RESUMO
The effects of olanzapine (OLZ) on the viability and functioning of human polymorphonuclearcells (PMNs) are clearly opposite to those previously reported forclozapine (CLZ). In fact, after 4- or 24-h-treatment with 20-50 μM OLZ, a significant inhibition of the respiratory burst in PMNs activated with opsonized zimosanor phorbol myristate acetate (PMA) was observed, whereas the burst provoked byformyl-methionyl-leucyl-phenylalanine (fMLP) was only inhibited at 50 μM OLZ.Under the same conditions, spontaneous apoptosis was accelerated at 20-50 μMOLZ, while the exogenous addition of H2O2 resulted in the PMN apoptosis beingdose-dependently inhibited by OLZ in the entire range of concentrations. However,when H2O2 was intracellularly generated by treatment with PMA, the induced apoptosis was only decreased at 2 μM OLZ. Absorbance scans revealed that OLZis able to react with equimolar quantities of either H2O2 or HOCl. These results suggest that OLZ inhibits both ROS-induced PMN apoptosis and respiratory burst due to extracellular scavenging of released ROS.
Los efectos de olanzapine (olz) sobre la viabilidad y el funcionamiento de células humanas polimorfonucleares (pmn, por sus siglas en inglés) claramente son opuestosa los señalados para la clozapine (clz). En efecto, después de 4-24 h de tratamiento con 20-50 μM olz, se observó una inhibición significativa del estallido respiratorio en pmn activados con zimosan o con forbol acetato miristato, mientras que la inhibición provocada por el formil-metionil-leucil-fenilalanina fue sólo inhibida a 50μM de olz. En las mismas condiciones, la apoptosis espontánea se aceleró con 20-50μM olz, mientras que la adición exógena de H2O2 dio lugar a la apoptosis de pmn en dosis dependiente inhibida por olz en el rango entero de concentraciones. Sin embargo, cuando se generó H2O2 intracelular por tratamiento con pma, la apoptosis inducida se disminuyó solamente con 2 μM olz. Las exploraciones de los espectros de absorbancia revelaron que olz puede reaccionar con cantidades equimolares de H2O2 o de HOCL. Estos resultados sugieren que olz inhibe ambos tipos de apoptosis de pmn (la inducida por especies reactivas oxigenadas y por estallido respiratorio debido a atrapadores extracelulares de estas especies reactivas oxigenadas).
Assuntos
Humanos , Apoptose , Neutrófilos , Estresse Oxidativo , Explosão RespiratóriaRESUMO
The ability of dipyridamole (DIP) to scavenge oxygen metabolites generated by either activated human neutrophils (PMNs) or cell-free systems using luminol(s)- and lucigenin-enhanced chemiluminescence was investigated. In the presence of DIP (15-50 microM) a dose-dependent inhibition period was seen in phorbol myristate acetate (PMA)-stimulated PMNs as assayed by isoluminol-enhanced chemiluminescence (ILCL) with horseradish peroxidase (HRP). Although such a lag period was not observed in the absence of HRP, 50 microM DIP inhibited extracellular ILCL by more than 50%. Intracellular luminol-enhanced chemiluminescence (LCL) as assayed in either PMA- or in ionomycin-activated PMNs was not affected by dipyridamole (15-50 microM). In cell-free systems, DIP produced concentration-dependent inhibition in H2O2-(45% at 50 microM), OH- (40%, at 0.1 microM) and HOCl-(20% at 10 microM). Both absorbance and fluorescence scans revealed that DIP is able to react with equimolar quantities of either H202 or HOCl. These results suggest that DIP scavenges reactive oxygen species (ROS) presumably secreted by activated human PMNs in the following decreasing order: *OH > HOCl > H2O2 >> O2-.
Assuntos
Antioxidantes/química , Dipiridamol/química , Acridinas/química , Citocromos c/metabolismo , Espaço Extracelular/química , Espaço Extracelular/metabolismo , Humanos , Peróxido de Hidrogênio/química , Ácido Hipocloroso/química , Técnicas In Vitro , Medições Luminescentes , N-Formilmetionina Leucil-Fenilalanina , Neutrófilos/efeitos dos fármacos , Neutrófilos/metabolismo , Neutrófilos/efeitos da radiação , Oxidantes/química , Oxirredução , Espécies Reativas de Oxigênio/químicaRESUMO
The phototoxic drug thiocolchicoside (2-dimethoxy-2-glucosidoxythiocolchicine, 1), is photolabile under irradiation with UV-A light from TL 100 W-P Philips bulbs (at lambda max 355 nm) light and also with a N2 laser (at 337 nm) in aerobic and anaerobic conditions. Irradiation of a methanol solution of 1 produces two photoproducts without a glucoside group. One of these lost the methylthio-group, while the other is oxidized (only under aerobic conditions) to sulfoxide. The formation of singlet oxygen by photolysis of 1 was evidenced by trapping with 2,5-dimethylfuran (GC-MS), furfuryl alcohol, 1,3-cyclohexadiene-1,4-diethanoate (HPLC) and by the histidine test as 1O2 scavengers. Thiocolchicoside has been shown to photosensitize the reduction of nitro blue tetrazolium by direct electron transfer mechanism, when irradiated under the same conditions as for photolysis. Oxygen may also be involved in this electron transfer reaction to form the superoxide anion radical. Thiocolchicoside was screened in vitro in different concentrations for UV-Vis-induced phototoxic effects in a photohemolysis test, in the presence and absence of different radical scavengers, singlet oxygen and superoxide radical quenchers. In addition, 1 photosensitized the peroxidation of linoleic acid, monitored by the UV-detection of dienic hydroperoxides. Studies on peripheral blood mononuclear cells (lymphocytes) demonstrated phototoxic effects on them. Protection by GSH, DABCO, sodium azide and SOD are indicative of both Type I and II photosensitization pathways mediated by free radicals and singlet molecular oxygen.
Assuntos
Colchicina/farmacologia , Luz , Fármacos Fotossensibilizantes/farmacologia , Colchicina/análogos & derivados , Colchicina/química , Colchicina/toxicidade , Transporte de Elétrons , Eritrócitos/efeitos dos fármacos , Eritrócitos/efeitos da radiação , Radicais Livres , Hemólise/efeitos dos fármacos , Hemólise/efeitos da radiação , Humanos , Técnicas In Vitro , Peroxidação de Lipídeos/efeitos dos fármacos , Peroxidação de Lipídeos/efeitos da radiação , Linfócitos/efeitos dos fármacos , Linfócitos/efeitos da radiação , Nitroazul de Tetrazólio/química , Oxirredução , Fotoquímica , Fotólise , Fármacos Fotossensibilizantes/química , Fármacos Fotossensibilizantes/toxicidade , Espectrofotometria Ultravioleta , Raios UltravioletaRESUMO
The in vitro antioxidant and photo-oxidant activity of dipyridamole was studied by its effect on superoxide- and singlet oxygen-mediated photohemolysis and viability of neutrophils. Dipyridamole was found to be phototoxic when examined by the photohemolysis on human erythrocytes and on linoleic acid as lipid peroxidation model at concentrations above 3.0 x 10(-5) M. On the contrary, when lower concentrations (1.0 x 10(-5) to 1.0 x 10(-6) M) were used, dipyridamole showed a protector action against singlet oxygen-mediated photohemolysis by other phototoxic compounds like triamterene. This antioxidant property is proposed to result from quenching of triamterene mediated by fluorescence energy transfer. Auto-oxidation and fluorescence-energy transfer is clearly an important mechanism for protection for this drug.
Assuntos
Antioxidantes/farmacologia , Dipiridamol/farmacologia , Dipiridamol/toxicidade , Hemólise/efeitos dos fármacos , Fotólise , Inibidores da Agregação Plaquetária/farmacologia , Inibidores da Agregação Plaquetária/toxicidade , Sequestradores de Radicais Livres/farmacologia , Hemólise/efeitos da radiação , Humanos , Técnicas In Vitro , Cinética , Ácido Linoleico , Oxigênio Singlete , Espectrofotometria Ultravioleta , Superóxidos , Triantereno/toxicidade , Raios UltravioletaRESUMO
The phototoxic anti-cancer drug flutamide is photolabile under UV-B light in either aerobic or anaerobic conditions. Irradiation of a methanol solution of this drug produces several photoproducts, one by photoreduction of the nitro group, one by rupture of the aromatic-NO2 bond of the parent compound, two as a result of the rupture of the CO-NH bond and one derived from the photoreduction product by scission of the aromatic-NH2 bond. Flutamide shows a photohemolytic effect on human erythrocytes and photoinduces lipid peroxidation. Studies on peripheral blood polymorphonuclear cells (neutrophils) demonstrated the phototoxicity of flutamide as well as inhibition of the cytotoxicity respiratory burst by the photoproduct derived from its photoreduction. The results suggest that the inhibition of the respiratory burst observed in phorbol myristate acetate (PMA)-activated cells is mediated by photosensitization and concomitant singlet oxygen production and/or formation of toxic photoproducts.
Assuntos
Antineoplásicos Hormonais/efeitos da radiação , Antineoplásicos Hormonais/toxicidade , Flutamida/efeitos da radiação , Flutamida/toxicidade , Antineoplásicos Hormonais/química , Flutamida/química , Hemólise/efeitos dos fármacos , Hemólise/efeitos da radiação , Humanos , Técnicas In Vitro , Peroxidação de Lipídeos/efeitos da radiação , Medições Luminescentes , Neutrófilos/efeitos dos fármacos , Neutrófilos/efeitos da radiação , Fotobiologia , Fotoquímica , Fotólise , Albumina Sérica/efeitos da radiação , Raios UltravioletaRESUMO
The phototoxic diuretic drug furosemide (1), a 5-(aminosulfonyl)-4-chloro-2-[(2-furanylmethyl)-amino] benzoic acid is photolabile under aerobic and anaerobic conditions. Irradiation of a methanol solution of 1 under oxygen produces photoproducts 2, 3, 4 and singlet oxygen, while under argon the photoproducts 2 and 4 were isolated. A peroxidic unstable photoproduct was detected during the photolysis under oxygen atmosphere. The formation of singlet oxygen by photolysis of 1 was evidenced by trapping with 2,5-dimethylfuran (GC-mass), furfuryl alcohol and 1,3-cyclohexadiene-1,4-diethanoate (HPLC) as 1O2 scavengers and by the histidine test. Furosemide was screened in vitro at different concentrations for UV-Vis-induced phototoxic effects in a photohemolysis test, in the presence and absence of different radical scavengers, singlet oxygen and hydroxyl radical quenchers. However, furosemide photosensitized the peroxidation of linoleic acid, as monitored by the UV-detection of dienic hydroperoxides and it also photosensitized the oxidation of histidine. The photodegradation was catalyzed in the presence of human serum albumin. Studies on peripheral blood mononuclear and polymorphonuclear cells (lymphocytes and neutrophils) demonstrated no phototoxicity on these cell lines.
Assuntos
Diuréticos/metabolismo , Eritrócitos/efeitos dos fármacos , Furosemida/metabolismo , Oxigênio/metabolismo , Fotoquímica , Dermatite Fototóxica , Diuréticos/toxicidade , Furosemida/toxicidade , Hemólise , Humanos , Peroxidação de Lipídeos , Oxigênio SingleteRESUMO
Our approach to assessing the photobiological risk of drugs combines photochemical and physicochemical studies with in vitro testing on human serum, human erythrocytes, lymphocytes and neutrophils. We have used this approach to investigate the photobiological risk of the diuretic drug triamterene, which has shown phototoxic effects in vivo. Photodecomposition studies of triamterene in methanol, phosphate buffered saline (PBS) solution and human serum, in the presence of oxygen, was followed by UV spectrophotometry and HPLC analysis using a sensitive HPLC method. Its photodegradation was observed only under irradiation with UV-B (290-320nm) light to produce the photoproduct 2. No photodecomposition was detected under UV-A (320-400nm) irradiation, yet singlet oxygen was generated. Triamterene shows a photohaemolytic effect and photoinduced lipid peroxidation. In the presence of oxygen, triamterene was able to induce photohaemolysis of human erythrocytes. The same process was observed under an inert atmosphere, although at a significantly lower rate. Studies on peripheral blood mononuclear and polymorphonuclear cells (lymphocytes and neutrophils) demonstrated phototoxicity on these cell lines.
RESUMO
The cytotoxicity of a low mol. wt fraction (LMWF) obtained from Aloe vera gel was determined by two different assays. Firstly, exposure of monolayers of chicken fibroblasts to LMWF induced disruption of intercellular junctions and detachment of individual cells from the bottom of the flask, with formation of cell-free gaps in the monolayer. Secondly, LMWF inhibited the production of reactive oxygen species by human polymorphonuclear leukocytes stimulated by zymosan, as followed by luminol-dependent chemiluminescence. The toxic activity of LMWF was compared to that of sodium dodecyl sulfate (a well-known toxic substance), aloe-emodin and aloin (an anthraquinone and its precursor present in Aloe vera cortex) using the chemilumescence assay, and was found to be of similar potency to these toxic substances on a weight-to-weight basis. These results confirm that Aloe vera gel contains toxic low mol. wt compounds, and every effort must be made to limit the amount of these toxins in the commercially prepared Aloe vera gel products.
Assuntos
Aloe/toxicidade , Plantas Medicinais , Animais , Células Cultivadas , Galinhas , Feminino , Géis , Humanos , Junções Intercelulares/efeitos dos fármacos , Peso Molecular , Espécies Reativas de OxigênioRESUMO
A yeast 50-kDa mRNA-binding protein (50mRNP) is found selectively associated with the 48S and 80S initiation complexes. This protein is structurally related to the translational elongation factor EF-1alpha. The protein reacts with antibodies directed against EF-1alpha and, similarly, EF-1alpha recognizes antibodies against the 50mRNP protein. This is evidence that they share at least one epitope which allows a similar antigenic behavior. In addition, both proteins show similar cleavage patterns upon treatment with the endoproteinase Lys-C. A murine antibody raised against 50mRNP inhibits both 48S and 80S initiation complex formation. The inhibitory effect is relieved by preincubating anti-50mRNP with EF-1alpha. Antibody to EF-1alpha manifests a similar inhibitory pattern for the formation of 48S and 80S complexes. These data strongly suggest that 50mRNP is an EF-1alpha-like polypeptide essential for the formation of the above complexes.
Assuntos
RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/metabolismo , Ribonucleoproteínas/metabolismo , Ribossomos/metabolismo , Saccharomyces cerevisiae/metabolismo , Anticorpos/imunologia , Centrifugação com Gradiente de Concentração , Eletroforese em Gel de Poliacrilamida , Epitopos/imunologia , Proteínas Fúngicas/química , Proteínas Fúngicas/metabolismo , Metaloendopeptidases/metabolismo , Iniciação Traducional da Cadeia Peptídica , Fator 1 de Elongação de Peptídeos , Fatores de Alongamento de Peptídeos/química , Fatores de Alongamento de Peptídeos/imunologia , Fatores de Alongamento de Peptídeos/fisiologia , Fragmentos de Peptídeos/isolamento & purificação , Peptídeos/antagonistas & inibidores , Peptídeos/metabolismo , Polirribossomos/química , Polirribossomos/metabolismo , Ligação Proteica , RNA Mensageiro/genética , Proteínas de Ligação a RNA/química , Ribonucleoproteínas/química , Ribonucleoproteínas/imunologia , Saccharomyces cerevisiae/químicaRESUMO
A yeast ribosomal subunit association factor (AF) has been purified from a high-salt ribosomal wash. The purified enzyme is a thermostable protein that associates ribosomal subunits at low Mg2+ concentration without requiring energy. It appears to be an aggregate of trimers or dimers (molecular mass 125 or 79 kDa) which on sodium dodecyl sulfate gels shows the presence of a major protein band whose estimated molecular mass is 43 kDa. Evidence also indicates the existence of a 50-kDa polypeptide which seems to be unstable since with freezing and thawing it gives rise to the 43-kDa polypeptide. It was shown that the labelled factor interacts with 80S ribosomes and with 40S ribosomal subunits. The purified polypeptide reacts with antibodies directed against EF-1 alpha, this last protein recognizing the antibodies raised against AF. Likewise, both EF-1 alpha and AF associate ribosomal subunits in the same way. When EF-1 is heated, it not only maintains its association activity, but also behaves like a 43-kDa polypeptide in an SDS electrophoresis run. These observations strongly suggest that AF originates from EF-1 alpha, which implies that the well-known elongation factor may also play a role in the initiation step of protein synthesis.