RESUMEN
Erythrocyte membrane potential was recorded via measurement of pH of the incubation medium in presence ofprothonophore. The increase of intracellular calcium concentration in presence of calcium ionophore A23187 and addition of the artificial redox-system ascorbate-phenazine methosulfate led to membrane hyperpolarization due to opening of Ca(2+)-activated potassium channels that are regulated by multiple signaling pathways. The opening of the Ca(2+)-activated potassium channels in presence of artificial redox-system ascorbate-phenazine methosulfate is mediated at least by two mechanisms including an increase in affinity of channels to calcium ions and involvement of the protein SH-groups and the components of the respiratory circuit which have beer found in erythrocyte membrane.
Asunto(s)
Ácido Ascórbico/metabolismo , Membrana Eritrocítica/metabolismo , Metosulfato de Metilfenazonio/metabolismo , Canales de Potasio Calcio-Activados/metabolismo , Calcimicina/farmacología , Humanos , Ionóforos/farmacología , Potenciales de la Membrana/efectos de los fármacosRESUMEN
Basal and insulin-induced production of NO by monocytes significantly increased in patients with metabolic cardiovascular syndrome. Plasma insulin concentration in these patients was below the control. No intergroup differences were found in C-peptide concentration. A negative correlation was revealed between insulin-induced NO production by monocytes and C-peptide/insulin ratio in patients. The role of monocytes on the regulation of vascular tone via NO production in patients with metabolic cardiovascular syndrome is discussed.
Asunto(s)
Insulina/farmacología , Síndrome Metabólico/metabolismo , Monocitos/metabolismo , Óxido Nítrico/biosíntesis , Humanos , Masculino , Persona de Mediana EdadRESUMEN
Insulin decreased A23187-induced hyperpolarization of the erythrocyte membrane in healthy donors. These data indicate that insulin plays a role in the regulation of Ca(2+)-activated potassium channels in human erythrocytes. However, insulin had little effect on hyperpolarization response of cells induced by artificial ascorbate--phenazine methosulfate donor-acceptor system. Addition of insulin to cell suspension from patients with type 2 diabetes mellitus did not modulate hyperpolarization of the erythrocyte membrane induced by A23187 or ascorbate-phenazine methosulfate, which reflects impairment of regulatory mechanisms for Ca(2+)-activated potassium channels in erythrocytes.
Asunto(s)
Arterias/patología , Calcio/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Eritrocitos/metabolismo , Hipertensión/patología , Insulina/farmacología , Ácido Ascórbico/química , Calcimicina/farmacología , Carbonil Cianuro m-Clorofenil Hidrazona/farmacología , Estudios de Casos y Controles , Femenino , Humanos , Concentración de Iones de Hidrógeno , Insulina/metabolismo , Ionóforos/farmacología , Masculino , Metosulfato de Metilfenazonio/farmacología , Oxidación-ReducciónRESUMEN
Increase in intracellular Ca2+ concentration caused by calcium ionophore A23187 or ascorbate+phenazine methosulphate electron donor system added to erythrocyte suspension induced similar shifts in erythrocyte membrane potential. These processes are most likely mediated by Ca2+-activated potassium channels. Changes in the osmolarity of the incubation medium produced opposite effects on membrane hyperpolarization induced by A23187 or ascorbate+phenazine methosulphate in erythrocyte isolated from healthy donors, which attests to the existence of different mechanisms of regulation of Ca2+-activated potassium channels. There was no difference in the volume-dependent changes of potassium permeability in cells from patients with type II diabetes mellitus combined with arterial hypertension induced by application A23187 or electron-donor system.
Asunto(s)
Diabetes Mellitus Tipo 2/fisiopatología , Eritrocitos/fisiología , Hipertensión/complicaciones , Canales de Potasio Calcio-Activados/fisiología , Calcimicina/farmacología , Diabetes Mellitus Tipo 2/complicaciones , Membrana Eritrocítica/efectos de los fármacos , Humanos , Potenciales de la Membrana/efectos de los fármacos , Concentración OsmolarRESUMEN
We studied the effect of changes in erythrocyte volume and irreversible thermal denaturation of cytoskeleton proteins and lipid matrix on activity of Ca(2+)-activated K+ channels in erythrocytes of alcoholic and patients with II type diabetes mellitus. Changes in Ca(2+)-dependent potassium permeability of erythrocyte membrane in alcoholic patients and patients with II type diabetes mellitus are related to modification of cytoskeleton, rather than to changes in lipid matrix.
Asunto(s)
Alcoholismo/sangre , Calcio/sangre , Proteínas del Citoesqueleto/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Eritrocitos/metabolismo , Lípidos/sangre , Canales de Potasio/metabolismo , Permeabilidad de la Membrana Celular , Diabetes Mellitus Tipo 2/sangre , Membrana Eritrocítica/metabolismo , Volumen de Eritrocitos , Humanos , Concentración de Iones de Hidrógeno , Concentración Osmolar , Desnaturalización ProteicaRESUMEN
Rates of basal and glucose-stimulated insulin and glutathione secretion were studied in experiments with isolated rat pancreas, as were prooxidant effects on these values. The rate of oxidized and recovered glutathione release was found increased at glucose concentration increase to 16.7 mmoles in perfusion solution. Addition of prooxidants (tert-butyl hydroperoxide and Fe2+) in concentrations 10(-4) mole did not change basal insulin secretion but resulted in reduction of glucose-stimulated hormone release. Under such conditions a reduction of the rate of oxidized and recovered glutathione release by the pancreas was observed which was adequate to changed GSH/GSSG ratio in isolated Langerhans' islets. It may be supposed that lipid peroxidation results in changed thiol-disulfide ratio in Langerhans' islets B cells and in reduction of their sensitivity to secretogen effect.
Asunto(s)
Glutatión/metabolismo , Insulina/metabolismo , Peroxidación de Lípido/efectos de los fármacos , Oxidantes/farmacología , Páncreas/metabolismo , Animales , Metabolismo Basal , Compuestos Ferrosos/farmacología , Glucosa/farmacología , Técnicas In Vitro , Secreción de Insulina , Masculino , Perfusión , Peróxidos/farmacología , Ratas , Ratas Wistar , terc-ButilhidroperóxidoRESUMEN
The effect of pro-oxidants (tert-butyl peroxide and FeSO4) on rats perfusing isolated pancreas insulin secretion was studied. Pro-oxidants (10(-4) M) preperfusion during 20 minutes did not change significantly the basal insulin secretion and decreased glucose-stimulated secretion of the one. The content of pancreas TBA-active products increased twice after pro-oxidants effect and ratio of reduced and oxidized glutathiones liberation rates decreased from 2 to 0.5. The disturbance of hormone secretion as a result of decrease of Langerhans islets B-cells reduced glutathione pool is suppose to be one of the mechanisms of hypoinsulinemia development at host's states characterizing with lipids peroxidation activation.
Asunto(s)
Insulina/metabolismo , Islotes Pancreáticos/efectos de los fármacos , Islotes Pancreáticos/metabolismo , Especies Reactivas de Oxígeno/farmacología , Animales , Compuestos Ferrosos/farmacología , Técnicas In Vitro , Secreción de Insulina , Peroxidación de Lípido/efectos de los fármacos , Masculino , Oxidantes/farmacología , Perfusión/métodos , Peróxidos/farmacología , Ratas , Ratas Wistar , Factores de Tiempo , terc-ButilhidroperóxidoRESUMEN
The activity of the Ca-pump in inside-out oriented human erythrocyte membrane vesicles was studied with the use of 45Ca and membrane filters. It was found that trifluoroperazine fully inhibits the calmodulin-induced increase in the maximal activity of the Ca-pump without affecting the calmodulin-stimulated increase in the Ca-pump affinity for Ca2+. The dependence of calcium concentrations of calmodulin-stimulated components of the Ca-pump activity, both inhibited and noninhibited by trifluoroperazine, as well as the dependence on calcium concentrations of the fluorescence intensity of N-phenyl-1-naphthylamine were analyzed. This analysis revealed essential differences between the mechanisms of calmodulin action on the maximal activity of the Ca-pump and its affinity for Ca2+. The maximal activity was elevated by addition of the Ca-calmodulin complex, whereas the increase in the affinity for Ca2+ was induced by calmodulin alone. These findings were supported by data on the dependence of the Ca-pump activity on calmodulin concentrations at low and saturating concentrations of Ca2+ as well as by data obtained in the study on moderate treatment of erythrocyte membranes with trypsin.
Asunto(s)
Calcio/metabolismo , Calmodulina/fisiología , Membrana Eritrocítica/metabolismo , Transporte Biológico/efectos de los fármacos , Colorantes Fluorescentes , Humanos , Técnicas In Vitro , Cinética , Trifluoperazina/farmacología , Tripsina/farmacologíaRESUMEN
The activity of Ca-ATPases of erythrocyte ghosts and sarcoplasmic reticulum and the rate of ATP-dependent uptake of 45Ca by erythrocyte membrane inside-out oriented vesicles, sarcoplasmic reticulum vesicles and mitochondria were investigated. It was found that in all cases studied the addition of EGTA to the incubation mixture caused an increase in the affinity of Ca-pumps for Ca2+; their maximal activity remained thereby unaffected. A similar effect of EGTA was observed when the affinity for Ca2+ of calmodulin, troponin C and the fluorescent dye quin 2 were determined. It was assumed that the effect of EGTA on the affinity for Ca2+ of Ca-transporting and Ca-binding systems of a cell is due to a removal of admixtures of bi- and trivalent cations.
Asunto(s)
Proteínas de Unión al Calcio/metabolismo , ATPasas Transportadoras de Calcio/metabolismo , Calcio/metabolismo , Ácido Egtácico/farmacología , Glicoles de Etileno/farmacología , Animales , Transporte Biológico/efectos de los fármacos , Calmodulina/metabolismo , Membrana Eritrocítica/enzimología , Membrana Eritrocítica/metabolismo , Colorantes Fluorescentes , Técnicas In Vitro , Cinética , Masculino , Mitocondrias Hepáticas/enzimología , Mitocondrias Hepáticas/metabolismo , Conejos , Ratas , Ratas Endogámicas WKY , Retículo Sarcoplasmático/enzimología , Retículo Sarcoplasmático/metabolismo , Troponina/metabolismo , Troponina CRESUMEN
The activity of Ca-pump in inside-out oriented vesicles obtained from erythrocyte membranes after their 30 min treatment with EGTA at 20 degrees C (membranes A) and 37 degrees C (membranes B) was investigated. It was shown that in membranes A placed into an incubation medium containing 0.1 mM EGTA (pH 7.4) the overall effect of exogenous calmodulin is due to the increase in the maximal activity of the enzyme, its affinity for Ca2+ being unaffected thereby. In membranes B placed into the same medium (pH 6.75) the activation of the Ca-pump by calmodulin is due to the increased affinity for Ca2+ at a constant maximal activity of the enzyme. The dependencies of the value of the calmodulin-stimulated component of membranes A and the Ca2+-binding capacity of calmodulin measured by the intensity of N-phenyl-1-naphthylamine fluorescence on the concentration of free Ca2+ are coincident. In the case of membranes B, the stimulation of Ca-pump by calmodulin occurs at much lower Ca2+ concentrations than the Ca2+ binding-induced conformational shifts in calmodulin. The experimental results suggest that the affinity of the Ca-pump for Ca2+ may affect calmodulin existing in a Ca2+-independent state. The hydrophobic interactions between the Ca-calmodulin complex and the Ca-ATPase molecule are apparently essential for the regulation of the maximal enzyme activity.