RESUMEN
We study the wetting critical behavior of the three-state (s=±1,0) Blume-Emery-Griffiths model using numerical simulations. This model provides a suitable scenario for the study of the role of vacancies on the wetting behavior of a thin magnetic film. To this aim we study a system confined between parallel walls with competitive short-range surface magnetic fields (h_{L}=-|h_{1}|). We locate relevant critical curves for different values of the biquadratic interaction and use a thermodynamic integration method to calculate the surface tension as well as the interfacial excess energy and determine the wetting transition. Furthermore, we also calculate the local position of the interface along the film and its fluctuations (capillary waves), which are a measure of the interface width. To characterize the role played by vacancies on the interfacial behavior we evaluate the excess density of vacancies, i.e., the density difference between a system with and without interface. We also show that the temperature dependence of both the local position of the interface and its width can be rationalized in term of a finite-size scaling description, and we propose and successfully test the same scaling behavior for the average position of the center of mass of the vacancies and its fluctuations. This shows that the excess of vacancies can be associated to the presence of the interface that causes the observed segregation. This segregation phenomena is also evidenced by explicitly evaluating the interfacial free energy.
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Circulating auto-antibodies against cytochrome P4502E1 (CYP2E1) have been observed in a significant fraction of patients with chronic hepatitis C (CHC). This study investigated the clinical significance of these auto-antibodies in relation to their antigen specificity. The presence of anti-CYP2E1 IgG was investigated in 137 consecutive patients with biopsy-proven CHC. Anti-CYP2E1 IgG above control threshold levels was detected in 52 (38%) subjects. By combined immunoprecipitation and western blotting, we observed that among anti-CYP2E1 IgG-positive sera, 23 (44%) were unreactive towards denaturated CYP2E1, indicating a prevalent recognition of conformational CYP2E1 antigens. Conformational anti-CYP2E1 auto-antibodies were unrelated to circulating gamma-globulins, alcohol intake or infection by specific HCV genotypes. The presence of anti-CYP2E1 auto-antibodies was associated with an 11-fold (OR 10.9 95%CI 1.4-86.6 P = 0.008) increased prevalence of necro-inflammatory grading ≥ 4 (Ishack's criteria) and 4-fold (OR 4.0; 95%CI 1.3-11-7: P = 0.014) increased prevalence of fibrosis staging ≥ 2, respectively. Multivariate analysis confirmed conformational anti-CYP2E1 IgG (P = 0.005) and age (P = 0.033) as independent predictors of necro-inflammatory grading ≥ 4. The development of anti-CYP2E1 auto-antibodies targeting conformational CYP2E1 epitopes is associated with more severe liver damage in CHC.
Asunto(s)
Autoanticuerpos/inmunología , Citocromo P-450 CYP2E1/inmunología , Anticuerpos Antihepatitis/inmunología , Hepatitis C Crónica/inmunología , Hepatitis C Crónica/patología , Hígado/inmunología , Hígado/patología , Adulto , Anciano , Autoanticuerpos/sangre , Western Blotting/métodos , Femenino , Anticuerpos Antihepatitis/sangre , Humanos , Inmunoglobulina G/sangre , Inmunoprecipitación/métodos , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Índice de Severidad de la EnfermedadRESUMEN
We previously reported that autoantibodies against cytochrome P4502E1 (CYP2E1) are frequent in patients with chronic hepatitis C. As autoimmune reactions are increasingly detected after orthotopic liver transplantation (OLT), this study investigates prevalence and significance of anti-CYP2E1 autoantibodies in 46 patients with post-OLT recurrent hepatitis C. IgG against recombinant human CYP2E1 above the control threshold was detected in 19 out 46 (41%) sera collected immediately before OLT and in 15 out 46 (33%) sera collected at the time of the 12 months follow-up liver biopsy. Although anti-CYP2E1 reactivity was not modified by OLT, the patients with persistently elevated anti-CYP2E1 IgG (n = 12; 26%) showed significantly higher prevalence of recurrent hepatitis with severe necroinflammation and fibrosis than those persistently negative or positive only either before or after OLT. Moreover, the probability of developing severe necroinflammation was significantly higher in persistently anti-CYP2E1-positive subjects. Multivariate regression and Cox analysis confirmed that the persistence of anti-CYP2E1 IgG, together with a history of acute cellular rejection and donor age >50 years, was an independent risk factor for developing recurrent hepatitis C with severe necroinflammation. We propose that autoimmune reactions involving CYP2E1 might contribute to hepatic damage in a subgroup of transplanted patients with recurrent hepatitis C.
Asunto(s)
Autoanticuerpos/sangre , Autoanticuerpos/inmunología , Citocromo P-450 CYP2E1/inmunología , Citocromo P-450 CYP2E1/metabolismo , Hepatitis C/enzimología , Hepatitis C/patología , Femenino , Estudios de Seguimiento , Hepatitis C/sangre , Hepatitis C/inmunología , Humanos , Inflamación/sangre , Inflamación/inmunología , Inflamación/patología , Cirrosis Hepática/etiología , Cirrosis Hepática/patología , Cirrosis Hepática/cirugía , Masculino , Necrosis/sangre , Necrosis/inmunología , Necrosis/patología , Recurrencia , Factores de RiesgoRESUMEN
In a recent paper [S. B. Lee, Physica A 387, 1567 (2008)] the epidemic spread of the contact process (CP) in deterministic fractals, already studied by I. Jensen [J. Phys. A 24, L1111 (1991)], has been investigated by means of computer simulations. In these previous studies, epidemics are started from randomly selected sites of the fractal, and the obtained results are averaged all together. Motivated by these early works, here we also studied the epidemic behavior of the CP in the same fractals, namely, a Sierpinski carpet and the checkerboard fractals but averaging epidemics started from the same site. These fractal media have spatial discrete scale invariance symmetry, and consequently the dynamic evolution of some physical observables may become coupled to the topology, leading to the logarithmic-oscillatory modulation of the corresponding power laws. In fact, by means of extensive simulations we shown that the topology of the substrata causes the oscillatory behavior of the epidemic observables. However, in order to observe these oscillations, which have not been reported in earlier works, the interference effect arising during the averaging of epidemics started from nonequivalent sites should be eliminated. Finally, by analyzing our data and those available on the literature for the dependence of the exponents eta and delta on the dimensionality of substrata, we conjectured that for integer dimensions (2
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The dynamical behavior of ferromagnetic Ising films confined in a DxLxL geometry (D<
RESUMEN
The nonequilibrium critical dynamics of the Ising magnet on a fractal substrate, namely the Sierpinski carpet with Hausdorff dimension d(H)=1.7925, has been studied within the short-time regime by means of Monte Carlo simulations. The evolution of the physical observables was followed at criticality, after both annealing ordered spin configurations (ground state) and quenching disordered initial configurations (high temperature state), for three segmentation steps of the fractal. We have obtained evidence showing that during these relaxation processes both the growth and the fragmentation of magnetic domains become influenced by the hierarchical structure of the substrate. In fact, the interplay between the dynamic behavior of the magnet and the underlying fractal leads to the emergence of a logarithmic-periodic oscillation, superimposed to a power law, which has been observed in the time dependence of both the decay of the magnetization and its logarithmic derivative. These oscillations have been carefully characterized in order to determine the critical temperature of the second-order phase transition and the critical exponents corresponding to the short-time regime. The effects of the substrate can also be observed from the dependence of the effective critical exponents on the segmentation step. The exponent theta of the initial increase of the magnetization has also been obtained and the results suggest that it would be almost independent of the fractal dimension of the substrate, provided that d(H) is close enough to d=2. The oscillations have been discussed within the framework of the discrete scale invariance of the substrate.
RESUMEN
Copper (II) is one of the most widely employed experimental models to induce in vitro low density lipoprotein (LDL) oxidation. It is generally assumed that Cu(I), generated from active reduction of Cu(II), is the real trigger for the peroxidation of polyunsaturated fatty acids in LDL. We have employed isolated human LDL challenged with Cu(II) and the Cu(I) chelator bathocuproine disulfonic acid (BC) to test the validity of this hypothesis. At lower Cu(II)/LDL molar ratios, BC completely inhibited copper-mediated LDL oxidation evaluated either as thiobarbituric acid reactive substances (TBARS) production or changes in apo B100 electrophoretic mobility. On the contrary, at higher Cu(II)/LDL molar ratios, BC did not prevent LDL oxidation but rather markedly stimulated both the generation of TBARS and the increase of apo B100 electronegativity. These oxidative modifications were completely prevented by the Cu(II) chelator EDTA. Furthermore, the BC-Cu(I) complex alone was neither redox active nor active inducer of TBARS generation. These findings indicate that, under these experimental conditions, [1] Cu(I) is not necessary to promote LDL oxidation, [2] the availability of Cu(II) is a prerequisite and [3] some of the reaction(s) involved in Cu(II) reduction may play an essential role in initiating LDL oxidation.
Asunto(s)
Cobre/química , Lipoproteínas LDL/química , Adulto , Quelantes , Sulfato de Cobre , Humanos , Lipoproteínas LDL/aislamiento & purificación , Oxidación-Reducción , Fenantrolinas , Sustancias Reactivas al Ácido Tiobarbitúrico/análisis , Factores de TiempoRESUMEN
Lipid peroxidation induced in isolated rat hepatocytes by FeCl3 (0.1 mM) was associated with an increase in the cytosolic free Ca2+ and in the ionophore-mobilizable Ca2+ content of both mitochondrial and extramitochondrial (endoplasmic reticular) pools. Ca2+ accumulation was completely prevented by the antioxidants promethazine and vitamin E succinate and was not linked to the inhibition of plasma membrane (Ca2+ + Mg2+)-ATPase and Ca2+ transport or to the depletion of intracellular ATP content. Moreover, preincubation of the hepatocytes with the Ca2+ channel blockers verapamil and nifedipine inhibited the elevation of cytosolic Ca2+, as well as the ion accumulation without interfering with the stimulation of lipid peroxidation by iron. These results suggest that peroxidative alterations of the hepatocyte plasma membranes might perturb the functions of verapamil- and nifedipine-sensitive Ca2+ channels resulting in a net influx of Ca2+, which is subsequently sequestrated in the intracellular compartments.
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Calcio/metabolismo , Compuestos Férricos/farmacología , Peroxidación de Lípido/efectos de los fármacos , Hígado/metabolismo , Animales , Calcimicina/farmacología , Carbonil Cianuro m-Clorofenil Hidrazona/farmacología , Membrana Celular/efectos de los fármacos , Membrana Celular/fisiología , Células Cultivadas , Cloruros , Citosol/metabolismo , Digitonina/farmacología , Retículo Endoplásmico/metabolismo , Cinética , Hígado/citología , Hígado/efectos de los fármacos , Masculino , Potenciales de la Membrana/efectos de los fármacos , Mitocondrias Hepáticas/metabolismo , Prometazina/farmacología , Ratas , Ratas Endogámicas , Tocoferoles , Vanadatos/farmacología , Vitamina E/análogos & derivados , Vitamina E/farmacologíaRESUMEN
Centrilobular hypoxia has been suggested to contribute to hepatic damage caused by alcohol intoxication. However, the mechanisms involved are still poorly understood. We have investigated whether alterations of Na(+) homeostasis might account for ethanol-mediated increase in hepatocyte sensitivity to hypoxia. Addition of ethanol (100 mmol/l) to isolated rat hepatocytes incubated under nitrogen atmosphere greatly stimulated cell death. An increase in intracellular Na(+) levels preceded cell killing and Na(+) levels in hepatocytes exposed to the combination of ethanol and hypoxia were almost twice those in hypoxic cells without ethanol. Na(+) increase was also observed in hepatocytes incubated with ethanol in oxygenated buffer. Ethanol addition significantly lowered hepatocyte pH. Inhibiting ethanol and acetaldehyde oxidation with, respectively, 4-methylpyrazole and cyanamide prevented this effect. 4-methylpyrazole, cyanamide as well as hepatocyte incubation in a HCO(3)(-)-free buffer or in the presence of Na(+)/H(+) exchanger blocker 5-(N,N-dimethyl)-amiloride also reduced Na(+) influx in ethanol-treated hepatocytes. 4-methylpyrazole and cyanamide similarly prevented ethanol-stimulated Na(+) accumulation and hepatocyte killing during hypoxia. Moreover, ethanol-induced Na(+) influx caused cytotoxicity in hepatocytes pre-treated with Na(+), K(+)-ATPase inhibitor ouabain. Also in this condition 4-methylpyrazole and 5-(N,N-dimethyl)-amiloride decreased cell killing. These results indicate that ethanol can promotes cytotoxicity in hypoxic hepatocytes by enhancing Na(+) accumulation.
Asunto(s)
Etanol/toxicidad , Isquemia/metabolismo , Hepatopatías/metabolismo , Hígado/efectos de los fármacos , Sodio/metabolismo , Adenosina Trifosfato/metabolismo , Animales , Hipoxia de la Célula , Separación Celular , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Cianamida/farmacología , Etanol/antagonistas & inhibidores , Fomepizol , Concentración de Iones de Hidrógeno , Técnicas In Vitro , Isquemia/patología , Hígado/metabolismo , Hígado/patología , Hepatopatías/patología , Masculino , Perfusión , Pirazoles/farmacología , Ratas , Ratas Wistar , Sodio/análisisRESUMEN
Reperfusion injury represents an important cause of primary graft non-function during liver transplantation. However, the mechanism responsible for cellular damage during reoxygenation has not yet been completely understood. We have investigated whether changes in intracellular Na(+) distribution might contribute to cause hepatocyte damage during reoxygenation buffer after 24 h of cold storage. Hepatocyte reoxygenation resulted in a rapid increase in cellular Na(+) content that was associated with cytotoxicity. Na(+) accumulation and hepatocyte death were prevented by the omission of Na(+) from the incubation medium, but not by the addition of antioxidants. Blocking Na(+)/H(+) exchanger and Na(+)/HCO(3)(-) co-transporter by, respectively, 5-(N,N-dimethyl)-amiloride or omitting HCO(3)(-) from the reoxygenation medium significantly decreased Na(+) overload and cytotoxicity. Stimulation of ATP re-synthesis by the addition of fructose also lowered Na(+) accumulation and cell death during reoxygenation. A significant protection against Na(+)-mediated reoxygenation injury was evident in hepatocytes maintained in an acidic buffer (pH 6.5) or in the presence of glycine. The cytoprotective action of glycine or of the acidic buffer was reverted by promoting Na(+) influx with the Na(+)/H(+) ionophore monensin. Altogether, these results suggest that Na(+) accumulation during the early phases of reoxygenation might contribute to liver graft reperfusion injury.
Asunto(s)
Hígado/metabolismo , Oxígeno/metabolismo , Sodio/metabolismo , Animales , Hipoxia de la Célula , Supervivencia Celular , Homeostasis , Técnicas In Vitro , Hígado/citología , Masculino , Ratas , Ratas Wistar , Daño por Reperfusión/metabolismo , Factores de TiempoRESUMEN
Electron spin resonance (EPR) spectroscopy analysis using the spin trapping agent 4-pyridyl-1-oxide-t-butyl nitrone (4-POBN) was used to measure the formation of free radical intermediates during NADPH-dependent oxidation of 1-methyl-, 1-ethyl-, and 1-isopropylhydrazine in rat liver microsomes and in reconstituted enzyme systems. The experiments in microsomes revealed that the specific activation of the hydrazines, as measured by the EPR signal intensities, was about two-fold higher, when expressed per nmol of P-450, in microsomes from rats treated with ethanol (EtOH) as compared to membranes isolated from either phenobarbital (PB)-, beta-naphthoflavone (beta-NF)-treated or control rats. Furthermore, kinetic experiments revealed that EtOH-microsomes had an apparent affinity for 1-ethylhydrazine about one order of magnitude higher than PB-microsomes. In reconstituted vesicular systems composed of phospholipids, NADPH cytochrome P-450 reductase and P-450, the intensities of EPR signals produced by the formation of the methyl-, ethyl- and isopropyl-free radicals, were 3- to 5-fold more intense in membrane vesicles containing ethanol-inducible CYP2E1 than phenobarbital-inducible CYP2B1. By contrast, CYP1A2, CYP2B4 and CYP2C4 were inefficient catalysts of radical formation. Desferrioxamine, catalase and superoxide dismutase did not influence the extent of ethyl radicals formed in EtOH-microsomes, indicating that hydroxyl radicals are not involved in the CYP2E1-dependent activation of 1-ethylhydrazine. Addition of cytochrome b5, an efficient donor of the second electron to P-450 and hence an inhibitor of the formation of the oxy-cytochrome P-450 complex, increased to be consistent with the results, did not influence the amount of ethyl radicals trapped. In liver microsomes from untreated rats selective substrates of CYP2E1, such as diethyl-dithiocarbamate and p-nitrophenol, as well as anti-CYP2E1-IgG, inhibited the free radical formation from 1-ethylhydrazine by about 60%. The anti-CYP2E1 IgG used significantly inhibited ethyl radical production also in human liver microsomes incubated with 1-ethylhydrazine and 4-POBN. Taken together, these results indicate that CYP2E1, as compared to other rat liver cytochromes P-450, is an efficient catalyst of transformation of alkylhydrazines to free radical intermediates, a finding that might be of importance in the development of the toxicity of these compounds.
Asunto(s)
Sistema Enzimático del Citocromo P-450/metabolismo , Etanol/farmacología , Hidrazinas/metabolismo , Microsomas Hepáticos/enzimología , Oxidorreductasas N-Desmetilantes/metabolismo , Animales , Benzoflavonas/farmacología , Citocromo P-450 CYP2E1 , Inhibidores Enzimáticos del Citocromo P-450 , Sistema Enzimático del Citocromo P-450/biosíntesis , Espectroscopía de Resonancia por Spin del Electrón , Inducción Enzimática/efectos de los fármacos , Radicales Libres , Humanos , Masculino , Microsomas Hepáticos/efectos de los fármacos , NADP/farmacología , Óxidos de Nitrógeno , Oxidorreductasas N-Desmetilantes/antagonistas & inhibidores , Oxidorreductasas N-Desmetilantes/biosíntesis , Fenobarbital/farmacología , Piridinas , Ratas , Ratas Sprague-Dawley , Marcadores de Spin , beta-naftoflavonaRESUMEN
The mechanisms responsible for the progression of nonalcoholic fatty liver disease (NAFLD) to more severe liver injury are still poorly understood. Data from animal models suggest that oxidative stress contributes to steatohepatitis and an increase of lipid peroxidation has been documented in human NAFLD. By measuring the titers of circulating antibodies against lipid peroxidation products as markers of oxidative stress we have observed that NAFLD patients have titers of these antibodies significantly higher than in controls. Moreover, the titers of lipid peroxidation-related antibodies are associated with a 3-fold increase in the risk of developing advanced fibrosis/cirrhosis. Although the mechanisms causing oxidative stress in NAFLD have not been elucidated, these results support the involvement of lipid peroxidation in the processes leading to liver fibrosis associated with NAFLD.
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Hígado Graso/metabolismo , Estrés Oxidativo/fisiología , Anticuerpos/sangre , Progresión de la Enfermedad , Hígado Graso/complicaciones , Hígado Graso/inmunología , Humanos , Peroxidación de Lípido/inmunología , Peroxidación de Lípido/fisiología , Cirrosis Hepática/etiología , Cirrosis Hepática/metabolismoRESUMEN
The short-time dynamic evolution of an Ising model embedded in an infinitely ramified fractal structure with noninteger Hausdorff dimension was studied using Monte Carlo simulations. Completely ordered and disordered spin configurations were used as initial states for the dynamic simulations. In both cases, the evolution of the physical observables follows a power-law behavior. Based on this fact, the complete set of critical exponents characteristic of a second-order phase transition was evaluated. Also, the dynamic exponent theta of the critical initial increase in magnetization, as well as the critical temperature, were computed. The exponent theta exhibits a weak dependence on the initial (small) magnetization. On the other hand, the dynamic exponent z shows a systematic decrease when the segmentation step is increased, i.e., when the system size becomes larger. Our results suggest that the effective noninteger dimension for the second-order phase transition is noticeably smaller than the Hausdorff dimension. Even when the behavior of the magnetization (in the case of the ordered initial state) and the autocorrelation (in the case of the disordered initial state) with time are very well fitted by power laws, the precision of our simulations allows us to detect the presence of a soft oscillation of the same type in both magnitudes that we attribute to the topological details of the generating cell at any scale.
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The alterations of mitochondrial membrane potential during the development of irreversible cell damage were investigated by measuring rhodamine-123 uptake and distribution in primary cultures as well as in suspensions of rat hepatocytes exposed to different toxic agents. Direct and indirect mechanisms of mitochondrial damage have been identified and a role for Ca2+ in the development of this type of injury by selected compounds was assessed by using extracellular as well as intracellular Ca2+ chelators. In addition, mitochondrial uncoupling by carbonylcyanide-m-chloro-phenylhydrazone (CCCP) resulted in a marked depletion of cellular ATP that was followed by an increase in cytosolic Ca2+ concentration, immediately preceding cell death. These results support the existence of a close relationship linking, in a sort of reverberating circuit, the occurrence of mitochondrial dysfunction and the alterations in cellular Ca2+ homeostasis during hepatocyte injury.
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Calcio/metabolismo , Enfermedad Hepática Inducida por Sustancias y Drogas , Mitocondrias Hepáticas/metabolismo , Adenosina Trifosfato/metabolismo , Animales , Carbonil Cianuro m-Clorofenil Hidrazona , Células Cultivadas , Colorantes Fluorescentes/metabolismo , Ionomicina , Cinética , Hepatopatías/metabolismo , Potenciales de la Membrana , Ratas , Ratas Endogámicas , Rodamina 123 , Rodaminas/metabolismo , Vitamina KRESUMEN
The formation of carbon centered free radicals, identified as methylcarbonyl species, was observed using ESR spectroscopy and the spin trapping agent 4-pyridyl-1-oxide-N-t-butyl nitrone (4-POBN) during the oxidation of acetaldehyde by xanthine oxidase. The reaction was dependent upon the presence of OH. radicals and was inhibited by the addition of superoxide dismutase, catalase or OH. radical scavengers. The generation of methylcarbonyl radicals was associated with a doubling of stable acetaldehyde adducts with serum albumin, and 4-POBN or superoxide dismutase and catalase, completely blocked this effect. Thus, methylcarbonyl radicals contributed to acetaldehyde-mediated protein alkylation which is involved in causing toxic as well as immunological reactions ascribed to acetaldehyde.
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Acetaldehído/metabolismo , Proteínas/metabolismo , Alquilación , Animales , Espectroscopía de Resonancia por Spin del Electrón , Radicales Libres/metabolismo , Oxidación-Reducción , Xantina Oxidasa/metabolismoRESUMEN
Ischemic preconditioning has been shown to improve liver resistance to hypoxia/reperfusion damage. A signal pathway involving A(2A)-adenosine receptor, G(i)-proteins, protein kinase C and p38 MAP kinase is responsible for the development of hypoxic preconditioning in hepatocytes. However, the coupling of this signal pathway with the mechanisms responsible for cytoprotection is still unknown. We have observed that stimulation of A(2A)-adenosine receptors or of p38 MAPK by CGS21680 or anisomycin, respectively, appreciably reduced intracellular acidosis and Na(+) accumulation developing during hypoxia. These effects were reverted by p38 MAPK inhibitor SB203580 as well as by blocking vacuolar proton ATPase with bafilomycin A(1). SB203580 and bafilomycin A(1) also abolished the cytoprotective action exerted by both CGS21680 and anisomycin. We propose that the stimulation of p38 MAPK by preconditioning might increase hepatocyte resistance to hypoxia by activating proton extrusion through vacuolar proton ATPase, thus limiting Na(+) overload promoted by Na(+)-dependent acid buffering systems.
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Acidosis/metabolismo , Adenosina/análogos & derivados , Hepatocitos/metabolismo , Precondicionamiento Isquémico , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Sodio/metabolismo , ATPasas de Translocación de Protón Vacuolares , Adenosina/farmacología , Animales , Anisomicina/farmacología , Hipoxia de la Célula/efectos de los fármacos , Células Cultivadas , Inhibidores Enzimáticos/farmacología , Hepatocitos/citología , Hepatocitos/efectos de los fármacos , Concentración de Iones de Hidrógeno/efectos de los fármacos , Líquido Intracelular/metabolismo , Masculino , Proteínas Quinasas Activadas por Mitógenos/antagonistas & inhibidores , Fenetilaminas/farmacología , Inhibidores de la Síntesis de la Proteína/farmacología , ATPasas de Translocación de Protón/antagonistas & inhibidores , ATPasas de Translocación de Protón/metabolismo , Agonistas del Receptor Purinérgico P1 , Ratas , Ratas Wistar , Receptor de Adenosina A2A , Receptores Purinérgicos P1/metabolismo , Transducción de Señal/efectos de los fármacos , ATPasa Intercambiadora de Sodio-Potasio/antagonistas & inhibidores , Proteínas Quinasas p38 Activadas por MitógenosRESUMEN
Isolated hepatocytes and liver microsomes incubated with monomethyl-1,1 dimethyl- and 1,2 dimethyl-hydrazines produced free radical intermediates which were detected by ESR spectroscopy by using 4-pyridyl-1-oxide-t-butyl nitrone (4-POBN) as spin trapping agent. The spectral features of the spin adducts derived from all three hydrazine derivatives corresponded to the values reported for the methyl free radical adduct of 4-POBN. In the microsomal preparations inhibitors of the mixed function oxidase system and the destruction of cytochrome P450 by pretreating the rats with CoCl2 all decreased the free radical formation. Methimazole, an inhibitor of FAD-containing monoxygenase system, similarly decreased the activation of 1,1 dimethyl-hydrazine, but not that of monomethyl- and 1,2 dimethyl-hydrazines. The addition to liver microsomes of physiological concentrations of glutathione (GSH) lowered by approx. 80% the intensities of the ESR signals. Consistently, incubation of isolated hepatocytes with methyl-hydrazines decreased the intracellular GSH content, suggesting that GSH can effectively scavenge the methyl free radicals. The results obtained suggest that methyl free radicals could be the alkylating species responsible for the toxic and/or carcinogenic effect of methyl-hydrazines.
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Dimetilhidrazinas/farmacocinética , Hígado/metabolismo , Metilhidrazinas/farmacocinética , Microsomas Hepáticos/metabolismo , Monometilhidrazina/farmacocinética , Animales , Biotransformación , Inhibidores Enzimáticos del Citocromo P-450 , Espectroscopía de Resonancia por Spin del Electrón , Radicales Libres , Glutatión/farmacología , Hígado/efectos de los fármacos , Masculino , Microsomas Hepáticos/efectos de los fármacos , Oxigenasas de Función Mixta/antagonistas & inhibidores , Óxidos de Nitrógeno , Piridinas , Ratas , Ratas Endogámicas , Marcadores de SpinRESUMEN
CCl4-induced impairment of the lipoprotein secretion pathway of intact rat hepatocytes was carried out using 14CCl4 to check the possibility of binding to lipoproteins by CCl4 metabolites. After separation of different cell suspension fractions by means of ultracentrifugation and chemical precipitation procedures, a significant amount of the radioisotope was found covalently bound to the lipid and protein components of low density lipoproteins. Suitable experiments demonstrated that the bound radioisotope was represented by CCl4 metabolites and not by unactivated CCl4.
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Tetracloruro de Carbono/metabolismo , Coloides , Lipoproteínas/metabolismo , Hígado/metabolismo , Micelas , Animales , Radioisótopos de Carbono , Lipoproteínas LDL/aislamiento & purificación , Lipoproteínas LDL/metabolismo , Masculino , Unión Proteica , Ratas , Ratas EndogámicasRESUMEN
Incubation of isolated hepatocytes with CCl4 results in early reduction of the intracellular calcium content, mostly due to loss from the mitochondrial compartment. CCl4 treatment directly affects mitochondrial functions as indicated by the inhibition of Ca2+ uptake in cells permeabilized to the ion by digitonin exposure and by the reduction of intracellular ATP content in hepatocytes incubated in a glucose-free medium. Such mitochondrial damage is not caused by CCl4-induced stimulation of lipid peroxidation since it is not prevented by alpha-tocopherol, used at a concentration able to inhibit completely peroxidative reactions without interfering with CCl4 activation. All data together are in favour of a direct action of CCl4-reactive metabolites on liver cell calcium homeostasis.
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Calcio/metabolismo , Intoxicación por Tetracloruro de Carbono/metabolismo , Homeostasis/efectos de los fármacos , Mitocondrias Hepáticas/metabolismo , Adenosina Difosfato/metabolismo , Animales , Tetracloruro de Carbono/farmacología , Carbonil Cianuro m-Clorofenil Hidrazona/farmacología , Cinética , Peróxidos Lipídicos/metabolismo , Masculino , Mitocondrias Hepáticas/efectos de los fármacos , Ratas , Vitamina E/farmacologíaRESUMEN
Antibodies against malondialdehyde (MDA)-modified proteins are often increased in patients with diseases related to oxidative stress. However, the clinical significance of these antibodies is hampered by their frequent presence also in healthy controls. Aim of this work has been to characterize the immune reactivity against MDA-derived antigens in healthy subjects. The sera of 120 healthy subjects contained IgG and IgM targeting MDA-modified human albumin (HSA), fibrinogen, and LDL. These sera also displayed weak reactivity with oxidized LDL and HSA complexed with oxidized arachidonic acid. Conversely, oxidized HSA or HSA complexed with other aldehydic lipid peroxidation products was not recognized. Control sera also did not recognize cyclic dihydropyridine-MDA products, while HSA-MDA reactivity was associated (r > 0.9; p <.0005) with the presence of fluorescent lysine-conjugated-imine cross-links. In Western blots both IgG and IgM recognized high molecular weight HSA-MDA aggregates, but not monomeric HSA-MDA adducts. The addition of sodium cyanoborohydride, that prevented conjugated-imine fluorescence and protein aggregation during HSA-MDA preparation, abolished the antibody binding. This suggested that the plasma of healthy subjects contained IgG and IgM recognizing protein aggregates linked through 1-amino-3-imino-propene bridges. The function of these antibodies is at the moment unknown, but they might contribute to scavenging MDA cross-linked proteins.