RESUMEN
BACKGROUND: The endothelial nitric oxide synthase (eNOS) inhibitor asymmetric dimethylarginine (ADMA) is a well-established risk factor for oxidative stress, vascular dysfunction, and congestive heart failure. The aim of the present study was to determine the impact of rapid atrial pacing (RAP) on ADMA levels and eNOS expression. METHODS AND RESULTS: ADMA levels were studied in 60 age- and gender-matched patients. Thirty five patients had persistent atrial fibrillation (AF)≥ 4months. In AF-patients, parameters were studied before and 24h after electrical cardioversion. Moreover, ADMA, eNOS expression, and calcium-handling proteins were studied in pigs subjected to RAP as well as in endothelial cell (EC) cultures. ADMA level was significantly higher in AF compared to sinus rhythm patients (p=0.024). ADMA was highest in AF-patients, who also showed elevated troponin T (TnT) levels. Moreover, ADMA showed a significant linear correlation to TnT (r=0.47; p<0.01). After electrical cardioversion ADMA returned to normal within 24h. In pigs, RAP for 7h increased ADMA levels (p=0.018) and TnI (p<0.05), and reduced mRNA expression of ventricular and aortic eNOS (-80%; p<0.05) compared to sham-control. However, ADMA per se did not affect eNOS mRNA level in EC cultures. CONCLUSION: The current study shows that acute and persistent episodes of atrial tachyarrhythmia are associated with elevated ADMA levels accompanied by increased ischemic myocardial markers. Moreover, RAP increases ADMA and down-regulates eNOS expression in an ADMA-independent manner. We conclude that the combination of these two separate and potentially synergistic mechanisms may contribute to long-term vascular injury during atrial tachyarrhythmia.
Asunto(s)
Arginina/análogos & derivados , Fibrilación Atrial/metabolismo , Endotelio Vascular/metabolismo , Óxido Nítrico Sintasa de Tipo III/sangre , Marcapaso Artificial , Taquicardia/metabolismo , Anciano , Animales , Arginina/sangre , Fibrilación Atrial/epidemiología , Fibrilación Atrial/terapia , Proteínas de Unión al Calcio/metabolismo , Modelos Animales de Enfermedad , Femenino , Frecuencia Cardíaca/fisiología , Células Endoteliales de la Vena Umbilical Humana , Humanos , Masculino , Persona de Mediana Edad , Contracción Miocárdica/fisiología , Estrés Oxidativo/fisiología , Factores de Riesgo , Porcinos , Taquicardia/epidemiología , Taquicardia/terapia , Función Ventricular Izquierda/fisiologíaRESUMEN
OBJECTIVE: Progenitor cells (PC) are thought to induce angiogenesis, and thereby, PC may help to improve ventricular performance in patients with ischemic heart disease (IHD). However, mobilization of progenitor cells by application of G-CSF gives inconsistent clinical effects. The aim of the present study was to assess pathophysiologic effects of progenitor cell mobilization. METHODS AND RESULTS: PC levels (CD34+/CD117+) were counted in 8 patients with severe coronary heart disease and angina pectoris symptoms refractory to conventional therapy during G-CSF treatment (5 µg/kg/d) on days 2, 5, 8, at the end of hospitalization (day 10-12) and after 142±33 days of follow-up. Levels of asymmetric dimethylarginine (ADMA; inhibitor of eNOS) and symmetric dimethylarginine (SDMA) were determined at each occasion and correlated with leukocyte count, systemic nitrite levels, myeloperoxidase (MPO) expression in leukocytes, and urine levels of 8-iso-prostaglandin F2α. Isolated CD34+ cells and endothelial cell cultures were used for functional experiments. G-CSF therapy induced leukocytosis and a rise in CD34+ cell levels. Amounts of MPO positive leukocytes and ADMA levels increased significantly during the treatment phase. ADMA levels correlated to MPO activity (r=0.78; p=0.001) and were inversely related to nitrate levels. In contrast, 8-iso-prostaglandin F2α and amounts of SDMA did not change. Culturing endothelial cells in the presence of myeloperoxidase caused an increase in endothelial ADMA synthesis, which was prevented by application of the antioxidant trolox. CONCLUSIONS: Leukocytosis associated with increased MPO activity during G-CSF therapy appears to be responsible for the systemic release of ADMA, which impairs eNOS activity. Thus, increased MPO and ADMA levels seem to counteract the potential beneficial effects of PC mobilization.
Asunto(s)
Arginina/análogos & derivados , Enfermedad Coronaria/terapia , Factor Estimulante de Colonias de Granulocitos/administración & dosificación , Movilización de Célula Madre Hematopoyética/métodos , Células Madre Hematopoyéticas/citología , Anciano , Anciano de 80 o más Años , Angina de Pecho/metabolismo , Angina de Pecho/fisiopatología , Angina de Pecho/terapia , Antioxidantes/administración & dosificación , Arginina/sangre , Células Cultivadas , Cromanos/administración & dosificación , Enfermedad Coronaria/metabolismo , Enfermedad Coronaria/fisiopatología , Células Endoteliales/citología , Células Endoteliales/efectos de los fármacos , Células Endoteliales/metabolismo , Femenino , Humanos , Leucocitosis/inducido químicamente , Leucocitosis/patología , Masculino , Persona de Mediana Edad , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/fisiología , Peroxidasa/metabolismo , Peroxidasa/farmacología , Venas Umbilicales/citologíaRESUMEN
To investigate the effect of three red wines (RWs) from different growing areas and made from different grapes on asymmetric dimethylarginine (ADMA), an endogenous inhibitor of nitric oxide synthase, in young and senescent human endothelial cells (ECs). All RWs decreased ADMA levels, but 2-fold concentration of German RW was necessary to reach the same effect on ADMA compared to Italian RW and French RW without affecting the cell viability and morphology. The ADMA-lowering effect of RW was increased in senescent compared to young cells, accompanied by enhanced activity of the metabolizing enzyme: dimethylarginine dimethylaminohydrolase (DDAH) II, whereas the same amount in the upregulated protein expression of DDAH II and the downregulated protein expression of the synthesizing enzyme: protein arginine methyltransferase 1 was revealed. These effects were associated with decreased 8-iso-prostaglandin F(2alpha) and peroxynitrite formation, enhanced protein expression of NAD(+)-dependent class III histone deacetylase sirtuin (SIRT) 1, and downregulated protein expression of histone senescence factor p53. Blockade of SIRT1 activity abolished the effect of red wine on ADMA. These data are the first demonstration that RW by activating SIRT1 impairs synthesis and increases metabolism of ADMA. This effect of RW is accentuated in senescent cells probably due to enhanced DDAH activity.
Asunto(s)
Arginina/análogos & derivados , Células Endoteliales/enzimología , Óxido Nítrico Sintasa/metabolismo , Sirtuina 1/biosíntesis , Vino , Factores de Edad , Envejecimiento/metabolismo , Amidohidrolasas/biosíntesis , Arginina/metabolismo , Supervivencia Celular , Células Cultivadas , Senescencia Celular , Humanos , Metiltransferasas/metabolismo , Óxido Nítrico Sintasa/antagonistas & inhibidores , Regulación hacia ArribaRESUMEN
We have recently shown that inhibition of nitric oxide (NO) synthesis by asymmetrical dimethylarginine (ADMA) accelerated endothelial cell (EC) senescence which was prevented by coincubation with L-arginine; however the effect of long-term treatment of l-arginine alone on senescence of ECs have not been investigated. Human ECs were cultured in medium containing different concentrations of L-arginine until senescence. L-Arginine paradoxically accelerated senescence indicated by inhibiting telomerase activity. Moreover, L-arginine decreased NO metabolites, increased peroxynitrite, and 8-iso-prostaglandin F(2alpha) formation. In old cells, the mRNA expression of human amino acid transporter (hCAT)2B, the activity and protein expression of arginase II were upregulated indicated by enhanced urea, L-ornithine, and L-arginine consumption. Inhibition of arginase activity, or transfection with arginase II siRNA prevented L-arginine-accelerated senescence. The most possible explanation for the paradoxical acceleration of senescence by L-arginine so far may be the translational and posttranslational activation of arginase II.
Asunto(s)
Arginina/análogos & derivados , Arginina/metabolismo , Senescencia Celular/fisiología , Células Endoteliales/fisiología , Endotelio Vascular/fisiología , Arginasa/antagonistas & inhibidores , Arginasa/biosíntesis , Arginasa/genética , Arginina/farmacología , Transportador de Aminoácidos Catiónicos 1/biosíntesis , Transportador de Aminoácidos Catiônicos 2/biosíntesis , Células Cultivadas , Senescencia Celular/efectos de los fármacos , Células Endoteliales/efectos de los fármacos , Células Endoteliales/metabolismo , Endotelio Vascular/efectos de los fármacos , Endotelio Vascular/metabolismo , Humanos , Óxido Nítrico/biosíntesis , Estrés OxidativoRESUMEN
The concentration of asymmetric dimethylarginine (ADMA), an endogenous inhibitor of nitric oxide synthase, is increased in patients with coronary heart disease (CHD). The potential effect of percutaneous coronary intervention (PCI) with stent placement on ADMA plasma level in CHD patients has not yet been investigated. Concentrations of ADMA, L-arginine, symmetric dimethylarginine (SDMA), and L-ornithine were measured in the plasma of 30 CHD patients 24 h before, and 1 h, 5 days, and 30 days following PCI with bare-metal stent or drug-eluting stent placement (stent group) and in the plasma of 20 patients without CHD who underwent angiography alone (control group). A repeated measures ANOVA revealed the significant time by group interaction for ADMA (F=12.8, p<0.0001), SDMA (F=5.5, p=0.013), L-ornithine (F=12.5, p<0.0001), L-aginine (F=4.7, p=0.013) and L-arginine/ADMA ratio (F=7.1, p<0.001). Post-hoc ANOVAs showed that this interaction was due to the fact that control patients without stent placement responded to the coronary angiography with a significant increase in ADMA (F=4.4, p=0.009), SDMA (F=4.7, p=0.007) and L-ornithine (F=28.3, p<0.0001) levels, whereas the stent implantation independent of the stent type used significantly reduced the cardiovascular risk factor ADMA (F=10.8, p<0.0001). Thus, the current study demonstrates that in patients with CHD, PCI stent placement markedly decreases the plasma level of cardiovascular risk factor ADMA. Coronary angiography alone results in an increase of ADMA. We conclude that the stent effect on ADMA level cannot be explained by unspecific effects of the coronary angiography and is independent of the stent type used.
Asunto(s)
Arginina/análogos & derivados , Enfermedad Coronaria/sangre , Enfermedad Coronaria/cirugía , Óxido Nítrico Sintasa/antagonistas & inhibidores , Stents , Anciano , Arginina/sangre , Procedimientos Quirúrgicos Cardiovasculares/rehabilitación , Estudios de Casos y Controles , Regulación hacia Abajo , Inhibidores Enzimáticos/sangre , Femenino , Estudios de Seguimiento , Humanos , Masculino , Persona de Mediana Edad , Factores de TiempoRESUMEN
Telmisartan, in addition to blocking angiotensin (Ang) II type 1 receptor (AT(1)R), activates peroxisome proliferator activated receptor gamma (PPARgamma) signaling that interferes with nitric oxide (NO) system. Because aging of endothelial cells (ECs) is hallmarked by a reduction in NO synthesis, we hypothesized that telmisartan increases NO formation by regulated asymmetrical dimethylarginine (ADMA)-dimethylarginine dimethylaminohydrolase (DDAH)-system through blocking AT(1)R and activating PPARgamma signaling. To test this hypothesis, ECs were cultured with telmisartan, eprosartan, Ang II, and GW9662 (PPARgamma antagonist) until the twelfth passage. During the process of aging, PPARgamma protein expression decreased significantly, whereas the expression of AT(1)R increased. Telmisartan reversed these effects and dose-dependently decreased reactive oxygen species and 8-iso-prostaglandin (PG) F(2alpha) formation. This effect was associated with an upregulated activity and protein expression of DDAH, accompanied by a decrease in ADMA concentration, an increase in NO metabolites, and delayed senescence. Blockade of PPARgamma signaling by GW9662 or PPARgamma small-interference RNA prevented the effect of telmisartan on ADMA-DDAH-NO system. Coincubation with Ang II did not affect the effect of telmisartan-delayed senescence, whereas Ang II itself accelerated endothelial aging. Moreover, AT(1)R blocker eprosartan that did not influence PPARgamma protein expression had no effect on ADMA system and senescence. We have demonstrated that telmisartan mainly by activating PPARgamma signaling can alter the catabolism and release of ADMA as an important cardiovascular risk factor. We therefore propose that telmisartan translationally and posttranslationally upregulated DDAH expression via activation of PPARgamma signaling, causing ADMA to diminish and increase NO synthesis sufficient to delay senescence.
Asunto(s)
Bloqueadores del Receptor Tipo 1 de Angiotensina II/farmacología , Arginina/análogos & derivados , Bencimidazoles/farmacología , Benzoatos/farmacología , Senescencia Celular/efectos de los fármacos , Óxido Nítrico/metabolismo , PPAR gamma/fisiología , Receptor de Angiotensina Tipo 1/fisiología , Acrilatos/farmacología , Amidohidrolasas/genética , Amidohidrolasas/metabolismo , Angiotensina II/farmacología , Anilidas/farmacología , Arginina/metabolismo , Células Cultivadas , Senescencia Celular/fisiología , Regulación hacia Abajo/efectos de los fármacos , Endotelio Vascular/citología , Endotelio Vascular/efectos de los fármacos , Endotelio Vascular/fisiología , Humanos , Imidazoles/farmacología , Estrés Oxidativo/efectos de los fármacos , PPAR gamma/antagonistas & inhibidores , PPAR gamma/genética , Receptor de Angiotensina Tipo 1/genética , Transducción de Señal/fisiología , Telmisartán , Tiofenos/farmacología , Regulación hacia Arriba/efectos de los fármacosRESUMEN
Cardiovascular diseases (CVD) are still the most frequent cause of death in Western Europe. Pathophysiological experiments revealed in the last years that the vascular endothelium, as well as a result of the synthesis of nitric oxide (NO), is a crucial regulator of vascular function and homeostasis. The vascular endothelium plays a key role in cardiovascular physiology and pathophysiology, largely via NO-dependent processes. L-Arginine is the substrate for the endothelial NO synthase (eNOS) to generate NO. Endothelial dysfunction is caused by various cardiovascular risk factors. In most studies, acute and chronic administration of L-arginine has been shown to improve endothelial function in animal models of hypercholesterolemia and atherosclerosis. Therefore, numerous studies have been conducted to elucidate whether dietary L-arginine supplementation can augment NO production in humans and thereby improve endothelium-dependent vasodilatation. The most likely mechanism that explains the occurrence of endothelial dysfunction and the effect of L-arginine is that application of L-arginine antagonizes asymmetric dimethylarginine (ADMA), the endogenous NO synthase (NOS) inhibitor. This could solve the L-arginine paradox namely that L-arginine improves NO-mediated vascular function in vivo, although its baseline plasma concentration is about 25- to 30-fold higher than the Michaelis-Menten constant Km of the isolated, purified eNOS in vitro. Recent findings suggest that large, prospective, randomized clinical trials might be needed to identify those patients who are the most likely to benefit from L-arginine. Testing patients for ADMA and L-arginine plasma levels for calculating the L-arginine/ADMA ratio might be an adequate strategy.
Asunto(s)
Arginina/análogos & derivados , Animales , Arginina/metabolismo , Arginina/fisiología , Suplementos Dietéticos , Humanos , Infarto del Miocardio/fisiopatología , Óxido Nítrico Sintasa de Tipo III/fisiologíaRESUMEN
BACKGROUND AND PURPOSE: Preclinical studies have revealed that the endogenous nitric oxide synthase inhibitor, asymmetric dimethylarginine (ADMA), increases vascular tone in cerebral blood vessels. Marked elevations of ADMA blood levels were found in patients with diseases characterized by decreased cerebral perfusion, such as ischemic stroke. Arterial stiffness is an independent predictor of stroke and other adverse cardiovascular events. The aim of this study was to investigate the influence of a systemic subpressor dose of ADMA on arterial stiffness and cerebral perfusion in humans. METHODS: Using a double-blind, vehicle-controlled study design, we allocated 20 healthy men in random order to infusion of either ADMA (0.10 mg ADMA/kg per min) or vehicle over a period of 40 minutes. Arterial stiffness was assessed noninvasively by pulse wave analysis. All volunteers underwent measurement of cerebral perfusion by dynamic contrast-enhanced perfusion magnetic resonance imaging of the brain. RESULTS: Infusion of ADMA significantly decreased total cerebral perfusion by 15.1+/-4.5% (P=0.007), whereas blood flow in the vehicle group increased by 7.7+/-2.8% (P=0.02). ADMA also increased arterial stiffness as assessed by measurement of the augmentation index (-12.6+/-1.9 to -9.6+/-1.5, P=0.007). CONCLUSIONS: Our results document for the first time that subpressor doses of ADMA increase vascular stiffness and decrease cerebral perfusion in healthy subjects. Thus, ADMA is an important endogenous modulator of cerebral vascular tone and may be involved in the pathogenesis of cerebrovascular disease.
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Arginina/análogos & derivados , Arterias Cerebrales/efectos de los fármacos , Arterias Cerebrales/fisiología , Circulación Cerebrovascular/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Sistema Vasomotor/efectos de los fármacos , Adulto , Arginina/administración & dosificación , Arginina/farmacología , Adaptabilidad , Relación Dosis-Respuesta a Droga , Método Doble Ciego , Humanos , Inyecciones Intravenosas , Imagen por Resonancia Magnética , Masculino , Óxido Nítrico Sintasa/antagonistas & inhibidores , Valores de ReferenciaRESUMEN
We investigated here the effect of l-arginine on asymmetric dimethylarginine (ADMA) or homocysteine-accelerated endothelial aging. Endothelial cells were cultured in medium containing 70micromol/L arginine until fourteenth passage. ADMA, dl-homocysteine, and l-arginine were replaced every 48h starting at the fourth passage. ADMA or homocysteine inhibited significantly the population doublings (PD) and accelerated the process of aging. Co-incubation with l-arginine enhanced PD, inhibited senescence associated beta-galactosidase activity, and increased telomerase activity. This effect was associated with an increase in NO synthesis and NO synthase protein expression. Furthermore, l-arginine-induced NO formation was accompanied by a reduction in oxidative stress and an increase in protein expression and enzyme activity of heme oxygenase (HO)-1. The NO synthase inhibitor l-NAME completely abolished the effect of l-arginine on ADMA or homocysteine-accelerated aging. These findings demonstrate that l-arginine prevents the onset of endothelial aging in ADMA or homocysteine-treated cells by increasing NO formation and consequently the induction of HO-1. This might provide a new strategy to delay ADMA or homocysteine-accelerated aging.
Asunto(s)
Arginina/análogos & derivados , Arginina/farmacología , Células Endoteliales/metabolismo , Homocisteína/metabolismo , Envejecimiento , Arginina/metabolismo , Células Cultivadas , Senescencia Celular , Endotelio Vascular/citología , Humanos , NG-Nitroarginina Metil Éster/farmacología , Estrés Oxidativo , Telomerasa/metabolismo , Factores de TiempoRESUMEN
Symmetrical dimethylarginine (SDMA) is the structural isomer of the endogenous nitric oxide synthase (NOS) inhibitor asymmetric dimethylarginine. Whereas the major route of asymmetric dimethylarginine elimination is the hydrolytic degradation by dimethylarginine dimethylaminohydrolase, SDMA is eliminated by renal excretion. SDMA does not directly inhibit NOS but is a competitor of arginine transport. This study showed for the first time that measurement of SDMA can be a marker of estimated GFR and extent of coronary artery disease (CAD). In 97 patients with CAD, SDMA was a marker of estimated GFR. On multiple regression analysis of the CAD parameter stenosis score, SDMA was the only parameter retained. In addition, endothelial cells from the third passage were cultured in medium that contained 70 micromol/L arginine and was incubated for 24 h in the presence of various concentration of SDMA (0, 2, 5, 10, and 100 micromol/L). The levels of nitrate and nitrite in conditioned media, the protein expression of NOS, and the content of reactive oxygen species in endothelial cells were determined. SDMA inhibited dose dependently the NO synthesis in intact endothelial cells, whereas it had no effect on protein expression of NOS. This effect was associated with an increase in reactive oxygen species. Co-incubation with L-arginine but not D-arginine reversed the effect of SDMA on NOS pathway. Our data suggest that SDMA reduced the endothelial NO synthesis, probably by limiting L-arginine supply to NOS. It is concluded that SDMA might be a useful parameter for detecting patients in very early stages of chronic kidney disease and for determining their risk for developing cardiovascular disease.
Asunto(s)
Arginina/análogos & derivados , Enfermedad de la Arteria Coronaria/diagnóstico , Pruebas de Función Renal/métodos , Anciano , Arginina/sangre , Arginina/química , Arginina/metabolismo , Biomarcadores/sangre , Biomarcadores/química , Biomarcadores/metabolismo , Estudios de Casos y Controles , Células Cultivadas , Enfermedad de la Arteria Coronaria/metabolismo , Células Endoteliales/efectos de los fármacos , Células Endoteliales/metabolismo , Inhibidores Enzimáticos/sangre , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/metabolismo , Humanos , Isomerismo , Fallo Renal Crónico/diagnóstico , Fallo Renal Crónico/metabolismo , Persona de Mediana Edad , Óxido Nítrico/biosíntesis , Óxido Nítrico Sintasa de Tipo III/antagonistas & inhibidores , Factores de RiesgoRESUMEN
We report here the effect of aspirin on the onset of replicative senescence. Endothelial cells that were cultured until cumulative population doublings 40 showed clear signs of aging. Incubation with aspirin inhibited senescence-associated beta-galactosidase activity and increased telomerase activity. Along with the delayed onset of senescence, aspirin decreased reactive oxygen species and increased nitric oxide (NO) and cGMP levels. Furthermore, aspirin reduced the elaboration of asymmetric dimethylarginine (ADMA), an endogenous inhibitor of NO synthase, and up-regulated the activity of dimethylarginine dimethylaminohydrolase, the enzyme that degrades ADMA. These effects were specific in that other nonsteroidal anti-inflammatory drugs, such as ibuprofen or acetaminophen, did not prevent the onset of endothelial senescence. The NO synthase inhibitor l-NAME, but not its inactive d-enantiomer, led to complete inhibition of aspirin-delayed senescence. These findings demonstrate that aspirin delays the onset of endothelial senescence by preventing a decrease in NO formation/generation. This might provide a therapeutic strategy aimed at blocking aging-induced NO inhibition.
Asunto(s)
Aspirina/administración & dosificación , Senescencia Celular/fisiología , GMP Cíclico/metabolismo , Células Endoteliales/citología , Células Endoteliales/metabolismo , Óxido Nítrico/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Células Cultivadas , Senescencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Células Endoteliales/efectos de los fármacos , Humanos , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/fisiologíaRESUMEN
The present study was designed to examine the effects of ET-1 (endothelin-1) and serum from PE (pre-eclamptic), HP (healthy pregnant) and HNP (healthy non-pregnant) women on uterine arterial perfusion pressure and uterine contractility. Swine uteri (n = 25) were perfused for a period of up to 11 h, with the aim being to preserve a viable organ. Various concentrations of ET-1 as well as serum from PE, HP and HNP women (n = 10 per group) were administered to the perfused swine uteri and IUP (intrauterine pressure) and IAP (intra-arterial pressure) were recorded. ET-1 produced dose-dependent increases in IUP and IAP. The ET-1 concentration in serum was higher in serum from PE women than in HP and HNP women (P > 0.05). Administration of all serum samples had a contractile effect on the swine uterus, with the greatest effect being seen in HNP women (12.8 +/- 5.3 mmHg), followed by PE (9.06 +/- 4.2 mmHg) and HP (6.1 +/- 4.1 mmHg) women. Statistically significant differences were observed between HNP and PE women (P = 0.048), and PE and HP women (P = 0.021). Increases in IAP following administration of serum from PE women (48.8 +/- 20.0 mmHg) were significantly higher (P = 0.024) compared with the effect of serum from HP women (28.4 +/- 13.7 mmHg). In conclusion, the findings show that serum from PE women has significant vasoconstrictive and oxytocic effects compared with serum from HP women. In pre-eclampsia, the balance between vasorelaxing and vasoactive substances is disturbed.
Asunto(s)
Presión Sanguínea/efectos de los fármacos , Endotelina-1/farmacología , Preeclampsia/sangre , Contracción Uterina/efectos de los fármacos , Útero/fisiopatología , Sistema Vasomotor/fisiopatología , Animales , Arterias , Femenino , Humanos , Técnicas de Cultivo de Órganos , Perfusión , Preeclampsia/fisiopatología , Embarazo , Porcinos , Útero/irrigación sanguíneaRESUMEN
Recombinant human erythropoietin therapy frequently causes hypertension in humans and animals with chronic renal failure. Asymmetric dimethylarginine (ADMA) is an endogenous inhibitor of nitric oxide (NO) synthase, and its accumulation has been associated with reducing NO bioavailability and increasing superoxide generation. Whether epoetin beta (EPO) or darbepoetin alpha (NESP) can modify the levels of ADMA in endothelial cells was investigated. Endothelial cells from the third passage were incubated for 24 h in the presence of various concentrations of EPO or NESP (0, 0.1, 1, 10, 50, 100, and 200 U/ml). The levels of ADMA, allantoin, nitrate, and nitrite in conditioned media and the activity of dimethylarginine dimethylaminohydrolase (DDAH), the content of thiols and reactive oxygen species in endothelial cells, were determined. When endothelial cells were exposed to EPO or NESP, ADMA concentration in the cell culture medium increased significantly in a dose-dependent manner versus control. This effect was associated with a reduced activity of DDAH, the enzyme that degrades ADMA. Furthermore, EPO- or NESP-induced accumulation of ADMA was accompanied by a significant reduction of NO synthesis and an increase in oxidative stress. Both allantoin, a marker of oxygen free radical generation, and reactive oxygen species increased significantly after EPO or NESP treatment compared with control. The antioxidant pyrrolidine dithiocarbamate preserved DDAH activity and reduced ADMA accumulation in the same way as the co-incubation with anti-EPO neutralizing antibody. EPO and NESP posttranslationally impair DDAH activity via increased oxidative stress, causing ADMA as an important cardiovascular risk factor to accumulate and inhibit NO synthesis.
Asunto(s)
Amidohidrolasas/fisiología , Arginina/análogos & derivados , Células Endoteliales/efectos de los fármacos , Células Endoteliales/metabolismo , Eritropoyetina/análogos & derivados , Eritropoyetina/farmacología , Estrés Oxidativo/efectos de los fármacos , Amidohidrolasas/antagonistas & inhibidores , Antioxidantes/farmacología , Arginina/metabolismo , Células Cultivadas , Darbepoetina alfa , Humanos , Membranas Intracelulares/metabolismo , Óxido Nítrico/antagonistas & inhibidores , Óxido Nítrico/biosíntesis , Pirrolidinas/farmacología , Proteínas Recombinantes , Tiocarbamatos/farmacologíaRESUMEN
Asymmetric dimethylarginine (ADMA) is an endogenous inhibitor of nitric oxide synthase and its accumulation has been associated with cardiovascular disease. We aimed to investigate the role of ADMA in endothelial cell senescence. Endothelial cells were cultured until the tenth passage. ADMA was replaced every 48 hours starting at the fourth passage. ADMA significantly accelerated senescence-associated beta-galactosidase activity. Additionally, the shortening of telomere length was significantly speeded up and telomerase activity was significantly reduced. This effect was associated with an increase of oxidative stress: both allantoin, a marker of oxygen free radical generation, and intracellular reactive oxygen species increased significantly after ADMA treatment compared with control, whereas nitric oxide synthesis decreased. Furthermore, ADMA-increased oxidative stress was accompanied by a decrease in the activity of dimethylarginine dimethylaminohydrolase, the enzyme that degrades ADMA, which could be prevented by the antioxidant pyrrolidine dithiocarbamate. Exogenous ADMA also stimulated secretion of monocyte chemotactic protein-1 and interleukin-8. Co-incubation with the methyltransferase inhibitor S-adenosylhomocysteine abolished the effects of ADMA. These data suggest that ADMA accelerates senescence, probably via increased oxygen radical formation by inhibiting nitric oxide elaboration. This study provides evidence that modest changes of intracellular ADMA levels are associated with significant effects on slowing down endothelial senescence.
Asunto(s)
Arginina/análogos & derivados , Senescencia Celular , Células Endoteliales/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Amidohidrolasas/metabolismo , Arginina/farmacología , Células Cultivadas , Quimiocina CCL2 , Relación Dosis-Respuesta a Droga , Células Endoteliales/enzimología , Humanos , Interleucina-8/metabolismo , Óxido Nítrico Sintasa/antagonistas & inhibidores , Estrés Oxidativo , Especies Reactivas de Oxígeno/metabolismo , S-Adenosilhomocisteína/farmacología , Telomerasa/antagonistas & inhibidores , Telomerasa/metabolismoRESUMEN
OBJECTIVE: Asymmetrical dimethylarginine (ADMA) is an endogenous inhibitor of nitric oxide synthase (NOS), and its accumulation has been associated with cardiovascular disease. We aimed to investigate the role of ADMA in endothelial cell senescence. METHODS AND RESULTS: Endothelial cells were cultured until the tenth passage. ADMA was replaced every 48 hours starting at the fourth passage. ADMA significantly accelerated senescence associated beta-galactosidase activity. Additionally, the shortening of telomere length was significantly accelerated and the telomerase activity was significantly reduced. This effect was associated with an increase of oxidative stress: allantoin, a marker of oxygen free radical generation, and intracellular reactive oxygen species (ROS) increased significantly after ADMA treatment compared with control, whereas cellular thiol status and NOx synthesis decreased. Furthermore, ADMA-increased oxidative stress was accompanied by a decrease in the activity of dimethylarginine dimethylaminohydrolase (DDAH), the enzyme that degrades ADMA, which could be prevented by the antioxidant pyrrolidine dithiocarbamate. Exogenous ADMA also stimulated secretion of MCP-1 and interleukin-8. Coincubation with the methyltransferase inhibitor S-adenosylhomocysteine abolished the effects of ADMA. CONCLUSIONS: These data suggest that ADMA accelerates senescence, probably via increased oxygen radical formation by inhibiting nitric oxide elaboration. This study provides evidence that modest changes of intracellular ADMA levels are associated with significant effects on slowing endothelial senescence.
Asunto(s)
Arginina/análogos & derivados , Arginina/farmacología , Senescencia Celular/efectos de los fármacos , Células Endoteliales/efectos de los fármacos , Óxido Nítrico Sintasa/antagonistas & inhibidores , Alantoína/metabolismo , Amidohidrolasas/metabolismo , Células Cultivadas , Quimiocina CCL2/metabolismo , Células Endoteliales/metabolismo , Inhibidores Enzimáticos/farmacología , Citometría de Flujo/métodos , Humanos , Inmunofenotipificación/métodos , Interleucina-8/metabolismo , Estrés Oxidativo/efectos de los fármacos , S-Adenosilhomocisteína/farmacología , Telomerasa/metabolismo , Venas Umbilicales/química , Venas Umbilicales/citología , Venas Umbilicales/enzimología , Venas Umbilicales/metabolismoRESUMEN
OBJECTIVES: The purpose of this study was to examine neurokinin B levels in serum from preeclamptic and normotensive and to investigate the role of neurokinin B in preeclampsia. STUDY DESIGN: Peripheral and uterine venous blood neurokinin B levels were measured in 14 normotensive and 8 preeclamptic pregnant women by radioimmunoassay. RESULTS: Neurokinin B levels in normotensive women were 4.91 +/- 2.67 nmol/L in peripheral and 5.59 +/- 2.06 nmol/L in uterine blood. In pregnant women with preeclampsia, neurokinin B levels were 2.79 +/- 1.68 nmol/L and 3.20 +/- 1.55 nmol/L, respectively. Neurokinin B levels were significantly higher in normotensive women (P=.032 in peripheral and P=.006 in uterine blood). CONCLUSIONS: Neurokinin B serum levels were higher in normotensive women. Higher neurokinin B concentrations in normotensive pregnant women may be due to the advanced gestational age and/or the result of a negative interaction of other vasoactive substances. The role of neurokinin B in preeclampsia remains to be determined.
Asunto(s)
Neuroquinina B/sangre , Preeclampsia/sangre , Adulto , Estudios de Casos y Controles , Femenino , Edad Gestacional , Síndrome HELLP/sangre , Humanos , Concentración Osmolar , Embarazo , Útero/irrigación sanguíneaRESUMEN
This study was conducted to compare the effects of serum from healthy pregnant women and that from pregnant women with pre-eclampsia on oxidative stress in endothelial cells in culture. Human umbilical vein endothelial cells (HUVECs) were incubated with serum from 18 pre-eclamptic, 18 healthy pregnant and 18 healthy non-pregnant women for 24 h. The levels of reduced glutathione (GSH) and lipid peroxides (LPOs) were measured in endothelial cell lysates. Measurement of malondialdehyde in combination with 4-hydroxyalkenals has been used as an indicator of LPOs. Serum from healthy pregnant women decreased significantly the LPO content in HUVECs in comparison with serum from pre-eclamptic women and healthy non-pregnant women (30.7+/-6.6 compared with 39.3+/-10.9 and 41.0+/-12.7 pmol/mg of protein respectively; P<0.003 and P<0.01 respectively). No differences in GSH content between the three groups (18.3+/-2.1 nmol/mg of protein for healthy pregnant, 19.2+/-3.3 nmol/mg for pre-eclamptic and 18.3+/-2.0 nmol/mg for healthy non-pregnant women) were found. Thus serum from normal pregnant women contains a factor(s) that decreases oxidative stress in human endothelial cells. This mechanism might be altered in pre-eclampsia.