RESUMO
Gonadotropin stimulates protein synthesis and growth in ovarian oocytes. The hormone is also known to modify transfollicular K+ fluxes and is now shown to cause increased intraoocytic K+ activity (aK). The hormone's effect on aK was duplicated by microinjecting K+ salts into oocytes which were incubated in paraffin oil. This treatment mimicked the influence of gonadotropin on both the rate of protein synthesis and the synthesis of specific polypeptides. These findings suggest that gonadotropin-stimulated oocyte growth is attributable largely to the hormone's influence on transfollicular K+ fluxes. They support the hypothesis that the K+ flux and aK changes observed during cell activation are critical in causing subsequent increases in protein synthesis and growth.
Assuntos
Gonadotropina Coriônica/farmacologia , Oócitos/metabolismo , Cloreto de Potássio/farmacologia , Animais , Eletroforese em Gel de Poliacrilamida , Feminino , Leucina/metabolismo , Microinjeções , Oócitos/efeitos dos fármacos , Oócitos/crescimento & desenvolvimento , Potássio/metabolismo , Biossíntese de Proteínas , Xenopus laevisRESUMO
BACKGROUND: Although studies have demonstrated that sirtinol administration following adverse circulatory conditions is known to be protective, the mechanism by which sirtinol produces the salutary effects remains unknown. We hypothesized that sirtinol administration in male rats following trauma-hemorrhage decreases cytokine production and protects against hepatic injury. METHODS: Male Sprague-Dawley rats underwent trauma-hemorrhage (mean blood pressure 40 mmHg for 90 min, then resuscitation). A single dose of sirtinol (1 mg/kg of body weight) or vehicle was administered intravenously during resuscitation. Twenty-four hours thereafter, tissue myeloperoxidase (MPO) activity (a marker of neutrophil sequestration), cytokine-induced neutrophil chemoattractant (CINC)-1, CINC-3, intercellular adhesion molecule (ICAM)-1, and interleukin (IL)-6 levels in the liver and plasma alanine aminotransferase (ALT) concentrations were measured (n=6 Sprague-Dawley rats/group). RESULTS: Trauma-hemorrhage increased hepatic MPO activity, CINC-1, CINC-3, ICAM-1, and IL-6 levels and plasma ALT concentrations. These parameters were significantly improved in the sirtinol-treated rats subjected to trauma-hemorrhage. CONCLUSION: The salutary effects of sirtinol administration on attenuation of hepatic injury following trauma-hemorrhage are, at least in part, related to reduction of pro-inflammatory mediators.
Assuntos
Benzamidas/farmacologia , Citocinas/biossíntese , Hemorragia/tratamento farmacológico , Fígado/efeitos dos fármacos , Naftóis/farmacologia , Ferimentos e Lesões/tratamento farmacológico , Alanina Transaminase/sangue , Animais , Citocinas/análise , Hemorragia/metabolismo , Molécula 1 de Adesão Intercelular/análise , Fígado/metabolismo , Masculino , Peroxidase/metabolismo , Ratos , Ratos Sprague-Dawley , Resultado do Tratamento , Ferimentos e Lesões/metabolismoAssuntos
Doença de Fabry/fisiopatologia , Antebraço/irrigação sanguínea , Microcirculação , Tomografia Computadorizada por Raios X/métodos , Adolescente , Adulto , Idoso , Criança , Temperatura Baixa , Feminino , Antebraço/fisiopatologia , Humanos , Masculino , Pessoa de Meia-Idade , Linhagem , Perfusão , Adulto JovemRESUMO
The effects of antidiuretic hormone (ADH) on the rate of transepithelial active Na transport JaNa and the rate of suprabasal O2 consumption of Jsbr were studied in paired hemiskins of frog. Within some 30 min following administration of ADH both JaNa and Jsbr increased to near-maximal levels and then remained stable for at least an hour. On symmetric perturbation of the transepithelial electrical potential delta psi at 6-min intervals, the dependence of JaNa and Jsbr on delta psi was near-linear, both in control and experimental hemi-skins. The stability and near-linearity of the system permitted systematic analysis of the parameters of linear non-equilibrium thermodynamic (NET) and electrical equivalent circuit (EC) formulations. ADH (100 mU/ml) stimulated two of the three NET phenomenological L coefficients, as well as A, the affinity (negative Gibbs free energy) of a metabolic reaction driving transport. Observations at partially depressed levels of transport indicated that the effects of kinetic and energetic factors are to some extent discrete. EC analysis showed stimulation of the amiloride-sensitive conductance Ka, but not of the apparent electromitive force of Na transport 'ENa'. Similar effects were produced by 10 mU/ml of ADH or by 10 mM dibutyryl cyclic AMP, although less marked effects on the L coefficients were noted with the lower concentration of hormone. It is suggested that, in contrast to EC analysis, the NET formulation distinguishes between kinetic and energetic determinants of transport, supporting a dual mechanism of action of ADH.
Assuntos
Pele/metabolismo , Sódio/metabolismo , Vasopressinas/farmacologia , Animais , Transporte Biológico Ativo/efeitos dos fármacos , Bucladesina/farmacologia , Relação Dose-Resposta a Droga , Eletrofisiologia , Cinética , Potenciais da Membrana/efeitos dos fármacos , Consumo de Oxigênio , Rana pipiens , TermodinâmicaRESUMO
In the study of active transport it is important to distinguish between oxygen consumption sustaining transepithelial transport and that responsible for other tissue functions (basal metabolism). Since amiloride blocks transepithelial active sodium transport and the associated oxygen consumption in the frog skin and toad bladder, we and others have employed this agent to evaluate the rate of basal metabolism. This technique has recently been criticized in a report that amiloride (and ouabain) increased oxygen consumption when no sodium was available for transport. We have been unable to corroborate these observations. With magnesium-Ringer as external bathing solutions, amiloride and ouabain failed to stimulate oxygen consumption. With sodium-Ringer as external bathing solution amiloride reduced oxygen consumption about 30%, to a level indistinguishable from that found on external substitution of magnesium-Ringer for sodium-Ringer. We conclude that the use of amiloride permits evaluation of the rate of basal metabolism with acceptable accuracy; a possible slight depressant effect of ouabain on basal metabolism remains to be investigated.
Assuntos
Consumo de Oxigênio , Pele/metabolismo , Sódio/metabolismo , Bexiga Urinária/metabolismo , Amilorida/farmacologia , Animais , Anuros , Transporte Biológico Ativo/efeitos dos fármacos , Bufo marinus , Feminino , Canais Iônicos/efeitos dos fármacos , Magnésio/farmacologia , Ouabaína/farmacologia , Consumo de Oxigênio/efeitos dos fármacos , Rana pipiensRESUMO
Conflicting reports exist concerning the presence of a Na(+)-coupled amino acid transport system in cultured endothelial cells. We have employed a non-metabolizable analog, 2-aminoisobutyric acid (AIB), to investigate the activity of Na(+)-dependent amino acid transport in cultured human umbilical vein endothelial cells (HUVEC). We found a pronounced saturable, Na(+)-dependent component of AIB uptake in 'fresh' (non-starved) HUVEC. The Na(+)-dependent component accounted for 78% of total AIB uptake with a high sensitivity to external Na+. The accumulation of AIB was inhibited by ouabain preincubation, consistent with the energetics of Na(+)-coupled transport. Amiloride, an epithelial Na+ channel blocker, also inhibited AIB transport at high concentration. The results strongly support the presence of a Na(+)-coupled transport system of amino acid in HUVEC.
Assuntos
Ácidos Aminoisobutíricos/metabolismo , Endotélio Vascular/metabolismo , Amilorida/farmacologia , Transporte Biológico , Radioisótopos de Carbono , Células Cultivadas , Humanos , Cinética , Ouabaína/farmacologia , Cloreto de Sódio/farmacologiaRESUMO
A net inward fluid reabsorption (salt-linked flow) has been observed in isolated skin epithelium (split skin) with the same magnitude as in whole skin when identical NaCl Ringer solutions were used to bathe both sides. Split skins also respond to a hyperosmotic sucrose solution bathing the outer (epithelial) surface by generating an outward osmotic flow. A non-linear relationship between osmotic flow and the osmotic gradient has been found in split skin similar to that found in whole skin.
Assuntos
Pele/metabolismo , Água/metabolismo , Animais , Anuros , Transporte Biológico , Manitol/metabolismo , Concentração Osmolar , Permeabilidade , Rana pipiensRESUMO
OBJECTIVE: Death of vascular smooth muscle cell (VSMC) induced by oxidized LDL (oxLDL) can occur by both necrosis and apoptosis which may contribute to plaque instability and rupture. Reactive oxygen species (ROS) induces apoptosis in VSMC and is involved in oxLDL action, we tested the hypothesis here that a coupling exists between ROS generation and apoptosis of oxLDL-treated VSMC. METHODS: Cultured VSMC from rat aorta were treated with oxLDL, apoptosis and necrosis were distinguished by using FITC-annexin V label and propidium iodide stain, analyzed by flow cytometry. ROS generation of VSMCs was detected by the fluorescence intensity of DCF. Apoptosis was also determined by cleavage of procaspase-3. RESULTS: OxLDL induced apoptosis (3 h) in a dose-dependent manner and reached maximum (with near-basal necrosis) at a concentration of 300 microg/ml. At this and lower (100 microg/ml) concentration, oxLDL, but not native LDL, stimulated ROS production rapidly (< or =5 min) and ROS level remained elevated for at least 45 min. Catalase and deferoxamine reduced both oxLDL-induced apoptosis and ROS generation. Superoxide dismutase and benzoic acid neither reduced the oxLDL-induced ROS generation nor inhibited apoptosis. Since oxLDL-induced ROS generation were inhibited by nordihydroguaiaretic acid and rotenone, lipoxygenase and mitochondrial pathways could be involved. In addition, catalase, deferoxamine, and N-acetylcysteine inhibited oxLDL-induced cleavage of procaspase-3 as well. CONCLUSIONS: ROS generation and apoptosis are tightly coupled in oxLDL-treated VSMCs. Antioxidants that reduced ROS level inhibited apoptosis, those that did not reduce ROS level were ineffective. Both mitochondrial and lipoxygenase activities may be involved.
Assuntos
Apoptose/efeitos dos fármacos , Lipoproteínas LDL/farmacologia , Músculo Liso Vascular/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Acetilação , Animais , Caspase 3 , Caspases/metabolismo , Técnicas de Cultura de Células , Relação Dose-Resposta a Droga , Precursores Enzimáticos/metabolismo , Masculino , Oxirredução , Ratos , Ratos Endogâmicos WKYRESUMO
We have previously shown that cholesterol enrichment reduces 3H-ouabain binding in cultured vascular endothelial cells. The present study aimed to determine the effect of cholesterol enrichment on ouabain-sensitive 86Rb (as a substitute for K+) influx, i.e. K+ transport via the pump, and to examine whether cellular K+ content was affected in human umbilical vein endothelial cells. 86Rb influx was inhibited by both ouabain and bumetanide in a dose-dependent manner. Consistent with an earlier report [1], inhibition achieved was greater for ouabain (approximately 70% at mM range) than for bumetanide (approximately 55% at 0.1 mM), indicating that K+ influx via Na+/K+ pump was greater than that via Na(+)-K(+)-Cl- cotransport in these cells. After incubation of 18 h or more with cholesterol-enriched liposomes (2:1 cholesterol to phospholipid ratio), a significant reduction (> 20%) of the ouabain-sensitive K+ influx and an increase in cellular cholesterol content were observed. The inhibitory effect was observed only at liposome concentrations above 2 mg/ml. Following 18 h incubation with 2 mg/ml cholesterol-enriched liposomes, cellular K+ content was significantly decreased. The phospholipid liposome treatment did not alter K+ content, suggesting that the inhibitory effect on Na+/K+ pump and the cellular K+ content reduction was not due to liposome fusion alone or to the phospholipid present. These findings indicate that cholesterol enrichment inhibits Na+/K+ pump and thus reduces cellular K+ content in endothelial cells, and may play a role in the widely observed abnormal endothelium-dependent vascular response induced by hypercholesterolemia.
Assuntos
Colesterol/farmacologia , Endotélio Vascular/metabolismo , ATPase Trocadora de Sódio-Potássio/metabolismo , Bumetanida/farmacologia , Células Cultivadas , Relação Dose-Resposta a Droga , Humanos , Ouabaína/farmacologia , Potássio/metabolismo , ATPase Trocadora de Sódio-Potássio/antagonistas & inibidores , Veias UmbilicaisRESUMO
Oxidized low-density lipoprotein (oxLDL) plays a key role in the development of atherogenesis, partly by causing injury to vascular cells. However, different preparations of LDL, methods of oxidation, and/or active components often produce cellular effects of various degrees. To explore the quantitative relationship between dose and level of oxidation of the oxLDL utilized, we employed combinations of different levels of oxidation and concentrations of oxLDL to induce cell death in cultured vascular smooth muscle cells (VSMC). We also examined the effect of lysophosphatidylcholine (lysoPC), a putative active component of oxLDL, on VSMCs by determining, in parallel with a cytotoxicity test (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay), DNA fragmentation ([3H]thymidine release), and flow cytometric analyses. We found that oxLDL caused cytotoxicity in an oxidative level- and dose-dependent manner, lysoPC also caused dose-dependent cytotoxicity with or without serum. Fragmentation of DNA was observed in both oxLDL- and lysoPC-treated VSMCs. Furthermore, lysoPC-induced DNA ladder was also demonstrated by gel electrophoresis at a concentration of 25 micromol/l or higher. Flow cytometric analysis yielded similar results for oxLDL- and lysoPC-treated VSMC; namely, an accumulation in the fraction of cells in G(0)/G(1) phase with a reciprocal change in S-phase fraction. Membrane phosphatidylserine exposure, detected by annexin V staining, provided additional evidence that lysoPC induced significant apoptosis in VSMC. Taken together, the degree of oxLDL-induced cytotoxicity/apoptosis of VSMC depended on combined effects of oxLDL concentration and oxidative level. Moreover, lysoPC also elicited a dose-dependent apoptosis in addition to cytotoxicity.
Assuntos
Apoptose , Lipoproteínas LDL/farmacologia , Lisofosfatidilcolinas/farmacologia , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/fisiologia , Animais , Ciclo Celular/efeitos dos fármacos , Morte Celular , Membrana Celular/metabolismo , Células Cultivadas , Fragmentação do DNA , Relação Dose-Resposta a Droga , Músculo Liso Vascular/citologia , Fosfatidilserinas/metabolismo , Ratos , Ratos Endogâmicos WKYRESUMO
We describe a technique for isolating the nucleus of the giant amphibian oocyte under paraffin oil. The method precludes the losses of small solutes and proteins that accompany isolation of nuclei into aqueous media. An individual oocyte is blotted, placed under oil, punctured near the animal pole and then squeezed to gently extrude the nucleus into the oil, thereby avoiding exposure to any aqueous environment. Light and electron microscopy of the oil-isolated nucleus demonstrate that its in vivo morphology is preserved. We also describe techniques that facilitate the study of nuclear functions under oil. Oil-isolated oocyte nuclei retain many in vivo functions for several hours, including size-selective envelope permeability, RNA synthesis and the ability to break down in response to cdc2/cyclin meiotic maturation promoting factor.
Assuntos
Fracionamento Celular/métodos , Núcleo Celular , Óleos , Oócitos/ultraestrutura , Animais , Núcleo Celular/metabolismo , Núcleo Celular/fisiologia , Núcleo Celular/ultraestrutura , Proteínas do Ovo/análise , Feminino , Fator Promotor de Maturação/metabolismo , Micromanipulação , Parafina , Permeabilidade , Potássio/metabolismo , RNA/biossíntese , Xenopus laevisRESUMO
Inositol 1,4,5-triphosphate has been proposed as a second messenger for calcium mobilization. The addition of inositol 1,4,5-triphosphate at a low concentration has been shown to cause calcium release from intracellular microsomal stores in rat hepatocytes. The effects of sepsis on the inositol 1,4,5-triphosphate binding from microsomal fractions of rat liver were investigated. Sepsis was induced by cecal ligation and puncture (CLP). Control rats were sham operated. Three microsomal fractions (rough, intermediate, and smooth I) were isolated from the rat liver. The study of inositol 1,4,5-triphosphate receptor binding was performed with tritium-labeled inositol 1,4,5-triphosphate. Our results showed that the Bmax of inositol 1,4,5-triphosphate binding in early septic, late septic, and control groups was 14.9 +/- .9 fmol/mg, 9.8 +/- 1.0 fmol/mg, and 17.2 +/- 1.3 fmol/mg, respectively. The binding activity was unaffected during early sepsis but was significantly depressed by 40-50% (p < .05, vs. control) during late sepsis (18 h after CLP) in all three subfractions of endoplasmic reticulum. Because the inositol 1,4,5-triphosphate binding plays an important role in the regulation of intra-cellular calcium homeostasis in hepatocytes, an impairment of the calcium release due to depressed inositol 1,4,5-triphosphate binding in the endoplasmic reticulum may have a pathophysiological significance in contributing to altered hepatic metabolism during septic shock.
Assuntos
Inositol 1,4,5-Trifosfato/metabolismo , Microssomos Hepáticos/metabolismo , Sepse/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Cálcio/metabolismo , Retículo Endoplasmático Rugoso/metabolismo , Líquido Intracelular/metabolismo , Transporte de Íons , Cinética , Masculino , Ratos , Ratos Sprague-Dawley , Sistemas do Segundo Mensageiro , Fatores de TempoRESUMO
The role of nitric oxide (NO) in microcirculation during the development of acute pancreatitis was not clear. An in vivo microscopic technique was used for evaluating leukocyte-endothelial adherence in the pancreatic microcirculation after induction (cerulein) of acute pancreatitis. Microdialysis was performed to detect pancreatic nitrate concentration (NO level) by high-performance liquid chromatography. Cerulein caused significantly reduced flow velocity in 1 h (31 %) and increased the number of sticking leukocytes in 2 h; both persisted for at least 3 h. Pancreatic NO level was found to be significantly elevated (2.5-fold) in 1 h and also persisted for 3 h. Both microcirculatory changes and NO elevation were significantly alleviated in cerulein-induced animals pretreated with NO synthase inhibitor (NG-nitro-L-arginine), indicating that elevation of NO could precede and account for a major portion of the observed microcirculatory changes. Furthermore, there was a strong positive correlation between numbers of adherent leukocytes and pancreatic NO level, suggesting that during the development of acute pancreatitis, NO could play an adverse role in microcirculation.
Assuntos
Ceruletídeo/toxicidade , Endotélio Vascular/metabolismo , Óxido Nítrico/metabolismo , Pancreatite/metabolismo , Pancreatite/patologia , Animais , Inibidores Enzimáticos/farmacologia , Ácido Glicodesoxicólico/farmacologia , Leucócitos/patologia , Masculino , Microcirculação , NG-Nitroarginina Metil Éster/farmacologia , Nitratos/análise , Nitratos/metabolismo , Pâncreas/irrigação sanguínea , Pancreatite/induzido quimicamente , Ratos , Ratos Sprague-DawleyRESUMO
BACKGROUND: Intracellular calcium concentration is an important regulator of cellular metabolism. Endoplasmic reticulum membranes play an important role in the regulation of cytoplasmic calcium in the mammalian liver. The characterization of the changes of calcium uptake in endoplasmic reticulum may contribute to the potential intracellular mechanisms for cellular dysfunction during sepsis. METHODS: The effects of sepsis on the calcium uptake in rough endoplasmic reticulum of rat liver were studied. Sepsis was induced by means of cecal ligation and puncture (CLP). The control rats underwent sham operation. Microsomal fractions were isolated from the liver with differential centrifugation. RESULTS: The calcium uptake by liver endoplasmic reticulum was decreased by 30% to 35% (p < 0.05) during early sepsis (9 hours after CLP) and by 38% to 43% (p < 0.05) during late sepsis (18 hours after CLP), respectively. The maximum velocity values for adenosine triphosphate (ATP) and for Ca2+ were also decreased by 25% to 37% (p < 0.05) during early sepsis and by 35% to 42% (p < 0.05) during late sepsis. The Michaelis-Menten constant for ATP and Ca2+ transport had no difference among three groups. The magnesium stimulation and vanadate inhibitory activity were also decreased by 17% to 38% (p < 0.05) during early sepsis and by 34% to 50% (p < 0.05) during late sepsis. CONCLUSIONS: These data demonstrate that ATP-dependent calcium uptake in rough endoplasmic reticulum of rat liver was impaired during early and late sepsis. Because the low intracellular calcium concentration plays an important role in the regulation of cellular function, an impairment in the ATP-dependent calcium uptake by endoplasmic reticulum during early and late sepsis may have a pathophysiologic significance in contributing to the development of altered hepatic metabolism during sepsis.
Assuntos
Trifosfato de Adenosina/farmacologia , Cálcio/metabolismo , Microssomos Hepáticos/metabolismo , Sepse/metabolismo , Animais , Retículo Endoplasmático/metabolismo , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
Vasoconstrictor response (VR) induced by inspiratory gasp exhibited a strong positive correlation with vital capacity (VC) which reflects the magnitude of the input stimulus for VR (Lau et al., Clin. Sci. 89:233-237, 1995). Whether a stoichiometric relationship existed between VC and VR is not known. We examined this question in two studies by determining VC and microcirculatory blood flow with laser Doppler flowmetry in healthy subjects. We first studied 40 non-smokers of different gender and age and found that the variation in VR cannot be eliminated by normalization with VC. In the second study we examined 10 young male smokers as well as matched non-smokers of identical VC, we found that smokers had reduced VR. Taking together, the present studies demonstrated that vasoconstrictor response (VR) was not determined by vital capacity alone and that smoking adversely affected VR in the absence of altered VC.
Assuntos
Pele/irrigação sanguínea , Fumar/efeitos adversos , Vasoconstrição , Capacidade Vital , Adulto , Humanos , Fluxometria por Laser-Doppler , Masculino , Fumar/fisiopatologiaRESUMO
Increase of intracellular reactive oxygen species (ROS) has been proposed to cause endothelial injury, and oxidized LDL (oxLDL) actions are associated with an early increase of ROS. Estrogen protects vascular cells partly via its antioxidant effects and by preventing lipid peroxidation. However, whether it can inhibit oxLDL-induced stimulation of ROS generation in endothelial cells is unknown. We utilized the fluorescent dye (DCFH-DA) to measure ROS generation and compared the stimulant effect of tert-butylhydroperoxide (TBH) and oxLDL in human umbilical vein endothelial cells (HUVECs). We found that TBH, H2O2, and oxLDL rapidly stimulated ROS generation, and in a dose-dependent manner with TBH. A concentration of estrogen effective in preventing lipid peroxidation was employed either by pretreatment of cells 18 h prior to or by direct co-incubation (30 min) with HUVEC and oxLDL. Estrogen (54 microM) pretreatment significantly suppressed both TBH- and oxLDL- induced stimulation of ROS generation. Both 1 and 54 microM concentration of estrogen could directly inhibit oxLDL-induced ROS production in HUVECs. Thus, either 18 h pretreatment or 30 min co-incubation with estrogen reduced stimulated ROS generation, suggesting that both cellular and direct actions of estrogen may be involved.
Assuntos
Antioxidantes/farmacologia , Endotélio Vascular/metabolismo , Estradiol/farmacologia , Lipoproteínas LDL/antagonistas & inibidores , Espécies Reativas de Oxigênio/metabolismo , Células Cultivadas , Relação Dose-Resposta a Droga , Endotélio Vascular/efeitos dos fármacos , Fluoresceínas/metabolismo , Lipoproteínas LDL/farmacologia , Veias Umbilicais/citologia , terc-Butil Hidroperóxido/farmacologiaRESUMO
Binding experiments were performed with [3H]ouabain on cultured human umbilical vein endothelial cells (huvEC). Saturation studies yielded a binding capacity (Bmax) of 820 +/- 81 fmole/mg pr.(n = 4) and dissociation constant (KD) of 11.7 +/- 2.1nM (n = 4) in K(+)-free buffer for specific [3H] ouabain binding on these cells. External K+ inhibited this binding in a dose-dependent manner. The mean value of Bmax is equivalent to about 4 x 10(5) sites per cell, comparable with that of smooth muscle cell. These data demonstrated the presence of specific [3H]ouabain binding linked to Na+/K+ pump, consistent with the observations of ouabain-sensitive 86Rb uptake in huvEC. Effect of cholesterol enrichment was also studied. Incubation in media supplemented with cholesterol-phospholipid liposomes of molar ratio of 2:1 for 18 hours reduced the Bmax by 31% (P < 0.05) without significantly changed the value of KD. This reduction of [3H]ouabain binding appeared to be specific for cholesterol since liposome made with pure phospholipid did not alter binding. Recent findings indicate that cholesterol-enrichment and plasma lipoproteins enhance vascular contractile response, our results suggest that the cholesterol-enrichment of endothelial cells may also indirectly affect the vascular response via disturbing the function of Na+/K+ pump.
Assuntos
Colesterol/farmacologia , Endotélio Vascular/metabolismo , Ouabaína/metabolismo , Sítios de Ligação , Células Cultivadas , Endotélio Vascular/citologia , Endotélio Vascular/efeitos dos fármacos , Humanos , Lipossomos , Potássio/farmacologia , ATPase Trocadora de Sódio-Potássio/efeitos dos fármacos , Veias UmbilicaisRESUMO
We studied the rate of 3H-leucine incorporation and the acute effect of ethanol on rat jejunum employing a newly-developed method originally for determining rapid solute uptake in vitro. 3H-leucine incorporation was linear up to 12 min and significantly inhibited by 10 microM of cycloheximide, a specific inhibitor for protein synthesis. The inhibition was 86% at 100 microM indicating that the rate of 3H-leucine incorporation was a good index for protein synthetic activity in the intestinal cells. Incorporation was not changed significantly by 8% dilution (with water) but was inhibited by 8% ethanol to less than 1/4 of the control level. Lower concentrations of ethanol had no consistent effects. The action of ethanol was rapid, independent of the precursor level, and was not readily reversible. Although the nature of ethanol's action was not clear, sucrose, in a dose-dependent manner, also significantly inhibited incorporation rapidly, indicated that the action of media hyperosmolality could not be eliminated as part of the cause of inhibition induced by ethanol.
Assuntos
Etanol/farmacologia , Mucosa Intestinal/metabolismo , Biossíntese de Proteínas , Animais , Cicloeximida/farmacologia , Feminino , Técnicas In Vitro , Leucina/metabolismo , Ratos , Ratos EndogâmicosRESUMO
Erythrocytes of normotensive and hypertensive humans, as well as Sprague-Dawley, Wistar-Kyoto and spontaneously hypertensive rats, were prepared to have similar ionic compositions adequate for determining the activities of lithium-sodium (Li-Na) countertransport and sodium (Na) pump. The rate of Li-Na countertransport was significantly higher in erythrocytes of hypertensive subjects. This activity was not detected in rat erythrocytes, at two different ages, and over a six-fold of lithium (Li) content. Activities of Na pump were not significantly different among various groups. Human cells, in general, had a higher activity than rat cells; among rat cells, spontaneously hypertensive rats had a higher rate of sodium pump. Both Na-independent Li efflux and ouabain-insensitive Na efflux were significantly higher in rat erythrocytes than in human cells. Thus, employing the same methods, we have determined that while the properties of Na pump were similar in human and rat erythrocytes, Li-Na countertransport did not operate in the erythrocytes of either normotensive or hypertensive rats.
Assuntos
Antiporters , Proteínas de Transporte/fisiologia , Eritrócitos/metabolismo , Hipertensão/sangue , Sódio/sangue , Equilíbrio Hidroeletrolítico/fisiologia , Adulto , Animais , Feminino , Humanos , Hipertensão/genética , Masculino , Fenótipo , Ratos , Ratos Endogâmicos , Fatores de RiscoRESUMO
The rate of erythrocyte Li-Na countertransport and cellular Na+ and K+ contents have been determined in normotensive (NT) and hypertensive (HT, essential hypertension) subjects in the presence and absence of atrial natriuretic peptide (ANP). The rate of Li-Na countertransport was significantly higher in erythrocytes of HT subjects, while the Na+ and K+ contents were not different between the NT and HT groups. We found that ANP (10(-9) and 10(-7) M) had no effect on either the rate of Li-Na countertransport or the cellular Na+ and K+ contents. Since ANP does not influence erythrocyte Na pump and Na-K-Cl cotransport, our results suggest that the Na transport systems of human erythrocytes do not respond to ANP and this lack of response in Li-Na countertransport is independent of the status of hypertension. These findings are consistent with the view that the rate of Li-Na countertransport of erythrocytes may serve as a useful genetic marker for essential hypertension in Chinese. However, the erythrocyte transport systems cannot provide further differentiation utilizing ANP response for essential hypertension.