RESUMO
Chronic renal insufficiency (CRI) is characterized by increased endothelin 1 (ET-1) synthesis. We studied rat kidney endothelin receptor A (ETA) and receptor B (ETB) expressions after 12 and 27 weeks of 5/6 nephrectomy, and after 12 weeks of 0.3% adenine diet, representing proteinuric and interstitial inflammation models of CRI, respectively. Uric acid and calcium-phosphate metabolism were modulated after 5/6 nephrectomy, while ETA blocker and calcimimetic were given with adenine. Endothelin receptor mRNA levels were measured using RT-qPCR and protein levels using autoradiography (5/6 nephrectomy) or ELISA (adenine model). Both 12 and 27 weeks after 5/6 nephrectomy, kidney cortex ETA protein was increased by ~60% without changes in ETB protein, and the ETB:ETA ratio was reduced. However, the ETB:ETA mRNA ratio did not change. In the adenine model, kidney ETA protein was reduced by ~70%, while ETB protein was suppressed by ~95%, and the ETB:ETA ratio was reduced by ~85%, both at the protein and mRNA levels. The additional interventions did not influence the observed reductions in the ETB:ETA ratio. To conclude, unfavorable reduction in the ETB:ETA protein ratio was observed in two different models of CRI. Therefore, ETA blockade may be beneficial in a range of diseases that cause impaired kidney function.
Assuntos
Adenina/efeitos adversos , Receptor de Endotelina A/genética , Receptor de Endotelina B/genética , Insuficiência Renal Crônica/genética , Animais , Cálcio/metabolismo , Modelos Animais de Doenças , Regulação da Expressão Gênica/efeitos dos fármacos , Córtex Renal/metabolismo , Masculino , Nefrectomia/efeitos adversos , Fosfatos/metabolismo , Ratos , Receptor de Endotelina A/metabolismo , Receptor de Endotelina B/metabolismo , Insuficiência Renal Crônica/induzido quimicamente , Insuficiência Renal Crônica/metabolismo , Ácido Úrico/metabolismoRESUMO
BACKGROUND: Disturbed calcium-phosphorus metabolism is associated with increased kidney angiotensin-converting enzyme (ACE) in experimental chronic renal insufficiency (CRI). However, information about the effects of phosphate binding and loading on vascular ACE is lacking. METHODS: Fifteen weeks after 5/6 nephrectomy (NX), rats were placed on a phosphate-binding (NX+Ca, 3.0% Ca), phosphate-loading (NX+Pi, 1.5% Pi), or control diet for 12 weeks (NX and sham). RESULTS: Aortic ACE, blood pressure, plasma phosphate, and parathyroid hormone were increased in the NX and NX+Pi groups, but were reduced with phosphate binding. Endothelium-mediated relaxations of isolated mesenteric conduit artery rings to acetylcholine were impaired in the NX and NX+Pi groups, but did not differ from sham in NX+Ca rats. Experiments with nitric oxide (NO) synthase inhibition in vitro suggested that the NO-mediated component of acetylcholine response was lower in the NX and NX+Pi groups, but did not differ from sham in NX+Ca rats. In all NX groups, aortic endothelial NO synthase (eNOS) was reduced, while plasma and urine concentrations of NO metabolites were increased. Aortic nitrated proteins and calcification were increased in the NX and NX+Pi groups when compared with the NX+Ca and sham groups. CONCLUSION: Hypertension in the NX model of CRI was associated with reduced vasorelaxation, decreased eNOS, and increased ACE and nitrated proteins in the aorta. Phosphate binding with calcium carbonate enhanced vasorelaxation via endogenous NO and suppressed elevation of ACE and nitrated proteins, suggesting reduced vascular oxidative stress. Our findings support the view that correction of the calcium-phosphorus balance prevents CRI-induced vascular pathophysiology.
Assuntos
Aorta/enzimologia , Óxido Nítrico Sintase Tipo III/metabolismo , Óxido Nítrico/metabolismo , Peptidil Dipeptidase A/metabolismo , Fosfatos/sangue , Insuficiência Renal Crônica/metabolismo , Acetilcolina/farmacologia , Animais , Doenças da Aorta , Pressão Sanguínea , Calcinose , Carbonato de Cálcio/administração & dosagem , Endotélio/fisiopatologia , Artérias Mesentéricas/fisiologia , Nefrectomia , Óxido Nítrico Sintase Tipo II/metabolismo , Óxido Nítrico Sintase Tipo III/antagonistas & inibidores , Norepinefrina/farmacologia , Estresse Oxidativo , Hormônio Paratireóideo/sangue , Fosfatos/administração & dosagem , Ratos , Vasoconstrição/efeitos dos fármacos , Vasodilatação/efeitos dos fármacosRESUMO
BACKGROUND: Cardiovascular diseases are the most important cause of death in patients with impaired kidney function. Left ventricular hypertrophy (LVH), cardiac interstitial fibrosis and cardiovascular calcifications are characteristic of chronic renal insufficiency (CRI). Periostin is a fibrogenesis- and calcification-related matricellular protein re-expressed in adult tissues undergoing remodelling in response to pathological stimuli. The role of periostin in CRI-induced LVH is unknown. METHODS: Rats were 5/6-nephrectomized (NX), and after 15 weeks of disease progression high-calcium, high-phosphate or paricalcitol treatment was given for 12 weeks. Cardiac tissue and blood samples were taken to study periostin gene expression and to determine factors contributing to its reactivation, respectively. Left ventricular (LV) periostin expression was also examined in response to angiotensin II or arginine(8)-vasopressin (AVP)-induced pressure overload and in spontaneously hypertensive rats. RESULTS: CRI resulted in a 6.5-fold increase in LV periostin messenger RNA (mRNA) levels. Positive extracellular immunostaining for periostin was detected in areas of infiltrated inflammatory cells and fibrotic lesions. There was a significant correlation between LV periostin mRNA levels and plasma biomarkers of impaired kidney function, LVH, fibrogenesis-related proteins osteopontin and osteoactivin, and anti-calcific matrix Gla protein. Moreover, LV periostin gene expression in CRI correlated positively with systolic blood pressure (BP) and was activated rapidly in response to angiotensin II or AVP infusions. CONCLUSIONS: Periostin is involved in fibrotic cardiac remodelling in CRI. The re-expression of periostin is localized to the fibrotic and inflammatory lesions and is most likely the consequence of elevated BP.
Assuntos
Biomarcadores/metabolismo , Moléculas de Adesão Celular/metabolismo , Fibrose/etiologia , Fibrose/patologia , Expressão Gênica , Ventrículos do Coração/metabolismo , Insuficiência Renal/patologia , Animais , Western Blotting , Conservadores da Densidade Óssea/farmacologia , Calcinose , Moléculas de Adesão Celular/genética , Modelos Animais de Doenças , Ergocalciferóis/farmacologia , Fibrose/metabolismo , Hipertensão/genética , Hipertensão/metabolismo , Hipertensão/patologia , Técnicas Imunoenzimáticas , Testes de Função Renal , Infarto do Miocárdio/genética , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/patologia , Nefrectomia/efeitos adversos , RNA Mensageiro/genética , Ratos , Ratos Endogâmicos SHR , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase em Tempo Real , Insuficiência Renal/complicações , Insuficiência Renal/metabolismoRESUMO
We evaluated the effects of the bisphosphonate pamidronate on bone histomorphometry, structure and strength in male rats with uninephrectomy or with chronic renal disease induced by 5/6 nephrectomy. In rats with chronic renal disease the plasma urea, phosphate and parathyroid hormone levels were significantly increased compared to rats with a uninephroctomy and none of these parameters was affected by pamidronate treatment. In the femoral midshaft, chronic renal disease reduced cortical bone mineral density and content. No difference was observed in the breaking load of the femoral midshaft. In the distal femur, a high-turnover renal osteodystrophy was found but pamidronate suppressed this bone turnover and increased bone mineral content. Treatment had no effect on chronic disease-induced augmentation of osteoid volume or fibroblast surface. These studies show that in this model of stage 3 renal disease, pamidronate increased mineral content in the femoral midshaft and distal metaphysis primarily by adding bone to endocortical and trabecular surfaces but did not reduce osteitis fibrosa.
Assuntos
Distúrbio Mineral e Ósseo na Doença Renal Crônica/tratamento farmacológico , Difosfonatos/farmacologia , Animais , Desmineralização Patológica Óssea/tratamento farmacológico , Desmineralização Patológica Óssea/etiologia , Densidade Óssea/efeitos dos fármacos , Remodelação Óssea/efeitos dos fármacos , Distúrbio Mineral e Ósseo na Doença Renal Crônica/etiologia , Difosfonatos/uso terapêutico , Fêmur/patologia , Humanos , Falência Renal Crônica/complicações , Falência Renal Crônica/tratamento farmacológico , Masculino , Nefrectomia , Pamidronato , Ratos , Ratos Sprague-DawleyRESUMO
BACKGROUND: Hyperuricemia is associated with renal insufficiency and may predispose to Na retention and hypertension. Whether hyperuricemia plays a causal role in the pathogenesis of cardiovascular disease remains controversial. OBJECTIVE: We examined the effects of hyperuricemia on circulating and renal components of the renin-angiotensin-aldosterone system in experimental renal insufficiency. METHODS: Three weeks after 5/6 nephrectomy or sham-operation, rats were put on 2.0% oxonic acid diet for 9 weeks. Blood pressure was monitored using tail-cuff, and blood, urine, and kidney samples were taken, as appropriate. Kidney angiotensin-converting enzyme, angiotensin-converting enzyme 2 and angiotensin II receptors (AT1R, AT2R) were examined using real-time reverse transcriptase-PCR and autoradiography. RESULTS: Oxonic acid diet increased plasma uric acid by 80-90 micromol/l, while blood pressure was elevated only in hyperuricemic 5/6 nephrectomy rats (18 mmHg). Creatinine clearance was reduced by 60% in both 5/6 nephrectomy groups and by 25% in hyperuricemic Sham rats. The 5/6 nephrectomy group showed over 90% suppression of plasma renin activity, whereas the Sham + oxonic acid diet group showed 1.2 and 1.4-fold, and 5/6 nephrectomy + oxonic acid diet group 2.5 and 2.3-fold increases in plasma renin activity and plasma aldosterone, respectively. Hyperuricemia increased K and decreased Na excretion in Sham and 5/6 nephrectomy rats, leading to a more than 1.6-fold increase in urine K to Na ratio. No changes in kidney angiotensin-converting enzyme, angiotensin-converting enzyme 2, AT1R or AT2R were detected that could explain the hyperuricemia-induced alteration in Na-K balance. CONCLUSION: As oxonic acid diet increased plasma renin activity, plasma aldosterone, and urine K to Na ratio, these changes may play a significant role in the harmful cardiovascular actions of hyperuricemia.
Assuntos
Aldosterona/sangue , Hipertensão Renal/metabolismo , Hiperuricemia/metabolismo , Ácido Oxônico/toxicidade , Insuficiência Renal Crônica/metabolismo , Ração Animal , Animais , Autorradiografia , Pressão Sanguínea , Modelos Animais de Doenças , Hipertensão Renal/induzido quimicamente , Hiperuricemia/induzido quimicamente , Masculino , Nefrectomia , Ratos , Ratos Sprague-Dawley , Receptor Tipo 1 de Angiotensina/genética , Receptor Tipo 1 de Angiotensina/metabolismo , Receptor Tipo 2 de Angiotensina/genética , Receptor Tipo 2 de Angiotensina/metabolismo , Insuficiência Renal Crônica/induzido quimicamente , Sistema Renina-Angiotensina/fisiologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Ácido Úrico/sangueRESUMO
AIM: To examine whether treatment of secondary hyperparathyroidism with paricalcitol provides benefits to arteries in uremic rats. METHODS: 5/6-nephrectomized rats were treated (NX+Pari) or not treated (NX) with paricalcitol (200 ng/kg, thrice weekly) for 12 weeks. Aortic histology and isolated segments of the main and 2nd-order mesenteric arterial branches were studied. RESULTS: Creatinine clearance was reduced by 54-61%, plasma phosphate increased 2.1- to 2.5-fold, and blood pressure by 40 mm Hg in both NX groups. PTH increased 13-fold in NX and 5-fold in NX+Pari rats. Calcification in aortic cross-sections increased from 2.1 to 7.1% after paricalcitol. In the large mesenteric artery, vasoconstriction to noradrenaline was reduced in NX+Pari rats. In the large and small arteries, vasorelaxation to acetylcholine was impaired in NX rats and unaffected by paricalcitol. In the small artery, paricalcitol increased nitric oxide synthase inhibition-resistant relaxation to acetylcholine, and maximal relaxation to levcromakalim. The small arteries of NX rats featured increased wall cross-sectional area, while paricalcitol further increased wall thickness and the wall:lumen ratio. CONCLUSION: Paricalcitol treatment showed both benefits and harmful effects in uremic rats: in the large artery vasoconstriction was reduced but calcification increased, while in the small artery vasorelaxation via potassium channels was moderately improved but hypertrophic remodeling was aggravated.
Assuntos
Ergocalciferóis/uso terapêutico , Insuficiência Renal/tratamento farmacológico , Animais , Artérias/efeitos dos fármacos , Artérias/patologia , Artérias/fisiopatologia , Ergocalciferóis/farmacologia , Masculino , Ratos , Ratos Sprague-Dawley , Insuficiência Renal/fisiopatologiaRESUMO
Purpose: Oxygen-induced retinopathy (OIR) is the most widely used model for ischemic retinopathies such as retinopathy of prematurity (ROP), proliferative diabetic retinopathy (PDR), and retinal vein occlusion (RVO). The purpose of this study was to perform the most comprehensive characterization of OIR by a recently developed technique, sequential window acquisition of all theoretical mass spectra (SWATH-MS) proteomics. Methods: Control and OIR retina samples collected from various time points were subjected to SWATH-MS and detailed data analysis. Immunohistochemistry from mouse retinas as well as neovascular membranes from human PDR and RVO patients were used for the detection of the localization of the proteins showing altered expression in the retina and to address their relevance to human ischemic retinopathies. Results: We report the most extensive proteomic profiling of OIR to date by quantifying almost 3000 unique proteins and their expression differences between control and OIR retinas. Crystallins were the most prominent proteins induced by hypoxia in the retina, while angiogenesis related proteins such as Filamin A and nonmuscle myosin IIA stand out at the peak of angiogenesis. Majority of the changes in protein expression return to normal at P42, but there is evidence to suggest that proteins involved in neurotransmission remain at reduced level. Conclusions: The results reveal new potential therapeutic targets to address hypoxia-induced pathological angiogenesis taking place in number of retinal diseases. The extensive proteomic profiling combined with pathway analysis also identifies novel molecular networks that could contribute to the pathogenesis of retinal diseases.
Assuntos
Proteínas do Olho/metabolismo , Espectrometria de Massas/métodos , Oxigênio/toxicidade , Proteoma/metabolismo , Proteômica/métodos , Retina/metabolismo , Retinopatia da Prematuridade/metabolismo , Adulto , Animais , Western Blotting , Retinopatia Diabética/metabolismo , Humanos , Imuno-Histoquímica , Camundongos Endogâmicos C57BL , Microscopia Confocal , Pessoa de Meia-Idade , Neovascularização Retiniana/metabolismo , Oclusão da Veia Retiniana/metabolismo , Retinopatia da Prematuridade/induzido quimicamenteRESUMO
UNLABELLED: Paricalcitol is a less hypercalcemic vitamin D analog that has been shown to suppress secondary hyperparathyroidism and to prevent the associated histomorphometric changes in bone. In this study, we show that paricalcitol also ameliorates the renal insufficiency-induced loss of bone mineral and the mechanical competence of bone. INTRODUCTION: Renal bone disease is a common consequence of chronic renal insufficiency and the associated secondary hyperparathyroidism (SH). Paricalcitol [19-nor-1,25(OH)(2)D(2)] has been shown to ameliorate SH and prevent renal failure-induced histomorphometric changes in bone with minimal calcemic and phosphatemic activity. However, information about its efficacy on restoration of bone structural strength is lacking. In this study, we explored the effects of paricalcitol treatment on bone structure and strength in a model of advanced renal disease. MATERIALS AND METHODS: Forty-five 8-week-old rats were randomly assigned to either surgical 5/6 nephrectomy (NTX) or Sham-operation. After a 15-week postoperative disease progression period, the NTX rats were further allocated to uremic control (NTX) and treatment (NTX + paricalcitol) groups, the latter of which received paricalcitol for the subsequent 12 weeks. After 27 weeks, the animals were killed, plasma samples were collected, and both femora were excised for comprehensive analysis of the femoral neck and midshaft (pQCT and biomechanical testing). RESULTS: High mortality that exceeded 30% was observed in both NTX groups. NTX induced over a 13-fold increase in plasma PTH, whereas this increase was only 5-fold after paricalcitol treatment. At the femoral neck, NTX was associated with an 8.1% decrease (p < 0.05) in vBMD and a 16% decrease in breaking load (p < 0.05) compared with the Sham group, whereas paricalcitol treatment completely prevented these changes. At the femoral midshaft, the NTX resulted in a 6.6% decrease in cortical BMD (p < 0.01 versus Sham), and this change was also prevented by paricalcitol. CONCLUSIONS: Paricalcitol administration prevented renal insufficiency-associated decreases in BMD in the femoral neck and the femoral midshaft and restored bone strength in the femoral neck. Therefore, paricalcitol can efficiently ameliorate renal insufficiency-induced loss of bone mineral and mechanical competence of bone.
Assuntos
Doenças Ósseas/tratamento farmacológico , Ergocalciferóis/uso terapêutico , Nefropatias/tratamento farmacológico , Animais , Aorta/patologia , Fenômenos Biomecânicos , Densidade Óssea , Doenças Ósseas/complicações , Doenças Ósseas/diagnóstico por imagem , Doenças Ósseas/patologia , Calcinose , Rim/patologia , Nefropatias/complicações , Nefropatias/diagnóstico por imagem , Nefropatias/patologia , Ratos , Tomografia Computadorizada por Raios XRESUMO
Chronic renal insufficiency (CRI) results in phosphate retention and secondary hyperparathyroidism, the treatment of which is largely based on the use of calcium salts as phosphate binders. Advanced CRI causes bone fragility, but information about bone geometry and strength in moderate CRI is scarce. We assigned 39 8-week-old male Sprague-Dawley rats to sham-operation (Sham) or 5/6 nephrectomy (NTX). Four weeks later, the rats were randomized to 0.3% calcium (Sham, NTX) or 3.0% calcium diet (Sham + Calcium, NTX + Calcium). After 8 weeks, the animals were sacrificed, plasma samples collected, and femora excised for neck and midshaft analyses: dual-energy X-ray absorptiometry, peripheral quantitative computed tomography, and biomechanical testing. The NTX increased plasma urea and PTH 1.6-fold and 3.6-fold, respectively, whereas high calcium intake suppressed PTH to 30% of controls. Total femoral bone mineral content decreased (-6.3%) in the NTX group, while this deleterious effect was reversed by high calcium diet. In the site-specific analysis of the femoral neck, the volumetric bone density (-6.5%) was decreased in the NTX group but not NTX + Calcium group. However, in the nephrectomized rats, there was also a concomitant increase in the cross-sectional area (+15%), and, despite the decrease in bone density, the mechanical strength of the femoral neck was maintained. In the midshaft, NTX decreased cortical volumetric bone density (-1.2%), but similar to the femoral neck, no differences were found in the mechanical strength. In conclusion, a decrease in bone mass in moderate experimental CRI was associated with a concomitant increase in bone size, and maintenance of mechanical competence. Although high calcium diet suppressed plasma PTH to under normal physiological levels, it prevented the CRI-induced loss of bone mass without an adverse influence on bone strength.
Assuntos
Reabsorção Óssea/etiologia , Osso e Ossos/anatomia & histologia , Cálcio da Dieta/farmacologia , Fêmur/efeitos dos fármacos , Insuficiência Renal Crônica/etiologia , Absorciometria de Fóton , Animais , Fenômenos Biomecânicos , Nitrogênio da Ureia Sanguínea , Reabsorção Óssea/diagnóstico por imagem , Osso e Ossos/diagnóstico por imagem , Cálcio/sangue , Cálcio da Dieta/administração & dosagem , Creatinina/sangue , Fêmur/diagnóstico por imagem , Fêmur/fisiologia , Concentração de Íons de Hidrogênio , Hiperparatireoidismo Secundário/sangue , Hiperparatireoidismo Secundário/etiologia , Hiperparatireoidismo Secundário/fisiopatologia , Nefrectomia , Hormônio Paratireóideo/sangue , Fosfatos/sangue , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Insuficiência Renal Crônica/fisiopatologia , Tomografia Computadorizada por Raios XRESUMO
Essential hypertension is associated with several alterations in arterial function. A wealth of information from animal models is available concerning hypertensive changes in the mesenteric circulation, while only few studies have examined human mesenteric arterial function. The tone of isolated mesenteric arterial segments (outer diameter 0.7-0.9 mm) was examined from individuals with high (n=17) or normal (n=22) blood pressure, grouped using the current definition of elevated blood pressure (140/90 mmHg). Since the majority of them were operated because of malignancies, we evaluated whether functional vascular properties provided information about patient prognosis. Wall tension development (mN/mm) in response to vasoconstrictors (noradrenaline, 5-hydroxy tryptamine, potassium chloride) was higher in mesenteric arterial rings from patients with high than normal blood pressure. There was no difference in vasoconstrictor sensitivity, or endothelium-dependent and endothelium-independent vasorelaxation. Arterial segment weight was higher in hypertensive subjects, suggesting vascular wall hypertrophy. The 10-year follow-up showed no differences in the control of arterial tone between the surviving (n=14) or deceased (n=25) patients. In conclusion, isolated mesenteric arterial segments from hypertensive patients showed increased wall tension in response to vasoconstrictors. Since the mesenteric circulation is an important regulator of peripheral arterial resistance, possible functional alterations in this vascular bed should be further investigated in hypertensive patients.
Assuntos
Hipertensão/fisiopatologia , Artérias Mesentéricas/efeitos dos fármacos , Vasoconstritores/farmacologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Diclofenaco/farmacologia , Feminino , Humanos , Técnicas In Vitro , Masculino , Artérias Mesentéricas/fisiologia , Artérias Mesentéricas/fisiopatologia , Pessoa de Meia-Idade , NG-Nitroarginina Metil Éster/farmacologia , Nifedipino/farmacologia , Norepinefrina/farmacologia , Cloreto de Potássio/farmacologia , Serotonina/farmacologia , Vasoconstrição/efeitos dos fármacosRESUMO
INTRODUCTION: Phosphate binding with sevelamer can ameliorate detrimental histomorphometric changes of bone in chronic renal insufficiency (CRI). Here we explored the effects of sevelamer-HCl treatment on bone strength and structure in experimental CRI. METHODS: Forty-eight 8-week-old rats were assigned to surgical 5/6 nephrectomy (CRI) or renal decapsulation (Sham). After 14 weeks of disease progression, the rats were allocated to untreated and sevelamer-treated (3% in chow) groups for 9 weeks. Then the animals were sacrificed, plasma samples collected, and femora excised for structural analysis (biomechanical testing, quantitative computed tomography). RESULTS: Sevelamer-HCl significantly reduced blood pH, and final creatinine clearance in the CRI groups ranged 30%-50% of that in the Sham group. Final plasma phosphate increased 2.4- to 2.9-fold, and parathyroid hormone 13- to 21-fold in CRI rats, with no difference between sevelamer-treated and untreated animals. In the femoral midshaft, CRI reduced cortical bone mineral density (-3%) and breaking load (-15%) (p<0.05 for all versus Sham), while sevelamer increased bone mineral density (+2%) and prevented the deleterious changes in bone. In the femoral neck, CRI reduced bone mineral density (-11%) and breaking load (-10%), while sevelamer prevented the decrease in bone mineral density (+6%) so that breaking load did not differ from controls. CONCLUSIONS: In this model of stage 3-4 CRI, sevelamer-HCl treatment ameliorated the decreases in femoral midshaft and neck mineral density, and restored bone strength despite prevailing acidosis. Therefore, treatment with sevelamer can efficiently preserve mechanical competence of bone in CRI.
RESUMO
In this study, we investigated the suitability of ultrathin and porous polyimide (PI) membrane as a carrier for subretinal transplantation of human embryonic stem cell (hESC) -derived retinal pigment epithelial (RPE) cells in rabbits. The in vivo effects of hESC-RPE cells were analyzed by subretinal suspension injection into Royal College of Surgeons (RCS) rats. Rat eyes were analyzed with electroretinography (ERG) and histology. After analyzing the surface and permeability properties of PI, subretinal PI membrane transplantations with and without hESC-RPE were performed in rabbits. The rabbits were followed for three months and eyes analyzed with fundus photography, ERG, optical coherence tomography (OCT), and histology. Animals were immunosuppressed with cyclosporine the entire follow-up time. In dystrophic RCS rats, ERG and outer nuclear layer (ONL) thickness showed some rescue after hESC-RPE injection. Cells positive for human antigen were found in clusters under the retina 41 days post-injection but not anymore after 105 days. In rabbits, OCT showed good placement of the PI. However, there was loss of pigmentation on the hESC-RPE-PI over time. In the eyes with PI alone, no obvious signs of inflammation or retinal atrophy were observed. In the presence of hESC-RPE, mononuclear cell infiltration and retinal atrophy were observed around the membranes. The porous ultrathin PI membrane was well-tolerated in the subretinal space and is a promising scaffold for RPE transplantation. However, the rejection of the transplanted cells seems to be a major problem and the given immunosuppression was insufficient for reduction of xenograft induced inflammation.
Assuntos
Células Epiteliais/citologia , Células Epiteliais/transplante , Células-Tronco Embrionárias Humanas/citologia , Epitélio Pigmentado da Retina/citologia , Epitélio Pigmentado da Retina/metabolismo , Alicerces Teciduais , Animais , Linhagem Celular , Modelos Animais de Doenças , Eletrorretinografia , Humanos , Ratos , Degeneração Retiniana/patologia , Degeneração Retiniana/terapia , Tomografia de Coerência Óptica , Resultado do TratamentoRESUMO
The effects of chronic nitric oxide deficiency on prostacyclin and thromboxane A(2) production in vivo are unknown. Therefore, we treated rats with N(G)-nitro-L-arginine methyl ester (L-NAME), and used losartan and high calcium diet as antihypertensive treatments. Forty eight Wistar rats were divided into six groups: control; losartan (20mgkg(-1)day(-1)); high calcium diet (dietary calcium elevated from 1.1% to 3%); L-NAME (20mgkg(-1)day(-1)); losartan+L-NAME and high calcium diet+L-NAME. Prostacyclin and thromboxane A(2) production were measured after eight weeks as urinary 2,3-dinor-6-keto-PGF(1alpha) and 11-dehydro-TXB(2), respectively. Both the high calcium diet and losartan reduced blood pressure in L-NAME hypertension. Chronic nitric oxide deficiency did not modulate prostacyclin production but it nearly doubled thromboxane A(2) production in vivo. This effect was not influenced by lowering of blood pressure by blockade of angiotensin II type 1 receptors. Independent of the level of blood pressure and blockade of nitric oxide synthesis the high calcium diet decreased prostacyclin production by one third and increased thromboxane A(2) production almost two-fold in vivo.
Assuntos
Cálcio da Dieta/metabolismo , Epoprostenol/metabolismo , Hipertensão/metabolismo , Óxido Nítrico/metabolismo , Receptores de Angiotensina/metabolismo , Tromboxano A2/metabolismo , Antagonistas de Receptores de Angiotensina , Animais , Anti-Hipertensivos/metabolismo , Anti-Hipertensivos/uso terapêutico , Inibidores Enzimáticos/metabolismo , Hipertensão/tratamento farmacológico , Losartan/metabolismo , Losartan/uso terapêutico , Masculino , NG-Nitroarginina Metil Éster/metabolismo , Ratos , Ratos WistarRESUMO
BACKGROUND: Angiotensin II type 1 (AT1) receptor antagonists provide end-organ protection and enhance resistance artery relaxation in uremia. The effect of AT1 blockade on conduit artery function in renal failure is unknown. METHODS: The influence of 8-week losartan therapy (20 mg/kg/day) on tone of isolated main branch mesenteric arterial rings was studied in 5/6 nephrectomized (NX) rats. Blood and urine chemistry were examined, and AT1 receptors quantified using autoradiography. RESULTS: NX rats showed decreased creatinine clearance without change in blood pressure. Losartan did not influence these variables, although [125I]-Sar1,Ile8-angiotensin II binding to renal AT1 receptors was significantly prevented. Vasoconstriction to endothelin-1 was reduced by losartan in NX and Sham rats. Vasorelaxation to acetylcholine was attenuated in untreated but not in losartan-treated NX rats, and experiments with Ca2+-activated K+ channel blockers suggested that impaired endothelium-mediated response after NX was due to deficient relaxation via K+ channels. Endothelium-independent relaxation to levcromakalim, adenosine triphosphate-sensitive K+ channel agonist, was impaired in untreated but not in losartan-treated NX rats. CONCLUSION: Losartan reduced conduit artery vasoconstriction to endothelin-1 and augmented vasorelaxation via K+ channels in NX rats, although blood pressure and renal function were unchanged. Therefore, AT1 blockade confers functional benefits to large arteries in renal failure.
Assuntos
Bloqueadores do Receptor Tipo 1 de Angiotensina II , Falência Renal Crônica/fisiopatologia , Losartan/farmacologia , Artérias Mesentéricas/fisiopatologia , Animais , Técnicas de Cultura , Endotélio Vascular/fisiopatologia , Rim/química , Falência Renal Crônica/etiologia , Masculino , Artérias Mesentéricas/efeitos dos fármacos , Nefrectomia , Ratos , Ratos Sprague-Dawley , Receptor Tipo 1 de Angiotensina/análise , Vasoconstrição/efeitos dos fármacos , Vasodilatação/efeitos dos fármacosRESUMO
BACKGROUND: Disturbed calcium-phosphorus balance significantly contributes to uraemic changes in large arteries. We examined the influences of high-calcium and high-phosphate intake on small artery tone in experimental renal insufficiency. METHODS: Sixty-five rats were assigned to 5/6 nephrectomy (NTX) or sham operation. After 15 week disease progression, NTX rats were given high-calcium (3%), high-phosphate (1.5%) or control diet (0.3% calcium, 0.5% phosphate) for 12 weeks. Then isolated segments of small mesenteric arteries were studied using wire and pressure myographs. RESULTS: Subtotal nephrectomy reduced creatinine clearance by 60% and increased parathyroid hormone (PTH) and phosphate 12-fold and 2.7-fold, respectively. High-phosphate intake further elevated PTH and phosphate (33-fold and 5.5-fold, respectively), while the calcium diet suppressed them (to 3.5 and 62% vs sham, respectively). Ventricular B-type natriuretic peptide synthesis was increased, and blood pressure was 27 and 18 mmHg higher in NTX rats on control and phosphate diet, respectively, than in calcium-fed rats. Vasorelaxation to acetylcholine was impaired by approximately 50% in uraemic rats, and was further deteriorated by high-phosphate intake, whereas the calcium diet improved endothelium-mediated relaxation via nitric oxide and potassium channels. Small arteries of all NTX groups featured eutrophic inward remodelling: wall-to-lumen ratio was increased 1.3-fold without change in cross-sectional area. CONCLUSION: High-phosphate intake had a detrimental influence on secondary hyperparathyroidism and vasodilatation, whereas high-calcium intake reduced blood pressure and PTH, alleviated volume overload and improved vasorelaxation in experimental renal insufficiency. Therefore, alterations in the calcium-phosphorus balance can significantly modulate small artery tone during impaired kidney function.
Assuntos
Artérias/efeitos dos fármacos , Cálcio da Dieta/farmacologia , Fósforo na Dieta/farmacologia , Insuficiência Renal/fisiopatologia , Ácido 8,11,14-Eicosatrienoico/análogos & derivados , Ácido 8,11,14-Eicosatrienoico/farmacologia , Acetilcolina/farmacologia , Animais , Artérias/fisiopatologia , Pressão Sanguínea/efeitos dos fármacos , Cálcio/metabolismo , Cálcio da Dieta/administração & dosagem , Creatina/metabolismo , Relação Dose-Resposta a Droga , Endotélio/metabolismo , Hiperparatireoidismo/induzido quimicamente , Hiperparatireoidismo/metabolismo , Técnicas In Vitro , Masculino , Artérias Mesentéricas/efeitos dos fármacos , Artérias Mesentéricas/fisiopatologia , Nitroprussiato/farmacologia , Hormônio Paratireóideo/metabolismo , Fosfatos/metabolismo , Fósforo na Dieta/administração & dosagem , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Insuficiência Renal/metabolismo , Insuficiência Renal/patologia , Fatores de Tempo , Vasodilatação/efeitos dos fármacosRESUMO
The early detection and appropriate treatment of brain ischemia is of paramount importance. The interstitial concentrations of neurotransmitter amino acids are often used as an index of neuronal injury. However, monitoring of non-neurotransmitter amino acids may be equally important. We have studied the behavior of 10 amino acids during K(+)-induced spreading depression (application of 70 mM KCl during 40 min) and global forebrain ischemia (two-vessel occlusion with hypotension during 20 min). The concentrations of glutamate, aspartate, taurine, GABA, glycine, and alanine, measured in the rat striatum by microdialysis, increased during both ischemia and spreading depression, whereas glutamine concentrations decreased in both cases. Only ischemia, but not spreading depression, led to enhanced release of serine, threonine, and asparagine. We thus conclude that an elevation in the interstitial concentrations of non-neurotransmitter amino acids is specific to deep ischemic injury to nervous tissue. We propose the monitoring of serine, asparagine, and threonine, together with excitatory amino acids, as an index of the degree of ischemic brain injury.
Assuntos
Aminoácidos/farmacologia , Isquemia Encefálica/fisiopatologia , Corpo Estriado/fisiologia , Depressão Alastrante da Atividade Elétrica Cortical/fisiologia , Potássio/farmacologia , Prosencéfalo/fisiologia , Animais , Corpo Estriado/efeitos dos fármacos , Corpo Estriado/fisiopatologia , Depressão Alastrante da Atividade Elétrica Cortical/efeitos dos fármacos , Prosencéfalo/fisiopatologia , RatosRESUMO
BACKGROUND: Vasorelaxation is impaired in renal failure (RF) and hypertension. A high calcium diet enhances vasodilatation and reduces blood pressure in experimental hypertension. Oral calcium salts are used as phosphate binders in RF. However, the effect of increased calcium intake on arterial tone in RF is unknown. METHODS: We investigated the influence of an 8-week high calcium diet (0.3 vs 3.0%) on resistance artery tone in 5/6 nephrectomized (NTX) rats. Calcium was supplemented as carbonate salt, blood pressure measured by tail-cuff, urine collected in metabolic cages, and samples taken for blood chemistry and parathyroid hormone (PTH). Functional studies of isolated third-order branches of the mesenteric artery in vitro were performed using the Mulvany multimyograph. RESULTS: Plasma urea was elevated 1.6-fold and systolic blood pressure by 10 mmHg after NTX, while increased calcium intake was without effect on these variables. Plasma PTH and phosphate were raised following NTX, and suppressed by high calcium diet. Vasorelaxations induced by K(+) channel agonists 11,12-epoxyeicosatrienoic acid and levcromakalim were impaired after NTX. Vasorelaxation induced by acetylcholine was also reduced following NTX, and experiments with N(G)-nitro-L-arginine methyl ester, diclofenac and charybdotoxin + apamin suggested that the K(+) channel-mediated component of endothelium-dependent relaxation was deficient after NTX. Increased calcium intake corrected all impairments of vasodilatation in NTX rats. CONCLUSIONS: Deficient vasorelaxation via K(+) channels was normalized by high calcium diet in experimental RF. This effect was independent of the degree of renal impairment and blood pressure, but was associated with improved calcium metabolism: plasma levels of PTH and phosphate were decreased and ionized calcium was increased.
Assuntos
Artérias/efeitos dos fármacos , Carbonato de Cálcio/farmacologia , Cálcio da Dieta/farmacologia , Hiperparatireoidismo/dietoterapia , Insuficiência Renal/fisiopatologia , Vasodilatação/efeitos dos fármacos , Animais , Pressão Sanguínea/efeitos dos fármacos , Pressão Sanguínea/fisiologia , Hiperparatireoidismo/etiologia , Masculino , Nefrectomia , Distúrbios do Metabolismo do Fósforo/dietoterapia , Distúrbios do Metabolismo do Fósforo/etiologia , Canais de Potássio/metabolismo , Ratos , Ratos Sprague-Dawley , Insuficiência Renal/complicaçõesRESUMO
Chronic renal failure (CRF) is associated with abnormal lipid metabolism and high prevalence of vascular complications. Calcium salts are commonly used in CRF as phosphate binders. Increased calcium intake may also lower plasma cholesterol and beneficially influence vascular tone. Therefore, we investigated the influence of increasing dietary calcium from 0.3% to 3.0% for 8 wk after 5/6 nephrectomy (NTX) on plasma cholesterol and mesenteric resistance vessel tone in male Sprague-Dawley rats. The groups were Sham, Sham-Calcium, NTX, and NTX-Calcium (n = 10-11). Blood pressure was modestly elevated after NTX, whereas the plasma creatinine, urea nitrogen, phosphate, and parathyroid hormone levels were clearly increased. The high-calcium diet suppressed plasma phosphate and parathyroid hormone but was without effect on blood pressure. The NTX resulted in 1.6-fold elevation in plasma total cholesterol and 40% reduction in high density-to-low density lipoprotein ratio (HDL/LDL). However, the lipid profile in NTX rats on the high-calcium diet did not differ from sham-operated controls. The endothelium-mediated relaxations induced by acetylcholine were impaired in NTX rats, whereas the response was normalized by a high-calcium diet. No differences in vasorelaxations by the endothelium-independent vasodilator nitroprusside were detected. In conclusion, improved vasorelaxation after a high-calcium diet could be due to reduced plasma total cholesterol and ameliorated HDL/LDL ratio, although decreased plasma phosphate and parathyroid hormone may also play a significant role in the vascular effects of increased calcium intake.
Assuntos
Cálcio/farmacologia , Colesterol/sangue , Falência Renal Crônica/tratamento farmacológico , Falência Renal Crônica/fisiopatologia , Vasodilatação/efeitos dos fármacos , Animais , Aorta Torácica/patologia , Pressão Sanguínea/efeitos dos fármacos , Peso Corporal/efeitos dos fármacos , Ingestão de Líquidos/efeitos dos fármacos , Rim/patologia , Falência Renal Crônica/sangue , Masculino , Miocárdio/patologia , Tamanho do Órgão/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Urina , Resistência Vascular/efeitos dos fármacosRESUMO
Chronic renal failure is associated with increased cardiovascular morbidity and reduced arterial elasticity. Only little information is available on the functional effects of uraemia on resistance arteries. Therefore, we studied the influence of renal failure on rat small mesenteric vessels. The responses of arterial rings were investigated in a Mulvany myograph 6 weeks after 5/6 nephrectomy or sham operation. The subtotal nephrectomy resulted in a 1.9-fold elevation of plasma urea nitrogen but was without significant effect on blood pressure. Endothelium-dependent relaxations, largely mediated via arterial K(+) channels, were preserved in the resistance vessels of uraemic rats. Endothelium-independent vasorelaxations, mediated via exogenous nitric oxide and the opening of ATP-sensitive K(+) channels, were also unchanged. However, the responses induced by isoprenaline were slightly reduced, indicating impaired relaxation via beta-adrenoceptors in experimental renal failure.
Assuntos
Endotélio Vascular/fisiopatologia , Artérias Mesentéricas/fisiopatologia , Receptores Adrenérgicos beta/metabolismo , Insuficiência Renal/fisiopatologia , Resistência Vascular , Vasodilatação/fisiologia , Animais , Técnicas In Vitro , Masculino , Ratos , Ratos Endogâmicos WKYRESUMO
BACKGROUND: Calcium salts are used as phosphate binders in renal failure, while high calcium diet also improves vasorelaxation and enhances natriuresis. The influences of calcium intake on renal renin-angiotensin system (RAS) are largely unknown. METHODS: Four weeks after NTX, rats were put on 3.0% or 0.3% calcium diet for 8 weeks (12-week study). In additional experiments, 15 weeks after NTX, rats were put on similar diets for 12 weeks (27-week study). Appropriate blood, urine, and kidney samples were taken. Renal angiotensin-converting enzyme (ACE) and angiotensin II receptors (AT1, AT2) were examined using autoradiography, ACE also using Western blotting, and connective tissue growth factor (CTGF) using immunohistochemistry. RESULTS: In the 12-week study, albuminuria increased 5-fold in NTX rats, but only 2-fold in calcium NTX rats on 3.0% calcium. In the 27-week study, high calcium intake decreased blood pressure, retarded progression of renal failure, reduced glomerulosclerosis, interstitial damage, and aortic calcifications, and improved survival from 50% to 92% in NTX rats. In both experiments plasma parathyroid hormone and phosphate were elevated after NTX, and suppressed by high calcium diet, while kidney ACE was down-regulated by 40% or more after increased calcium intake. In the 27-week study renal CTGF was decreased and cortical AT1 receptor density reduced after high calcium diet. CONCLUSION: High calcium diet down-regulated kidney ACE, reduced albuminuria and blood pressure, and favorably influenced kidney morphology in experimental renal failure. These findings suggest a link between calcium metabolism and kidney ACE expression, which may play a role in the progression of renal damage.