RESUMEN
Nucleotide metabolism plays a major role in a number of vital cellular processes such as energetics. This, in turn, is important in pathologies such as atherosclerosis. Three month old atherosclerotic mice with knock outs for LDLR and apolipoprotein E (ApoE) were used for the experiments. Activities of AMP-deaminase (AMPD), ecto5'-nucleotidase (e5NT), adenosine deaminase (ADA), purine nucleoside phosphorylase (PNP) were measured in heart, liver and kidney cortex and medulla by analysing conversion of substrates into products using HPLC. The activity of ecto5'-nucleotidase differ in hearts of LDLR-/- and ApoE-/- mice with no differences in ADA and AMPD activity. We noticed highest activity of e5NT in kidney medulla of the models. This model of atherosclerosis characterize with an inhibition of enzyme responsible for production of protective adenosine in heart but not in other organs and different metabolism of nucleotides in kidney medulla.
Asunto(s)
Apolipoproteínas E/genética , Riñón/enzimología , Hígado/enzimología , Miocardio/enzimología , Nucleótidos de Purina/metabolismo , Receptores de LDL/genética , Adenosina/metabolismo , Animales , Apolipoproteínas E/metabolismo , Humanos , Riñón/metabolismo , Hígado/metabolismo , Ratones Endogámicos C57BL , Ratones Noqueados , Miocardio/metabolismo , Receptores de LDL/metabolismoRESUMEN
Little is known about the effects of coffee that are not related to the presence of caffeine. The aim of the study was to analyse changes in kidney function and nucleotide metabolism related to high intake of decaffeinated coffee. Mice consumed decaffeinated coffee extract for two weeks. Activities of AMP deaminase, ecto5'-nucleotidase, adenosine deaminase, purine nucleoside phosphorylase were measured in kidney cortex and medulla by analysis of conversion of substrates into products using HPLC. Concentration of nucleotides in kidney cortex, kidney medulla and serum were estimated by HPLC. Activity of ecto5'-nucleotidase increased from 0.032 ± 0.006 to 0.049 ± 0.014 nmol/mg tissue/min in kidney cortex of mice administered high-dose decaffeinated coffee (HDC) together with increase in cortex adenosine concentration and decrease in plasma creatinine concentration. HDC leads to increased activity of ecto5'-nucleotidase in kidney cortex that translates to increase in concentration of adenosine. Surprisingly this caused improved kidney excretion function.
Asunto(s)
Café , Corteza Renal/metabolismo , Médula Renal/metabolismo , Nucleótidos/metabolismo , Animales , RatonesRESUMEN
During pregnancy the isoform composition of human placental AMP-deaminase changes. This may reflect the adaptation of enzyme to changing metabolic requirements of the growing fetus. In this paper kinetic and regulatory properties of AMP-deaminase purified from human preterm (â¼ 25 week of gestation) placenta were described and compared with these of the enzyme purified from term placenta. AMP-deaminase from preterm placenta was less sensitive to pH changes and in contrast to the enzyme from the term organ, at low range of substrate concentrations was not inhibited but activated by physiological concentrations of orthophosphate. This may significantly improve the catalytic efficiency of enzyme at early phase of the pregnancy.
Asunto(s)
AMP Desaminasa/metabolismo , Placenta/enzimología , Segundo Trimestre del Embarazo/fisiología , AMP Desaminasa/antagonistas & inhibidores , Femenino , Humanos , Concentración de Iones de Hidrógeno , Recién Nacido , Recien Nacido Prematuro , Isoenzimas/metabolismo , Cinética , Fosfatos/farmacología , Embarazo , Tercer Trimestre del EmbarazoRESUMEN
A C34T mutation in the AMPD1 gene is proposed to cause local or systemic augmentations in blood adenosine level and improvement of prognoses in heart diseases like congestive heart failure or heart ischemic disease. This study examines some physico-chemical properties of AMP-deaminase isolated from cardiac muscle of a 10-year-old boy heterozygote for this mutation.
Asunto(s)
AMP Desaminasa/genética , Miocardio/enzimología , AMP Desaminasa/metabolismo , Niño , Heterocigoto , Humanos , Masculino , MutaciónRESUMEN
During pregnancy the activity of AMP-deaminase in developing human placenta gradually decreases, being in homogenates of mature, term placenta (approximately 40 week of gestation) one fourth to one third of that in homogenates of immature (approximately 25 week of gestation) organ. The gradual decrease of activity correlates inversely with the increasing presence of the form of enzyme predominating in homogenates of the mature placenta. The discrepancy observed indicate that isozymic pattern of AMP-deaminase in developing human placenta changes.
Asunto(s)
AMP Desaminasa/metabolismo , Placenta/enzimología , Adulto , Western Blotting , Electroforesis en Gel de Poliacrilamida , Femenino , Edad Gestacional , Humanos , Placentación , EmbarazoRESUMEN
A short chain synthetic analogue of lipid hydroperoxides was used to overload glutathione peroxidase (GPx) in human choriocarcinoma cell line JAR cells. Cells exposed to 100 microM tBuOOH displayed a 40% reduction in ATP level and significantly increased in membrane permeability, visualised by the lactate dehydrogenase (LDH) release into the extracellular medium. The intracellular level of oxygen free radicals measured as an oxidation of the dichlorodihydro-fluorescein diacetate (H2DCF-DA) significantly increased after 2 hours of cell exposition to 100 microM tBuOOH. Concomitantly MDA, 4-HNE level increased to 2 nmol/mg of cell protein after 2 hours. Mitochondria stained with MitoTracker Red CMXRos displayed a filamentous appearance in control cells but changed into granular less energised organelles after exposition to tBuOOH. Collectively, the above results indicate the importance of the contribution of oxidative stress in the development of pre-eclampsia.
Asunto(s)
Línea Celular Tumoral/efectos de los fármacos , Glutatión Peroxidasa/deficiencia , terc-Butilhidroperóxido/farmacología , Adenosina Trifosfato/metabolismo , Línea Celular Tumoral/metabolismo , Forma de la Célula , Coriocarcinoma , Radicales Libres/metabolismo , Humanos , L-Lactato Deshidrogenasa/metabolismo , Peroxidación de Lípido , Peróxidos Lipídicos/química , Mitocondrias/metabolismo , Mitocondrias/ultraestructura , Oxidación-Reducción , Especies Reactivas de Oxígeno/metabolismoRESUMEN
The effects of ascorbate and alpha-tocopherol as antioxidants and as co-operative factors against NADPH-dependent lipid peroxidation in human placental mitochondria have been studied. The addition of ascorbate at low concentration (up to 50 microM) to the NADPH-generating system resulted in increasing lipid peroxidation and Fe3+ to Fe2+ reduction. High concentration of ascorbate (150 microM), which produced maximal rate of ascorbate-dependent lipid peroxidation, was found to inhibit almost completely NADPH-dependent lipid peroxidation by maintaining too much iron in its reduced form. Either stimulatory or inhibitory effect of ascorbate on NADPH-dependent lipid peroxidation depends on the appropriate Fe3+/Fe2+ ratio. Alpha-tocopherol caused a decrease of NADPH-dependent lipid peroxidation, inhibiting completely this process at 150 microM concentration. The inhibitory effect of alpha-tocopherol increased rapidly with the increasing ascorbate concentration, almost complete inhibition of NADPH-dependent lipid peroxidation being obtained at 25 microM alpha-tocopherol and 50 microM ascorbate. This strong inhibitory combined effect of alpha-tocopherol and ascorbate was independent of the Fe3+/Fe2+ ratio, as alpha-tocopherol is not able to reduce Fe3+ to Fe2+ under the conditions employed. These findings suggest that antioxidant effects of ascorbate in placental mitochondria are mediated by recycling of alpha-tocopherol rather than by strong reduction of Fe3+ to Fe2+. On the basis of the results obtained, we assume that adequate concentrations of alpha-tocopherol and ascorbate in placental tissue may prevent the release of lipid peroxide from placental mitochondria and therefore could be protective against the development of preeclampsia.