RESUMO
Cognitive impairment (CI) is a complication of chronic kidney disease (CKD) that is frequently observed among patients. The aim of this study was to evaluate the potential crosstalk between changes in cognitive function and the levels of Klotho in the brain cortex in an experimental model of CKD. To induce renal damage, Wistar rats received a diet containing 0.25% adenine for six weeks, while the control group was fed a standard diet. The animals underwent different tests for the assessment of cognitive function. At sacrifice, changes in the parameters of mineral metabolism and the expression of Klotho in the kidney and frontal cortex were evaluated. The animals with CKD exhibited impaired behavior in the cognitive tests in comparison with the rats with normal renal function. At sacrifice, CKD-associated mineral disorder was confirmed by the presence of the expected disturbances in the plasma phosphorus, PTH, and both intact and c-terminal FGF23, along with a reduced abundance of renal Klotho. Interestingly, a marked and significant decrease in Klotho was observed in the cerebral cortex of the animals with renal dysfunction. In sum, the loss in cerebral Klotho observed in experimental CKD may contribute to the cognitive dysfunction frequently observed among patients. Although further studies are required, Klotho might have a relevant role in the development of CKD-associated CI and represent a potential target in the management of this complication.
Assuntos
Córtex Cerebral , Disfunção Cognitiva , Glucuronidase , Proteínas Klotho , Insuficiência Renal Crônica , Animais , Masculino , Ratos , Córtex Cerebral/metabolismo , Disfunção Cognitiva/metabolismo , Disfunção Cognitiva/etiologia , Modelos Animais de Doenças , Fator de Crescimento de Fibroblastos 23/metabolismo , Fatores de Crescimento de Fibroblastos/metabolismo , Glucuronidase/metabolismo , Rim/metabolismo , Proteínas Klotho/metabolismo , Ratos Wistar , Insuficiência Renal Crônica/metabolismoRESUMO
The aim of this work was to compare the potential induction of oxidative stress and the antioxidant enzymatic response after a short-term waterborne exposure to copper (Cu) and arsenic (As) with that of the nanoparticles (NPs) of these elements (Cu-NPs and As-NPs) in fish larvae of the species Dicentrarchus labrax. Larvae were grouped in several tanks and exposed to different concentrations of contaminants (0 to 10 mg/L) for 24 or 96 h under laboratory conditions. Copper and arsenic concentrations were analysed in larval tissues using ICP-MS. A set of oxidative stress biomarkers, including the levels of hydroperoxides (HPs), and superoxide dismutase (SOD) and catalase (CAT) activities were assessed. The trace element concentrations (mg/kg d.w.) in larvae ranged as follows: 3.28-6.67 (Cu at 24 h) and 2.76-3.42 (Cu at 96 h); 3.03-8.31 (Cu-NPs at 24 h) and 2.50-4.86 (Cu-NPs at 96 h); 1.92-3.45 (As at 24 h) and 2.22-4.71 (As at 96 h); and 2.19-8.56 (As-NPs at 24 h) and 1.75-9.90 (As-NPs at 96 h). In Cu tests, the oxidative damage (ROOH levels) was induced from 0.1 mg/L at both exposure times, while for Cu-NPs, this damage was not observed until 1 mg/L, which was paralleled by concomitant increases in SOD activity. The CAT activity was also increased but at lower metal concentrations (0.01 mg/L and 0.1 mg/L for both chemical forms). No oxidative damage was observed for As or As-NPs after 24 h, but it was observed for As after 96 h of treatment with 0.01 mg/L. A decrease in SOD activity was observed for As after 24 h, but it turned out to be increased after 96 h. However, As-NPs did not alter SOD activity. The CAT activity was stimulated only at 96 h by As and at 24 h by As-NPs. Therefore, the two chemical forms of Cu exhibited a higher bioaccumulation and toxicity potential as compared to those of As. Importantly, the association of both Cu and As in NPs reduced the respective trace metal bioaccumulation, resulting also in a reduction in the toxic effects (mortality and biochemical). Furthermore, the assessment of oxidative stress-related biomarkers in seabass larvae appears to be a useful tool for biomonitoring environmental-occurring trace elements.
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BACKGROUND: Metabolic syndrome (MetS) and chronic kidney disease (CKD) are commonly associated with cardiovascular disease (CVD) and in these patients Mg concentration is usually decreased. This study evaluated whether a dietary Mg supplementation might attenuate vascular dysfunction through the modulation of oxidative stress and inflammation in concurrent MetS and CKD. METHODS: A rat model of MetS (Zucker strain) with CKD (5/6 nephrectomy, Nx) was used. Nephrectomized animals were fed a normal 0.1%Mg (MetS+Nx+Mg0.1%) or a supplemented 0.6%Mg (MetS+Nx+Mg0.6%) diet; Sham-operated rats with MetS receiving 0.1%Mg were used as controls. RESULTS: As compared to controls, the MetS+Nx-Mg0.1% group showed a significant increase in oxidative stress and inflammation biomarkers (lipid peroxidation and aortic interleukin-1b and -6 expression) and Endothelin-1 levels, a decrease in nitric oxide and a worsening in uremia and MetS associated pathology as hypertension, and abnormal glucose and lipid profile. Moreover, proteomic evaluation revealed changes mainly related to lipid metabolism and CVD markers. By contrast, in the MetS+Nx+Mg0.6% group, these parameters remained largely similar to controls. CONCLUSION: In concurrent MetS and CKD, dietary Mg supplementation reduced inflammation and oxidative stress and improved vascular function.
RESUMO
Although phosphorus is an essential element for life, it is not found in nature in its native state but rather combined in the form of inorganic phosphates (PO43-), with tightly regulated plasma levels that are associated with deleterious effects and mortality when these are out of bounds. The growing interest in the accumulation of PO43- in human pathophysiology originated in its attributed role in the pathogenesis of secondary hyperparathyroidism (SHPT) in chronic kidney disease. In this article, we review the mechanisms by which this effect was justified and we commemorate the important contribution of a Spanish group led by Dr. M. Rodríguez, just 25 years ago, when they first demonstrated the direct effect of PO43- on the regulation of the synthesis and secretion of parathyroid hormone by maintaining the structural integrity of the parathyroid glands in their original experimental model. In addition to demonstrating the importance of arachidonic acid (AA) and the phospholipase A2-AA pathway as a mediator of parathyroid gland response, these findings were predecessors of the recent description of the important role of PO43- on the activity of the calcium sensor-receptor, and also fueled various lines of research on the importance of PO43- overload not only for the pathophysiology of SHPT but also in its systemic pathogenic role.
Assuntos
Hiperparatireoidismo Secundário , Insuficiência Renal Crônica , Humanos , Glândulas Paratireoides , Fosfatos , Hormônio Paratireóideo , Hiperparatireoidismo Secundário/complicações , Insuficiência Renal Crônica/complicaçõesRESUMO
BACKGROUND: Inflammation is a common feature in chronic kidney disease (CKD) that appears specifically associated with cardiovascular derangements in CKD patients. Observational studies have revealed a link between low Mg levels and inflammation. In this study, we hypothesize that Mg might have a modulatory effect on the inflammation induced under the uraemic milieu. METHODS: In vivo studies were performed in a 5/6 nephrectomized rat model of CKD. Furthermore, a possible direct effect of Mg was addressed through in vitro studies with vascular smooth muscle cells (VSMCs). RESULTS: Uraemic rats fed a normal (0.1%) Mg diet showed a systemic inflammatory response evidenced by the elevation in plasma of the pro-inflammatory cytokines TNF-α, IL-1ß and IL-6, and GPx activity, a marker of oxidative stress. Importantly, an increased expression of these cytokines in the aortic tissue was also observed. In contrast, a dietary Mg supplementation (0.6%) greatly prevented the oxidative stress and the pro-inflammatory response. In vitro, in VSMCs cultured in a pro-inflammatory high phosphate medium, incubation with Mg 1.6 mM inhibited the increase in the production of ROS, the rise in the expression of TNF-α, IL-1ß, IL-6 and IL-8 and the activation of NF-κB signalling that was observed in cells incubated with a normal (0.8 mM) Mg. CONCLUSION: Mg supplementation reduced inflammation associated with CKD, exerting a direct effect on vascular cells. These findings support a possible beneficial effect of Mg supplementation along the clinical management of CKD patients.
Assuntos
Suplementos Nutricionais , Inflamação/prevenção & controle , Magnésio/uso terapêutico , Insuficiência Renal Crônica/tratamento farmacológico , Animais , Células Cultivadas , Citocinas/sangue , Magnésio/administração & dosagem , Masculino , Miócitos de Músculo Liso/efeitos dos fármacos , Estresse Oxidativo , Ratos , Ratos Wistar , Espécies Reativas de Oxigênio , Transdução de SinaisRESUMO
BACKGROUND: Fibroblast growth factor 23 (FGF23) is a major determinant of mineral metabolism derangements and emerges as a possible risk factor underlying the negative cardiovascular outcome in CKD patients. However, its contribution in non-CKD individuals is less clear. This cross-sectional study investigated the associations between FGF23 and mineral metabolism parameters and with carotid atherosclerosis in a population at high cardiovascular risk with preserved renal function. METHODS: We employed 939 subjects with coronary heart disease enrolled in the CORDIOPREV study (mean eGFR=93.0⯱â¯0.7â¯ml/min/1.73 m2 and median FGF23=44.9 (IQR=13.1) pg/ml), in which intima-media thickness of both common carotid arteries (IMT-CC) was measured. RESULTS: Adjusted for anthropometric factors, FGF23 associated positively with creatinine, phosphate, calcium and 25(OH)-vitaminD and negatively with eGFR and calcitriol. In multivariable-adjusted models all of them were independent contributors to FGF23 levels. FGF23 showed a positive relationship with IMT-CC; both the higher third and fourth quartiles associated significantly with IMT-CC (Beta= 0.135 and 0.187, respectively) and after additional adjustment for established cardiovascular risk factors and morbidities FGF23 remained as a significant contributor to IMT-CC. Logistic regression analysis confirmed its predictive ability to differentiate patients at higher atherosclerotic risk defined by an IMT-CC≥0.7â¯mm (OR for FGF23 quartiles 3 and 4â¯vs. 1: 1.860; 95%CI 1.209-2.862 and 2.114; 95%CI 1.339-3.337, respectively). CONCLUSION: Even in the setting of a normally functioning phosphate-FGF23-calcitriol system, FGF23 independently associated with IMT-CC, a surrogate of atherosclerotic vascular dysfunction. This supports the notion of FGF23 as a predictor of cardiovascular risk independent of renal failure.
Assuntos
Doenças das Artérias Carótidas , Nefropatias , Doenças das Artérias Carótidas/diagnóstico por imagem , Doenças das Artérias Carótidas/epidemiologia , Espessura Intima-Media Carotídea , Estudos Transversais , Fator de Crescimento de Fibroblastos 23 , Fatores de Crescimento de Fibroblastos , Humanos , Fatores de RiscoRESUMO
This study aimed to ascertain whether there is an independent association between serum magnesium (Mg) and the Carotid Intima-Media Thickness (IMT-CC), a well-accepted atherosclerotic-biomarker surrogate of cardiovascular disease (CVD), in a population with high cardiovascular risk. Serum Mg and traditional atherosclerotic risk factors were recorded in 939 patients (mean age, 59.6 ± 0.3 years, 83.2% men) with coronary heart disease (CHD) enrolled in the CORDIOPREV trial. Serum Mg strongly associated with IMT-CC. Before adjusting for potential confounding factors, IMT-CC decreased by 0.111 ± 0.011 mm per mg/dl increase in serum Mg (p < 0.001). After adjustment, the effect of Mg did not appear mediated through factors related to glucose metabolism, the lipid profile or the mineral metabolism and renal function. Multivariate models showed the lower Mg levels (quartile 1) as a strong independent factor contributing to IMT-CC along with age, sex, SBP, HDL-C, and diuretic use. Logistic regression analysis confirmed the predictive ability of serum Mg to differentiate patients at higher atherosclerotic risk as defined by an IMT-CC ≥ 1.0 mm, yielding a OR for the lower quartile of 10.623 (95%CI 2.311-48.845; P = 0.002) and a ROC-derived cutoff of 1.61 mg/dl. Therefore, our findings outline low serum magnesium as a possible independent risk factor for carotid atherosclerosis.
Assuntos
Doenças das Artérias Carótidas/sangue , Doença das Coronárias/sangue , Magnésio/sangue , Placa Aterosclerótica/sangue , Idoso , Doenças Cardiovasculares/sangue , Doenças Cardiovasculares/patologia , Artérias Carótidas/patologia , Doenças das Artérias Carótidas/patologia , Espessura Intima-Media Carotídea , Doença das Coronárias/patologia , Feminino , Humanos , Lipídeos/sangue , Masculino , Pessoa de Meia-Idade , Placa Aterosclerótica/patologia , Fatores de RiscoRESUMO
Although magnesium has been shown to prevent vascular calcification in vitro, controlled in vivo studies in uremic animal models are limited. To determine whether dietary magnesium supplementation protects against the development of vascular calcification, 5/6 nephrectomized Wistar rats were fed diets with different magnesium content increasing from 0.1 to 1.1%. In one study we analyzed bone specimens from rats fed 0.1%, 0.3%, and 0.6% magnesium diets, and in another study we evaluated the effect of intraperitoneal magnesium on vascular calcification in 5/6 nephrectomized rats. The effects of magnesium on established vascular calcification were also evaluated in uremic rats fed on diets with either normal (0.1%) or moderately increased magnesium (0.6%) content. The increase in dietary magnesium resulted in a marked reduction in vascular calcification, together with improved mineral metabolism and renal function. Moderately elevated dietary magnesium (0.3%), but not high dietary magnesium (0.6%), improved bone homeostasis as compared to basal dietary magnesium (0.1%). Results of our study also suggested that the protective effect of magnesium on vascular calcification was not limited to its action as an intestinal phosphate binder since magnesium administered intraperitoneally also decreased vascular calcification. Oral magnesium supplementation also reduced blood pressure in uremic rats, and in vitro medium magnesium decreased BMP-2 and p65-NF-κB in TNF-α-treated human umbilical vein endothelial cells. Finally, in uremic rats with established vascular calcification, increasing dietary magnesium from 0.1% magnesium to 0.6% reduced the mortality rate from 52% to 28%, which was associated with reduced vascular calcification. Thus, increasing dietary magnesium reduced both vascular calcification and mortality in uremic rats.
Assuntos
Osso e Ossos/metabolismo , Suplementos Nutricionais , Magnésio/administração & dosagem , Fosfatos/metabolismo , Uremia/complicações , Calcificação Vascular/dietoterapia , Animais , Quelantes/administração & dosagem , Modelos Animais de Doenças , Células Endoteliais da Veia Umbilical Humana , Humanos , Magnésio/sangue , Masculino , Nefrectomia , Ratos , Ratos Wistar , Uremia/sangue , Uremia/dietoterapia , Calcificação Vascular/sangue , Calcificação Vascular/mortalidadeRESUMO
In chronic kidney disease patients, high phosphate (HP) levels are associated with cardiovascular disease, the major cause of morbidity and mortality. Since serum phosphate has been independently correlated with inflammation, the present study aimed to investigate an independent direct effect of HP as a pro-inflammatory factor in VSMCs. A possible modulatory effect of vitamin D (VitD) was also investigated. The study was performed in an in vitro model of human aortic smooth muscle cells (HASMCs). Incubation of cells in an HP (3.3 mM) medium caused an increased expression of the pro-inflammatory mediators intercellular adhesion molecule 1 (ICAM-1), interleukins (ILs) IL-1ß, IL-6, IL-8 and tumour necrosis factor α (TNF-α) (not corroborated at the protein levels for ICAM-1), as well as an increase in reactive oxygen/nitrogen species (ROS/RNS) production. This was accompanied by the activation of nuclear factor κ-light-chain-enhancer of activated B cells (NF-κB) signalling as demonstrated by the increase in the nuclear translocation of nuclear factor κ-light-chain-enhancer of activated B cells protein 65 (p65-NF-κΒ) assessed by Western blotting and confocal microscopy. Since all these events were attenuated by an antioxidant pre-incubation with the radical scavenger Mn(III)tetrakis (4-benzoic acid) porphyrin (MnTBAP), it is suggested that the inflammatory response is upstream mediated by the ROS/RNS-induced activation of NF-κΒ. Addition of paricalcitol (PC) 3·10-8 M to cells in HP prevented the phosphate induced ROS/RNS increase, the activation of NF-κΒ and the cytokine up-regulation. A bimodal effect was observed, however, for different calcitriol (CTR) concentrations, 10-10 and 10-12 M attenuated but 10-8 M stimulated this phosphate induced pro-oxidative and pro-inflammatory response. Therefore, these findings provide novel mechanisms whereby HP may directly favour vascular dysfunctions and new insights into the protective effects exerted by VitD derivatives.
Assuntos
Mediadores da Inflamação/metabolismo , Músculo Liso Vascular/efeitos dos fármacos , Miócitos de Músculo Liso/efeitos dos fármacos , Fosfatos/farmacologia , Aorta/citologia , Aorta/metabolismo , Calcitriol/administração & dosagem , Calcitriol/farmacologia , Núcleo Celular/metabolismo , Células Cultivadas , Citocinas/metabolismo , Relação Dose-Resposta a Droga , Ergocalciferóis/farmacologia , Humanos , Molécula 1 de Adesão Intercelular/metabolismo , Músculo Liso Vascular/citologia , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Espécies Reativas de Nitrogênio/biossíntese , Espécies Reativas de Oxigênio/metabolismo , Fator de Transcrição RelA/metabolismoRESUMO
Cu, Pb, and As, which are among the most abundant metals in the aquatic environment, are also among the most health-threatened by causing diverse cellular injuries. The aim of this study was to assess and compare the potential early induction of genotoxic effects after waterborne Cu, Pb, and As exposure in European seabass, Dicentrarchus labrax, a commercial widely cultured fish, using the micronucleus (MN) assay in peripheral blood erythrocytes. Fish were exposed under laboratory conditions to nominal solutions ranging 0-10 mg/L for 24 and 96 h. Furthermore, actual metal ion concentrations were measured by inductively coupled plasma atomic emission spectroscopy (ICP-AES) or differential pulse anodic stripping voltammetry (DPASV) in water and four fish tissues differentially related to environmental exposition and metal accumulation, i.e. the gills, liver, muscle, and brain. Dose-dependent increases of micronuclei (MNi) frequency were observed after these very short exposures; based on measured metal concentrations in water, the genotoxic effect ordered as Cu > As > Pb. Significant genotoxic effect at 0.009 mg/L Cu, 0.57 mg/L Pb, and 0.01 mg/L As was seen. For Cu and Pb these are only slightly higher, but for As it is notably lower than the USEPA criteria of maximum concentration to prevent acute toxicity in aquatic organisms. Furthermore, genotoxicity was differentially related to metal accumulation. MNi frequency correlated positively with the content of Pb in all the organs, with the content of As in liver and gills and only with the content of Cu in the brain. In conclusion, our findings raised environmental concerns because these depicted a genotoxic potential of Cu, Pb, and As after a very short exposure to low but environmentally relevant concentrations, too close to regulatory thresholds. In addition, the MN test in D. labrax could be considered an early biomarker of genotoxicity induced by these metals in fish.
Assuntos
Arsênio/toxicidade , Bass , Cobre/toxicidade , Chumbo/toxicidade , Mutagênicos/toxicidade , Poluentes Químicos da Água/toxicidade , Animais , Arsênio/análise , Cobre/análise , Dano ao DNA , Testes para Micronúcleos , Espectrofotometria Atômica , Poluentes Químicos da Água/análiseRESUMO
Clinical and epidemiologic studies reveal an association between vitamin D deficiency and increased risk of cardiovascular disease. Because vascular smooth muscle cell (VSMC)-derived tissue factor (TF) is suggested to be critical for arterial thrombosis, we investigated whether the vitamin D molecules calcitriol and paricalcitol could reduce the expression of TF induced by the proinflammatory cytokine TNF-α in human aortic VSMCs. We found that, compared with controls, incubation with TNF-α increased TF expression and procoagulant activity in a NF-κB-dependent manner, as deduced from the increased nuclear translocation of nuclear factor κ-light-chain-enhancer of activated B cells protein 65 (p65-NF-κB) and direct interaction of NF-κB to the TF promoter. This was accompanied by the up-regulation of TF signaling mediator protease-activated receptor 2 (PAR-2) expression and by the down-regulation of vitamin D receptor expression in a miR-346-dependent way. However, addition of calcitriol or paricalcitol blunted the TNF-α-induced TF expression and activity (2.01 ± 0.24 and 1.32 ± 0.14 vs. 3.02 ± 0.39 pmol/mg protein, P < 0.05), which was associated with down-regulation of NF-κB signaling and PAR-2 expression, as well as with restored levels of vitamin D receptor and enhanced expression of TF pathway inhibitor. Our data suggest that inflammation promotes a prothrombotic state through the up-regulation of TF function in VSMCs and that the beneficial cardiovascular effects of vitamin D may be partially due to decreases in TF expression and its activity in VSMCs.
Assuntos
Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Receptor PAR-2/metabolismo , Tromboplastina/metabolismo , Vitamina D/metabolismo , Calcitriol/farmacologia , Células Cultivadas , Regulação para Baixo/efeitos dos fármacos , Ergocalciferóis/farmacologia , Humanos , Inflamação/metabolismo , Músculo Liso Vascular/efeitos dos fármacos , Miócitos de Músculo Liso/efeitos dos fármacos , NF-kappa B/metabolismo , Receptores de Calcitriol/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fator de Necrose Tumoral alfa/metabolismo , Regulação para Cima/efeitos dos fármacosRESUMO
BACKGROUND: Vascular calcification (VC) is highly prevalent in patients with chronic kidney disease (CKD). Low magnesium levels are associated with VC, and recent in vitro studies confirm a protective role of magnesium, which is mediated by its entry into the VSMCs through the Transient Receptor Potential Melastatin 7 (TRPM7) channel. The role of Angiotensin II (Ang II) on VC is still unclear. As Ang II is able to stimulate TRPM7 activity, we hypothesize that it might prevent VC. Thus, the aim of this study was to dissect the direct effect of Ang II on VC. MATERIALS AND METHODS: We worked with a model of high phosphate (HP)-induced calcification in human aortic smooth muscle cells, which resembles the CKD-related VC. RESULTS: Addition of Ang II to cells growing in HP decreased calcification, which was associated with the upregulation of the osteogenic factors BMP2, Runx2/Cbfa1, Osterix and ALP. A reduction of magnesium entry into the HP-calcifying cells was found. The treatment with Ang II avoided this reduction, which was reversed by the cotreatment with the TRPM7-inhibitor 2-APB. The protective effect of Ang II was related to AT1R-induced ERK1/2 MAPKinase activation. HP-induced calcification was also associated with the upregulation of the canonical Wnt/beta-catenin pathway, while its downregulation was related to attenuation of calcification by Ang II. CONCLUSION: As hypothesized, Ang II prevented phosphate-induced calcification in VSMCs, which appears mediated by the increase of magnesium influx and by the activation of the ERK1/2 and the inhibition of the canonical Wnt/beta-catenin signalling pathways.
Assuntos
Angiotensina II/farmacologia , Magnésio/metabolismo , Músculo Liso Vascular/efeitos dos fármacos , Miócitos de Músculo Liso/efeitos dos fármacos , Proteínas Serina-Treonina Quinases/efeitos dos fármacos , Canais de Cátion TRPM/efeitos dos fármacos , Calcificação Vascular/metabolismo , Vasoconstritores/farmacologia , Fosfatase Alcalina/efeitos dos fármacos , Fosfatase Alcalina/metabolismo , Proteína Morfogenética Óssea 2/efeitos dos fármacos , Proteína Morfogenética Óssea 2/metabolismo , Compostos de Boro/farmacologia , Células Cultivadas , Subunidade alfa 1 de Fator de Ligação ao Core/efeitos dos fármacos , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Músculo Liso Vascular/citologia , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/metabolismo , Fator de Transcrição Sp7 , Canais de Cátion TRPM/antagonistas & inibidores , Canais de Cátion TRPM/metabolismo , Fatores de Transcrição/efeitos dos fármacos , Fatores de Transcrição/metabolismo , Regulação para Cima , Via de Sinalização Wnt/efeitos dos fármacosRESUMO
BACKGROUND: Transforming growth factor-ß (TGF-ß) is a key cytokine during differentiation of mesenchymal stem cells (MSC) into vascular smooth muscle cells (VSMC). High phosphate induces a phenotypic transformation of vascular smooth muscle cells (VSMC) into osteogenic-like cells. This study was aimed to evaluate signaling pathways involved during VSMC differentiation of MSC in presence or not of high phosphate. RESULTS: Our results showed that TGF-ß induced nuclear translocation of Smad3 as well as the expression of vascular smooth muscle markers, such as smooth muscle alpha actin, SM22α, myocardin, and smooth muscle-myosin heavy chain. The addition of high phosphate to MSC promoted nuclear translocation of Smad1/5/8 and the activation of canonical Wnt/ß-catenin in addition to an increase in BMP-2 expression, calcium deposition and alkaline phosphatase activity. The administration of TGF-ß to MSC treated with high phosphate abolished all these effects by inhibiting canonical Wnt, BMP and TGF-ß pathways. A similar outcome was observed in high phosphate-treated cells after the inhibition of canonical Wnt signaling with Dkk-1. Conversely, addition of both Wnt/ß-catenin activators CHIR98014 and lithium chloride enhanced the effect of high phosphate on BMP-2, calcium deposition and alkaline phosphatase activity. CONCLUSIONS: Full VSMC differentiation induced by TGF-ß may not be achieved when extracellular phosphate levels are high. Moreover, TGF-ß prevents high phosphate-induced osteogenesis by decreasing the nuclear translocation of Smad 1/5/8 and avoiding the activation of Wnt/ß-catenin pathway.
Assuntos
Proteínas Morfogenéticas Ósseas/metabolismo , Osteogênese/efeitos dos fármacos , Osteogênese/fisiologia , Transdução de Sinais/efeitos dos fármacos , Fator de Crescimento Transformador beta/farmacologia , Proteínas Wnt/metabolismo , beta Catenina/metabolismo , Animais , Proteína Morfogenética Óssea 2/genética , Proteína Morfogenética Óssea 2/metabolismo , Diferenciação Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Perfilação da Expressão Gênica , Imunofenotipagem , Masculino , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/metabolismo , Músculo Liso Vascular/citologia , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/citologia , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/metabolismo , Fosfatos/metabolismo , Transporte Proteico , Ratos , Proteína Smad3/metabolismoRESUMO
BACKGROUND: The interest on magnesium (Mg) has grown since clinical studies have shown the efficacy of Mg-containing phosphate binders. However, some concern has arisen for the potential effect of increased serum Mg on parathyroid hormone (PTH) secretion. Our objective was to evaluate the direct effect of Mg in the regulation of the parathyroid function; specifically, PTH secretion and the expression of parathyroid cell receptors: CaR, the vitamin D receptor (VDR) and FGFR1/Klotho. METHODS: The work was performed in vitro by incubating intact rat parathyroid glands in different calcium (Ca) and Mg concentrations. RESULTS: Increasing Mg concentrations from 0.5 to 2 mM produced a left shift of PTH-Ca curves. With Mg 5 mM, the secretory response was practically abolished. Mg was able to reduce PTH only if parathyroid glands were exposed to moderately low Ca concentrations; with normal-high Ca concentrations, the effect of Mg on PTH inhibition was minor or absent. After 6-h incubation at a Ca concentration of 1.0 mM, the expression of parathyroid CaR, VDR, FGFR1 and Klotho (at mRNA and protein levels) was increased with a Mg concentration of 2.0 when compared with 0.5 mM. CONCLUSIONS: Mg reduces PTH secretion mainly when a moderate low calcium concentration is present; Mg also modulates parathyroid glands function through upregulation of the key cellular receptors CaR, VDR and FGF23/Klotho system.
Assuntos
Cálcio/farmacologia , Compostos de Magnésio/farmacologia , Glândulas Paratireoides/efeitos dos fármacos , Hormônio Paratireóideo/metabolismo , Fosfatos/farmacologia , Receptor Tipo 1 de Hormônio Paratireóideo/metabolismo , Animais , Cálcio/sangue , Células Cultivadas , Modelos Animais de Doenças , Expressão Gênica , Imuno-Histoquímica , Glândulas Paratireoides/metabolismo , RNA Mensageiro/genética , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase em Tempo Real , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/genética , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/metabolismo , Receptor Tipo 1 de Hormônio Paratireóideo/genética , Receptores de Calcitriol/genética , Receptores de Calcitriol/metabolismo , Receptores de Detecção de Cálcio/genética , Receptores de Detecção de Cálcio/metabolismo , Regulação para CimaRESUMO
The present study investigates the differential effect of two vitamin D receptor agonists, calcitriol and paricalcitol, on human aortic smooth muscle cells calcification in vitro. Human vascular smooth muscle cells were incubated in a high phosphate (HP) medium alone or supplemented with either calcitriol 10(-8)M (HP + CTR) or paricalcitol 3·10(-8) M (HP + PC). HP medium induced calcification, which was associated with the upregulation of mRNA expression of osteogenic factors such as bone morphogenetic protein 2 (BMP2), Runx2/Cbfa1, Msx2, and osteocalcin. In these cells, activation of Wnt/ß-catenin signaling was evidenced by the translocation of ß-catenin into the nucleus and the increase in the expression of direct target genes as cyclin D1, axin 2, and VCAN/versican. Addition of calcitriol to HP medium (HP + CTR) further increased calcification and also enhanced the expression of osteogenic factors together with a significant elevation of nuclear ß-catenin levels and the expression of cyclin D1, axin 2, and VCAN. By contrast, the addition of paricalcitol (HP + PC) not only reduced calcification but also downregulated the expression of BMP2 and other osteoblastic phenotype markers as well as the levels of nuclear ß-catenin and the expression of its target genes. The role of Wnt/ß-catenin on phosphate- and calcitriol-induced calcification was further demonstrated by the inhibition of calcification after addition of Dickkopf-related protein 1 (DKK-1), a specific natural antagonist of the Wnt/ß-catenin signaling pathway. In conclusion, the differential effect of calcitriol and paricalcitol on vascular calcification appears to be mediated by a distinct regulation of the BMP and Wnt/ß-catenin signaling pathways.
Assuntos
Ergocalciferóis/farmacologia , Músculo Liso Vascular/efeitos dos fármacos , Miócitos de Músculo Liso/efeitos dos fármacos , Fosfatos/farmacologia , Calcificação Vascular/metabolismo , Proteínas Wnt/metabolismo , beta Catenina/metabolismo , Aorta/citologia , Aorta/efeitos dos fármacos , Aorta/metabolismo , Calcificação Fisiológica/efeitos dos fármacos , Calcitriol/farmacologia , Linhagem Celular , Células Cultivadas , Humanos , Músculo Liso Vascular/citologia , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/citologia , Miócitos de Músculo Liso/metabolismo , Transdução de Sinais/efeitos dos fármacosRESUMO
Fibroblast growth factor (FGF) 23 inhibits calcitriol production, which could exacerbate calcium deficiency or hypocalcemia unless calcium itself modulates FGF23 in this setting. In Wistar rats with normal renal function fed a diet low in both calcium and vitamin D, the resulting hypocalcemia was associated with low FGF23 despite high parathyroid hormone (PTH) and high calcitriol levels. FGF23 correlated positively with calcium and negatively with PTH. Addition of high dietary phosphorus to this diet increased FGF23 except in rats with hypocalcemia despite high PTH levels. In parathyroidectomized rats, an increase in dietary calcium for 10 days increased serum calcium, with an associated increase in FGF23, decrease in calcitriol, and no change in phosphorus. Also in parathyroidectomized rats, FGF23 increased significantly 6 hours after administration of calcium gluconate. Taken together, these results suggest that hypocalcemia reduces the circulating concentrations of FGF23. This decrease in FGF23 could be a response to avoid a subsequent reduction in calcitriol, which could exacerbate hypocalcemia.
Assuntos
Cálcio/deficiência , Cálcio/metabolismo , Fatores de Crescimento de Fibroblastos/sangue , Hipocalcemia/metabolismo , Animais , Calcitriol/metabolismo , Cálcio/farmacologia , Gluconato de Cálcio/farmacologia , Fatores de Crescimento de Fibroblastos/efeitos dos fármacos , Masculino , Modelos Animais , Hormônio Paratireóideo/metabolismo , Paratireoidectomia , Fósforo na Dieta/farmacologia , Ratos , Ratos Wistar , Vitamina D/metabolismoRESUMO
Manganese is a relatively common, yet poorly studied element in freshwater ecosystems, where it can be significantly bioconcentrated. The knowledge about the mechanisms of Mn toxicity on fish health is still limited. The aim of the present study was to assess the potential induction of oxidative stress and the antioxidant response after a 96 h waterborne Mn-exposure (at 0.1 and 1mM) in gill, kidney, liver and brain of goldfish (Carassius auratus). Mn 1mM induced an increase of lipid hydroperoxides, superoxide dismutase (SOD) and glutathione peroxidase (GPx) activities in all tissues with the exception of SOD inhibition in the brain. Particular response of catalase (CAT) was indicated-its inhibition in the liver and kidney, but activation in the gill. Exposure to Mn 0.1mM provoked most prominent changes in the liver and did not change the indexes in brain. These results strongly suggest that Mn exposure caused a generalized oxidative stress in the fish and revealed an organ specific antioxidant response involving a differential modulation of the SOD, CAT and GPx activities.
Assuntos
Carpa Dourada/fisiologia , Manganês/toxicidade , Poluentes Químicos da Água/toxicidade , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Catalase/metabolismo , Brânquias/efeitos dos fármacos , Brânquias/metabolismo , Rim/efeitos dos fármacos , Rim/metabolismo , Peróxidos Lipídicos/metabolismo , Fígado/efeitos dos fármacos , Fígado/metabolismo , Manganês/metabolismo , Estresse Oxidativo , Superóxido Dismutase/metabolismo , Poluentes Químicos da Água/metabolismoRESUMO
BACKGROUND: The involvement of bone marrow hematopoietic stem cells (BMHSC) mobilization during liver regeneration from hepatectomized patients is under debate. The main aim of this study was to investigate the role of BMHSC mobilization after hepatic resection in 33 patients with liver disease. METHODS AND RESULTS: Mobilization of CD34(+) BMHSC after 72 h of surgery was found in peripheral blood of some, but not all, of the hepatectomized patients. These CD34(+) cells co-expressed other stem cells markers. The patients without BMHSC mobilization showed high levels of circulating and liver tissue BMHSC (CD34(+) cells) previous to surgery. Therefore, two types of patients: "mobilizers" and "non-mobilizers" were distinguished based on the values of CD34(+) cells before and after surgery. Changes in cytokines involved in the hepatic regeneration (HGF and TGF-ß), and in BMHSC mobilization process (SCF, SDF-1, IL-12, or MMP-2), were detected in both groups. In addition, a higher activation previous to surgery of the SDF-1/CXCR4 axis in liver tissue was observed in non mobilizers patients compared to mobilizer patients. CONCLUSION: BMHSC mobilization seems to be associated with variations in the levels of cytokines and proteolytic enzymes involved in hepatic regeneration and bone marrow matrix degradation. Hepatectomy may be an insufficient stimulus for BMSHC mobilization. The pre-hepatectomy higher levels CD34(+) cells in peripheral blood and liver, associated to the activation of hepatic SDF-1/CXCR4 axis, suggest a BMHSC mobilization process previous to surgery in non mobilizer patients.
Assuntos
Antígenos CD34/metabolismo , Mobilização de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/metabolismo , Regeneração Hepática/fisiologia , Transdução de Sinais/fisiologia , Adolescente , Adulto , Idoso , Quimiocina CXCL12/sangue , Feminino , Células-Tronco Hematopoéticas/fisiologia , Hepatectomia , Fator de Crescimento de Hepatócito/sangue , Humanos , Interleucina-12/sangue , Antígeno Ki-67/metabolismo , Fígado/metabolismo , Fígado/patologia , Fígado/cirurgia , Masculino , Metaloproteinase 2 da Matriz/sangue , Pessoa de Meia-Idade , Receptores CXCR4/sangue , Fator de Células-Tronco/sangue , Fator de Crescimento Transformador beta/sangueRESUMO
Fibroblast growth factor 23 (FGF23) modulates mineral metabolism by promoting phosphaturia and decreasing the production of 1,25-dihydroxyvitamin D(3). FGF23 decreases parathyroid hormone (PTH) mRNA and secretion, but despite a marked elevation in FGF23 in uremia, PTH production increases. Here, we investigated the effect of FGF23 on parathyroid function in normal and uremic hyperplastic parathyroid glands in rats. In normal parathyroid glands, FGF23 decreased PTH production, increased expression of both the parathyroid calcium-sensing receptor and the vitamin D receptor, and reduced cell proliferation. Furthermore, FGF23 induced phosphorylation of extracellular signal-regulated kinase 1/2, which mediates the action of FGF23. In contrast, in hyperplastic parathyroid glands, FGF23 did not reduce PTH production, did not affect expression of the calcium-sensing receptor or vitamin D receptor, and did not affect cell proliferation. In addition, FGF23 failed to activate the extracellular signal-regulated kinase 1/2-mitogen-activated protein kinase pathway in hyperplastic parathyroid glands. We observed very low expression of the FGF23 receptor 1 and the co-receptor Klotho in uremic hyperplastic parathyroid glands, which may explain the lack of response to FGF23 in this tissue. In conclusion, in hyperparathyroidism secondary to renal failure, the parathyroid cells resist the inhibitory effects of FGF23, perhaps as a result of the low expression of FGF23 receptor 1 and Klotho in this condition.
Assuntos
Fatores de Crescimento de Fibroblastos/farmacologia , Glândulas Paratireoides/metabolismo , Hormônio Paratireóideo/metabolismo , Uremia/metabolismo , Animais , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Glucuronidase/metabolismo , Hiperplasia/metabolismo , Hiperplasia/patologia , Proteínas Klotho , Masculino , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Glândulas Paratireoides/efeitos dos fármacos , Glândulas Paratireoides/patologia , Ratos , Ratos Wistar , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/metabolismo , Receptores de Calcitriol/metabolismo , Receptores de Detecção de Cálcio/metabolismo , Técnicas de Cultura de Tecidos , Uremia/patologiaRESUMO
We have previously demonstrated that the activation of rat parathyroid calcium-sensing receptor (CaSR) upregulates VDR expression in vivo (Garfia B, Cañadillas S, Luque F, Siendones E, Quesada M, Almadén Y, Aguilera-Tejero E, Rodríguez M. J Am Soc Nephrol 13: 2945-2952, 2002; Rodriguez ME, Almaden Y, Cañadillas S, Canalejo A, Siendones E, Lopez I, Aguilera-Tejero E, Martin D, Rodriguez M. Am J Physiol Renal Physiol 292: F1390-F1395, 2007). The present study was designed to characterize the signaling system that mediates the stimulation of parathyroid VDR gene expression by extracellular calcium. Experiments were performed in vitro by the incubation of rat parathyroid glands and in vivo with normal and uremic (Nx) rats receiving injections of CaCl(2) or EDTA to obtain hypercalcemic or hypocalcemic clamps. A high calcium concentration increased VDR expression. The addition of arachidonic acid (AA) to the low-calcium medium produced an increase in VDR mRNA of the same magnitude as that observed with high calcium. The addition of ionophore to the low-calcium medium also increased VDR mRNA expression. High calcium or the addition of AA to the low-calcium medium induced the activation (phosphorylation) of ERK1/2-MAPK. The specific inhibition of the ERK1/2-MAPK activity prevented the stimulation of VDR expression by high calcium or AA. These results suggest that AA regulates parathyroid VDR gene expression through the activation of the ERK1/2-MAPK. CaSR activation induced the activation of transcription factor Sp1, but not of NF-κB p50 or p65 or activator protein-1. The addition of AA to the low-calcium medium increased specific DNA-binding activity of Sp1 to almost the same level as high calcium, which was prevented by the inhibition of ERK1/2. Furthermore, mithramycin A (a Sp1 inhibitor) prevented the upregulation of VDR mRNA by high calcium. Finally, both sham and Nx hypercalcemic rats showed similar increased levels of VDR mRNA compared with sham and Nx hypocalcemic rats. Our results demonstrate that extracellular calcium stimulates VDR expression in parathyroid glands through the elevation of the cytosolic calcium level and the stimulation of the PLA(2)-AA-dependent ERK1/2-pathway. Furthermore, the transcription factor Sp1 mediates this effect.