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M T Santini

Publications and source records attributed to M T Santini.

At least 37 records · Page 2Linked to original sources
PubMed1995

Veratryl alcohol oxidase from Pleurotus ostreatus participates in lignin biodegradation and prevents polymerization of laccase-oxidized substrates.

Oxidative enzymes (laccases and peroxidases) isolated from the culture media of different fungi are involved in the basic mechanism of ligninolysis via radical intermediates. However, experiments aimed at reproducing natural biodegradation in vitro have been unsuccessful so far since the single biocatalysts alone are not able to solubilize lignins because of the simultaneous recondensation of these intermediates. FAD oxidases can prevent this side reaction in lignin depolymerization by reducing quinonoids and radical compounds. This study investigates the possible role of a laccase and a FAD-dependent aryl alcohol oxidase (veratryl alcohol oxidase, VAO) excreted by the basidiomycete Pleurotus ostreatus. In fact, we found that VAO is able to reduce synthetic quinones, laccase-generated quinonoids, and phenoxy radicals with concomitant oxidation of veratryl alcohol to veratryl aldehyde. This cooperative action of laccase and VAO also prevented the polymerization of phenolic compounds and reduced the molecular weight of soluble lignosulfonates to a significant extent.

Alcohol Oxidoreductases↗
PubMed1995

Fusinite as a specific probe for the determination of molecular oxygen concentration in cells.

The feasibility of using EPR and the paramagnetic derivative of coal 'fusinite' to measure intracellular oxygen concentration in cultured cells in which this substance was internalized in the cytoplasm was examined. First, the possible cytotoxic effects of fusinite on cultured cells were ruled out by both morphological as well as by growth characteristics analyses. After construction of a calibration curve in which the EPR spectral linewidth of this substance was measured in response to known oxygen concentrations, the efficacy of using fusinite in the determination of intracellular oxygen concentration in cells was also tested by flowing different known oxygen gas mixtures outside cultured cells. The results indicate that fusinite is able of measuring the variations in cytoplasmic oxygen concentration that exist in response to the different gas mixtures. In addition, as an example of a possible use of fusinite, data are also presented demonstrating a decrease in cytoplasmic oxygen concentration during respiration in cells with a limited supply of oxygen. In fact, as the oxygen is consumed by the cells, the linewidth of fusinite narrows giving an intracellular oxygen concentration corresponding to zero. From the results obtained, fusinite appears to represent a new extremely precise biophysical cellular oxygen probe which may prove useful in the understanding of the complex interrelationships between oxygen and normal cell physiology and/or pathology.

Carbon↗
PubMed1995

Vitamin E prevents UVB-induced cell blebbing and cell death in A431 epidermoid cells.

Cultured A431 epidermoid cells exposed to UVB (120-2400 J/m2) develop numerous blebs on their surface, detach from the plastic dish, and undergo injury and death. Numerous detached cells display fragmented nuclei, typical of apoptotic cells. Since bleb formation also occurs after oxidative stress it was assumed that the morphological variations observed are the consequence of free radical-mediated insult. In order to test this hypothesis, the antioxidant alpha-tocopherol (vitamin E) was added to cell cultures at different times, before or after irradiation. The results indicate that vitamin E inhibits UVB-induced surface blebbing as well as cell detachment from the substrate. Moreover, vitamin E is most effective in stimulating cell recovery when it is added after the end of UVB irradiation. Finally, vitamin E treatment also seems to reduce the fraction of cells undergoing death (probably those which will undergo apoptosis) after exposure to UVB radiation.

Antioxidants↗
PubMed1994

Different susceptibilities to cell death induced by t-butylhydroperoxide could depend upon cell histotype-associated growth features.

The effects of the oxidizing agent t-butylhydroperoxide (t-BHP) were investigated on three human cell lines of different origin and growth features (A431 epithelial cells, ADF astrocytoma cells and U937 leukemic cells) using electron microscopy and electron paramagnetic resonance spectroscopy. The results indicate that important biophysical and ultrastructural modifications are induced in the plasma and mitochondrial membranes of these cells and that these changes can ultimately lead to cell death. In addition, the cell cytoskeleton also appears to be a target of hydroperoxide-mediated stress. In particular, all three cell types undergo cytoskeletal alterations leading to surface blebbing, a typical characteristic of cell damage. However, the timing and extent of this damage as well as that occurring at the mitochondrial and plasma membrane levels seems to be different: cells with weak (ADF) or absent (U937) cell-to-cell and cell-substrate contacts and a poorly developed cytoskeleton appear to be more susceptible than other cell types (e.g., A431) to t-BHP-mediated injury. These diverse cell susceptibilites to hydroperoxide-mediated oxidative stress could thus depend upon cell histotype-associated growth features.

Actin Cytoskeleton↗
PubMed1994

Both UVA and UVB induce cytoskeleton-dependent surface blebbing in epidermoid cells.

Data on the morphological changes induced by UVA or UVB irradiation of A431 epidermoid cells in culture are presented. After irradiation with different doses of UVB (120-2400 J m-2) or UVA (10(4)-10(5) J m-2), the membrane and cytoskeleton of these cells were analysed by immunofluorescence and scanning electron microscopy at different times after exposure (0-48 h). Both UVA and UVB alter microtubules and microfilaments and surface blebs are formed after UV irradiation. In particular, UVB induces multiple small blebs on the cells, while UVA induces one single large bleb on each cell. Since cytoskeletal damage and surface blebbing of this type are also induced by oxidative stress, these results add to the body of evidence indicating that UV radiation is capable of pro-oxidant behaviour. Specifically, the morphological changes described in this paper are reminiscent of the modifications which accompany epidermal keratinocytes during their transformation to sunburn cells after UV irradiation. The physiological implications of these findings are discussed.

Actins↗
PubMed1994

A static magnetic field does not affect the dielectric properties of chick embryo myoblast membranes.

We have recently demonstrated using dielectic relaxation studies in the radiowave frequency range that sinusoidal 50 Hz magnetic fields (with intensities ranging from 1 to 10 mT) induce a nonlinear change in both membrane conductivity and permittivity of primary chick embryo myoblasts in vitro. It was the aim of the present study to determine if a DC-induced static magnetic field is capable of generating similar variations in the membrane conductivity and/or the membrane permittivity of chick embryo myoblasts. The results indicate that when the myogenic cells are exposed to a static magnetic field of either 1, 3 or 5 mT (values comparable with the previous extremely low frequency study), no changes in the membrane electrical parameters can be observed with respect to controls. Differences in the characteristics of static and extremely low frequency fields as well as the possible mechanisms underlying the contrasting results with these two types of magnetic fields are discussed.

Animals↗
PubMed1994

Membrane electrical properties associated with insulin receptor downregulation in human erythrocytes.

This study investigated the alterations in the electrical properties of human erythrocytes that occur during the insulin-induced internalization of insulin receptors (downregulation). Using a dielectric relaxation technique based on the Maxwell-Wagner effect, the data indicate a steady decrease over time in membrane conductivity, membrane permittivity, and conductivity of the cytosol in erythrocytes that have been stimulated to undergo downregulation by the addition of insulin. We hypothesize that variations in active transport properties as well as changes in membrane lipids and/or proteins occur during the process of insulin-induced downregulation of insulin receptors.

Cell Membrane↗
PubMed1994

Cytoskeletal oxidative changes lead to alterations of specific cell surface receptors.

Electron acceptors in biological systems, termed "oxidants", have been associated with numerous cytopathic conditions. In particular, oxygen-derived free radicals have been shown to induce various subcellular oxidative injuries which can lead to different types of pathologies. It has also been suggested that oxidative stress induced by xenobiotics can determine specific subcellular alterations in in vitro models. An oxidative imbalance in cells seems in fact to affect intracellular functions including the expression of some growth factor surface receptors. In this work, we observed a specific early alteration of the microfilament system and the microtubular network induced in K562 cells by oxidative stress. In particular, we hypothesize that oxidative imbalance could lead to an impairment of the expression of these receptors, such as transferrin receptors, via a modification of the cytoskeleton. This could represent a general mechanism by which a modification of receptor regulation can lead to cell aging, injury or death.

Cell Membrane↗
PubMed1993

Down-modulation of CD4 antigen during programmed cell death in U937 cells.

It has been hypothesized that programmed cell death (PCD), an active cell suicide process occurring in place of necrosis, can be associated with the pathogenesis of acquired immunodeficiency syndrome (AIDS). The entry of human immunodeficiency virus (HIV) into competent cells is mediated by the CD4 molecule present on the surface of certain lymphocyte subpopulations as well as on some cultured cell lines, e.g. U937 myelomonocytic cells. The present paper focuses on some specific aspects of PCD induced by the cytokine tumor necrosis factor (TNF). The results obtained indicate that the exposure of U937 cells to cycloheximide facilitates TNF-mediated PCD via a short term cell death program and modifies the expression of CD4 surface molecules. This change in surface antigen expression, manifested by internalization of the CD4 molecule, occurs in cells in which apoptosis has been triggered, but not in cells undergoing necrosis. These results indicate that the progression of cell death could be associated with specific alterations of certain surface molecules and could have a role in the entry of HIV into cells.

Apoptosis↗
PubMed1993

N-acetylcysteine inhibits apoptosis and decreases viral particles in HIV-chronically infected U937 cells.

Apoptosis or programmed cell death (PCD) is a type of death occurring in various physiological processes. Several data suggest that: (1) apoptosis may play a critical role in AIDS pathogenesis; (2) an increase of endocellular free radical levels can be associated with activation of previously latent HIV virus. Tumor necrosis factor (TNF), a cytokine capable of inducing oxygen free radicals and apoptosis, appears also to be involved in HIV activation. The present findings, which elucidate a relationship between the percentage of apoptotic cells, reduced glutathione (GSH) depletion and an increase of p24 antigenemia, suggest that pretreatment with N-acetylcysteine (NAC) is capable of decreasing the above-mentioned phenomena in HIV-infected U937 cells.

Acetylcysteine↗
PubMed1993

A 50-Hz magnetic field induces structural and biophysical changes in membranes.

The effects of a 50-Hz extremely low frequency magnetic field on cultured K562 cells growing in suspension were studied by means of scanning electron microscopy and electron paramagnetic resonance spectroscopy. Exposure of K562 cells at 2.5 mT for periods to 96 hours induced significant changes in cell-surface structure and physiology without modification of proliferative capability as indicated by quantitative analysis. Thus extremely low frequency fields seem able to induce injurious, sublethal cell alterations, and the plasma membrane seems to play an important role in this effect.

Cell Division↗
PubMed1993

Human erythrocyte insulin receptor processing is affected by the oxidizing agent menadione.

Insulin-induced down-regulation of erythrocyte insulin receptors is a simplified model that can provide useful information on the cell surface regulative phenomena and on role of the plasma membrane and cytoskeleton in such physiological processes. Oxidative imbalance was examined since it was shown to play an important role in numerous cellular pathologies as well as in cell aging. Specifically, the free radical inducer menadione was used in order to evaluate if this compound is able to modify (and in which manner) the down-regulation process. Biochemical, biophysical, and ultrastructural approaches were used. The results obtained seem to indicate that menadione-induced oxidative damage was able to decrease the insulin-induced down-regulation process, as measured by binding assays. This effect was accompanied by slight alterations in plasma membrane ultrastructure and insignificant variations in plasma membrane lipid composition. In addition, the decrease in membrane order, measured by electron paramagnetic resonance, which was shown to usually occur during the process of down-regulation, was not observed. In contrast, cytoskeletal protein assembly, as previously shown in other in vitro systems, appeared to be remarkably altered. Such changes in specific cytoskeletal elements could lead to the decrease of down-regulation phenomenon induced by menadione. Changes in electrophoretic pattern of some cytoskeletal proteins (e.g., spectrin) reinforce this hypothesis. Considering the importance of free radicals in cell injury, data reported here could represent a specific example of a general mechanism by which cell surface receptor expression and recycling can be modified by changes in some intracellular molecule redox status and cell ionic homeostasis.

Actin Cytoskeleton↗
PubMed1993

The fusion radiosensitivity of differentiating chick embryo myoblasts in vitro is not determined by the plasma membrane.

We have recently demonstrated by dielectric relaxation studies in the radiofrequency range that the sharp drop in the conductivity and permittivity of the membranes of chick embryo myoblasts in vitro, representative of fusion, is either delayed or completely blocked by sublethal doses of ionizing radiation (Santini et al. 1990a). The lowest of the doses investigated (3.25 Gy) caused a 10 h delay in myoblast membrane fusion when the cells were exposed at 24 h of culture, indicating that radiation-induced membrane injury had occurred. The purpose of this study was to determine if the myoblast system under investigation shows the same radiosensitive characteristics if irradiated with 3.25 Gy at various stages of differentiation. Consequently, the myoblasts were exposed to this dose at two different stages of differentiation (12 h or 48 h of culture). We show here that the time at which the myogenic cells are irradiated (state of differentiation) does not seem to affect the magnitude of the fusion delay (which was used as a measure of radiosensitivity of the myoblasts). In fact, the sharp drop in both membrane conductivity and membrane permittivity occurs with the same 10 h delay independent of the time of exposure. The role played by the plasma membrane in determining myoblast response to radiation damage is discussed.

Animals↗
PubMed1993

Menadione-induced oxidative stress leads to a rapid down-modulation of transferrin receptor recycling.

It has been demonstrated that perturbation of oxidative balance plays an important role in numerous pathological states as well as in physiological modifications leading to aging. In order to evaluate the role of the oxidative state in cells, biochemical and ultrastructural studies were carried out on K562 and HL-60 cell cultures. Particular attention was given to the transferrin receptor, which plays an important role in cellular iron metabolism. In order to evaluate if oxidative stress influences the transferrin receptor regulation process, the free-radical inducer menadione was used. The results obtained seem to indicate that oxidative stress is capable of inducing a rapid and specific down-modulation of the membrane transferrin receptor due to a block of receptor recycling on the cell surface, without affecting ligand-binding affinity. These effects were observed in the early stages of menadione treatment and before any typical signs of subcellular damage, including surface blebbing, a well-known cytopathological marker of menadione-induced injury. The mechanisms underlying such phenomena appear to be related to cytoskeletal protein thiol group oxidation as well as to the perturbation of calcium homeostasis, both induced by menadione. It is thus hypothesized that the data reported here represent a specific example of a general mechanism by which cell surface receptor expression and recycling can be influenced by oxidative balance.

Actin Cytoskeleton↗
PubMed1993

Oxidative stress and transferrin receptor recycling.

Perturbation of the oxidative balance in biological systems plays an important role in numerous pathological states as well as in many physiological processes such as receptor activity. In order to evaluate if oxidative stress induced by menadione influences membrane receptor processes, a study was conducted on the transferrin receptor. Consequently, biochemical, biophysical and ultrastructural studies were carried out on different cell lines. The results obtained seem to indicate that oxidative stress is able of inducing a rapid and specific down-modulation of membrane transferrin receptor due to a block of receptor recycling on the cell surface without affecting binding affinity.

Biotechnology↗
PubMed1992

Changes in erythrocyte membrane lipid composition affect the transient decrease in membrane order which accompanies insulin receptor down-regulation.

We have recently demonstrated, using electron paramagnetic resonance (EPR) spectroscopy, that insulin receptor internalization in response to insulin incubation (down-regulation) in human erythrocytes is accompanied by a transient decrease in membrane order, as measured by the 2T' parallel order parameter. Since membrane lipids play such an important role in receptor internalization, we investigated the possible effects that an alteration of the normally-occurring lipid profile might have on down-regulation and the concomitant transient decrease in membrane order. Consequently, human erythrocytes enriched with cholesterol and erythrocytes from cirrhotic patients were examined, because both of these groups of cells have a higher cholesterol/phospholipid molar ratio (CH/PL) than controls. The 5-nitroxystearate spin label, which inserts into the lipid bilayer of cell membranes, was used to monitor changes in 2T' parallel for a 3-h period at 37 degrees C. We report here that both cholesterol-enriched and cirrhotic erythrocytes do not down-regulate, as demonstrated by binding assays, and that they do not show the typical transient decrease in membrane order observed in controls. The results seem to indicate that a more ordered membrane inhibits internalization of the insulin receptor in erythrocytes, and that an increase in membrane disorder is necessary for insulin receptor down-regulation.

Cholesterol↗
PubMed1992

Differences in membrane electrical properties between C3H 10T1/2 mouse embryo fibroblasts and their ionizing radiation and chemically transformed counterparts.

Membrane electrical properties of mouse embryo fibroblasts and their ionizing radiation and chemically transformed counterparts were investigated using dielectric relaxation measurements in the radio frequency range. This determination is possible because, in the radio frequency range, suspensions of cells in an electrolyte buffer show a conductivity dispersion due to interfacial polarization. An analysis of the experimental data based on a "single-shell" model showed that conductivity and permittivity of the membranes of both radiation and chemically transformed fibroblasts were lower than in normal cells. In addition, the conductivity of the cytoplasm was higher in both transformed cell types than in the normal mouse fibroblasts. We discuss the significance of these findings in view of the possible structural and functional modifications brought about by the process of neoplastic transformation.

Animals↗