & NOTEWORTHY Our findings suggest that the mitochondrial Ca2+ uniporter may be an upstream regulator of manganese superoxide dismutase and indicate a biochemical basis for the interaction between the ATP-sensitive K+ channel and Ca2+ signaling. We believe that our study makes a significant contribution to the literature because our results indicate that the ATP-sensitive K+ channel may be a potential therapeutic target for podocyte injury and nephrotic syndrome.
Asunto(s)
Hipoalbuminemia , Nefrosis , Síndrome Nefrótico , Nicorandil , Podocitos , Animales , Ratas , Adenosina Trifosfato/metabolismo , Antioxidantes/metabolismo , Nefrosis/inducido químicamente , Nefrosis/prevención & control , Síndrome Nefrótico/inducido químicamente , Síndrome Nefrótico/tratamiento farmacológico , Síndrome Nefrótico/prevención & control , Nicorandil/uso terapéutico , Proteinuria/inducido químicamente , Proteinuria/prevención & control , Puromicina Aminonucleósido/toxicidad , Superóxido DismutasaRESUMEN
We report a case of a patient who developed dialysis-requiring acute kidney injury (AKI) after the use of canagliflozin. A 66-year-old man with type 2 diabetes who was recovering from left knee septic arthritis at a rehabilitation facility was admitted with oliguric AKI 5 days after starting treatment with canagliflozin, an inhibitor of sodium/glucose cotransporter 2 (SGLT2). The patient presented with hematuria, non-nephrotic-range proteinuria, and serum creatinine level of 6.8 (baseline, 1.1-1.3) mg/dL. There was no recent use of radiocontrast agents or exposure to other nephrotoxins. The patient subsequently required hemodialysis. Due to recent antibiotic use (ampicillin-sulbactam), acute interstitial nephritis was considered in the differential diagnosis. Kidney biopsy was performed, which showed the presence of osmotic nephropathy. The patient's kidney function returned to baseline after 2 weeks of hemodialysis. This case provides evidence of an association of osmotic nephropathy with the use of canagliflozin and discusses potential mechanisms. We recommend kidney biopsy for cases of severe AKI associated with SGLT2 inhibitors to better understand the relationship of this complication with the use of this class of medications.
Asunto(s)
Lesión Renal Aguda/inducido químicamente , Lesión Renal Aguda/diagnóstico por imagen , Canagliflozina/efectos adversos , Nefrosis/inducido químicamente , Nefrosis/diagnóstico por imagen , Inhibidores del Cotransportador de Sodio-Glucosa 2/efectos adversos , Lesión Renal Aguda/metabolismo , Anciano , Diuréticos Osmóticos/efectos adversos , Humanos , Masculino , Nefrosis/metabolismoRESUMEN
Ste20-like kinase SLK is critical for embryonic development and may play an important role in wound healing, muscle homeostasis, cell migration, and tumor growth. Mice with podocyte-specific deletion of SLK show albuminuria and damage to podocytes as they age. The present study addressed the role of SLK in glomerular injury. We induced adriamycin nephrosis in 3- to 4-mo-old control and podocyte SLK knockout (KO) mice. Compared with control, SLK deletion exacerbated albuminuria and loss of podocytes, synaptopodin, and podocalyxin. Glomeruli of adriamycin-treated SLK KO mice showed diffuse increases in the matrix and sclerosis as well as collapse of the actin cytoskeleton. SLK can phosphorylate ezrin. The complex of phospho-ezrin, Na+/H+ exchanger regulatory factor 2, and podocalyxin in the apical domain of the podocyte is a key determinant of normal podocyte architecture. Deletion of SLK reduced glomerular ezrin and ezrin phosphorylation in adriamycin nephrosis. Also, deletion of SLK reduced the colocalization of ezrin and podocalyxin in the glomerulus. Cultured glomerular epithelial cells with KO of SLK showed reduced ezrin phosphorylation and podocalyxin expression as well as reduced F-actin. Thus, SLK deletion leads to podocyte injury as mice age and exacerbates injury in adriamycin nephrosis. The mechanism may at least in part involve ezrin phosphorylation as well as disruption of the cytoskeleton and podocyte apical membrane structure.
Asunto(s)
Citoesqueleto de Actina/enzimología , Doxorrubicina , Glomeruloesclerosis Focal y Segmentaria/enzimología , Nefrosis/enzimología , Podocitos/enzimología , Proteínas Serina-Treonina Quinasas/deficiencia , Citoesqueleto de Actina/patología , Actinas/metabolismo , Albuminuria/inducido químicamente , Albuminuria/enzimología , Albuminuria/genética , Animales , Células Cultivadas , Proteínas del Citoesqueleto/metabolismo , Modelos Animales de Enfermedad , Técnicas de Silenciamiento del Gen , Glomeruloesclerosis Focal y Segmentaria/inducido químicamente , Glomeruloesclerosis Focal y Segmentaria/genética , Glomeruloesclerosis Focal y Segmentaria/patología , Ratones Noqueados , Proteínas de Microfilamentos/metabolismo , Nefrosis/inducido químicamente , Nefrosis/genética , Nefrosis/patología , Fosfoproteínas/metabolismo , Fosforilación , Podocitos/patología , Proteínas Serina-Treonina Quinasas/genética , Proteínas/metabolismo , Intercambiadores de Sodio-Hidrógeno/metabolismoRESUMEN
Genetic ablation of calcium-independent phospholipase A2γ (iPLA2γ) in mice results in marked damage of mitochondria and enhanced autophagy in glomerular visceral epithelial cells (GECs) or podocytes. The present study addresses the role of iPLA2γ in glomerular injury. In adriamycin nephrosis, deletion of iPLA2γ exacerbated albuminuria and reduced podocyte number. Glomerular LC3-II increased and p62 decreased in adriamycin-treated iPLA2γ knockout (KO) mice, compared with treated control, in keeping with increased autophagy in KO. iPLA2γ KO GECs in culture also demonstrated increased autophagy, compared with control GECs. iPLA2γ KO GECs showed a reduced oxygen consumption rate and increased phosphorylation of AMP kinase (pAMPK), consistent with mitochondrial dysfunction. Adriamycin further stimulated pAMPK and autophagy. After co-transfection of GECs with mito-YFP (to label mitochondria) and RFP-LC3 (to label autophagosomes), or RFP-LAMP1 (to label lysosomes), there was greater colocalization of mito-YFP with RFP-LC3-II and with RFP-LAMP1 in iPLA2γ KO GECs, compared with WT, indicating enhanced mitophagy in KO. Adriamycin increased mitophagy in WT cells. Thus, iPLA2γ has a cytoprotective function in the normal glomerulus and in glomerulopathy, as deletion of iPLA2γ leads to mitochondrial damage and impaired energy homeostasis, as well as autophagy and mitophagy.
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Calcio/metabolismo , Doxorrubicina/farmacología , Técnicas de Inactivación de Genes , Fosfolipasas A2 Grupo IV/deficiencia , Fosfolipasas A2 Grupo IV/genética , Glomérulos Renales/efectos de los fármacos , Nefrosis/genética , Proteínas Quinasas Activadas por AMP/metabolismo , Animales , Estrés del Retículo Endoplásmico/efectos de los fármacos , Estrés del Retículo Endoplásmico/genética , Glomérulos Renales/lesiones , Glomérulos Renales/patología , Ratones , Ratones Endogámicos C57BL , Proteínas Asociadas a Microtúbulos/metabolismo , Mitocondrias/efectos de los fármacos , Mitocondrias/patología , Mitofagia/efectos de los fármacos , Mitofagia/genética , Nefrosis/inducido químicamente , Nefrosis/enzimología , Nefrosis/patologíaAsunto(s)
Enfermedades de los Perros/inducido químicamente , Eméticos/efectos adversos , Enfermedades de los Caballos/virología , Nefrosis/veterinaria , Phascolarctidae , Fiebre del Nilo Occidental/veterinaria , Animales , Antiinflamatorios/uso terapéutico , Asma/tratamiento farmacológico , Asma/veterinaria , Dexametasona/uso terapéutico , Enfermedades de los Perros/tratamiento farmacológico , Perros , Acaparamiento , Caballos , Indoles/efectos adversos , Nebulizadores y Vaporizadores/veterinaria , Nefrosis/inducido químicamente , Nueva Gales del Sur , Oxalatos/efectos adversos , Pneumocystis , Infecciones por Pneumocystis/inducido químicamente , Infecciones por Pneumocystis/veterinaria , Polvos , Pirroles/efectos adversos , Vómitos , Virus del Nilo Occidental/aislamiento & purificaciónRESUMEN
Glomerular diseases are the leading cause of chronic kidney disease, and mesangial cells (MCs) have been demonstrated to be involved in the pathogenesis. Puromycin aminonucleoside (PAN) is a nephrotoxic drug that induces glomerular injury with elusive mechanisms. The present study was undertaken to investigate the role of PAN in MC apoptosis, as well as the underlying mechanism. Here we found that PAN induced MC apoptosis accompanied by declined cell viability and enhanced inflammatory response. The apoptosis was further evidenced by increments of apoptosis regulator BAX (BAX) and caspase-3 expression. In line with the apoptotic response in MCs following PAN treatment, we also found a remarkable induction of estrogen-related receptor-α (ERRα), an orphan nuclear receptor, at both mRNA and protein levels. Interestingly, ERRα silencing by an siRNA approach resulted in an attenuation of the apoptosis and inflammatory response caused by PAN. More importantly, overexpression of ERRα in MCs significantly triggered MC apoptosis in line with increased BAX and caspase-3 expression. In PAN-treated MCs, ERRα overexpression further aggravated PAN-induced apoptosis. In agreement with the in vitro study, we also observed increased ERRα expression in line with enhanced apoptotic response in renal cortex from PAN-treated rats. These data suggest a detrimental effect of ERRα on PAN-induced MC apoptosis and inflammatory response, which could help us to better understand the pathogenic mechanism of MC injury in PAN nephropathy.
Asunto(s)
Apoptosis , Receptor alfa de Estrógeno/metabolismo , Nefrosis/metabolismo , Podocitos/metabolismo , Puromicina Aminonucleósido , Animales , Proteínas Reguladoras de la Apoptosis/metabolismo , Línea Celular , Modelos Animales de Enfermedad , Receptor alfa de Estrógeno/genética , Masculino , Ratones , Nefrosis/inducido químicamente , Nefrosis/patología , Podocitos/patología , Ratas Sprague-Dawley , Transducción de SeñalRESUMEN
BACKGROUND: Foot process effacement is one of the pathologic indicators of podocyte injury. However, the morphologic changes associated with it remain unclear. METHODS: To clarify the developmental process, we analyzed puromycin nephrotic podocytes reconstructed from serial focused-ion beam/scanning electron microscopy (FIB/SEM) images. RESULTS: Intact podocytes consisted of four subcellular compartments: cell body, primary process, ridge-like prominence (RLP), and foot process. The RLP, a longitudinal protrusion from the basal surface of the cell body and primary process, served as an adhesive apparatus for the cell body and primary process to attach to the glomerular basement membrane. Foot processes protruded from both sides of the RLP. In puromycin nephrotic podocytes, foot process effacement occurred in two ways: by type-1 retraction, where the foot processes retracted while maintaining their rounded tips; or type-2 retraction, where they narrowed across their entire lengths, tapering toward the tips. Puromycin nephrotic podocytes also exhibited several alterations associated with foot process effacement, such as deformation of the cell body, retraction of RLPs, and cytoplasmic fragmentation. Finally, podocytes were reorganized into a broad, flattened shape. CONCLUSIONS: The three-dimensional reconstruction of podocytes by serial FIB/SEM images revealed the morphologic changes involved in foot process effacement in greater detail than previously described.
Asunto(s)
Membrana Basal Glomerular/patología , Imagenología Tridimensional , Nefrosis/patología , Podocitos/patología , Puromicina Aminonucleósido/farmacología , Tomografía Computarizada por Rayos X/métodos , Animales , Células Cultivadas , Modelos Animales de Enfermedad , Inyecciones Intraperitoneales , Masculino , Microscopía Electrónica de Rastreo/métodos , Nefrosis/inducido químicamente , Podocitos/citología , Podocitos/efectos de los fármacos , Distribución Aleatoria , Ratas , Ratas Wistar , Valores de ReferenciaRESUMEN
Recent studies showed that JAK/STAT pathway plays role in glomerular damages. The fact that STAT3 could be activated also by oxidative stress make Puromycin Aminonucleoside (PAN) Nephrosis model very appropriate for examination of STAT3 expression changes in glomerular pathology. Along with a control group, three PAN groups sacrificed on different days were formed by the i.p. injection of PAN for 5 consecutive days. Throughout the experiment, 24-hour-urines were collected on specific days and proteinuria levels were monitored. At the end of the experiments, tissue specimens were stained immunohistochemically for both total and phosphorylated STAT3 and evaluated subjectively. They were also examined ultrastructurally in transmission electron microscope. The proteinuria levels did not increase significantly on 5th day but showed a dramatic increase on 10th and 15th days. On 20th and 25th days, urinary protein levels gradually decreased. Ultrastructural examinations showed glomerular damages such as significant decrease in slit pore number, a significant gradual increase in glomerular basement membrane thickness and podocyte hypertrophy on 5th and 15th days; besides significant increase in mesangial matrix. The first significant increases in phosphorylated and total STAT3 levels occurred in 5th day and 15th day groups respectively. These increases diminished in 25th day group. Regarding all the findings, it was deduced that STAT3 is one of the active factors in glomerular pathologies.
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Regulación de la Expresión Génica/efectos de los fármacos , Glomérulos Renales/metabolismo , Nefrosis/inducido químicamente , Nefrosis/metabolismo , Puromicina Aminonucleósido/efectos adversos , Factor de Transcripción STAT3/metabolismo , Animales , Glomérulos Renales/ultraestructura , Masculino , Nefrosis/patología , Fosforilación/efectos de los fármacos , Puromicina Aminonucleósido/farmacología , Ratas , Ratas WistarRESUMEN
Rac1, a Rho family member, is ubiquitously expressed and participates in various biological processes. Rac1 expression is induced early in podocyte injury, but its role in repair is unclear. To investigate the role of Rac1 expression in podocytes under pathological conditions, we used podocyte-specific Rac1 conditional knock-out (cKO) mice administered adriamycin (ADR), which causes nephrosis and glomerulosclerosis. Larger areas of detached podocytes, more adhesion of the GBM to Bowman's capsule, and a higher ratio of sclerotic glomeruli were observed in Rac1 cKO mice than in control mice, whereas no differences were observed in glomerular podocyte numbers in both groups after ADR treatment. The mammalian target of rapamycin (mTOR) pathway, which regulates the cell size, was more strongly suppressed in the podocytes of Rac1 cKO mice than in those of control mice under pathological conditions. In accordance with this result, the volumes of podocytes in Rac1 cKO mice were significantly reduced compared with those of control mice. Experiments using in vitro ADR-administered Rac1 knockdown podocytes also supported that a reduction in Rac1 suppressed mTOR activity in injured podocytes. Taken together, these data indicate that Rac1-associated mTOR activation in podocytes plays an important role in preventing the kidneys from developing glomerulosclerosis.
Asunto(s)
Glomeruloesclerosis Focal y Segmentaria/genética , Nefrosis/genética , Neuropéptidos/genética , Podocitos/metabolismo , Serina-Treonina Quinasas TOR/genética , Proteína de Unión al GTP rac1/genética , Animales , Apoptosis/genética , Movimiento Celular/genética , Modelos Animales de Enfermedad , Células Epiteliales/metabolismo , Células Epiteliales/patología , Regulación de la Expresión Génica/genética , Glomeruloesclerosis Focal y Segmentaria/inducido químicamente , Glomeruloesclerosis Focal y Segmentaria/patología , Humanos , Riñón/metabolismo , Riñón/patología , Glomérulos Renales/metabolismo , Glomérulos Renales/patología , Ratones , Ratones Noqueados , Nefrosis/inducido químicamente , Nefrosis/patología , Podocitos/patología , Transducción de Señal/genéticaRESUMEN
BackgroundThe pathogenesis of idiopathic nephrotic syndrome (INS) remains unclear, although recent studies suggest endothelin 1 (ET-1) and CD80 of podocytes are involved. We investigated the potential of antagonist to ET-1 receptor type A (ETRA) as therapeutic agent through the suppression of CD80 in a rat model of INS.MethodsPuromycin aminonucleoside (PAN) was injected to Wister rats to induce proteinuria: some were treated with ETRA antagonist and others were treated with 0.5% methylcellulose. Blood and tissue samples were collected. Quantitative PCR was used to determine the expression of Toll-like receptor-3 (TLR-3), nuclear factor-κB (NF-κB), CD80, talin, ETRA, and ET-1 in the kidney. To confirm the level of CD80 protein expression, immunofluorescence staining and western blot analysis of the renal tissue were performed.ResultsAmount of proteinuria in the treatment group was significantly lower than the other groups. The same-day body weight, serum creatinine values, and blood pressure were not significantly different. ETRA antagonist restores podocyte foot process effacement as well as the aberrant expression of TLR-3, nuclear factor-κB (NF-κB), and CD80 in PAN-injured kidneys.ConclusionsThe ETRA antagonist may be promising drug for INS as it showed an antiproteinuric effect. Its action was considered to be through suppression of CD80 expression on podocytes.
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Antagonistas de los Receptores de la Endotelina A/farmacología , Nefrosis/inducido químicamente , Proteinuria/tratamiento farmacológico , Puromicina Aminonucleósido/efectos adversos , Animales , Antígeno B7-1/metabolismo , Presión Sanguínea , Peso Corporal , Creatinina/sangre , Modelos Animales de Enfermedad , Endotelina-1/metabolismo , Femenino , Riñón/metabolismo , Enfermedades Renales/patología , Glomérulos Renales/metabolismo , FN-kappa B/metabolismo , Síndrome Nefrótico , Fenilpropionatos/farmacología , Podocitos/metabolismo , Piridazinas/farmacología , Ratas , Ratas Wistar , Receptor de Endotelina A/metabolismo , Receptor Toll-Like 3/metabolismoRESUMEN
Nephrin is a key structural component of the podocyte slit diaphragm, and proper expression of nephrin on the cell surface is critical to ensure integrity of the blood filtration barrier. Maintenance of nephrin within this unique cell junction has been proposed to require dynamic phosphorylation events and endocytic recycling, although the molecular mechanisms that control this interplay are poorly understood. Here, we investigated the possibility that the phosphotyrosine adaptor protein ShcA regulates nephrin turnover. Western blotting and immunostaining analysis confirmed that ShcA is expressed in podocytes. In immunoprecipitation and pulldown assays, ShcA, via its SH2 domain, was associated with several phosphorylated tyrosine residues on nephrin. Overexpression of ShcA promoted nephrin tyrosine phosphorylation and reduced nephrin signaling and cell surface expression in vitro In a rat model of reversible podocyte injury and proteinuria, phosphorylated nephrin temporally colocalized with endocytic structures coincident with upregulation of ShcA expression. In vivo biotinylation assays confirmed that nephrin expression decreased at the cell surface and correspondingly increased in the cytosol during the injury time course. Finally, immunostaining in kidney biopsy specimens demonstrated overexpression of ShcA in several human proteinuric kidney diseases compared with normal conditions. Our results suggest that increases in ShcA perturb nephrin phosphosignaling dynamics, leading to aberrant nephrin turnover and slit diaphragm disassembly.
Asunto(s)
Endocitosis , Enfermedades Renales/metabolismo , Proteínas de la Membrana/metabolismo , Podocitos/metabolismo , Proteinuria/metabolismo , Proteína Transformadora 1 que Contiene Dominios de Homología 2 de Src/metabolismo , Animales , Biotinilación , Membrana Celular/metabolismo , Citosol/metabolismo , Células HEK293 , Humanos , Enfermedades Renales/patología , Masculino , Nefrosis/inducido químicamente , Fosforilación , Ratas , Ratas Sprague-Dawley , Transducción de Señal , Proteína Transformadora 1 que Contiene Dominios de Homología 2 de Src/genética , Tirosina/metabolismo , Regulación hacia ArribaRESUMEN
The highly conserved spalt (sal) gene family members encode proteins characterized by multiple double zinc finger motifs of the C2H2 type. Humans and mice each have four known Sal-like genes (SALL1-4 in humans and Sall1-4 in mice). Sall1 is known to have a crucial role in kidney development. To explore the significance of Sall1 in differentiated podocytes, we investigated podocyte-specific Sall1-deficient mice (Sall1 KOp°d°/p°d°) using a podocin-Cre/loxP system and siRNA Sall1 knockdown (Sall1 KD) podocytes. Under physiological conditions, Sall1 KOp°d°/p°d° mice exhibited no proteinuria during their lifetime, but foot-process effacement was detected in some of the podocytes. To elucidate the role of Sall1 in injured podocytes, we used an adriamycin (ADR)-induced model of nephrosis and glomerulosclerosis. Surprisingly, the expression of Sall1 was elevated in control mice on day 14 after ADR injection. On day 28 after ADR injection, Sall1 KOp°d°/p°d° mice exhibited significantly higher levels of proteinuria and higher numbers of sclerotic glomeruli. Differentiated Sall1 KD podocytes showed a loss of synaptopodin, suppressed stress fiber formation, and, ultimately, impaired directed cell migration. In addition, the loss of Sall1 increased the number of apoptotic podocytes following ADR treatment. These results indicated that Sall1 has a protective role in podocytes; thus, we investigated the endoplasmic reticulum stress marker GRP78. GRP78 expression was higher in ADR-treated Sall1 KOp°d°/p°d° mice than in control mice. Sall1 appeared to influence the expression of GRP78 in injured podocytes. These results suggest that Sall1 is associated with actin reorganization, endoplasmic reticulum stress, and apoptosis in injured podocytes. These protective aspects of Sall1 re-expression in injured podocytes may have the potential to reduce apoptosis and possibly glomerulosclerosis.
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Antibióticos Antineoplásicos/efectos adversos , Doxorrubicina/efectos adversos , Riñón/efectos de los fármacos , Nefrosis/prevención & control , Podocitos/metabolismo , Inhibidores de Topoisomerasa II/efectos adversos , Factores de Transcripción/metabolismo , Citoesqueleto de Actina/efectos de los fármacos , Citoesqueleto de Actina/metabolismo , Citoesqueleto de Actina/patología , Animales , Apoptosis/efectos de los fármacos , Biomarcadores , Línea Celular Transformada , Movimiento Celular/efectos de los fármacos , Cruzamientos Genéticos , Chaperón BiP del Retículo Endoplásmico , Estrés del Retículo Endoplásmico/efectos de los fármacos , Regulación de la Expresión Génica/efectos de los fármacos , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Riñón/metabolismo , Riñón/patología , Ratones Noqueados , Ratones Transgénicos , Proteínas de Microfilamentos/genética , Proteínas de Microfilamentos/metabolismo , Nefrosis/inducido químicamente , Nefrosis/metabolismo , Nefrosis/patología , Podocitos/efectos de los fármacos , Podocitos/patología , Interferencia de ARN , Proteínas Recombinantes/metabolismo , Factores de Transcripción/antagonistas & inhibidores , Factores de Transcripción/genéticaRESUMEN
PURPOSE: Podocyte injury is a key event in proteinuric kidney disease and eventually glomerular scarring. While adrenomedullin (AM), a potent vasodilatory peptide, has been reported to confer renoprotection in several experimental models of kidney diseases, its effect on injured podocytes and the related mechanism is still largely unknown. METHODS: Employing Western blotting analysis, immunoprecipitation and immunofluorescence, we investigated the effects of AM on the expressions of podocyte cytoskeletal proteins and Rho-family small GTPases (Rho GTPases) in puromycin aminonucleoside (PAN)-induced podocyte injury, both in cultured podocytes and in PAN nephrosis rats. Urinary protein excretion and the morphologic changes of kidney in PAN nephrosis rats were evaluated. Glutathione-S-transferase pull-down assay was applied for Rho GTPases activity. RESULTS: PAN induced massive albuminuria and morphologic injury, which were significantly mitigated by AM administration. AM significantly antagonized not only the PAN-decreased expressions of synaptopodin, nephrin, CD2AP and podocin, but also the PAN-disrupted interactions between synaptopodin-RhoA, nephrin-CD2AP, and CD2AP-Rac1-cortactin. These effects of AM in cultured podocytes were mostly significantly blocked by H89, a PKA inhibitor. AM dramatically upregulated the PAN-induced Rho GTPases protein expressions and their activities. PAN increased the expressions of endogenous AM and its receptor RAMP2 which was furthermore upregulated by AM administration. CONCLUSIONS: AM alleviated podocyte injury induced by PAN both in cell culture and in PAN nephrosis. The beneficial effects of AM on podocytes can be attributable to direct modulation of podocyte cytoskeletal proteins and Rho GTPases, mainly via a PKA-dependent pathway.
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Adrenomedulina/uso terapéutico , Nefrosis/tratamiento farmacológico , Nefrosis/metabolismo , Podocitos/ultraestructura , Vasodilatadores/farmacología , Proteínas de Unión al GTP rho/metabolismo , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Adrenomedulina/metabolismo , Adrenomedulina/farmacología , Albuminuria/tratamiento farmacológico , Albuminuria/etiología , Animales , Línea Celular , Cortactina/metabolismo , Proteínas del Citoesqueleto/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Glomérulos Renales/patología , Masculino , Proteínas de la Membrana/metabolismo , Ratones , Proteínas de Microfilamentos/metabolismo , Nefrosis/inducido químicamente , Nefrosis/patología , Neuropéptidos/metabolismo , Podocitos/efectos de los fármacos , Puromicina Aminonucleósido , Ratas , Ratas Sprague-Dawley , Proteína 2 Modificadora de la Actividad de Receptores/metabolismo , Vasodilatadores/uso terapéutico , Proteína de Unión al GTP cdc42/metabolismo , Proteína de Unión al GTP rac1/metabolismo , Proteína de Unión al GTP rhoARESUMEN
PURPOSE: The purpose of this study was to investigate the effects of bone marrow-derived mesenchymal stem cells (BMSC) on podocytes of puromycin amino nuclear glucoside (PAN) -induced nephrosis in mice. METHODS: Mice were randomly divided into Control, PAN and BMSC groups. Mice were injected with PAN (0.5 mg/g weight) via the tail vein. The 24-h urinary protein was obtained after modelling, and urinary protein excretion was determined. The blood and kidney specimens were isolated after the tenth day of modelling. Blood samples were collected for measuring serum creatinine (SCr) and blood urea nitrogen (BUN). A sample of kidney was taken for observing pathological changes through hematoxylin-eosin staining and electron microscopy, and the rest of the kidney was used for detecting the protein and mRNA expression of nephrin, CD2AP, synaptopodin, TRPC6 by real-time quantitative PCR, Western-blot and immunohistochemistry. RESULTS: After PAN injection, podocyte foot process fusion was detected by electron microscopy, and the 24 h urinary protein excretion increased compared with control mice on days 3, 7 and 10 post-PAN injection (P.
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Células de la Médula Ósea/metabolismo , Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas/metabolismo , Nefrosis/terapia , Podocitos/metabolismo , Aloinjertos , Animales , Células de la Médula Ósea/patología , Células Madre Mesenquimatosas/patología , Ratones Endogámicos BALB C , Nefrosis/inducido químicamente , Nefrosis/metabolismo , Nefrosis/patología , Podocitos/patología , Puromicina Aminonucleósido/efectos adversos , Puromicina Aminonucleósido/farmacologíaRESUMEN
We previously described albumin endocytosis through caveolae in human renal glomerular endothelial cells (HRGECs). This suggested a new albumin transcytosis pathway, in addition to the fenestral pathway. As a next step, we investigated albumin transcytosis in HRGECs after caveolar endocytosis. HRGECs were incubated with Alexa Fluor 488-labeled bovine serum albumin from 0 to 360 min. Next, markers for endosomes, endoplasmic reticulum (ER), golgi apparatus (GA), lysosomes, and proteasomes and Fc receptors, microtubules, and actin were monitored by immunofluorescence. Labeled albumin co-localization with endosomes was gradually and significantly increased and it was significantly higher than with the other markers at any timepoint. Albumin, placed on inside of the Transwell membrane, diffused through HRGEC monolayers during a 360 min incubation period. This transportation of albumin through HRGECs was inhibited by methyl beta cyclodextrin (MBCD), a caveolae disrupting agent. MBCD also decreased albuminuria, causing decreased caveolin-1 (Cav-1) expression on glomerular capillaries, in puromycin aminonucleoside induced nephrotic mice. Albumin transcytosis depends on early endosomes, but not on other organelles, Fc receptors, or cytoskeletal components. Caveolae disruption prevented albumin transportation through HRGECs and decreased albuminuria in nephrotic mice. This newly described caveolae-dependent albumin pathway through glomerular endothelial cells is a potential pathogenetic mechanism for albuminuria, independent of the fenestrae.
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Albuminuria/metabolismo , Caveolas/metabolismo , Endocitosis , Endosomas/metabolismo , Células Endoteliales/metabolismo , Glomérulos Renales/irrigación sanguínea , Albúmina Sérica Bovina/metabolismo , Transcitosis , Albuminuria/inducido químicamente , Albuminuria/prevención & control , Animales , Caveolas/efectos de los fármacos , Caveolina 1/metabolismo , Células Cultivadas , Modelos Animales de Enfermedad , Endocitosis/efectos de los fármacos , Endosomas/efectos de los fármacos , Células Endoteliales/efectos de los fármacos , Humanos , Masculino , Ratones Endogámicos C57BL , Nefrosis/inducido químicamente , Nefrosis/metabolismo , Puromicina Aminonucleósido , Factores de Tiempo , Transcitosis/efectos de los fármacos , beta-Ciclodextrinas/farmacologíaRESUMEN
BACKGROUND: Sodium bicarbonate and ascorbic acid have been proposed to prevent contrast-induced nephropathy (CIN). The present study evaluated the effect of their combined use on CIN incidence.MethodsâandâResults:We prospectively enrolled 429 patients with chronic kidney disease (CKD: baseline estimated glomerular filtration rate <60 mL/min/1.73 m2) prior to elective coronary catheterization. CIN was defined as absolute (≥0.5 mg/dL) or relative (≥25%) increase in serum creatinine within 72 h. In the saline hydration (n=218) and combined sodium bicarbonate+ascorbic acid (n=211) groups, a total of 1,500-2,500 mL 0.9% saline was given before and after the procedure. In addition, the combination group received 20 mEq sodium bicarbonate and 3 g ascorbic acid i.v. before the procedure, followed by 2 g ascorbic acid after the procedure and a further 2 g after 12 h. There were no significant differences between the basic characteristics and contrast volume in the 2 groups. CIN occurred in 19 patients (8.7%) in the saline group, and in 6 patients (2.8%) in the combined treatment group (P=0.008). CONCLUSIONS: Combined sodium bicarbonate and ascorbic acid could prevent CIN following catheterization in CKD patients.
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Ácido Ascórbico/uso terapéutico , Medios de Contraste/efectos adversos , Nefrosis/prevención & control , Insuficiencia Renal Crónica/complicaciones , Bicarbonato de Sodio/uso terapéutico , Anciano , Anciano de 80 o más Años , Cateterismo Cardíaco , Creatinina/sangre , Quimioterapia Combinada , Femenino , Humanos , Masculino , Persona de Mediana Edad , Nefrosis/inducido químicamente , Insuficiencia Renal Crónica/terapiaRESUMEN
Acute kidney injury (AKI) is an abrupt reduction in kidney function caused by different pathological processes. It is associated with a significant morbidity and mortality in the acute phase and an increased risk of developing End Stage Renal Disease. Despite the progress in the management of the disease, mortality rates in the last five decades remain unchanged at around 50%. Therefore there is an urgent need to find new therapeutic strategies to treat AKI. Lysosomal proteases, particularly Cathepsin D (CtsD), play multiple roles in apoptosis however, their role in AKI is still unknown. Here we describe a novel role for CtsD in AKI. CtsD expression was upregulated in damaged tubular cells in nephrotoxic and ischemia reperfusion (IRI) induced AKI. CtsD inhibition using Pepstatin A led to an improvement in kidney function, a reduction in apoptosis and a decrease in tubular cell damage in kidneys with nephrotoxic or IRI induced AKI. Pepstatin A treatment slowed interstitial fibrosis progression following IRI induced AKI. Renal transplant biopsies with acute tubular necrosis demonstrated high levels of CtsD in damaged tubular cells. These results support a role for CtsD in apoptosis during AKI opening new avenues for the treatment of AKI by targeting lysosomal proteases.
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Lesión Renal Aguda/metabolismo , Catepsina D/metabolismo , Túbulos Renales/citología , Nefrosis/complicaciones , Daño por Reperfusión/complicaciones , Lesión Renal Aguda/tratamiento farmacológico , Lesión Renal Aguda/etiología , Lesión Renal Aguda/fisiopatología , Animales , Apoptosis , Línea Celular , Modelos Animales de Enfermedad , Ácido Fólico/efectos adversos , Humanos , Pruebas de Función Renal , Túbulos Renales/efectos de los fármacos , Túbulos Renales/enzimología , Masculino , Ratones , Nefrosis/inducido químicamente , Nefrosis/tratamiento farmacológico , Nefrosis/enzimología , Pepstatinas/administración & dosificación , Pepstatinas/farmacología , Daño por Reperfusión/tratamiento farmacológico , Regulación hacia ArribaRESUMEN
Bile cast nephropathy is a tubulo-interstitial nephropathy. Its diagnosis may be under-estimated. It develops in patients who have cholestatic jaundice, with high bilirubinemia. Bile salts are freely filtered through glomerulus. Under certain circumstances, it forms casts into the tubule and cause an acute tubular necrosis. The diagnosis evidence is histologic, but fulfilment of renal biopsy is often made difficult, because of the hemostatic abnormalities that patients with hepatocellular injury develop. The treatment is supportive and etiological. We report here the case of a patient who presented a drug-induced hepatic jaundice, complicated with acute kidney failure secondary to bile cast nephropathy. We present the histological diagnosis evidence.
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Ictericia Obstructiva/diagnóstico , Ictericia Obstructiva/terapia , Nefrosis/diagnóstico , Nefrosis/terapia , Diálisis Renal , Lesión Renal Aguda/diagnóstico , Lesión Renal Aguda/terapia , Anciano , Antioxidantes/metabolismo , Ácidos y Sales Biliares/metabolismo , Bilirrubina/sangre , Biomarcadores/sangre , Biopsia , Fármacos Gastrointestinales/metabolismo , Humanos , Ictericia Obstructiva/sangre , Ictericia Obstructiva/inducido químicamente , Túbulos Renales/efectos de los fármacos , Túbulos Renales/patología , Masculino , Necrosis , Nefrosis/sangre , Nefrosis/inducido químicamente , Diálisis Renal/métodos , Factores de Riesgo , Resultado del TratamientoRESUMEN
Drugs containing adrenocorticotropic hormone have been used as therapy for patients with nephrotic syndrome. We have previously shown that adrenocorticotropic hormone and a selective agonist for the melanocortin 1 receptor (MC1R) exert beneficial actions in experimental membranous nephropathy with reduced proteinuria, reduced oxidative stress, and improved glomerular morphology and function. Our hypothesis is that MC1R activation in podocytes elicits beneficial effects by promoting stress fibers and maintaining podocyte viability. To test the hypothesis, we cultured podocytes and used highly specific agonists for MC1R. Podocytes were subjected to the nephrotic-inducing agent puromycin aminonucleoside, and downstream effects of MC1R activation on podocyte survival, antioxidant defense, and cytoskeleton dynamics were studied. To increase the response and enhance intracellular signals, podocytes were transduced to overexpress MC1R. We showed that puromycin promotes MC1R expression in podocytes and that activation of MC1R promotes an increase of catalase activity and reduces oxidative stress, which results in the dephosphorylation of p190RhoGAP and formation of stress fibers through RhoA. In addition, MC1R agonists protect against apoptosis. Together, these mechanisms protect the podocyte against puromycin. Our findings strongly support the hypothesis that selective MC1R-activating agonists protect podocytes and may therefore be useful to treat patients with nephrotic syndromes commonly considered as podocytopathies.
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Catalasa/metabolismo , Podocitos/efectos de los fármacos , Receptor de Melanocortina Tipo 1/agonistas , Proteína de Unión al GTP rhoA/metabolismo , Animales , Antimetabolitos , Células Cultivadas , Activación Enzimática , Ratones , Nefrosis/inducido químicamente , Nefrosis/metabolismo , Estrés Oxidativo/efectos de los fármacos , Puromicina Aminonucleósido , Especies Reactivas de Oxígeno/metabolismo , Receptor de Melanocortina Tipo 1/genética , Fibras de Estrés/efectos de los fármacosRESUMEN
PodocyteTRPC6 channels have been implicated in glomerular diseases. Syndecan-4 (Sdc4) is a membrane proteoglycan that can be cleaved to release a soluble ectodomain capable of paracrine and autocrine signaling. We have confirmed that overexpression of Sdc4 core protein increases surface abundance of TRPC6 channels in cultured podocytes, whereas Sdc4 knockdown has the opposite effect. Exposure to soluble Sdc4 ectodomain also increased the surface abundance of TRPC6, and increased cationic currents evoked by a diacylglycerol analog in podocytes. Sdc4 ectodomain increased generation of reactive oxygen species (ROS), reduced activation of RhoA, increased activation of Rac1, increased nuclear abundance of NFATc1, and increased total ß3-integrin. The effects of Sdc4 ectodomain on cell-surface TRPC6 were blocked by the ROS quencher TEMPOL, and by the Rac1 inhibitor NSC-23766, but were not blocked by inhibition of calcineurin-NFATc1 signaling. The Sdc4 core protein co-immunoprecipitates with ß3-integrin in cultured podocytes. Moreover, effects of Sdc4 ectodomain on TRPC6, ROS generation, Rac1 and RhoA modulation, and NFATc1 activation were blocked by cilengitide, a selective inhibitor of outside-in signaling through αv-containing integrins. Exposure to TNF, or serum from three patients with recurrent FSGS in relapse, increased shedding of podocyte Sdc4 ectodomains into the surrounding medium. This was also observed after treating podocytes with the metalloproteinase ADAM17 or after overexpression of the Sdc4 core protein. Increased concentrations of Sdc4 ectodomain were detected in urine of rats during acute puromycin aminonucleoside nephrosis. Locally generated Sdc4 may play a role in regulating TRPC6 channels, and may contribute to glomerular pathology.