RESUMEN
Changes in the ecto-5'-nucleotidase activity-an extracellular nucleotide catabolic enzyme may lead to the inflammation and endothelial dysfunction. We investigated the effect of CD73 deletion on the endothelial function and L-arginine metabolism in various age groups of mice. 1-,3-,6-, and 12-month-old, male C57BL/6 J wild type (WT) and C57BL/6 J CD73-/- (CD73-/-) mice were used. Blood samples were used for the analysis of adenine nucleotide concentrations. Serum samples were analyzed for the concentration of amino acids, Interleukin 6 (IL-6), Intercellular Adhesion Molecule 1 (ICAM-1), Vascular Cell Adhesion Molecule 1 (VCAM-1), and endothelial nitric oxide synthase (eNOS) level. Serum and aortic nitrate/nitrite, as well as aortic arginase and NOS activity in endothelial cells (EC) were evaluated. CD73 deletion led to age-dependent increase in IL-6, ICAM-1, and VCAM-1 concentration compared to WT. All CD73-/- mice age groups were characterized by reduced L-Arginine concentration and eNOS level. Significantly lower NOS activity was noticed in EC isolated from CD73-/- mice lungs in comparison to EC isolated from WT lungs. The L-Arginine/ADMA ratio in the CD73-/- decreased in age-dependent manner in comparison to WT. The nitrate/nitrite ratio was reduced in serum and in aortas of 6-month-old CD73-/- mice as compared to WT. The ornithine/arginine and ornithine/citrulline ratios were increased in CD73-/- compared to controls. Blood (erythrocyte) Adenosine-5'-triphosphate and Adenosine-5'-diphosphate levels were reduced in favor to higher blood Adenosine-5'-monophosphate concentration in CD73-/- mice in comparison to WT. The CD73 deletion leads to the development of age-dependent endothelial dysfunction in mice, associated with impaired L-arginine metabolism. CD73 activity seems to protect endothelium.
Asunto(s)
5'-Nucleotidasa/deficiencia , Arginina/sangre , Endotelio Vascular/metabolismo , Adenosina Difosfato/sangre , Adenosina Difosfato/genética , Adenosina Trifosfato/sangre , Adenosina Trifosfato/genética , Animales , Arginina/genética , Endotelio Vascular/patología , Molécula 1 de Adhesión Intercelular/sangre , Molécula 1 de Adhesión Intercelular/genética , Interleucina-6/sangre , Interleucina-6/genética , Ratones , Ratones Noqueados , Óxido Nítrico Sintasa de Tipo III/sangre , Óxido Nítrico Sintasa de Tipo III/genética , Molécula 1 de Adhesión Celular Vascular/sangre , Molécula 1 de Adhesión Celular Vascular/genéticaRESUMEN
Aortic stenosis is known to involve inflammation and thrombosis. Changes in activity of extracellular enzyme - ecto-5'-nucleotidase (referred also as CD73) can alter inflammatory and thrombotic responses. This study aimed to evaluate the effect of CD73 deletion in mice on development of aortic valve dysfunction and to compare it to the effect of high-fat diet. Four groups of mice (normal-diet Wild Type (WT), high-fat diet WT, normal diet CD73-/-, high-fat diet CD73-/-) were maintained for 15weeks followed by echocardiographic analysis of aortic valve function, measurement of aortic surface activities of nucleotide catabolism enzymes as well as alkaline phosphatase activity, mineral composition and histology of aortic valve leaflets. CD73-/- knock out led to an increase in peak aortic flow (1.06±0.26m/s) compared to WT (0.79±0.26m/s) indicating obstruction. Highest values of peak aortic flow (1.26±0.31m/s) were observed in high-fat diet CD73-/- mice. Histological analysis showed morphological changes in CD73-/- including thickening and accumulation of dark deposits, proved to be melanin. Concentrations of Ca2+, Mg2+ and PO43- in valve leaflets were elevated in CD73-/- mice. Alkaline phosphatase (ALP) activity was enhanced after ATP treatment and reduced after adenosine treatment in aortas incubated in osteogenic medium. AMP hydrolysis in CD73-/- was below 10% of WT. Activity of ecto-adenosine deaminase (eADA), responsible for adenosine deamination, in the CD73-/- was 40% lower when compared to WT. Deletion of CD73 in mice leads to aortic valve dysfunction similar to that induced by high-fat diet suggesting important role of this surface protein in maintaining heart valve integrity.
Asunto(s)
5'-Nucleotidasa/deficiencia , Aorta , Válvula Aórtica , Eliminación de Gen , Enfermedades de las Válvulas Cardíacas , Animales , Aorta/metabolismo , Aorta/patología , Aorta/fisiopatología , Válvula Aórtica/metabolismo , Válvula Aórtica/patología , Válvula Aórtica/fisiopatología , Grasas de la Dieta/efectos adversos , Grasas de la Dieta/farmacología , Enfermedades de las Válvulas Cardíacas/genética , Enfermedades de las Válvulas Cardíacas/metabolismo , Enfermedades de las Válvulas Cardíacas/patología , Enfermedades de las Válvulas Cardíacas/fisiopatología , Ratones , Ratones NoqueadosRESUMEN
Obestatin is a 23 amino acid peptide derived from the preproghrelin precursor, and originally purified from the rat stomach mucosa. It was shown that obestatin may counteract the effects of its sister peptide, ghrelin, on food intake and gastrointestinal motility but the other roles in controlling the gastrointestinal function remain unknown. The aim of the present study was to determine the influence of exogenous obestatin on the secretion of pancreatic juice. In anesthetized male Wistar rats the external jugular vein was catheterized, and the common biliary-pancreatic duct was cannulated with polyethylene tubing for collection of pancreatic-biliary juice (P-BJ). Obestatin boluses (30, 100 and 300 nmol/kg b. wt.) were injected intravenously or intraduodenally every 30 min. Obestatin was also administered in vagotomized (subdiaphragmatic vagotomy) rats. In the examined rats, obestatin intravenous and intraduodenal boluses did not affect the P-BJ volume. On the other hand, obestatin boluses increased the protein output and trypsin activity. Vagotomy abolished the effects of exogenous obestatin administration. In conclusion, the present study demonstrates for the first time that exogenous obestatin may stimulate the secretion of pancreatic juice enzymes. The effect is dose-dependent and requires intact vagal supply.