RESUMO
Interleukin (IL)-17-secreting invariant natural killer T (NKT) cells are involved in several inflammatory diseases. However, their role in lupus nephritis (LN) has not been fully characterized. Samples from patients with LN or glomerulonephritis and healthy controls were obtained, and elevated IL-17+ NKT cell numbers and IL-17 expression were observed in blood cells and kidneys, respectively, in patients with LN. Comparison of a mouse model of experimental autoimmune LN with the parental strain (NKT-deficient B6.CD1d-/- mice) revealed improved proteinuria, disease severity, and histopathology and decreased levels of chemokine (C-X-C motif) ligand 16 and T cell receptor-α variable 14 expression. Spleens and kidneys of B6.CD1d-/- mice also showed downregulation of inflammatory markers and IL-17. In coculture with renal mesangial and NKT cells, inflammatory markers and IL-17 were upregulated following α-galactosylceramide treatment and downregulated after treatment with IL-17-blocking antibodies. This was most prominent with killer cell lectin-like receptor subfamily B member 1 C (NK1.1)- NKT cells. Thus, IL-17 is upregulated in LN. Activation of NKT cells regulates IL-17-related immune responses systemically and in the kidneys, primarily via NK1.1- NKT cells. IL-17-secreting NK1.1- NKT cells could serve as diagnostic and therapeutic targets for LN.NEW & NOTEWORTHY This study makes a significant contribution to the literature because our results indicate that IL-17 is upregulated in lupus nephritis and that natural killer T (NKT) cells are involved in its pathogenesis. Activation of NKT cells regulates IL-17-related immune responses, both systemically and in the kidney, and this mainly involves NK1.1- NKT cells. Furthermore, IL-17-secreting NK1.1- NKT cells could serve as a diagnostic and therapeutic target for lupus nephritis.
Assuntos
Interleucina-17/metabolismo , Rim/metabolismo , Nefrite Lúpica/metabolismo , Ativação Linfocitária , Células T Matadoras Naturais/metabolismo , Animais , Estudos de Casos e Controles , Comunicação Celular , Células Cultivadas , Técnicas de Cocultura , Modelos Animais de Doenças , Humanos , Rim/imunologia , Rim/patologia , Nefrite Lúpica/imunologia , Nefrite Lúpica/patologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Células T Matadoras Naturais/imunologia , Fenótipo , Índice de Gravidade de Doença , Células Th17/imunologia , Células Th17/metabolismo , Regulação para CimaRESUMO
BACKGROUNDS: IgA nephropathy (IgAN) is the most common primary glomerulonephritis causing end stage renal disease (ESRD), and vasculopathy is known to involve disease progression. Klotho, a gene related to aging, has been reported to play a role in atherosclerosis and endothelial dysfunction. We investigated whether klotho gene polymorphism affect clinical course of IgAN. METHODS: The data registered for PREMIER study which enrolled the patients with biopsy proven IgAN were analyzed. Two single nucleotide polymorphisms for klotho gene, G395A of promoter region and C1818T of exon 4, were examined, and investigated the association klotho genotypes with the progression of IgAN and patient survival. RESULTS: Clinical data from 973 patients confirmed about survival were analyzed. The allele frequency was 0.830 and 0.170 for allele G and A, and 0.816 and 0.184 for allele C and T, which were complied with Hardy-Weinberg equilibrium (p=0.996 and 0.531 respectively). Death was observed more frequently in A-allele carriers of G395A polymorphism (0.7 vs. 2.6 %, GG vs. GA+AA, p=0.022). Renal survival in Kaplan-Meier survival curve was also worse in same group (p=0.04). CONCLUSION: Klotho gene polymorphism was associated with patient survival and disease progression of IgAN.
Assuntos
Progressão da Doença , Glomerulonefrite por IGA/genética , Glomerulonefrite por IGA/mortalidade , Glucuronidase/genética , Polimorfismo de Nucleotídeo Único/genética , Adulto , Éxons/genética , Feminino , Frequência do Gene/genética , Genótipo , Glomerulonefrite por IGA/etnologia , Humanos , Estimativa de Kaplan-Meier , Proteínas Klotho , Coreia (Geográfico) , Masculino , Pessoa de Meia-Idade , Taxa de SobrevidaRESUMO
The most widespread dietary problem in the world is mineral deficiency. We used the nicotianamine synthase (NAS) gene to increase mineral contents in rice grains. Nicotianamine (NA) is a chelator of metals and a key component of metal homeostasis. We isolated activation-tagged mutant lines in which expression of a rice NAS gene, OsNAS3, was increased by introducing 35S enhancer elements. Shoots and roots of the OsNAS3 activation-tagged plants (OsNAS3-D1) accumulated more Fe and Zn. Seeds from our OsNAS3-D1 plants grown on a paddy field contained elevated amounts of Fe (2.9-fold), Zn (2.2-fold), and Cu (1.7-fold). The NA level was increased 9.6-fold in OsNAS3-D1 seeds. Analysis by size exclusion chromatography coupled with inductively coupled plasma mass spectroscopy showed that WT and OsNAS3-D1 seeds contained equal amounts of Fe bound to IP6, whereas OsNAS3-D1 had 7-fold more Fe bound to a low molecular mass, which was likely NA. Furthermore, this activation led to increased tolerance to Fe and Zn deficiencies and to excess metal (Zn, Cu, and Ni) toxicities. In contrast, disruption of OsNAS3 caused an opposite phenotype. To test the bioavailability of Fe, we fed anemic mice with either engineered or WT seeds for 4 weeks and measured their concentrations of hemoglobin and hematocrit. Mice fed with engineered seeds recovered to normal levels of hemoglobin and hematocrit within 2 weeks, whereas those that ate WT seeds remained anemic. Our results suggest that an increase in bioavailable mineral content in rice grains can be achieved by enhancing NAS expression.
Assuntos
Alquil e Aril Transferases/genética , Ração Animal , Regulação Enzimológica da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Ferro/administração & dosagem , Oryza/genética , Sementes/metabolismo , Adaptação Fisiológica , Anemia Ferropriva/terapia , Animais , Espectrometria de Massas , Metais Pesados/toxicidade , Camundongos , Mutação , Oryza/embriologia , Oryza/enzimologia , Oryza/fisiologiaRESUMO
There is a significant immune response to ischemia-reperfusion injury (IRI), but the role of immunomodulatory natural killer T (NKT) cell subtypes is not well understood. Here, we compared the severity of IRI in mice deficient in type I/II NKT cells (CD1d(-/-)) or type I NKT cells (Jα18(-/-)). The absence of NKT cells, especially type II NKT cells, accentuated the severity of renal injury, whereas repletion of NKT cells attenuated injury. Adoptively transferred NKT cells trafficked into the tubulointerstitium, which is the primary area of injury. Sulfatide-induced activation of type II NKT cells protected kidneys from IRI, but inhibition of NKT cell recruitment enhanced injury. In co-culture experiments, sulfatide-induced activation of NKT cells from either mice or humans attenuated apoptosis of renal tubular cells after transient hypoxia via hypoxia-inducible factor (HIF)-1α and IL-10 pathways. Renal tissue of patients with acute tubular necrosis (ATN) frequently contained NKT cells, and the number of these cells tended to negatively correlate with ATN severity. In summary, sulfatide-reactive type II NKT cells are renoprotective in IRI, suggesting that pharmacologic modulation of NKT cells may protect against ischemic injury.
Assuntos
Injúria Renal Aguda/imunologia , Células T Matadoras Naturais/fisiologia , Traumatismo por Reperfusão/imunologia , Animais , Citocinas/metabolismo , Células Epiteliais/metabolismo , Hipóxia/metabolismo , Fator 1 Induzível por Hipóxia/metabolismo , Interleucina-10/metabolismo , Túbulos Renais/metabolismo , Ativação Linfocitária , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Traumatismo por Reperfusão/metabolismo , Sulfoglicoesfingolipídeos/metabolismoRESUMO
We generated rice lines with increased content of nicotianamine (NA), a key ligand for metal transport and homeostasis. This was accomplished by activation tagging of rice nicotianamine synthase 2 (OsNAS2). Enhanced expression of the gene resulted in elevated NA levels, greater Zn accumulations and improved plant tolerance to a Zn deficiency. Expression of Zn-uptake genes and those for the biosynthesis of phytosiderophores (PS) were increased in transgenic plants. This suggests that the higher amount of NA led to greater exudation of PS from the roots, as well as stimulated Zn uptake, translocation and seed-loading. In the endosperm, the OsNAS2 activation-tagged line contained up to 20-fold more NA and 2.7-fold more zinc. Liquid chromatography combined with inductively coupled plasma mass spectrometry revealed that the total content of zinc complexed with NA and 2'-deoxymugineic acid was increased 16-fold. Mice fed with OsNAS2-D1 seeds recovered more rapidly from a zinc deficiency than did control mice receiving WT seeds. These results demonstrate that the level of bio-available zinc in rice grains can be enhanced significantly by activation tagging of OsNAS2.
Assuntos
Alquil e Aril Transferases/metabolismo , Regulação Enzimológica da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Oryza/enzimologia , Zinco/metabolismo , Alquil e Aril Transferases/genética , Animais , Ácido Azetidinocarboxílico/análogos & derivados , Ácido Azetidinocarboxílico/metabolismo , Endosperma/metabolismo , Genes de Plantas , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Músculos/metabolismo , Oryza/genética , Plantas Geneticamente Modificadas/enzimologia , Plantas Geneticamente Modificadas/genética , Plasma/metabolismo , Sideróforos/biossíntese , Aumento de Peso , Zinco/deficiênciaRESUMO
The TonE-binding protein (TonEBP) is a transcriptional activator in the Rel family that includes NFkappaB and NFAT. TonEBP is critical for the development and function of the renal medulla, which is a major regulator of water homeostasis. TonEBP is also implicated in diabetic nephropathy and inflammation. Established methods for biochemical and histochemical detection and functional analysis of TonEBP, including identification of novel TonEBP target genes, are described for those who are interested in investigating function and regulation of TonEBP.
Assuntos
Diferenciação Celular/fisiologia , Fatores de Transcrição NFATC/fisiologia , Sequência de Aminoácidos , Animais , Ensaio de Desvio de Mobilidade Eletroforética , Humanos , Soluções Hipertônicas/farmacologia , Medula Renal/fisiologia , Dados de Sequência Molecular , Fatores de Transcrição NFATC/análise , Fatores de Transcrição NFATC/imunologia , Interferência de RNARESUMO
BACKGROUND: The relationship between SLC12A3 mutations and actual sodium-chloride (Na-Cl) cotransporter (NCC) expression in patients with Gitelman syndrome (GS) was rarely evaluated. Detection of urinary thiazide-sensitive NCC was not tried in patients with GS. STUDY DESIGN: Case series. SETTING & PARTICIPANTS: 6 patients with GS and 1 patient with surreptitious vomiting. OUTCOMES & MEASUREMENTS: Renal clearance study, mutation analysis using reverse-transcription polymerase chain reaction and direct sequencing for the SLC12A3 gene, and immunohistochemical staining for NCC, Na-K-2Cl-cotransporter, alpha1-subunit of Na(+),K(+)-ATPase, and calbindin-D(28K) of the renal biopsy specimens were performed. Membrane fractions of urine were obtained by using differential centrifugation and probed with antibodies against human NCC and aquaporin 2. RESULTS: Results of clearance studies were consistent with GS, showing decreased distal fractional chloride reabsorption with only furosemide. SLC12A3 gene mutations were found in all patients with GS. Immunohistochemistry showed markedly decreased NCC expression in the distal convoluted tubule, whereas expression of other transporters remained intact. Urinary NCC excretion was markedly decreased in patients with GS, but not in the patient with surreptitious vomiting. LIMITATIONS: Small number of patients and lack of mutation analysis of CLCNKB. CONCLUSIONS: There were no relations between NCC expression and types of mutations. Detection of urinary NCC might be helpful for the differential diagnosis of GS.
Assuntos
Síndrome de Gitelman/urina , Receptores de Droga/genética , Simportadores de Cloreto de Sódio-Potássio/urina , Simportadores/genética , Adolescente , Adulto , Centrifugação , Creatinina/urina , Análise Mutacional de DNA , Feminino , Variação Genética , Síndrome de Gitelman/diagnóstico , Síndrome de Gitelman/genética , Humanos , Immunoblotting , Imuno-Histoquímica , Túbulos Renais Distais , Masculino , Concentração Osmolar , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Membro 3 da Família 12 de Carreador de SolutoRESUMO
Atherosclerosis is a chronic progressive inflammatory disorder and the leading cause of cardiovascular mortality. Here we assessed the dynamic changes of T-cell-derived cytokines, such as inteferon (IFN)-γ, interleukin (IL)-17 and IL-4, during the progression of atherosclerosis in apolipoprotein E-null (ApoE(-/-)) mice, to understand the role of immune responses in different stages of atherosclerosis. Male ApoE(-/-) mice were fed a high-fat, western-type diet (WD: 21% lipid, 1.5% cholesterol) after 5 weeks of age and were compared with C57BL/6 wild-type control mice fed a standard chow diet. Atherosclerotic lesions appeared in the aortic sinus of ApoE(-/-) mice 4 weeks after WD and the lesions progressed and occupied >50% of the total sinus area 16 weeks after WD. Aortic IL-17 mRNA and protein expression started to increase in ApoE(-/-) mice after 4 weeks on the WD and peaked at around 8-12 weeks on the WD. In terms of systemic expression of T-cell-derived cytokines, IL-17 production from splenocytes after anti-CD3/CD28 stimuli increased from 4 weeks on the WD, peaked at 12 weeks and returned to control levels at 16 weeks. The production of IFN-γ and IL-4 (Th1 and Th2 cytokines, respectively) from splenocytes was delayed compared with IL-17. Taken together, the present data indicate that Th17 cell response may be involved at an early stage in the development of atherosclerosis.
Assuntos
Aorta/patologia , Apolipoproteínas E/genética , Aterosclerose/genética , Aterosclerose/patologia , Interleucina-17/genética , Animais , Aorta/metabolismo , Aterosclerose/etiologia , Aterosclerose/imunologia , Dieta Hiperlipídica/efeitos adversos , Deleção de Genes , Interferon gama/genética , Interleucina-17/imunologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Linfócitos T/imunologia , Linfócitos T/metabolismo , Linfócitos T/patologia , Regulação para CimaRESUMO
The aquaporin-2 (AQP2) water channel is mainly located in the apical plasma membrane of collecting duct epithelial cells, but there has been some evidence of a moderate amount of basolateral localization of AQP2 at least in the inner medullary collecting duct (IMCD). Previous in vitro microperfusion studies showed that oxytocin has an antidiuretic action, most likely mediated by the vasopressin V2 receptor (V2R) in rat IMCD. By using immunohistochemistry in kidneys from male Sprague-Dawley rats, we observed acute effects of oxytocin on AQP2 localization which were prevented by a V2R antagonist. After intraperitoneal administration of oxytocin (10 U), immunohistochemistry of IMCD revealed that AQP2 was shifted from diffuse cytoplasmic localization in controls to the apical and basolateral membrane domains in oxytocin-treated rats. This pattern of AQP2 redistribution was noted in connecting tubule, cortical collecting duct and outer medullary collecting duct as well as in IMCD, although the tendency to basolateral localization was somewhat less. The pretreatment using a V2R antagonist blocked redistribution of AQP2 in response to oxytocin. We conclude that oxytocin induces a V2R-mediated redistribution of AQP2-containing cytoplasmic vesicles to both apical and basolateral plasma membrane domains in rat kidney. Oxytocin may be one of the factors that accounts for vasopressin-independent AQP2 targeting in the kidney.
Assuntos
Aquaporinas/metabolismo , Rim/efeitos dos fármacos , Ocitocina/farmacologia , Animais , Aquaporina 2 , Aquaporina 3 , Aquaporina 4 , Aquaporina 6 , Imuno-Histoquímica , Rim/química , Medula Renal/química , Medula Renal/efeitos dos fármacos , Túbulos Renais/química , Túbulos Renais/efeitos dos fármacos , Túbulos Renais Coletores/química , Túbulos Renais Coletores/efeitos dos fármacos , Masculino , Concentração Osmolar , Ratos , Ratos Sprague-Dawley , Proteínas Recombinantes/farmacologia , Micção/efeitos dos fármacos , Urina/química , Urodinâmica/efeitos dos fármacosRESUMO
Because micronutrients in human diets ultimately come from plant sources, malnutrition of essential minerals is a significant public health concern. By increasing the expression of nicotianamine synthase (NAS), we fortified the level of bioavailable iron in rice seeds. Activation of iron deficiency-inducible OsNAS2 resulted in a rise in Fe content (3.0-fold) in mature seeds. Its ectopic expression also increased that content. Enhanced expression led to higher tolerance of Fe deficiency and better growth under elevated pH. Mice fed with OsNAS2-D1 seeds recovered more rapidly from anemia, indicating that bioavailable Fe contents were improved by this increase in OsNAS2 expression.
Assuntos
Alquil e Aril Transferases/metabolismo , Ferro/metabolismo , Oryza/enzimologia , Proteínas de Plantas/metabolismo , Sementes/metabolismo , Alquil e Aril Transferases/genética , Anemia/sangue , Anemia/dietoterapia , Animais , Clorofila/metabolismo , Indução Enzimática , Feminino , Alimentos Geneticamente Modificados , Expressão Gênica , Deficiências de Ferro , Camundongos , Camundongos Endogâmicos BALB C , Oryza/crescimento & desenvolvimento , Oryza/metabolismo , Proteínas de Plantas/genética , Raízes de Plantas/enzimologia , Brotos de Planta/enzimologia , Brotos de Planta/crescimento & desenvolvimento , Sementes/enzimologiaRESUMO
The heterogeneous cellular composition of the mammalian renal collecting duct enables regulation of fluid, electrolytes, and acid-base homeostasis, but the molecular mechanism of its development has yet to be elucidated. The Notch signaling pathway is involved in cell fate determination and has been implicated in proximal-distal patterning in the mammalian kidney. To investigate the role of Notch signaling in renal collecting duct development, we generated mice in which Mind bomb-1 (Mib1), an E3 ubiquitin ligase required for the initiation of Notch signaling, was specifically inactivated in the ureteric bud of the developing kidney. Mice lacking Mib1 in the renal collecting duct displayed increased urinary production, decreased urinary osmolality, progressive hydronephrosis, sodium wasting, and a severe urinary concentrating defect manifested as nephrogenic diabetes insipidus. Histological analysis revealed a diminished number of principal cells and corresponding increase in the number of intercalated cells. Transgenic overexpression of Notch intracellular domain reversed the altered cellular composition of mutant renal collecting duct, with principal cells occupying the entire region. Our data demonstrate that Notch signaling is required for the development of the mammalian renal collecting duct and principal cell differentiation and indicate that pathway dysregulation may contribute to distal renal tubular disorders.
Assuntos
Diabetes Insípido Nefrogênico/fisiopatologia , Túbulos Renais Coletores/fisiopatologia , Receptores Notch/metabolismo , Transdução de Sinais , Animais , Diferenciação Celular , Expressão Gênica , Hidronefrose/fisiopatologia , Túbulos Renais Coletores/citologia , Túbulos Renais Coletores/embriologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Fenótipo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismoRESUMO
TRPV5 is believed to play an important role in the regulation of urinary calcium excretion. We assessed the effects of hydrochlorothiazide (HCTZ) on the expression of TRPV5, calbindin-D(28K), and several sodium transporters in hypercalciuric rats. Sprague-Dawley rats were divided into 4 groups; control, HCTZ, high salt, and high salt with HCTZ group in experiment 1; control, HCTZ, high calcium (Ca), and high Ca with HCTZ group in experiment 2. To quantitate the expression of TRPV5, calbindin-D(28K), and sodium transporters, western blotting was performed. In both experiments, HCTZ significantly decreased urinary calcium excretion. TRPV5 protein abundance decreased in all hypercalciuric rats, and restored by HCTZ in both high salt with HCTZ and high Ca with HCTZ group. Calbindin-D(28K) protein abundance increased in the high salt and high salt with HCTZ groups, but did not differ among groups in experiment 2. Protein abundance of NHE3 and NKCC2 decreased in all hypercalciuric rats, and were restored by HCTZ in only high Ca-induced hypercalciuric rats. In summary, protein abundance of TRPV5, NHE3, and NKCC2 decreased in all hypercalciuric rats. The hypocalciuric effect of HCTZ is associated with increased protein abundance of TRPV5 in high salt or calcium diet-induced hypercalciuric rats.
Assuntos
Hipercalciúria/terapia , Proteína G de Ligação ao Cálcio S100/biossíntese , Sódio/metabolismo , Canais de Cátion TRPV/biossíntese , Tiazidas/farmacologia , Animais , Transporte Biológico , Calbindina 1 , Calbindinas , Cálcio/urina , Canais de Cálcio/química , Hidroclorotiazida/farmacologia , Masculino , Modelos Biológicos , Ratos , Ratos Sprague-Dawley , Trocador 3 de Sódio-Hidrogênio , Trocadores de Sódio-Hidrogênio/química , Simportadores de Cloreto de Sódio-Potássio/metabolismo , Membro 1 da Família 12 de Carreador de Soluto , Canais de Cátion TRPV/químicaRESUMO
The induction of heme oxygenase-1 (HO-1) ameliorates oxidative stress and inflammatory process, which play important roles in IgA nephropathy. We hypothesized length polymorphism in the promoter region of the HO-1 gene, which is related to the level of gene transcription, is associated with disease severity of IgA nephropathy. The subjects comprised 916 patients with IgA nephropathy and gene data. Renal impairment was defined as an estimated glomerular filtration rate less than 60 mL/min/1.73 m(2) at diagnosis. The short (S: <23), medium (M: 23-28), and long (L: >28) (GT) repeats in the HO-1 gene was determined. The frequencies of S/S, S/M, M/M, S/L, L/M, and L/L genotypes were 7.2%, 6.9%, 3.1%, 30.8%, 22.7%, and 29.4%, respectively. The baseline characteristics were not different. In the S/S genotypic group, the renal impairment rate was 18.2%, which was lower than 32.2% in the group with M/M, L/M, or L/L genotype. The odds ratio of renal impairment in S/S genotype, compared to that in M/M, L/M, or L/L genotype, was 0.216 (95% confidence interval, 0.060-0.774, p=0.019). The HO-1 gene promoter length polymorphism was related to the renal impairment of IgA nephropathy at diagnosis, which is an important risk factor for mortality in IgA nephropathy patients.
Assuntos
Glomerulonefrite por IGA/genética , Glomerulonefrite por IGA/patologia , Heme Oxigenase-1/genética , Adulto , Progressão da Doença , Feminino , Frequência do Gene , Genótipo , Taxa de Filtração Glomerular , Glomerulonefrite por IGA/mortalidade , Humanos , Masculino , Pessoa de Meia-Idade , Razão de Chances , Polimorfismo Genético , Regiões Promotoras Genéticas , Fatores de RiscoRESUMO
We determined the relationship between the progression of immunoglobulin A nephropathy (IgAN) and the A1818T polymorphism in intron 2 of Angiotensin II type 2 receptor (AT2R) gene, which might play protective roles in the pathogenesis of IgAN. Patients with biopsy-proven IgAN were recruited from the registry of the Progressive REnal disease and Medical Informatics and gEnomics Research (PREMIER) which was sponsored by the Korean Society of Nephrology. A1818T polymorphism of AT2R gene was analyzed with PCR-RFLP method and the association with the progression of IgAN, which was defined as over 50% increase in baseline serum creatinine level, was analyzed with survival analysis. Among the 480 patients followed for more than 10 months, the group without T allele had significantly higher rates of progression of IgAN than the group with T allele (11.4% vs. 3.9%, p=0.024), although there were no significant differences in the baseline variables such as initial serum creatinine level, the degree of proteinuria, and blood pressure. In the Cox's proportional hazard model, the hazard ratio of disease progression in the patients with T allele was 0.221 (95% confidence interval for Exp(B): 0.052-0.940, p=0.041) compared to that of without T allele. In conclusion, A1818T polymorphism of AT2R gene was associated with the progression of IgAN.
Assuntos
Glomerulonefrite por IGA/genética , Polimorfismo Genético , Polimorfismo de Nucleotídeo Único , Receptor Tipo 2 de Angiotensina/genética , Alelos , Creatinina/sangue , Progressão da Doença , Genótipo , Glomerulonefrite por IGA/etnologia , Humanos , Coreia (Geográfico) , Modelos Genéticos , Modelos Estatísticos , Polimorfismo de Fragmento de Restrição , Fatores de Tempo , Resultado do TratamentoRESUMO
Plasma calcium concentration is maintained within a narrow range (8.5-10.5 mg/dL) by the coordinated action of parathyroid hormone (PTH), 1,25(OH)2D3, calcitonin, and ionized calcium (iCa(2+)) itself. The kidney plays a key role in this process by the fine regulation of calcium excretion. More than 95% of filtered calcium is reabsorbed along the renal tubules. In the proximal tubules, 60% of filtered calcium is reabsorbed by passive mechanisms. In the thick ascending limb, 15% of calcium is reabsorbed by paracellular diffusion through paracellin-1 (claudin-16). The calcium sensing receptor (CaSR) in the basolateral membrane of the thick ascending limb senses the change in iCa(2+) and inhibits calcium reabsorption independent to PTH and 1,25(OH)2D3. The fine regulation of calcium excretion occurs in the distal convoluted tubules and connecting tubules despite the fact that only 10-15% of filtered calcium is reabsorbed there. Transient receptor potential vanilloid 5 (TRPV5) and 6 (TRPV6) in the apical membrane act as the main portal of entry, calbindin-D28K delivers Ca(2+) in the cytoplasm, and then Na(2+)/Ca(2+) exchanger (NCX1) and plasma membrane Ca(2+)-ATPase in the basolateral membrane serve as an exit. In the cortical collecting duct, TRPV6 is expressed, but the role might be negligible. In addition to PTH and 1,25(OH)2D3, acid-base disturbance, diuretics, and estrogen affect on these calcium channels. Recently, klotho and fibroblast growth factor 23 (FGF23) are suggested as new players in the calcium metabolism. Klotho is exclusively expressed in the kidney and co-localized with TRPV5, NCX1, and calbindin-D28K. Klotho increases calcium reabsorption through trafficking of TRPV5 to the plasma membrane, and also converts FGF receptor to the specific FGF23 receptor. FGF23:klotho complex bound to FGF receptor inhibits 1α-hydroxylase of vitamin D, and contributes to calcium reabsorption and phosphate excretion in the kidney.
RESUMO
Renal tubulo-interstitial inflammation is frequently associated with polyuria and urine concentration defects. This led us to investigate the effects of the major pro-inflammatory nuclear factor kappaB (NF-kappaB) pathway on aquaporin 2 (AQP2) expression by the collecting duct. Using immortalized collecting duct principal cells (mpkCCDcl4), we found that, acting independently of vasopressin, activation of NF-kappaB by lipopolysaccharide (LPS) decreased AQP2 mRNA and protein levels in a time- and dose-dependent manner but did not decrease AQP2 mRNA stability. Consistently, constitutively active IkappaB kinase beta decreased AQP2 expression. The LPS-induced decrease in AQP2 mRNA levels was confirmed in rat kidney slices and was reproduced both under conditions of elevated cAMP concentration and V(2) receptor antagonism. Computer analysis of the AQP2 promoter revealed two putative kappaB elements. Mutation of either kappaB element abolished the LPS-induced decrease of luciferase activity in cells expressing AQP2 promoter-luciferase plasmid constructs. Chromatin immunoprecipitation revealed that LPS challenge decreased p65, increased p50 and p52, and had no effect on RelB and c-Rel binding to kappaB elements of the AQP2 promoter. RNA-mediated interference silencing of p65, p50, and p52 confirmed controlled AQP2 transcription by these NF-kappaB subunits. We additionally found that hypertonicity activated NF-kappaB in mpkCCDcl4 cells, an effect that may counteract the Tonicity-responsive enhancer binding protein (TonEBP)-dependent increase in AQP2 gene transcription. Taken together, these findings indicate that NF-kappaB is an important physiological regulator of AQP2 transcription.
Assuntos
Aquaporina 2/biossíntese , Regulação da Expressão Gênica , Túbulos Renais Coletores/metabolismo , NF-kappa B/metabolismo , Transcrição Gênica , Animais , Relação Dose-Resposta a Droga , Inflamação , Rim/metabolismo , Lipopolissacarídeos/metabolismo , Lipopolissacarídeos/farmacologia , Modelos Biológicos , Regiões Promotoras Genéticas , Ratos , Fatores de Tempo , TransfecçãoRESUMO
Tonicity-responsive enhancer binding protein (TonEBP) is a transcriptional activator that is regulated by ambient tonicity. TonEBP protects the renal medulla from the deleterious effects of hyperosmolality and regulates the urinary concentration by stimulating aquaporin-2 and urea transporters. The therapeutic use of cyclosporin A (CsA) is limited by nephrotoxicity that is manifested by reduced GFR, fibrosis, and tubular defects, including reduced urinary concentration. It was reported recently that long-term CsA treatment was associated with decreased renal expression of TonEBP target genes, including aquaporin-2, urea transporter, and aldose reductase. This study tested the hypothesis that long-term CsA treatment reduces the salinity/tonicity of the renal medullary interstitium as a result of inhibition of active sodium transporters, leading to downregulation of TonEBP. CsA treatment for 7 d did not affect TonEBP or renal function. Whereas expression of sodium transporters was altered, the medullary tonicity seemed unchanged. Conversely, 28 d of CsA treatment led to downregulation of TonEBP and overt nephrotoxicity. The downregulation of TonEBP involved reduced expression, cytoplasmic shift, and reduced transcription of its target genes. This was associated with reduced expression of active sodium transporters-sodium/potassium/chloride transporter type 2 (NKCC2), sodium/chloride transporter, and Na(+),K(+)-ATPase-along with increased sodium excretion and reduced urinary concentration. Infusion of vasopressin restored the expression of NKCC2 in the outer medulla as well as the expression and the activity of TonEBP. It is concluded that the downregulation of TonEBP in the setting of long-term CsA administration is secondary to the reduced tonicity of the renal medullary interstitium.
Assuntos
Ciclosporina/toxicidade , Desamino Arginina Vasopressina/farmacologia , Diurese/efeitos dos fármacos , Rim/fisiologia , Sódio/metabolismo , Simportadores/metabolismo , Animais , Transporte Biológico Ativo , Homeostase , Rim/efeitos dos fármacos , Rim/patologia , Medula Renal/efeitos dos fármacos , Medula Renal/patologia , Masculino , Modelos Animais , Ratos , Ratos Sprague-DawleyRESUMO
Hypokalemia causes a significant decrease in the tonicity of the renal medullary interstitium in association with reduced expression of sodium transporters in the distal tubule. We asked whether hypokalemia caused downregulation of the tonicity-responsive enhancer binding protein (TonEBP) transcriptional activator in the renal medulla due to the reduced tonicity. We found that the abundance of TonEBP decreased significantly in the outer and inner medullas of hypokalemic rats. Underlying mechanisms appeared different in the two regions because the abundance of TonEBP mRNA was lower in the outer medulla but unchanged in the inner medulla. Immunohistochemical examination of TonEBP revealed cell type-specific differences. TonEBP expression decreased dramatically in the outer and inner medullary collecting ducts, thick ascending limbs, and interstitial cells. In the descending and ascending thin limbs, TonEBP abundance decreased modestly. In the outer medulla, TonEBP shifted to the cytoplasm in the descending thin limbs. As expected, transcription of aldose reductase, a target of TonEBP, was decreased since the abundance of mRNA and protein was reduced. Downregulation of TonEBP appeared to have also contributed to reduced expression of aquaporin-2 and UT-A urea transporters in the renal medulla. In cultured cells, expression and activity of TonEBP were not affected by reduced potassium concentrations in the medium. These data support the view that medullary tonicity regulates expression and nuclear distribution of TonEBP in the renal medulla in cell type-specific manners.
Assuntos
Hipopotassemia/metabolismo , Rim/metabolismo , Fatores de Transcrição NFATC/biossíntese , Animais , Aquaporina 2/biossíntese , Aquaporina 2/genética , Western Blotting , Linhagem Celular , Células Cultivadas , Cães , Regulação para Baixo , Proteínas de Choque Térmico HSP70/biossíntese , Hipopotassemia/enzimologia , Rim/enzimologia , Medula Renal/metabolismo , Masculino , Proteínas de Membrana Transportadoras/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização/metabolismo , Potássio na Dieta/farmacologia , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Ratos , Ratos Sprague-Dawley , Simportadores de Cloreto de Sódio-Potássio/metabolismo , Membro 1 da Família 12 de Carreador de Soluto , Transportadores de UreiaRESUMO
Furosemide administration stimulates distal acidification. This has been attributed to the increased lumen-negative voltage in the distal nephron, but the aspect of regulatory mechanisms of H(+)-ATPase has not been clear. The purpose of this study is to investigate whether chronic administration of diuretics alters the expression of H(+)-ATPase and whether electrogenic Na(+) reabsorption is involved in this process. A 7-day infusion of furosemide or hydrochlorothiazide (HCTZ) lowered urine pH significantly. However, this effect of furosemide-induced distal acidification was not changed with amiloride-blocking electrogenic Na(+) reabsorption. On immunoblotting, a polyclonal antibody against the H(+)-ATPase B1 subunit recognized a specific approximately 56-kDa band in membrane fractions from the kidney. The protein abundance of H(+)-ATPase was significantly increased by furosemide and HCTZ infusion in both the cortex and outer medulla. Furosemide plus amiloride administration also increased the H(+)-ATPase protein abundance significantly. However, no definite subcellular redistribution of H(+)-ATPase was observed by furosemide +/- amiloride infusion with immunohistochemistry. Chronic furosemide +/- amiloride administration induced a translocation of pendrin to the apical membrane, while total protein abundance was not increased. The mRNA expression of H(+)-ATPase was not altered by furosemide +/- amiloride infusion. We conclude that chronic administration of diuretics enhances distal acidification by increasing the abundance of H(+)-ATPase irrespective of electrogenic Na(+) reabsorption. This upregulation of H(+)-ATPase in the intercalated cells may be the result of tubular hypertrophy by diuretics.
Assuntos
Diuréticos/farmacologia , Furosemida/farmacologia , Hidroclorotiazida/farmacologia , Rim/enzimologia , ATPases Translocadoras de Prótons/biossíntese , Animais , Western Blotting , Antiportadores de Cloreto-Bicarbonato/metabolismo , Eletroforese em Gel de Poliacrilamida , Imuno-Histoquímica , Rim/efeitos dos fármacos , Rim/patologia , Masculino , ATPases Translocadoras de Prótons/imunologia , RNA/biossíntese , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sódio/metabolismo , Transportadores de SulfatoRESUMO
Hemodynamic factors play an important role in the development and/or progression of diabetic nephropathy. We hypothesized that renal sodium transporter dysregulation might contribute to the hemodynamic alterations in diabetic nephropathy. Otsuka Long Evans Tokushima Fatty (OLETF) rats were used as an animal model for type 2 diabetes. Long Evans Tokushima (LETO) rats were used as controls. Renal sodium transporter regulation was investigated by semiquantitative immunoblotting and immunohistochemistry of the kidneys of 40-week-old animals. The mean serum glucose level in OLETF rats was increased to 235+/-25 mg/dL at 25 weeks, and the hyperglycemia continued up to the end of 40 weeks. Urine protein/ creatinine ratios were 10 times higher in OLETF rats than in LETO rats. At 40th week, the abundance of the epithelial sodium channel (ENaC) beta-subunit was increased in OLETF rats, but the abundance of the ENaC gamma-subunit was decreased. No significant differences were observed in the ENaC alpha-subunit or other major sodium transporters. Immunohistochemistry for the ENaC beta-subunit showed increased immunoreactivity in OLETF rats, whereas the ENaC gamma-subunit showed reduced immunoreactivity in these rats. In OLETF rats, ENaC beta-subunit upregulation and ENaC gamma-subunit downregulation after the development of diabetic nephropathy may reflect an abnormal sodium balance.