RESUMEN

The development of new antihypertensive agents is becoming even more important. We need better blood pressure control and also agents that treat hypertension as a disease of the vascular endothelium. Recently, it has been shown that blocking the renin-angiotensin system with angiotensin converting enzyme (ACE) inhibitors reduces blood pressure and decreases the incidence of vascular disease. Another peptide system, the natriuretic peptide system, has also been shown to be important in blood pressure control and volume homeostasis. Because ACE and neutral endopeptidase, the enzyme responsible for the degradation of the natriuretic peptides, are both zinc metalloproteases, new pharmaceuticals that inhibit both enzymes have been developed. The first of these, omapatrilat, has been shown to be an effective antihypertensive agent and to have great potential for treating congestive heart failure.

Asunto(s)

Antihipertensivos/uso terapéutico , Enfermedades Cardiovasculares/tratamiento farmacológico , Inhibidores de Proteasas/uso terapéutico , Piridinas/uso terapéutico , Tiazepinas/uso terapéutico , Inhibidores de la Enzima Convertidora de Angiotensina/uso terapéutico , Animales , Factor Natriurético Atrial/efectos de los fármacos , Enfermedades Cardiovasculares/fisiopatología , Endotelio Vascular/efectos de los fármacos , Humanos , Mesilatos/uso terapéutico , Metaloendopeptidasas/antagonistas & inhibidores , Péptido Natriurético Encefálico/fisiología , Sistema Renina-Angiotensina/fisiología , Tirosina/análogos & derivados , Tirosina/uso terapéutico

RESUMEN

Guanosine 3',5'-cyclic monophosphate (cGMP)-binding, cGMP-specific phosphodiesterase (PDE5) is abundant in vascular smooth muscle, and this enzyme is a potent substrate for cGMP-dependent protein kinase (PKG) in vitro. Binding of cGMP to the allosteric sites of PDE5 is required for this phosphorylation to occur. Vascular smooth muscle cells (VSMC) were used to determine if PDE5 is phosphorylated in intact cells when cGMP is increased. With the use of anti-PDE5 antibodies, a phosphorylated 93-kDa protein band was immunoprecipitated from early passaged primary cultures of VSMC that had been preincubated with 32(Pi) to label cellular ATP and then treated with atrial natriuretic factor (ANF). In the absence of ANF, there was no detectable incorporation of radiolabeled phosphate into this band. Phosphorylation of the 93-kDa protein was augmented by pretreating cells with 8-bromoguanosine 3',5'-cyclic monophosphate (8-BrcGMP) to activate PKG before addition of ANF. 8-BrcGMP, which interacts poorly with the allosteric sites of PDE5, had no effect on PDE5 phosphorylation in the absence of ANF. Phosphorylation of PDE5 in response to treatment of cells with ANF was associated with a two- to fourfold increase in PDE activity in immunoprecipitates. Multiple-passaged VSMC, which are deficient in PKG but retain PDE5, demonstrated no ANF-dependent increase in phosphorylation or catalytic activity of PDE5. However, incubation of immunoprecipitated PDE5 from these cells with purified PKG, cGMP, and a phosphorylation mixture containing [gamma-32P]ATP resulted in 32(Pi) incorporation into PDE5 that was correlated with increased catalytic activity. These studies are the first to demonstrate phosphorylation of PDE5 in intact cells, thus suggesting a physiological role for this enzyme in smooth muscle regulation.

Asunto(s)

3',5'-GMP Cíclico Fosfodiesterasas/metabolismo , Factor Natriurético Atrial/farmacología , Músculo Liso Vascular/enzimología , 3',5'-GMP Cíclico Fosfodiesterasas/análisis , Adenosina Trifosfato/metabolismo , Sitio Alostérico , Animales , Aorta/enzimología , Bovinos , Células Cultivadas , GMP Cíclico/análogos & derivados , GMP Cíclico/metabolismo , GMP Cíclico/farmacología , Activación Enzimática/efectos de los fármacos , Técnicas de Inmunoadsorción , Masculino , Fosforilación , Proteínas Quinasas/metabolismo , Ratas , Ratas Sprague-Dawley

RESUMEN

Previous work from this and other laboratories has demonstrated that the vasoconstrictor peptide angiotensin II results in hypertrophy of rat aortic smooth muscle cells that is associated with an increase in transcription of the early growth response gene c-fos. To explore the molecular mechanism responsible for c-fos induction in rat aortic smooth muscle cells, we used a series of reporter constructs linked to the chloramphenicol acetyl transferase gene in transient transfection experiments in rat aortic smooth muscle cells. Constructs containing both the serum response element and cAMP response element exhibited a 20-fold increase in chloramphenicol acetyl transferase activity in response to either serum or angiotensin II, whereas no increase was seen in vehicle-treated cells. Mutations in either the serum response element or cAMP response element alone, which have been demonstrated to inactivate these elements in other cell types, had no effect on chloramphenicol acetyl transferase inducibility. In contrast, if both elements were mutated, inducibility was almost abolished. Electrophoretic mobility shift assays with oligonucleotides corresponding to either serum response element or cAMP response element demonstrated that these oligonucleotides are capable of forming specific complexes with proteins from rat aortic smooth muscle cell nuclear extracts. One of the proteins binding to the serum response element is the previously described serum response factor, since it was supershifted by a monospecific antibody. These studies demonstrate that c-fox induction in smooth muscle occurs by a dual mechanism that can activate transcription via the serum response element or cAMP response element. These elements appear to act equally and independently, involving a distinct set of transacting factors.

Asunto(s)

Angiotensina II/fisiología , AMP Cíclico/fisiología , Regulación de la Expresión Génica/fisiología , Genes fos/genética , Músculo Liso Vascular/fisiología , Animales , Secuencia de Bases , Células Cultivadas , Cloranfenicol O-Acetiltransferasa/genética , Datos de Secuencia Molecular , Plásmidos/genética , Conejos , Ratas , Transcripción Genética , Transfección

RESUMEN

Elevated levels of endogenous angiotensin can cause hypertensive nephrosclerosis as a result of the potent vasopressor action of the peptide. We have produced by gene targeting mice homozygous for a null mutation in the angiotensinogen gene (Atg-1-). Postnatally, Atg-1- animals show a modest delay in glomerular maturation. Although Atg-1- animals are hypotensive by 7 wk of age, they develop, by 3 wk of age, pronounced lesions in the renal cortex, similar to those of hypertensive nephrosclerosis. In addition, the papillae of homozygous mutant kidneys are reduced in size. These lesions are accompanied by local up-regulation of PDGF-B and TGF-beta1 mRNA in the cortex and down-regulation of PDGF-A mRNA in the papilla. The study demonstrates an important requirement for angiotensin in achieving and maintaining the normal morphology of the kidney. The mechanism through which angiotensin maintains the volume homeostasis in mammals includes promotion of the maturational growth of the papilla.

Asunto(s)

Angiotensina II/metabolismo , Angiotensinógeno/deficiencia , Angiotensinógeno/genética , Expresión Génica , Sustancias de Crecimiento/biosíntesis , Riñón/crecimiento & desarrollo , Envejecimiento , Angiotensinógeno/sangre , Animales , Presión Sanguínea , Peso Corporal , Homeostasis , Riñón/metabolismo , Riñón/patología , Corteza Renal/crecimiento & desarrollo , Corteza Renal/patología , Glomérulos Renales/crecimiento & desarrollo , Glomérulos Renales/patología , Médula Renal/crecimiento & desarrollo , Médula Renal/patología , Ratones , Ratones Noqueados , Ratones Mutantes , Tamaño de los Órganos , Factor de Crecimiento Derivado de Plaquetas/biosíntesis , ARN Mensajero/análisis , ARN Mensajero/biosíntesis , Mapeo Restrictivo , Factor de Crecimiento Transformador beta/biosíntesis

RESUMEN

For both in vitro and in vivo experiments, especially in vascular cells, an efficient and easy method of gene transfer into this tissue would be extremely useful. Previous methods have either yielded low levels of expression or require complicated manipulation of viral vectors. The goal of this study was to develop an easy, efficient method to introduce unmodified plasmid DNA into vascular tissue. In this report it is demonstrated that complexing unmodified plasmid DNA with replication-deficient adenovirus (Ad5 dl312) via cationic lipids enhances gene transfer up to 1000-fold in cultured bovine aortic endothelial cells (BAECs). Further, utilizing a balloon-injured rabbit femoral artery model, intense nuclear staining in both the neointimal smooth muscle cell layer and the adventitia was seen following transfection with a plasmid containing the lacZ gene and the SV40 nuclear localization signal. Control arteries demonstrated no detectable staining. Our studies suggest that complexing plasmid DNA with adenovirus via lipids greatly enhances gene transfer both in vivo and in vitro. This method could have a wide range of applications for experiments in vascular tissue.

Asunto(s)

Endotelio Vascular/citología , Transfección/métodos , Adenovirus Humanos/genética , Adenovirus Humanos/fisiología , Animales , Aorta , Cateterismo/efectos adversos , Bovinos , Línea Celular , Replicación del ADN , Virus Defectuosos/genética , Virus Defectuosos/fisiología , Endotelio Vascular/lesiones , Endotelio Vascular/patología , Arteria Femoral/lesiones , Vectores Genéticos , Liposomas , Masculino , Músculo Liso Vascular/lesiones , Plásmidos , Conejos , Proteínas Recombinantes/biosíntesis , Replicación Viral , beta-Galactosidasa/biosíntesis , beta-Galactosidasa/genética

RESUMEN

BACKGROUND: Restenosis after catheter-based revascularization has been demonstrated to be primarily caused by medial and/or intimal smooth muscle cell (SMC) proliferation. The objective of this study was investigate the ability of local emission of beta-particles from a 32P-impregnated titanium "stent" wire source to inhibit vascular SMC and endothelial cell proliferation in cell culture and to determine the dose-response characteristics of this inhibition. METHODS AND RESULTS: A series of experiments were performed using 0.20-mm-diameter titanium wires that were impregnated with varying low concentrations of 32P (activity range, 0.002 to 0.06 microCi/cm wire, n = 47) or 31P (nonradioactive control, n = 28) in cultures of rat and human aortic SMCs and in cultured bovine aortic endothelial cells. The zone of complete cell growth inhibition (in millimeters from stent wire) was measured using light microscopy in the cultures exposed to the radioactive (32P) or control (31P) wires at 6 and 12 days after plating. In both rat and human SMC cultures there was a distinct 5.5- to 10.6-mm zone of complete SMC inhibition at wire activity levels > or = 0.006 microCi/cm. In contrast, there was no zone of inhibition surrounding the control (31P impregnated) wires (P < .001 versus 32P wires at all wire activities > or = 0.006 microCi/cm for human and rat SMCs). Proliferating bovine endothelial cells were more radioresistant than SMCs, with no zone of inhibition observed at wire activity levels up to 0.019 microCi/cm (P < .001 versus SMCs at 0.006 microCi/cm and 0.019 microCi/cm). CONCLUSIONS: We conclude that very low doses of beta-particle emission from a 32P-impregnated stent wire (activity levels as low as 0.006 microCi/cm of wire) completely inhibit the growth and migration of both rat and human SMCs within a range of 5.5 to 10.6 mm from the wire. Endothelial cells appear to be much more radioresistant than SMCs. These data suggest that an intra-arterial stent impregnated with a low concentration of 32P may have a salutary effect on the restenosis process. Whether this approach can be used successfully and safely to inhibit restenosis in vivo and in the clinical setting is under investigation.

Asunto(s)

Partículas beta , Músculo Liso Vascular/citología , Stents , Animales , Bovinos , División Celular/efectos de la radiación , Células Cultivadas , Relación Dosis-Respuesta en la Radiación , Humanos , Masculino , Ratas , Ratas Sprague-Dawley

RESUMEN

Focal adhesion kinase (FAK) is a structurally unique nonreceptor protein-tyrosine kinase that localizes to focal adhesion plaques. Regulation of its activity has been implicated in diverse signaling pathways, including those mediated by extracellular matrix/integrin interactions, G-protein coupled receptors for mitogenic neuropeptides, and certain oncogene products. To gain evidence for specific processes in which FAK may be involved in vivo, a study was initiated to determine its expression pattern during mouse development. FAK expression was detected in early embryos and appeared to be distributed throughout all cell types at about the time of neurulation. Subsequent to neural tube closure, expression became particularly abundant in the developing vasculature. This included expression in the medial layer of arteries populated by smooth muscle cells. In vitro studies using cultured rat aortic vascular smooth muscle cells demonstrate that FAK phosphotyrosine content is dramatically elevated in response to plating cells onto the adhesive glycoprotein, fibronectin. Also, enhanced tyrosine phosphorylation of FAK is observed in these cells upon stimulation with the vasoconstrictor angiotensin II. Thus, in vascular smooth muscle cells, like fibroblasts, FAK appears to play a role in signaling mechanisms induced by extracellular matrix components as well as G-protein coupled receptor agonists. The combined results of this study suggest that signaling through FAK may play an important role in blood vessel morphogenesis and function.

Asunto(s)

Angiotensina II/farmacología , Moléculas de Adhesión Celular/biosíntesis , Músculo Liso Vascular/enzimología , Proteínas Tirosina Quinasas/biosíntesis , Tirosina/análogos & derivados , Animales , Moléculas de Adhesión Celular/química , Moléculas de Adhesión Celular/genética , Desarrollo Embrionario y Fetal/fisiología , Inducción Enzimática/efectos de los fármacos , Femenino , Quinasa 1 de Adhesión Focal , Proteína-Tirosina Quinasas de Adhesión Focal , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos ICR , Músculo Liso Vascular/citología , Músculo Liso Vascular/embriología , Fosfotirosina , Proteínas Tirosina Quinasas/química , Proteínas Tirosina Quinasas/genética , Factores de Tiempo , Tirosina/metabolismo

RESUMEN

In recent years numerous data have given evidence that the tissue renin-angiotensin system may play an equal or perhaps an even more important role than the circulating renin-angiotensin system in numerous physiologic processes. This was first suggested by the observation that the blood pressure lowering effect of angiotension-converting enzyme (ACE) inhibitors correlates better with tissue ACE activity than with plasma ACE activity. In response to hypertension and arterial injury, vascular smooth muscle cells undergo three responses: hypertrophy, hyperplasia, and remodeling. The end result is a decrease in lumen diameter and an increase in peripheral vascular resistance. Blockade of angiotensin II formation inhibits these smooth muscle responses in a number of animal models. This review discusses the evidence supporting the existence of local tissue renin-angiotensin system in the vasculature and its physiologic effects. Inhibition of the vascular renin-angiotensin system may have important implications in the treatment of patients with hypertension, atherosclerosis, and restenosis following balloon coronary angioplasty.

Asunto(s)

Músculo Liso Vascular/metabolismo , Sistema Renina-Angiotensina/fisiología , Animales , División Celular , Sustancias de Crecimiento/fisiología , Humanos , Músculo Liso Vascular/citología , Músculo Liso Vascular/enzimología

RESUMEN

Our previous studies demonstrated that the sexually dimorphic pattern of hypertension in the spontaneously hypertensive rat is androgen dependent. Gonadectomy retards the development of hypertension in young males, but not in females, and administration of testosterone propionate to gonadectomized spontaneously hypertensive rats of both sexes confers a male pattern of blood pressure development. The current study tested the hypothesis that renal and hepatic renin and angiotensinogen gene expression are also androgen dependent in the spontaneously hypertensive rat. Male and female spontaneously hypertensive rats underwent gonadectomy or a sham operation at 4 weeks of age. Subgroups of gonadectomized rats of both sexes were implanted with a 15-mm or 30-mm Silastic capsule filled with testosterone at the same time the gonadectomy was performed; a third group received an empty Silastic capsule. Northern and slot blot analyses were used to characterize and quantitate renin and angiotensinogen messenger RNA (mRNA) in the kidney and liver 18 weeks after the gonadectomy. Blood pressure, plasma renin activity, and hepatic angiotensinogen mRNA levels were higher in intact males than in females. Orchidectomy retarded the development of hypertension and lowered plasma renin and renal and hepatic angiotensinogen mRNA levels, and testosterone replacement restored the male pattern of hypertension and plasma renin and increased renal and hepatic angiotensinogen mRNA. Ovariectomy did not alter blood pressure or plasma renin but did lower renal renin and renal and hepatic angiotensinogen mRNA; testosterone increased blood pressure, plasma renin, renal renin and angiotensinogen mRNA, and hepatic angiotensinogen mRNA levels in ovariectomized females.(ABSTRACT TRUNCATED AT 250 WORDS)

Asunto(s)

Angiotensinógeno/genética , Regulación de la Expresión Génica/efectos de los fármacos , Hipertensión/genética , ARN Mensajero/metabolismo , Renina/genética , Testosterona/farmacología , Animales , Secuencia de Bases , Peso Corporal/efectos de los fármacos , Femenino , Hipertensión/sangre , Hipertensión/enzimología , Riñón/efectos de los fármacos , Riñón/enzimología , Hígado/efectos de los fármacos , Hígado/enzimología , Masculino , Datos de Secuencia Molecular , Orquiectomía , Ovariectomía , Ratas , Ratas Endogámicas SHR , Renina/sangre , Caracteres Sexuales

RESUMEN

One of the major consequences of hypertension is an increase in the thickness of the arterial medial smooth muscle cell layer. This has been shown in both large and medium size resistance vessels caused by smooth muscle cell hypertrophy. Both in vivo and in vitro data suggest that the vasoconstrictor peptide angiotensin II (Ang II) may play an important role in the development of the smooth muscle hypertrophy. We have demonstrated that Ang II, when added to quiescence cultures of vascular smooth muscle cells, results in the rapid induction of the early growth response genes c-fos, c-myc, and c-jun. This is due to new transcription as demonstrated by nuclear runoff transcription assay, but is not dependent on new protein synthesis, as it is not blocked by the addition of cycloheximide. The effect is due, however, to an increase in intracellular calcium, suggesting that any vasoconstrictor which results in an increase in intracellular calcium may act in this manner. Following the induction of the early growth response genes there is delayed induction of the platelet derived growth factor A-chain gene. Data from our laboratory and from that of others has shown in preliminary studies that blockade of either the Ang II-induced increases in c-fos or in the platelet-derived growth factor A-chain increases smooth muscle cell protein synthesis. This suggests that Ang II and other vasoconstrictors may play an important role in vascular smooth muscle growth, in hypertension and also in atherosclerosis and following balloon injury of the arterial wall.(ABSTRACT TRUNCATED AT 250 WORDS)

Asunto(s)

Angiotensina II/fisiología , División Celular/efectos de los fármacos , Hipertensión/patología , Músculo Liso Vascular/citología , Angiotensina II/farmacología , Animales , Expresión Génica/efectos de los fármacos , Humanos , Modelos Biológicos , Músculo Liso Vascular/efectos de los fármacos , Músculo Liso Vascular/patología

Asunto(s)

ADP Ribosa Transferasas , Arteriosclerosis/tratamiento farmacológico , Factores de Virulencia , Angioplastia Coronaria con Balón/efectos adversos , Inhibidores de la Enzima Convertidora de Angiotensina/farmacología , Toxinas Bacterianas , Supervivencia Celular , Enfermedad Coronaria/terapia , Exotoxinas/genética , Técnicas Genéticas , Humanos , Músculo Liso Vascular/citología , Músculo Liso Vascular/efectos de los fármacos , Músculo Liso Vascular/metabolismo , Factor de Crecimiento Derivado de Plaquetas/metabolismo , Recurrencia , Factor de Crecimiento Transformador alfa/genética , Exotoxina A de Pseudomonas aeruginosa

RESUMEN

Recent data demonstrate the existence of a vascular renin angiotensin system. In this study we examine the localization of angiotensinogen mRNA in the blood vessel wall of two rat strains, the Wistar and Wistar Kyoto (WKY), as well as the regulation of vascular angiotensinogen mRNA expression by dietary sodium. Northern blot analysis and in situ hybridization histochemistry demonstrate that in both strains angiotensinogen mRNA is detected in the aortic medial smooth muscle layer as well as the periaortic fat. In WKY rats fed a 1.6% sodium diet, angiotensinogen mRNA concentration is 2.6-fold higher in the periaortic fat than in the smooth muscle, as analyzed by quantitative slot blot hybridization. Angiotensinogen mRNA expression in the medial smooth muscle layer is sodium regulated. After 5 d of a low (0.02%) sodium diet, smooth muscle angiotensinogen mRNA levels increase 3.2-fold (P less than 0.005) as compared with the 1.6% sodium diet. In contrast, angiotensinogen mRNA level in the periaortic fat is not influenced by sodium diet. In summary, our data demonstrate regional (smooth muscle vs. periaortic fat) differential regulation of angiotensinogen mRNA levels in the blood vessel wall by sodium. This regional differential regulation by sodium may have important physiological implications.

Asunto(s)

Angiotensinógeno/genética , Aorta/fisiología , Tejido Adiposo/fisiología , Animales , Aorta/anatomía & histología , Northern Blotting , ADN/genética , Expresión Génica , Masculino , Hibridación de Ácido Nucleico , ARN Mensajero/genética , Ratas , Ratas Endogámicas , Ratas Endogámicas WKY , Sodio en la Dieta/metabolismo

RESUMEN

Angiotensin (Ang) II causes hypertrophy of rat aortic smooth muscle cells in culture and results in the rapid activation of c-fos. This study demonstrated that Ang II also activated c-jun and, in addition, could activate the AP-1 enhancer element. These data add support for a role of Ang II as an important mediator of vascular smooth muscle cell growth.

Asunto(s)

Angiotensina II/farmacología , Proteínas de Unión al ADN/genética , Músculo Liso Vascular/fisiología , Factores de Transcripción/genética , Animales , Células Cultivadas , Elementos de Facilitación Genéticos , Expresión Génica/efectos de los fármacos , Técnicas In Vitro , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas c-fos , Proteínas Proto-Oncogénicas c-jun , Proteínas Proto-Oncogénicas c-myc , Proto-Oncogenes , Ratas , Secuencias Reguladoras de Ácidos Nucleicos , Saralasina/farmacología , Factores de Tiempo , Transcripción Genética/efectos de los fármacos

RESUMEN

Because renin is an important enzyme in blood pressure regulation, we studied the possibility that an alteration in the structure of the human renin gene is genetically linked to human essential hypertension or associated with levels of plasma renin activity or blood pressure. By using specific DNA probes, we have identified four polymorphisms in the human renin gene with the restriction enzymes Taq I, HindIII, Bgl I, and Bgl II. The gene location of all of these polymorphisms except for the Bgl II polymorphism has been determined, and their frequencies were initially estimated in a population of 50 random subjects. To test the clinical significance of these polymorphisms, we studied 68 persons from a large Utah pedigree with a high incidence of hypertension. Among nine relatives with hypertension, genetic linkage without recombination was ruled out by observing several obligate recombinants. We also found no significant association of the restriction fragment length polymorphisms with quantitative measurements of sitting or standing, systolic or diastolic blood pressures, or plasma renin activity in 59 untreated members of this pedigree. Although we found no genetic linkage in this set of study subjects, the characterization of the restriction fragment length polymorphisms for the renin gene may be useful in future studies of other selected pedigrees for the presence of one or more of these to be a genetic marker in hypertension.

Asunto(s)

Frecuencia de los Genes , Hipertensión/genética , Polimorfismo de Longitud del Fragmento de Restricción , Renina/genética , Genotipo , Humanos , Linaje

RESUMEN

Probes to hypervariable minisatellite regions of DNA identify multiple loci scattered over the autosomal chromosomes and produce a complex Southern blot pattern of fragments termed a DNA 'fingerprint'. As concern has been raised that different stocks of spontaneously hypertensive rats (SHR) and Wistar-Kyoto rats (WKY) may not be biologically identical, we have compared the DNA of SHR and WKY from several sources using two such probes which identify different sets of minisatellite sequences. While the DNA fingerprints of SHR from the various sources were identical, variability was observed in those of WKY, indicating genetic heterogeneity between different WKY stocks. In animals from one of the commercial suppliers even inter-rat variability in DNA fingerprints was seen, suggesting genetic heterogeneity within that single colony. These observations indicate that experimental results obtained using WKY from different sources may not be directly comparable and could provide an explanation for some of the conflicting data that exist on the comparative characteristics of SHR and WKY. In separate studies, direct comparisons both of the DNA fingerprints of SHR and WKY and of SHR and stroke-prone spontaneously hypertensive rats (SHRSP) showed multiple differences between the strains. The polymorphisms seen could provide useful linkage markers in locating the chromosomal sites of the genetic loci responsible for raised blood pressure in the SHR and the propensity to strokes in the SHRSP.

Asunto(s)

ADN/genética , Hipertensión/genética , Mapeo Nucleótido , Animales , Southern Blotting , Ligamiento Genético , Ratas , Ratas Endogámicas SHR , Ratas Endogámicas WKY

RESUMEN

Angiotensin II (Ang II) has been shown to cause hypertrophy of cultured quiescent rat aortic smooth muscle cells. This observation along with the recent demonstration of angiotensinogen messenger RNA (mRNA) in the vessel wall has led us to postulate a role for Ang II in hypertensive smooth muscle hypertrophy. One of the earliest responses in a wide variety of cells in response to a growth-promoting agent is the induction of the proto-oncogene c-fos. To investigate the mechanism of the action of Ang II, we investigated the effect of Ang II on the expression of the c-fos gene in rat aortic smooth muscle cells that were made quiescent by being grown in a defined serum-free media for 48 hours. Ang II (10(-6)-10(-10) M) resulted in a dose-dependent increase in c-fos mRNA expression. This induction was angiotensin-receptor specific since it was completely abolished by the competitive inhibitor saralasin. Inhibition of protein synthesis did not block the rise in c-fos mRNA expression; it resulted in a superinduction and stabilization of the c-fos mRNA. Using a nuclear runoff transcription assay, we demonstrated that Ang II stimulated the transcription rate of the c-fos gene. This activation of c-fos gene expression may be an important mechanism in the angiotensin-induced smooth muscle hypertrophy.

Asunto(s)

Angiotensina II/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Músculo Liso/fisiología , Proto-Oncogenes/efectos de los fármacos , Transcripción Genética/efectos de los fármacos , Animales , Cicloheximida/farmacología , ARN Mensajero/metabolismo

RESUMEN

Recently, angiotensin II (Ang II) has been shown to cause hypertrophy of cultured quiescent rat aortic smooth muscle (RASM) cells. This observation along with the demonstration of angiotensinogen mRNA in the vessel wall has led us to postulate a role for vascular angiotensin in hypertensive blood vessel hypertrophy. To investigate further the possible molecular mechanisms, we examined the effect of Ang II on the expression of two genes known to be involved with cellular growth response. Near-confluent RASM cells were made quiescent by 48-h exposure to a defined serum-free medium. Ang II (10(-6) to 10(-11) M) resulted in an induction of the protooncogene c-myc mRNA within 30 min which persisted for 6 h. Interestingly, 6 h after the addition of Ang II, platelet-derived growth factor (PDGF) A-chain mRNA expression was elevated, peaked in 9 h, and persisted for 11 h. This was accompanied with a 15-20-fold increase in PDGF concentration in the culture medium. These effects were dose-dependent and were blocked by saralasin. Whereas the inhibition of protein synthesis by cycloheximide resulted in a stabilization of c-myc mRNA, cycloheximide abolished the elevation of the PDGF A-chain mRNA. Taken together, our data show that exposure of RASM cells to Ang II results in the sequential activation of c-myc and PDGF A-chain mRNA expressions. This sequential activation of protooncogene and growth factor gene may be an important mechanism in angiotensin-induced smooth muscle growth and hypertrophy.

Asunto(s)

Angiotensina II/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Músculo Liso Vascular/metabolismo , Factor de Crecimiento Derivado de Plaquetas/biosíntesis , Proteínas Proto-Oncogénicas/biosíntesis , Animales , Células Cultivadas , Músculo Liso Vascular/efectos de los fármacos , Oncogenes/efectos de los fármacos , Factor de Crecimiento Derivado de Plaquetas/genética , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas c-myc , ARN Mensajero/metabolismo , Ratas

RESUMEN

The expression of the intrarenal renin angiotensin mRNAs in Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR) were investigated in rats fed a low (0.02%)- or normal (1.6%)-sodium diet for 5 days. Total RNA was isolated from the kidneys and analyzed by both Northern and slot blots. The results indicated that the kidneys of all four groups of rats containing readibly detectable levels of renin and angiotensinogen mRNAs. The kidneys of the WKY contained higher levels of angiotensinogen mRNA under normal-salt diet compared with the SHR (P less than 0.01). Mild sodium depletion stimulated angiotensinogen mRNA in WKY kidneys by almost 50% compared with normal salt diet (P less than 0.01). In contrast, there was no significant difference in the renal angiotensinogen mRNA levels in the kidneys of SHR fed a low- or normal-sodium diet. Renin mRNA concentrations were comparable in both strains. Mild sodium depletion failed to yield any detectable changes in renin mRNA levels in either strain. Thus the SHR exhibits an alteration in the sodium regulation of intrarenal angiotensinogen mRNA expression. These results may have implications in the renal physiology of these animals.

Asunto(s)

Angiotensinógeno/genética , Regulación de la Expresión Génica/efectos de los fármacos , Riñón/efectos de los fármacos , ARN Mensajero/genética , Sodio en la Dieta/farmacología , Animales , Northern Blotting , Riñón/enzimología , Masculino , ARN Mensajero/efectos de los fármacos , Ratas , Ratas Endogámicas SHR , Ratas Endogámicas WKY , Renina/metabolismo

RESUMEN

To test the hypothesis that structural abnormalities exist in the cell membrane in persons with essential hypertension and that these abnormalities affect membrane-related cellular functions, we examined several membrane-dependent phenomena and membrane lipid composition in the blood cells of subjects with essential hypertension. We analyzed platelet aggregability, membrane fluidity, membrane fatty acid composition, and erythrocyte deformability in four normolipidemic subjects with untreated essential hypertension and in five age-matched normotensive controls. As compared with the controls, the subjects with essential hypertension had platelets that aggregated at lower concentrations of adenosine 5'-diphosphate, platelet membranes that were less fluid, and erythrocytes that were more deformable. Lipid analysis of the membranes of platelets from the two study groups showed that although the cholesterol content was identical, the membranes from the essential hypertension group contained significantly less linoleic acid (18:2) than did those from the normotensive controls. Given the known effects of cis-unsaturated fatty acyl composition on membrane fluidity and membrane-related cellular functions, these data suggest that one factor contributing to essential hypertension is an inherent structural membrane abnormality that alters the physical and functional properties of the cell membrane.

Asunto(s)

Membrana Celular/fisiología , Hipertensión/sangre , Adulto , Plaquetas/análisis , Colesterol/sangre , Deformación Eritrocítica , Ácidos Grasos/sangre , Femenino , Humanos , Masculino , Fluidez de la Membrana , Lípidos de la Membrana/análisis , Agregación Plaquetaria , ATPasa Intercambiadora de Sodio-Potasio/sangre

RESUMEN

We previously observed that the renal nerves facilitate sodium retention and contribute to the development of DOCA-salt hypertension in the rat. To determine whether the renal nerves also participate in the maintenance of DOCA-salt hypertension, we studied the effects of renal denervation after 3 or 10 weeks of DOCA-salt treatment on systolic blood pressure, urinary sodium excretion, creatinine clearance, and precapillary arteriolar wall/lumen ratios of renal, hindlimb muscle, and cremaster muscle vascular beds. Systolic blood pressures of animals given DOCA-salt reached a plateau by 3 weeks of treatment at which time a sham operation or renal denervation was performed. Sham operation in hypertensive animals resulted in no change in systolic blood pressure and no change in percent sodium intake excreted. Wall/lumen ratio of the renal precapillary arteriole in sham-operated hypertensive animals was increased compared to similar sized vessels in hindlimb and cremaster muscle. In contrast, renal denervation resulted in a natriuresis and an attenuation of the hypertension (208 +/- 7 mmHg; p less than 0.01). Wall/lumen ratio of the renal capillary arterioles in renal denervated animals was no different than similar sized vessels in hindlimb and cremaster muscle and significantly less than that seen in sham-operated animals (0.85 +/- 0.05 vs 1.03 +/- 0.06; p less than 0.05). In another group of animals, sham operation or renal denervation was performed after 10 weeks of DOCA-salt treatment. At this time neither operation altered systolic blood pressure or sodium balance. In contrast to 3-week DOCA-salt-treated hypertensive sham-operated animals, renal precapillary arteriolar wall/lumen ratio of 10-week animals was no different than similar sized vessels in hindlimb and cremaster muscle. In addition, renal precapillary arteriolar wall/lumen ratio of 10-week DOCA-salt-treated renal-denervated animals was no different than that seen in 10-week DOCA-salt-treated sham-operated hypertensive animals. Creatinine clearance of the 10-week DOCA-salt-treated sham-operated or renal-denervated animals was significantly (p less than 0.01) lower than that of the 3-week DOCA-salt-treated groups (0.25 +/- 0.14 vs 1.03 +/- 0.10 ml/min). These data suggest that the renal nerves contribute to the early established phase of DOCA-salt hypertension by shifting the arterial pressure-renal sodium excretion curve to the right. With time, the renal nerves play a diminishing role in the maintenance of established DOCA-salt hypertension in the rat, while other renal factors, including decreased glomerular filtration rate and probable fixed renal vascular changes, play an increasingly important role.

Asunto(s)

Hipertensión/fisiopatología , Riñón/inervación , Natriuresis , Sistema Nervioso Simpático/fisiopatología , Animales , Arteriolas/patología , Presión Sanguínea/efectos de los fármacos , Creatinina/metabolismo , Desoxicorticosterona , Hipertensión/inducido químicamente , Hipertensión/metabolismo , Riñón/metabolismo , Masculino , Norepinefrina/metabolismo , Ratas , Ratas Endogámicas , Renina/sangre , Cloruro de Sodio , Bazo/inervación , Bazo/metabolismo , Simpatectomía , Factores de Tiempo