RESUMO
Research on biological influence of vanadium has gained major importance because it exerts potent toxic, mutagenic, and genotoxic effects on a wide variety of biological systems. However, hematological toxicity is one of the less studied effects. The lack of information on this issue prompted us to study the structural effects induced on the human erythrocyte membrane by vanadium (V). Sodium orthovanadate was incubated with intact erythrocytes, isolated unsealed human erythrocyte membranes (IUM) and molecular models of the erythrocyte membrane. The latter consisted of bilayers of dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylethanolamine (DMPE), phospholipid classes located in the outer and inner monolayers of the human erythrocyte membrane, respectively. This report presents evidence in order that orthovanadate interacted with red cell membranes as follows: a) in scanning electron microscopy (SEM) studies it was observed that morphological changes on human erythrocytes were induced; b) fluorescence spectroscopy experiments in isolated unsealed human erythrocyte membranes (IUM) showed that an increase in the molecular dynamics and/or water content at the shallow depth of the lipids glycerol backbone at concentrations as low as 50µM was produced; c) X-ray diffraction studies showed that orthovanadate 0.25-1mM range induced increasing structural perturbation to DMPE; d) somewhat similar effects were observed by differential scanning calorimetry (DSC) with the exception of the fact that DMPC pretransition was shown to be affected; and e) fluorescence spectroscopy experiments performed in DMPC large unilamellar vesicles (LUV) showed that at very low concentrations induced changes in DPH fluorescence anisotropy at 18°C. Additional experiments were performed in mice cholinergic neuroblastoma SN56 cells; a statistically significant decrease of cell viability was observed on orthovanadate in low or moderate concentrations.
Assuntos
Eritrócitos/metabolismo , Neuroblastoma/metabolismo , Sódio/farmacologia , Vanadatos/farmacologia , Acetilcoenzima A/química , Animais , Anisotropia , Varredura Diferencial de Calorimetria/métodos , Linhagem Celular Tumoral , Sobrevivência Celular , Dimiristoilfosfatidilcolina/química , Eritrócitos/efeitos dos fármacos , Humanos , Técnicas In Vitro , Lipídeos/química , Camundongos , Microscopia Eletrônica de Varredura/métodos , Fosfatidiletanolaminas/química , Espectrometria de Fluorescência/métodos , Temperatura , Lipossomas Unilamelares/química , Vanádio/farmacologiaRESUMO
While traces of manganese (Mn) take part in important and essential functions in biology, elevated exposures have been shown to cause significant toxicity. Chronic exposure to the metal leads to manganese neurotoxicity (or manganism), a brain disorder that resembles Parkinsonism. Toxic effect mechanisms of Mn is not understood, toxic concentrations of manganese are not well defined and blood manganese concentration at which neurotoxicity occurs has not been identified. There are reports indicating that the most abundant Mn-species in Mn carriers within blood is the Mn-citrate complex. Despite the well-documented information about the toxic effects of Mn, there are scarce reports concerning the effects of manganese compounds on both structure and functions of cell membranes, particularly those of human erythrocytes. With the aim to better understand the molecular mechanisms of the interaction of Mn with cell membranes, MnCl(2), and the Mn-citrate complex were incubated with intact erythrocytes, isolated unsealead human erythrocyte membranes (IUM), and molecular models of the erythrocyte membrane. These consisted in bilayers of dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylethanolamine (DMPE), phospholipid classes present in the outer and inner monolayers of the erythrocyte membrane, respectively. The capacity of the Mn compounds to perturb the bilayer structures of DMPC and DMPE was evaluated by X-ray diffraction, IUM were studied by fluorescence spectroscopy, and intact human erythrocytes were observed by scanning electron microscopy (SEM). In all these systems it was found that Mn(2+) exerted considerable higher structural perturbations than the Mn-citrate complex.
Assuntos
Citratos/toxicidade , Membrana Eritrocítica/química , Membrana Eritrocítica/efeitos dos fármacos , Manganês/toxicidade , Modelos Moleculares , Citratos/química , Dimiristoilfosfatidilcolina/química , Membrana Eritrocítica/ultraestrutura , Humanos , Manganês/química , Microscopia Eletrônica de Varredura , Fosfatidiletanolaminas/química , Difração de Raios XRESUMO
Zinc is an essential element for nutrition as well as for the proper development and function of brain cells, and its traces are present in a wide range of foods. It is a constituent of many enzyme systems and is an integral part of insulin and of the active site of intracellular enzymes. However, excessive accumulation of zinc or its release from the binding sites may become detrimental for neurons. With the aim to better understand the molecular mechanisms of the interaction of zinc ions with cell membranes, it was incubated with intact human erythrocytes, isolated unsealed human erythrocyte membranes (IUM), cholinergic murine neuroblastoma cells, and molecular models of cell membranes. These consisted in bilayers built-up of dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylethanolamine (DMPE), phospholipid classes present in the outer and inner monolayers of most plasmatic cell membranes, particularly that of human erythrocytes, respectively. The capacity of zinc ions to perturb the bilayer structures of DMPC and DMPE was assessed by X-ray diffraction, DMPC large unilamellar vesicles (LUV) and IUM were studied by fluorescence spectroscopy, intact human erythrocytes were observed with scanning electron microscopy (SEM), and neuroblastoma cell morphology was observed under inverted microscope. This study presents evidence that 0.1mM Zn and higher concentrations affect cell membrane and molecular models.
Assuntos
Membrana Celular/efeitos dos fármacos , Eritrócitos/efeitos dos fármacos , Bicamadas Lipídicas/química , Zinco/farmacologia , Animais , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Dimiristoilfosfatidilcolina/química , Eritrócitos/ultraestrutura , Humanos , Camundongos , Microscopia Eletrônica de Varredura , Fosfatidiletanolaminas/química , Espectrometria de Fluorescência , Difração de Raios XRESUMO
DPPC incorporation into egg-PC unilamellar vesicles reduces their oxidation rate beyond that expected from the unsaturated lipid dilution. Addition of the unsaturated lipids produces changes in the physical properties of the inner parts of the lipid bilayer, as sensed by fluorescence anisotropy of DPH, and in the hydrophilic/hydrophobic region, as sensed by the generalized polarization of laurdan. DPPC (30 mol%) incorporation into egg-PC vesicles produces a decrease in alkyl chain mobility in the inner part of the bilayer, evaluated by the increase of DPH fluorescence anisotropy, and a rise of the generalized polarization value of laurdan in the bilayer interface. It also leads to a decrease in the rate of water efflux promoted by a hypertonic shock. Oxidation of PC LUVs, promoted by AAPH, as sensed by oxygen uptake and MDA formation, leads to qualitatively similar results than DPPC addition: rigidification at the inner part and the surface of the liposomes, and a lower rate of water permeation. It is suggested that these changes could contribute to the observed decrease in oxidation rate with conversion.
Assuntos
1,2-Dipalmitoilfosfatidilcolina/química , Peroxidação de Lipídeos , Fosfatidilcolinas/química , Anisotropia , Polarização de Fluorescência , Bicamadas Lipídicas , Lipídeos/química , Lipossomos/química , Fluidez de Membrana , Microscopia de Fluorescência/métodos , Oxigênio/metabolismo , Consumo de Oxigênio , Permeabilidade , Peróxidos/metabolismo , Água/químicaRESUMO
Chromium exists in many oxidation states, of which only the hexavalent Cr(VI) and the trivalent Cr(III) ions are stable under environmental conditions. It is generally reported that Cr(VI) is highly toxic while Cr(III) is relatively innocuous, although others have reported just the opposite. On the other hand, despite the many studies on chromium toxicity, and particularly after the knowledge that Cr(VI) anions readily enter the erythrocytes where they are reduced to Cr(III), there are practically no reports on the structural effects induced by chromium compounds on the erythrocyte membrane. With the aim to better understand the molecular mechanisms of the interaction of Cr(III) and Cr(VI) with cell membranes, CrCl(3), and K(2)CrO(4) were incubated with intact erythrocytes, isolated unsealed human erythrocyte membranes (IUM), and molecular models of the erythrocyte membrane. These consisted in bilayers built-up of dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylcholine (DMPE), phospholipid classes present in the outer and inner monolayers of the erythrocyte membrane, respectively. The capacity of Cr(III) and Cr(VI) to perturb the bilayer structures of DMPC and DMPE was evaluated by X-ray diffraction, DMPC large unilamellar vesicles (LUV) and IUM were studied by fluorescence spectroscopy, and intact human erythrocytes were observed with scanning electron microscopy (SEM). In all these systems, it was found that Cr(III) induced considerably higher structural perturbations than Cr(VI).
Assuntos
Cromo/química , Membrana Eritrocítica/química , Membrana Eritrocítica/ultraestrutura , Humanos , Microscopia Eletrônica de Varredura , Modelos Moleculares , Espectrometria de Fluorescência , Difração de Raios XRESUMO
Ugni molinae Turcz, also known as "Murtilla", is a plant that grows in the south of Chile. Infusions of their leaves have long been used in traditional native herbal medicine. The chemical composition of the leaves indicates the presence of polyphenols, which have antioxidant properties. In order to evaluate the mechanisms of their antioxidant properties and the toxicity of the aqueous extracts of leaves, the extracts were induced to interact with human red cells, their isolated unsealed membranes (IUM) and large unilamellar vesicles (LUV) of dimyristoylphosphatidyltidylcholine (DMPC), representative of phospholipid classes located in the outer monolayer of the erythrocyte membrane. Scanning electron microscopy (SEM) observations indicated that the extracts achieved a significant alteration in the shape of the erythrocytes as they changed their discoid shape to echinocytes. According to the bilayer couple hypothesis, the shape change indicates that the polyphenols were located in the outer moiety of the red cell membrane. This conclusion was confirmed by the fluorescence experiments performed in IUM and DMPC LUV. In fact, the extracts produced slight initial increases followed by sharp decreases at higher concentrations in the anisotropy and general polarization parameters. These results imply that the extracts induced structural perturbations in the acyl chain and polar group packing arrangements of the erythrocyte IUM and DMPC LUV lipid bilayers: first ordering and afterwards disordering them as the extract concentration increased.
Assuntos
Membrana Eritrocítica/efeitos dos fármacos , Eritrócitos/efeitos dos fármacos , Myrtaceae/química , Extratos Vegetais/farmacologia , Antioxidantes/farmacologia , Dimiristoilfosfatidilcolina/química , Membrana Eritrocítica/metabolismo , Membrana Eritrocítica/ultraestrutura , Eritrócitos/metabolismo , Eritrócitos/ultraestrutura , Flavonoides/farmacologia , Polarização de Fluorescência , Humanos , Bicamadas Lipídicas/química , Microscopia Eletrônica de Varredura , Fenóis/farmacologia , Extratos Vegetais/química , Folhas de Planta/química , PolifenóisRESUMO
Chlordane is a widely used organochlorine insecticide. In order to evaluate its perturbing effect upon the morphology of human erythrocytes it was caused to interact with human red cells and molecular models of cell membranes. These consisted in bilayers of dimyristoylphosphatidylethanolamine (DMPE) and of dimyristoylphosphatidylcholine (DMPC), representative of phospholipid classes located in the inner and outer monolayers of the erythrocyte membrane, respectively. Scanning electron microscopy (SEM) observations indicated that this pesticide induced a significant alteration in the shape of the erythrocytes as they changed their discoid shape to spherocytes. According to the bilayer couple hypothesis, the shape changes induced in erythrocytes by foreign molecules are due to differential expansion of their two monolayers. The fact that chlordane produced spherocytes would indicate that the pesticide was equally located in the outer and the inner moieties of the red cell membrane. This conclusion was supported by the results obtained from X-ray diffraction studies. These showed that the hydrophobic and polar head regions of DMPC bilayers were perturbed when the insecticide was in a 1:10 molar ratio with respect to the lipid. These results were confirmed by the fluorescence experiments performed in DMPC large unilamellar vesicles (LUV). Chlordane produced a sharp decrease in the anisotropy and general polarization parameters in the 0-0.1 mM range, implying an increase in the fluidity at the acyl chain and polar region of DMPC. On the other hand, the bilayer structure of DMPE was perturbed in a fashion similar to that observed by X-ray diffraction in DMPC, a fact that explains the morphological change induced by chlordane to the human erythrocytes.
Assuntos
Clordano/toxicidade , Membrana Eritrocítica/efeitos dos fármacos , Eritrócitos/efeitos dos fármacos , Inseticidas/toxicidade , Bicamadas Lipídicas/química , Adulto , Clordano/efeitos adversos , Dimiristoilfosfatidilcolina/química , Membrana Eritrocítica/ultraestrutura , Eritrócitos/ultraestrutura , Humanos , Técnicas In Vitro , Inseticidas/efeitos adversos , Masculino , Microscopia Eletrônica de Varredura/métodos , Fosfatidiletanolaminas/química , Espectrometria de Fluorescência , Difração de Raios XRESUMO
The structural effects of titanium citrate on the human erythrocyte membrane were studied through its interaction with intact erythrocytes and isolated unsealed human erythrocyte membranes (IUM). The studies were carried out by scanning electron microscopy and fluorescence spectroscopy, respectively. Titanium citrate induced shape changes in erythrocytes, which were damaged and ruptured leaving empty and retracted membranes. Fluorescence spectroscopy measurements in IUM indicated a disordering effect at both the polar head group and the acyl chain packing arrangements of the membrane phospholipid bilayer. Titanium citrate also interacted with molecular models of the erythrocyte membrane consisting in bilayers of dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylethanolamine (DMPE), representing classes of phospholipids located in the outer and inner monolayers of the erythrocyte membrane, respectively. X-ray diffraction indicated that titanium citrate induced structural perturbation of the polar head group and of the hydrophobic acyl regions of DMPC, while the effects on DMPE bilayers were negligible. This conclusion is supported by fluorescence spectroscopy measurements on DMPC large unilamellar vesicles. All these findings indicate that the structural perturbations induced by titanium to human erythrocytes can be extended to other cells, thereby affecting their functions.
Assuntos
Ácido Cítrico/farmacologia , Membrana Eritrocítica/efeitos dos fármacos , Bicamadas Lipídicas/química , Acilação , Ácido Cítrico/química , Ácido Cítrico/metabolismo , Dimiristoilfosfatidilcolina/química , Relação Dose-Resposta a Droga , Membrana Eritrocítica/química , Membrana Eritrocítica/ultraestrutura , Humanos , Microscopia Eletrônica de Varredura , Fosfatidiletanolaminas/química , Espectrometria de Fluorescência , Difração de Raios XRESUMO
The structural effects of cadmium on cell membranes were studied through the interaction of Cd(2+) ions with human erythrocytes and their isolated unsealed membranes (IUM). Studies were carried out by scanning electron microscopy and fluorescence spectroscopy, respectively. Cd(2+) induced shape changes in erythrocytes, which took the form of echinocytes. According to the bilayer couple hypothesis, this result meant that Cd(2+) ions located in the outer monolayer of the erythrocyte membrane. Fluorescence spectroscopy measurements in IUM indicated a disordering effect at both the polar headgroup and the acyl chain packing arrangements of the membrane phospholipid bilayer. Cd(2+) ions also interacted with molecular models of the erythrocyte membrane consisting in bilayers of dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylethanolamine (DMPE), representing classes of phospholipids located in the outer and inner monolayers the erythrocyte membrane, respectively. X-ray diffraction indicated that Cd(2+) ions induced structural perturbation of the polar headgroup and of the hydrophobic acyl regions of DMPC, while the effects of cadmium on DMPE bilayers were much milder. This conclusion is supported by fluorescence spectroscopy measurements on DMPC large unilamellar vesicles (LUV). All these findings point to the important role of phospholipid bilayers in the interaction of cadmium on cell membranes.
Assuntos
Compostos de Cádmio/química , Membrana Eritrocítica/química , Bicamadas Lipídicas/química , Modelos Químicos , Membrana Eritrocítica/ultraestrutura , Humanos , Microscopia Eletrônica de Varredura , Fosfolipídeos/químicaRESUMO
Lead has no biological function; however, low, and particularly, high levels of exposure have a number of negative consequences for human health. Despite the number of reports about lead toxicity, very little information has been obtained regarding its effects on cell membranes. For this reason, the structural effects of lead on the human erythrocyte membranes were investigated. This aim was attained by making lead ions interact with intact erythrocytes, isolated unsealed erythrocyte membranes (IUM) and molecular models. The latter consisted of bilayers of dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylethanolamine (DMPE), representing phospholipid classes located in the outer and inner monolayers of the human erythrocyte membrane. The results, obtained by electron microscopy, fluorescence spectroscopy and X-ray diffraction, indicated that (a) lead particles adhered to the external and internal surfaces of the human erythrocyte membrane; (b) lead ions disturbed the lamellar organization of IUM and DMPC large unilamellar vesicles (LUV) and (c) induced considerable molecular disorder in both lipid multilayers, the effects being much more pronounced in DMPC.