RESUMEN
We reported that aspirin (ASA) abnormally prolongs bleeding time (BT) in uremia. The present study was designed to investigate whether the abnormally prolonged post-ASA BT in uremia is due to different ASA pharmacokinetics and bioavailability that might be a consequence of uremic condition, platelet cyclooxygenase is peculiarly sensitive to ASA in uremia, and ASA affects primary hemostasis in uremia by a mechanism independent of cyclooxygenase inhibition. Our results showed that in patients with uremia, but not in normal subjects, ASA markedly prolongs the BT. This effect is transient and depends on the presence of ASA in the blood. The observed differences in ASA kinetic parameters are not an explanation of the exaggerated effect of ASA on primary hemostasis in uremia. The sensitivity of platelet cyclooxygenase to ASA inhibition is comparable in uremics and in normal subjects. The temporal dissociation between ASA-induced prolongation of BT and the effect on platelet thromboxane A2 generation suggests that ASA inhibits platelet function in uremia by a mechanism distinct from cyclooxygenase blocking. This possibility is strengthened by the observation that ibuprofen at a dose that fully inhibits platelet cyclooxygenase activity does not significantly prolong BT.
Asunto(s)
Aspirina/farmacología , Tiempo de Sangría , Plaquetas/enzimología , Inhibidores de la Ciclooxigenasa , Pruebas de Función Plaquetaria , Uremia/sangre , Acetilación , Adulto , Anciano , Aspirina/metabolismo , Femenino , Trastornos Hemorrágicos/etiología , Humanos , Ibuprofeno/farmacología , Cinética , Masculino , Persona de Mediana Edad , Tromboxano A2/biosíntesis , Uremia/complicacionesRESUMEN
Good evidence exists that genetic predisposition is a major determinant in the development of renal and cardiovascular complications of diabetes. In particular, the role of familial predisposition is well established in diabetic nephropathy which may cluster within families, both in type I (IDDM) and in type II (NIDDM) diabetes. The genes responsible for predisposition to renal and cardiovascular complications are not known, but those of the renin-angiotensin system (RAS) are plausible candidates. Beside the large number of studies aimed at evaluating the role of polymorphisms in these genes, particularly in angiotensin-converting enzyme (ACE) gene, in development of renal disease, no clear-cut evidence has been provided until now. Furthermore, a number of trials have shown that ACE-inhibitors (ACEi) may reduce the rate of progression of renal failure. If the RAS genotype were able to foresee the response to ACEi it would provide new strategies for a specific treatment of subjects at higher risk.
Asunto(s)
Diabetes Mellitus Tipo 1/complicaciones , Diabetes Mellitus Tipo 2/complicaciones , Nefropatías Diabéticas/genética , Peptidil-Dipeptidasa A/genética , Polimorfismo Genético , Sistema Renina-Angiotensina/genética , Angiotensinógeno/genética , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 2/genética , Humanos , Receptores de Angiotensina/genéticaRESUMEN
Cyclosporine A (CsA) is a recently introduced immunosuppressive agent that represents a significant advance in the clinical control of graft rejection. However despite the remarkable effectiveness, one of the limiting factors to a more extensive use of CsA appears to be its nephrotoxicity. CsA is reported to induce a tubular epithelial damage whereas histological examination of glomeruli does not reveal abnormalities unless particularly high doses of the drug are used. On the other hand a decrease in glomerular filtration rate (GFR) has been reported in association with acute administration of CsA. Relatively few studies are available to compare structural and functional abnormalities in the same animals after an acute administration of CsA. Here we evaluated morphological abnormalities and renal function in the same animals treated for 20 days with CsA. Moreover in the attempt to find a link between tubular damage and decrease in GFR we studied the effect of CsA on the renin-angiotensin system as well as on glomerular synthesis of prostacyclin (PGI2) and prostaglandin E2 (PGE2). Our results confirmed that CsA induces a focal damage of tubular epithelial cells with isometric vacuolization of cytoplasm. No glomerular or vascular changes have been detected at histological examination. Serum creatinine was significantly elevated after 20 days of treatment, whereas creatinine clearance showed a progressive tendency to decrease without reaching a statistical significance. Plasma renin activity was found to progressively increase during CsA administration, whereas the synthesis of PGI2 and PGE2 by isolated glomeruli was not modified in CsA-treated animals in respect to animals receiving the solvent alone.(ABSTRACT TRUNCATED AT 250 WORDS)
Asunto(s)
Ciclosporinas/toxicidad , Glomérulos Renales/metabolismo , Riñón/efectos de los fármacos , Prostaglandinas/metabolismo , Sistema Renina-Angiotensina/efectos de los fármacos , 6-Cetoprostaglandina F1 alfa/biosíntesis , Animales , Creatinina/sangre , Dinoprostona , Glomérulos Renales/patología , Túbulos Renales/patología , Masculino , Prostaglandinas E/biosíntesis , Ratas , Ratas Endogámicas , Renina/sangre , Factores de TiempoRESUMEN
Recent in vitro and in vivo data suggest that endothelin, a potent vasoconstrictor peptide originally isolated from cultured porcine aortic endothelial cells, may be one of the factors accounting for progressive glomerulosclerosis in experimental and human glomerulopathies. Endothelin is expressed and produced by glomerular endothelial and mesangial cells in culture, has a mitogenic effect on mesangial cells and stimulates mesangial synthesis of extracellular matrix components such as type I, III and IV collagen and laminin. In rats with renal mass ablation, a model of chronic renal disease characterized by systemic hypertension, proteinuria and progressive glomerulosclerosis, a significant increase in urinary excretion of endothelin was documented 45 days after surgery as compared with basal values. Experiments involving the infusion of labeled endothelin into renal arteries of rats with renal mass ablation have suggested that the enhanced urinary excretion rate of endothelin could reflect an increased renal production of the peptide. Recent experiments done with Northern blot analysis have evidenced a 2.5 fold increase in pre-proendothelin transcript in kidney homogenates from rats with remnant kidney at 30 days after surgery. The increase averaged 4-5 fold at 60 and 120 days. In the same animals a significant correlation was found between urinary endothelin excretion and the percent of glomeruli affected by glomerulosclerosis. We conclude that the progression of renal disease after surgical ablation of renal mass is associated with an increased renal endothelin gene expression together with excessive urinary excretion of the corresponding protein. It is speculated that endothelin may mediate glomerular structural abnormalities associated with the progression of renal disease.
Asunto(s)
Modelos Animales de Enfermedad , Endotelinas/fisiología , Fallo Renal Crónico/etiología , Animales , Endotelinas/farmacología , Glomeruloesclerosis Focal y Segmentaria/etiología , Glomeruloesclerosis Focal y Segmentaria/fisiopatología , Riñón/efectos de los fármacos , Riñón/fisiopatología , Fallo Renal Crónico/fisiopatología , Ratas , Receptores de Endotelina/fisiologíaRESUMEN
The bleeding tendency associated with uremia is likely due to a qualitative platelet dysfunction. So far the data available on platelet aggregation are conflicting. Since platelet-activating factor (PAF) plays a role in primary hemostasis, we studied platelet aggregation in response to PAF in 40 patients with chronic uremia on regular hemodialysis and 12 control subjects. Our results showed that in 28 of 40 uremics, platelet aggregation response to PAF was normal, whereas in the remaining 12 it was defective in that no second wave of aggregation was elicited even if the PAF concentrations were increased by a factor of 10,000. This abnormal response was peculiar to PAF and only partially related to factor(s) of plasma origin. The number of platelet PAF receptors and their affinity for the agonist were comparable in controls and "PAF-unresponsive" patients. The defective platelet aggregation in response to PAF was associated with a statistically significant reduction (P less than 0.01) in thromboxane A2 (TxA2) generation in platelet-rich plasma (PRP) challenged with PAF (10 and 100 nmol/L). When PRPs from PAF-unresponsive patients were preincubated with a stable analogue of prostaglandin endoperoxides/TxA2 U-46619, an irreversible platelet aggregation in response to PAF was obtained. Thus in a subpopulation of uremics, platelet aggregation in response to PAF is selectively abnormal as a consequence of a reduced TxA2 generation.
Asunto(s)
Plaquetas/metabolismo , Factor de Activación Plaquetaria/farmacología , Agregación Plaquetaria/efectos de los fármacos , Tromboxano A2/biosíntesis , Uremia/fisiopatología , Adulto , Anciano , Femenino , Humanos , Masculino , Persona de Mediana Edad , Agregación Plaquetaria/fisiología , Unión Proteica , Radioinmunoensayo , Tromboxano B2/biosíntesisRESUMEN
We have studied the effect of human recombinant tumour necrosis factor-alpha (TNF-alpha) on gene expression and production of endothelin-1 in cultured bovine aortic endothelial cells. TNF-alpha (10 and 100 ng ml(-1)) increased in a time dependent manner the preproendothelin-1 mRNA levels in respect to unstimulated endothelial cells. TNF-alpha induced endothelin-1 gene expression was associated with a parallel increase in the release of the corresponding peptide in the culture medium. These findings suggest that the enhanced synthesis and release of endothelin-1 occurring in conditions of increased generation of TNF, may act as a modulatory factor that counteracts the hypotensive effect and the excessive platelet aggregation and adhesion induced by TNF.
RESUMEN
We have recently reported that renal preproendothelin-1 gene is up-regulated in rats with renal mass reduction (RMR) and that time-dependent increase in urinary excretion of the corresponding peptide correlates with renal disease progression. Here we evaluated whether a specific endothelin subtype A (ETA) receptor antagonist, FR139317, reduced signs of disease activity in this model. Two groups of rats were given FR139317 or its vehicle (saline) from day 7 to day 60 after the surgical procedure. Sham-operated animals were the control group. Blood pressure, urinary protein excretion and serum creatinine were evaluated at days 0, 7 (before FR139317 or saline administration), 30, 45 and 60. At sacrifice, histological evaluation of renal tissue was performed. The results showed that ETA receptor blocker reduced the abnormal permeability to proteins, limited glomerular injury and prevented renal function deterioration thus confirming the working hypothesis. These findings suggest that this class of compounds may eventually prove useful in the treatment of human progressive nephropathies.
Asunto(s)
Azepinas/farmacología , Antagonistas de los Receptores de Endotelina , Indoles/farmacología , Enfermedades Renales/patología , Animales , Presión Sanguínea/efectos de los fármacos , Creatinina/sangre , Expresión Génica/efectos de los fármacos , Genes fos , Riñón/efectos de los fármacos , Riñón/patología , Enfermedades Renales/fisiopatología , Masculino , Proteinuria/orina , Ratas , Ratas Sprague-DawleyRESUMEN
Systemic and renal hemodyanmic changes in normal pregnancy have been attributed in part to altered vascular synthesis of vasodilatory prostaglandins. Besides vasodilatory substances, endothelium also generates vasoconstrictors, including endothelin. We evaluated the capacity of placental tissue from normal pregnant women to express endothelin gene and to generate endothelin. Placental tissue expressed a single 2.3 kb preproendothelin messenger ribonucleic acid and produced comparable amounts of endothelin 3, Big endothelin 1, and endothelin 1 and a minor quantity of endothelin 2. To investigate the possible influence of placental endothelin production on plasma levels of the peptide, plasma endothelin concentrations were measured in normal pregnant women at delivery and were found to be numerically higher than those measured in nonpregnant subjects. Urinary excretion of endothelin, taken as a marker of the renal synthesis of the peptide, tended to increase, although not significantly, in the first 14 weeks of pregnancy. This trend continued throughout pregnancy, resulting in a significant increase from the second trimester to delivery.
Asunto(s)
Endotelinas/genética , Expresión Génica , Placenta/metabolismo , Embarazo/metabolismo , Adulto , Endotelinas/fisiología , Endotelinas/orina , Femenino , HumanosRESUMEN
Renal disease progression in the rat is associated with a time-dependent upregulation of renal endothelin-1 (ET-1) gene expression and synthesis. We have previously demonstrated that endothelin A receptor subtype (ETA) blockade in rats with remnant kidney reduced signs of disease activity, suggesting that ET-1 exerts part of its deleterious effects on the kidney through ETA. No data are available so far on the role of ETB receptor in progressive renal injury. We first studied renal ETA and ETB receptor gene expression in rats with remnant kidney on days 7, 30, and 120 after the surgical procedure. While renal expression of ETA was unaffected, ETB receptor gene was significantly upregulated with time in rats with remnant kidney, being 3.5-fold and sixfold higher than shamoperated rats at days 30 and 120. We also evaluated whether bosentan, a nonpeptidic ETA and ETB receptor antagonist, offered better protection against renal disease progression than reported for ETA-selective blockers and whether it improved survival in animals with renal ablation. Two groups of rats with renal mass reduction (n = 11 each) were given bosentan 100 mg/kg/d orally or its vehicle (carboxymethyl cellulose) beginning day 7 after the surgical procedure and were followed until the death of the vehicle-treated animals. Sham-operated animals comprised the control group. Bosentan partially prevented increases in blood pressure and proteinuria, but had a remarkable protective effect on renal function and significantly prolonged animal survival. These data suggest that blocking both renal ETA and ETB receptors might have major implications in the treatment of human progressive nephropathies.
Asunto(s)
Antagonistas de los Receptores de Endotelina , Enfermedades Renales/tratamiento farmacológico , Riñón/efectos de los fármacos , Sulfonamidas/uso terapéutico , Análisis de Varianza , Animales , Northern Blotting , Bosentán , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Evaluación Preclínica de Medicamentos , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Riñón/metabolismo , Enfermedades Renales/metabolismo , Enfermedades Renales/mortalidad , Masculino , Nefrectomía , Ratas , Ratas Sprague-Dawley , Receptor de Endotelina A , Receptor de Endotelina B , Receptores de Endotelina/análisis , Receptores de Endotelina/efectos de los fármacos , Sulfonamidas/farmacologíaRESUMEN
Type I diabetes is associated with the DQ loci of the MHC and to a lesser extent with the T cell antigen receptor (TcR) beta chain genes. The non-obese diabetic (NOD) mouse is an animal model of human diabetes, in which up to 90% of female mice develop overt insulin-dependent diabetes. Genetic studies in the NOD mouse suggest that there are at least three diabetogenic genes; one that maps to the MHC, another that may map to the mouse Thy-I locus, and a third that has still to be identified. We have investigated loci in the vicinity of the human Thy-I locus on chromosome 11q23 and report here the results of restriction fragment length polymorphism (RFLP) analysis of the CD3 epsilon locus of 168 Caucasoid patients with type I diabetes. While no association was found between this locus and type I diabetes, a significant difference in the frequency of the CD3 epsilon 8-kb allele was found between male and female patients (0.268 versus 0.430; P less than 0.0025, Pc = 0.02) and between female patients and healthy female controls (0.430 versus 0.267; P less than 0.015). These results suggest that a gene residing on chromosome 11q23 may confer susceptibility to type I diabetes in women.
Asunto(s)
Cromosomas Humanos Par 11 , Diabetes Mellitus Tipo 1/genética , Receptores de Antígenos de Linfocitos T/genética , Adulto , Alelos , Autoinmunidad/genética , Autoinmunidad/inmunología , Mapeo Cromosómico , Sondas de ADN , Diabetes Mellitus Tipo 1/inmunología , Femenino , Frecuencia de los Genes , Predisposición Genética a la Enfermedad , Genotipo , Antígenos HLA-DR/inmunología , Prueba de Histocompatibilidad , Humanos , Masculino , Hibridación de Ácido Nucleico , Polimorfismo de Longitud del Fragmento de RestricciónRESUMEN
Experimental evidence is available to indicate that intrarenal mechanisms play a role in the impaired salt excretion of nephrotic syndrome by multiple and still incompletely defined mediators. It is documented herein that the gene encoding for cyclophilin-like protein (Cy-LP) is up-regulated in renal medulla from adriamycin (ADR)-treated rats as compared with control animals. In the cortex of rats with ADR nephrosis, no change in Cy-LP as compared with that in controls was found for the entire observation period. By contrast, in the medulla of nephrotic rats, Cy-LP gene expression was significantly higher than in controls. Values of urinary Na excretion were inversely correlated to Cy-LP mRNA expression levels. Because in ADR nephrosis a blunted natriuretic response to ANP has been previously reported, it was investigated whether ANP infusion modulated Cy-LP mRNA in the renal medulla. ADR-treated rats, but not control rats, infused for 1 h with ANP (1 microgram/kg.min) had a significant (P < 0.05) increase in medullary Cy-LP mRNA as compared with nephrotic animals receiving the vehicle alone. These findings might be taken to suggest that renal Cy-LP gene expression is positively modulated in nephrotic syndrome and parallels changes in sodium excretion.
Asunto(s)
Factor Natriurético Atrial/farmacología , Proteínas Portadoras/genética , Regulación de la Expresión Génica , Natriuresis , Síndrome Nefrótico/metabolismo , Animales , Secuencia de Bases , Proteínas Portadoras/biosíntesis , Doxorrubicina/toxicidad , Regulación de la Expresión Génica/efectos de los fármacos , Médula Renal/efectos de los fármacos , Médula Renal/metabolismo , Masculino , Datos de Secuencia Molecular , Natriuresis/efectos de los fármacos , Síndrome Nefrótico/inducido químicamente , Ratas , Ratas Sprague-Dawley , Sodio/orinaRESUMEN
OBJECTIVE: To assess whether increased placental or systemic endothelin synthesis has a pathogenic role in pre-eclampsia (gestational proteinuric hypertension). DESIGN: Prospective observations study. SUBJECTS: 19 women with pre-eclampsia and 10 healthy pregnant women were studies. All were in the last trimester. MAIN OUTCOME MEASURES: Preproendothelin-1 gene expression by Northern blot analysis and generation of endothelin-1 precursor, big-endothelin-1, and endothelin isoforms, namely endothelin-1, 2 and 3, were assessed by specific radio-immunoassays, in placental tissue. Plasma endothelin-1 levels and urinary excretion of big-endothelin-1 and endothelin-1 were measured. RESULTS: Placental preproendothelin-1 gene expression and immunoreactive big-endothelin-1 and endothelin-1, 2 and 3, were comparable in placental tissue from pre-eclamptic and normal pregnant women. Plasma levels of endothelin-1 did not differ between pre-eclamptic and normal pregnancies. In contrast, urinary excretion of endothelin-1, which is likely to reflect the renal synthesis of the peptide, was significantly decreased in pre-eclamptic, as compared with normal pregnant women. This was not due to a decreased renal generation of endothelin-1 precursor, since urinary excretion of big-endothelin-1 did not differ between pre-eclamptic and normal pregnancies. These data suggest an increased renal endothelin-1 breakdown in pre-eclampsia. CONCLUSIONS: Endothelin is unlikely to play a role in the pathogenesis of pre-eclampsia. Instead, an increased renal breakdown may have a role in limiting the negative effects of other vasoactive factors on the renal circulation.
Asunto(s)
Endotelinas/metabolismo , Preeclampsia/metabolismo , Adulto , Endotelinas/genética , Endotelinas/orina , Femenino , Humanos , Riñón/metabolismo , Placenta/metabolismo , Preeclampsia/etiología , Preeclampsia/orina , Embarazo , Estudios Prospectivos , ARN/metabolismoRESUMEN
Changes in glomerular hemodynamics have been observed in animals and humans after a high-protein feeding. It has been postulated that these changes can induce progressive deterioration of renal function favoring loss of glomerular permselectivity properties and subsequent glomerulosclerosis, especially when the renal mass is already reduced surgically or by a disease process. We studied the consequence of long-term protein supplementation on renal function parameters in normal animals and in animals affected by adriamycin nephrosis, a model of renal damage that closely mimics human "minimal change". We also wanted to investigate whether vasodilatory prostaglandins (PGs) generated at the renal level are responsible for the adaptive hemodynamic changes that follow dietary manipulation in normal animals and in animals with experimental nephrosis. The model of glomerular damage we used is characterized by heavy and persistent proteinuria induced in the rat by adriamycin (ADR). Two isocaloric diets were selected containing 20% and 35% protein. High-protein feeding induced a significant increase in glomerular filtration rate in both normal and nephrotic animals. In normal animals the high-protein diet did not modify the urinary excretion of 6-keto-PGF1 alpha, the stable breakdown product of prostacyclin (PGI2), but significantly reduced urinary excretion of prostaglandin E2. In nephrotic rats, the high-protein diet increased urinary excretion of 6-keto-PGF1 alpha, without modifying urinary excretion of prostaglandin E2. Glomerular synthesis of vasodilatory prostaglandins paralleled the urinary excretion pattern. The cyclooxygenase inhibitor indomethacin effectively inhibited urinary excretion of vasodilatory PGs but did not prevent hyperfiltration in normal animals fed the high-protein diet. At variance, when given to nephrotic animals fed the high-protein diet, indomethacin at a dose that reduced 6-keto-PGF1 alpha and prostaglandin E2 urinary excretion by 84% and 93%, respectively, inhibited hyperfiltration. We conclude that the same hemodynamic changes that occur in normal animals given a high-protein diet also take place when glomeruli are uniformly damaged by a disease process as in ADR nephrosis. However, whereas hyperfiltration in normal animals appears to be independent of renal PGs, in nephrotic animals an enhanced renal synthesis of PGI2 appears to play a crucial role in the adaptive changes responsible for hyperfiltration.
Asunto(s)
Proteínas en la Dieta/administración & dosificación , Tasa de Filtración Glomerular , Riñón/metabolismo , Nefrosis Lipoidea/fisiopatología , Prostaglandinas/metabolismo , 6-Cetoprostaglandina F1 alfa/metabolismo , Animales , Dinoprostona , Doxorrubicina , Indometacina/farmacología , Masculino , Nefrosis Lipoidea/inducido químicamente , Nefrosis Lipoidea/metabolismo , Prostaglandinas E/metabolismo , Ratas , Ratas Endogámicas , Circulación RenalRESUMEN
We previously demonstrated that urinary endothelin excretion is increased in rats with extensive renal mass reduction, a model of progressive renal disease. Here we explored whether the increased urinary endothelin in this model were due to induction of renal pre-pro-endothelin-1 gene and whether changes in endothelin synthetic pathway correlated with the development of glomerulosclerosis. Four groups of rats with renal mass reduction and four groups of sham-operated control rats were studied 7, 30, 60 and 120 days after the surgical procedure. Urinary protein excretion in renal mass ablation animals did not differ from controls at seven days, but was already significantly elevated (P < 0.01) 30 days after surgery. Then proteinuria progressively increased in rats with remnant kidney at values above 400 mg/day at day 120. Serum creatinine concentration also progressively increased with time in renal mass ablation rats, unlike sham-operated animals, and values were significantly different (P < 0.01) at each of the points considered. Rats with renal mass reduction, unlike sham-operated animals, developed focal glomerulosclerosis that affected 8% of glomeruli at day 30, and 24% of glomeruli at day 120. Seven days after renal mass reduction renal pre-pro-endothelin-1 (pre-pro ET) mRNA was comparable to that of sham-operated rats, while a 2.5-, 5- and fourfold increase in 2.3 Kb pre-proET-1 transcript was observed at 30, 60 and 120 days, respectively. Urinary excretion of endothelin was significantly elevated (P < 0.01) in rats with renal mass reduction with respect to sham-operated rats, starting from 30 days after surgery and increased further thereafter.(ABSTRACT TRUNCATED AT 250 WORDS)
Asunto(s)
Endotelinas/genética , Glomeruloesclerosis Focal y Segmentaria/genética , Riñón/metabolismo , Animales , Secuencia de Bases , ADN/genética , Modelos Animales de Enfermedad , Endotelinas/orina , Expresión Génica , Glomeruloesclerosis Focal y Segmentaria/etiología , Glomeruloesclerosis Focal y Segmentaria/metabolismo , Masculino , Datos de Secuencia Molecular , Nefrectomía , Proteinuria/etiología , Ratas , Ratas Sprague-DawleyRESUMEN
Rats with extensive renal mass reduction develop hypertension, proteinuria and progressive glomerulosclerosis. Previous studies have demonstrated that these changes are associated with an increased urinary excretion of thromboxane compared with normal rats and that the administration of a thromboxane synthetase inhibitor prevents glomerulosclerosis and progressive renal function deterioration. On this basis it has been speculated that the thromboxane synthetase inhibitor, by inhibiting platelet thromboxane, reduces platelet aggregation and prevents the generation of substances that can influence glomerular functional properties. Because the thromboxane synthetase inhibitor also inhibits thromboxane synthesis by resident glomerular cells and lowers blood pressure in these animals, the question of whether platelet thromboxane is indeed the factor implicated in the development of renal disease after renal ablation remains unanswered. To address this issue the authors administered at different time intervals from the surgical procedure a low-dose of oral aspirin (ASA) to rats with remnant kidney. This approach resulted in selective inhibition of platelet cyclooxygenase leading to an almost complete prevention of platelet thromboxane generation. Low-dose ASA spared renal cyclooxygenase as documented by a lack of significant inhibition of glomerular and urinary 6-keto-PGF1 alpha and did not lower blood pressure. Renal function studies showed that low-dose ASA, despite inhibiting platelet aggregation, had no effect on proteinuria and progressive renal insufficiency irrespectively if administered late (ie, 80 days after surgery) and given daily for all the observation period (ie, 20 days) or earlier in the course of the disease (ie, 40 and 10 days after surgery). Histologic data showed that the degree of glomerulosclerosis and tubulo-interstitial damage was not significantly different in rats with reduction of renal mass alone compared with rats with remnant kidney given low-dose ASA. In conclusion, the present findings indicate that inhibition of platelet aggregation and thromboxane formation does not prevent the progressive glomerulosclerosis that develops in rats with surgical reduction of renal mass. It is suggested that the beneficial results obtained previously in the same model by the use of a thromboxane synthesis inhibitor must be attributed either to an effect on resident glomerular cell thromboxane synthesis or to lowering systemic blood pressure.
Asunto(s)
Aspirina/administración & dosificación , Enfermedades Renales/patología , Riñón/patología , Tromboxano B2/metabolismo , Animales , Aspirina/farmacología , Plaquetas/enzimología , Plaquetas/metabolismo , Riñón/enzimología , Riñón/fisiopatología , Enfermedades Renales/sangre , Enfermedades Renales/etiología , Enfermedades Renales/fisiopatología , Glomérulos Renales/ultraestructura , Masculino , Inhibidores de Agregación Plaquetaria/farmacología , Prostaglandina-Endoperóxido Sintasas/metabolismo , Proteinuria , Ratas , Ratas Endogámicas , Tromboxano B2/antagonistas & inhibidores , Tromboxano B2/fisiologíaRESUMEN
In the present study, we examined the hypothesis that dynamic characteristics of flow modulate the production of vasoactive mediators, namely nitric oxide (NO) and endothelin-1 (ET-1), by human umbilical vein endothelial cells (HUVECs). Cells were exposed for 6 hours in a cone-and-plate apparatus to different types of flow: steady laminar, with shear stresses of 2, 8, and 12 dyne/cm2, pulsatile laminar, with shear stress from 8.2 to 16.6 dyne/cm2 and a frequency of 2 Hz; periodic laminar, with square wave cycles of 15 minutes and shear stress from 2 to 8 dyne/cm2, and turbulent, with shear stress of 8 dyne/cm2 on average. A second culture dish was kept in a normal incubator as a static control for each experiment. Laminar flow induced synthesis of NO by HUVECs that was dependent on shear-stress magnitude. Laminar shear stress at 8 dyne/cm2 also upregulated the level of NO synthase mRNA. As observed with steady laminar flow, pulsatile flow also induced an increase in NO release by endothelial cells. When HUVECs were subjected to step-change increases of laminar shear, a further increase of NO synthesis was observed, compared with steady laminar shear of the same magnitude. Turbulent flow did not upregulate NO synthase mRNA or increase NO release. Both laminar and turbulent shear stress reduced, although not significantly, ET-1 mRNA and ET-1 production compared with the static condition. These results indicate that local blood flow conditions modulate the production of vasoactive substances by endothelial cells. This may affect vascular cell functions such as nonthrombogenicity, regulation of blood flow, and vascular tone.
Asunto(s)
Aminoácido Oxidorreductasas/genética , Arteriosclerosis/etiología , Endotelinas/biosíntesis , Endotelio Vascular/metabolismo , Óxido Nítrico/biosíntesis , Arteriosclerosis/metabolismo , Fenómenos Biomecánicos , Circulación Sanguínea , Northern Blotting , Células Cultivadas , Medios de Cultivo , ADN Complementario/análisis , Endotelio Vascular/citología , Expresión Génica , Humanos , NADPH Deshidrogenasa/genética , Óxido Nítrico/análisis , Óxido Nítrico Sintasa , Reacción en Cadena de la Polimerasa , Venas UmbilicalesRESUMEN
The present study was designed to assess whether human glomerular mesangial cells in culture express preproendothelin gene and whether endothelin gene expression in the mesangium is regulated by factors potentially released by inflammatory cells and platelets infiltrating the glomerular tuft during the course of various types of glomerulonephritis. For this purpose mesangial cells were incubated for 6 hours in the presence of absence of interleukin 1 beta (IL-1 beta), transforming growth factor-beta (TBF-beta), the thromboxane A2 analogue U-46619, and thrombin. Resting mesangial cells expressed a 2.3-kilobase mRNA on blot hybridization analysis with a human cDNA preproendothelin probe, indicating that this type of cells, in addition to glomerular endothelial cells, constitutively expresses endothelin gene. IL-1 beta did not change endothelin mRNA levels in respect to unstimulated mesangial cells. At variance, TGF-beta, U-46619, and thrombin had a marked effect on endothelin mRNA, stimulating a 3- to 8-fold increase over basal levels. Quantification of actin mRNA and analysis of the autoradiographic signals provided validation of the difference in the endothelin mRNA levels. Expression of preproendothelin mRNA in either resting or stimulated mesangial cells was associated with synthesis and release of the corresponding peptide in the cell supernatant as determined by a specific radioimmunoassay for endothelin. Endothelin production from IL-1 beta stimulated mesangial cells was not different from that of unstimulated cells, whereas a significant (p less than 0.01) increase in endothelin production was observed after cell stimulation with TGF-beta, U-46619, and thrombin. The demonstration that mesangial cells constitutively express mRNA for preproendothelin and release endothelin into culture medium, together with the finding that endothelin gene expression and production in mesangial cells are regulated by molecules potentially released at glomerular level during an inflammatory reaction may suggest that endothelin participates in the complex process of glomerular disease progression.
Asunto(s)
Endotelinas/genética , Mesangio Glomerular/metabolismo , Endoperóxidos de Prostaglandinas Sintéticos/farmacología , Precursores de Proteínas/genética , Trombina/farmacología , Tromboxano A2/farmacología , Factores de Crecimiento Transformadores/farmacología , Ácido 15-Hidroxi-11 alfa,9 alfa-(epoximetano)prosta-5,13-dienoico , Autorradiografía , Northern Blotting , Células Cultivadas , Clonación Molecular , ADN/genética , Endotelina-1 , Endotelinas/biosíntesis , Regulación de la Expresión Génica/efectos de los fármacos , Mesangio Glomerular/citología , Mesangio Glomerular/efectos de los fármacos , Humanos , Interleucina-1/farmacología , Precursores de Proteínas/biosíntesis , ARN/aislamiento & purificación , ARN Mensajero/genética , RadioinmunoensayoRESUMEN
Experimental and human proteinuric glomerulopathies are associated with tubulo-interstitial injury that correlates with the decline of renal function even better than glomerular lesions do. Mechanism(s) leading to tubulo-interstitial damage are unknown. It has been proposed that excessive reabsorption of filtered proteins activates renal cells to produce vasoactive and inflammatory molecules including endothelin-1. The aim of the present study was twofold: we first evaluated the cellular origin of excessive renal endothelin-1 production in the renal mass reduction model and then related endothelin-1 distribution to the development of kidney lesions. Four groups of renal mass reduction (n = 15) and four groups of control rats (n = 5) were studied at 7, 14, 21, and 28 days after surgery. Urinary proteins in renal mass reduction rats were comparable with controls at day 7 but became significantly higher thereafter. Renal mass reduction rats first developed tubulo-interstitial changes, which were already evident at day 14 in the majority of them. At 28 days, renal mass reduction rats also developed glomerulosclerosis. A parallel increase of renal endothelin-1 gene expression and synthesis of the corresponding peptide in renal mass reduction rats versus controls was observed from day 14. Nonradioactive in situ hybridization confirmed a pattern of endothelin-1 mRNA consistent with the distribution of lesions. At day 14, endothelin-1 staining was stronger in renal mass reduction than in control kidneys and mainly localized to the cytoplasm of tubular cells, whereas glomeruli were negative. At day 28, endothelin-1 expression further increased in renal mass reduction rats as compared with controls, and the staining was apparent also in glomeruli. Thus, in renal mass reduction, a progressive up-regulation of endothelin-1 occurs during the development of renal injury, that first involves the tubules and, only in a subsequent phase, the glomeruli.
Asunto(s)
Endotelina-1/genética , Expresión Génica/fisiología , Enfermedades Renales/genética , Enfermedades Renales/metabolismo , Animales , Progresión de la Enfermedad , Hibridación in Situ , Riñón/metabolismo , Enfermedades Renales/fisiopatología , Masculino , Nefrectomía/métodos , ARN Mensajero/metabolismo , Ratas , Ratas Sprague-Dawley , Valores de Referencia , Factores de Tiempo , Distribución TisularRESUMEN
There is evidence that aspirin in low doses favorably influences the course of pregnancy-induced hypertension, but the mechanism, although assumed to involve suppression of the production of thromboxane by platelets, has not been established. We performed a randomized study of the effect of the long-term daily administration of 60 mg of aspirin (n = 17) or placebo (n = 16) on platelet thromboxane A2 and vascular prostacyclin in women at risk for pregnancy-induced hypertension. Low doses of aspirin were associated with a longer pregnancy and increased weight of newborns. Serum levels of thromboxane B2, a stable product of thromboxane A2, were almost completely (greater than 90 percent) inhibited by low doses of aspirin. The urinary excretion of immunoreactive thromboxane B2 was significantly reduced without changes in the level of 6-keto-prostaglandin F1 alpha, a product of prostacyclin. Mass spectrometric analysis showed that aspirin reduced the excretion of the 2,3-dinor-thromboxane B2 metabolite--mainly of platelet origin--by 81 percent and of thromboxane B2, probably chiefly of renal origin, by 59 percent. The urinary excretion of 6-keto-prostaglandin F1 alpha and of its metabolite 2,3-dinor-6-keto-prostaglandin F1 alpha was not affected. Low doses of aspirin only partially (63 percent) reduced neonatal serum thromboxane B2. No hemorrhagic complications were observed in the newborns. Thus, in women at risk for pregnancy-induced hypertension, low doses of aspirin selectively suppressed maternal platelet thromboxane B2 while sparing vascular prostacyclin, but only partially suppressed neonatal platelet thromboxane B2, allowing hemostatic competence in the fetus and newborn.
Asunto(s)
Aspirina/administración & dosificación , Plaquetas/metabolismo , Feto/metabolismo , Hipertensión/sangre , Complicaciones Cardiovasculares del Embarazo/sangre , Tromboxanos/biosíntesis , 6-Cetoprostaglandina F1 alfa/sangre , Adulto , Aspirina/efectos adversos , Aspirina/uso terapéutico , Epoprostenol/biosíntesis , Femenino , Humanos , Hipertensión/tratamiento farmacológico , Hipertensión/metabolismo , Embarazo , Complicaciones Cardiovasculares del Embarazo/tratamiento farmacológico , Complicaciones Cardiovasculares del Embarazo/metabolismo , Distribución Aleatoria , Tromboxano A2/biosíntesis , Tromboxano B2/sangreRESUMEN
BACKGROUND: The development of strategies to enhance the survival of transplanted organs and to potentially lower or even discontinue immunosuppressive therapy would represent a significant advancement in post-transplant patient care. METHODS: We studied the effect of pretransplant infusion of donor leukocytes alone or in combination with a short course of cyclosporine on the long-term outcome of a rat model of kidney allograft. RESULTS: A single intravenous infusion of donor peripheral blood leukocytes (100x10(6) cells) from Brown-Norway (BN) rats into major histocompatibility complex (MHC) incompatible Lewis recipients largely failed to prolong kidney allograft viability from the same donor transplanted 60, 40, or 30 days after cell infusion. A short course of cyclosporine (per se, unable to prolong graft survival) was started at the same day of donor leukocyte infusion, but instead was able to prolong the survival of the BN kidney transplant-performed 40 days later-but not of a Wistar Furth (WF) third party, with some animals even developing tolerance. A mixed lymphocyte reaction of host cells from long-term surviving rats to BN stimulator cells was significantly reduced as compared with controls. Donor BN DNA was detected in the peripheral blood of Lewis rats until day 40 after BN leukocyte infusion. Microchimerism persisted (60 to 70 days post-transplant) in most long-term graft recipients. Reducing the time interval between donor leukocyte infusion and subsequent kidney transplant to 10 days still prolonged graft survival. Donor peripheral blood mononuclear cells, but not polymorphonuclear cells, in the leukocyte preparation contributed to prolong kidney allograft survival. CONCLUSIONS: Pretransplant donor leukocyte infusion under the appropriate conditions can tip the immune balance toward improved graft acceptance. This result could be relevant to the achievement of donor-specific tolerance of the graft with the maintenance of an intact response to third-party antigens.