PubMed Health⌕ Search

Biomedical subjects

M Fiorani

Publications and source records attributed to M Fiorani.

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

Hydrogen peroxide-and fetal bovine serum-induced DNA synthesis in vascular smooth muscle cells: positive and negative regulation by protein kinase C isoforms.

Hydrogen peroxide and fetal bovine serum stimulate DNA synthesis in growth-arrested smooth muscle cells with remarkably similar kinetics and cell density dependence. However, while stimulation with fetal bovine serum results in cell proliferation, that by H2O2 is followed by cell death. Depletion of conventional and novel protein kinase C isoforms, resulting from a long treatment with phorbol-12-myristate-13-acetate, further increases H2O2-induced DNA synthesis. On the other hand, the specific protein kinase C inhibitor calphostin C abolished the increased DNA synthesis promoted by fetal bovine serum or H2O2. H2O2 increases protein kinase C activity in smooth muscle cells. This effect is markedly reduced, but not abolished, by down-regulation of the alpha, delta and epsilon protein kinase C isoforms. Thus, the zeta isoform of protein kinase C, which is not down-regulated, may be responsible for the residual H2O2 stimulation of protein kinase C. In conclusion, the results obtained show that H2O2 stimulates protein kinase C activity and DNA synthesis in growth-arrested smooth muscle cells: these events are not followed by cell proliferation but rather by cell death. This H2O2 stimulated DNA synthesis appears to be negatively controlled by alpha, delta and epsilon isoforms and positively controlled by the zeta isoform of protein kinase C.

Cell Count↗
PubMed1995

High-performance capillary electrophoretic separation of proteins and peptides using a bonded hydrophilic phase capillary.

High performance capillary electrophoresis (HPCE) was applied to the separation of protein and peptide mixtures with molecular masses ranging from 1300 to 96000 Da using a new bonded hydrophilic phase capillary, CElect-P150. This coated capillary reduces the interaction between proteins and silanol groups in capillary walls, allowing a complete recovery of the proteins and peptides of interest. HPCE was also used for the analysis of a complex mixture of tryptic fragments and to monitor the process of enzymatic digestion. Moreover, using a CElect-P150 capillary, highly reproducible analysis was possible without preconditioning the capillary with acid or basic solutions before each new analysis.

Animals↗
PubMed1995

Mitochondria-bound hexokinase from rabbit reticulocytes is resistant to the inactivation induced by Fe(II)/ascorbate.

Exposure of rabbit reticulocytes to Fe(II)/ascorbate induced a pronounced decay in hexokinase activity. In reticulocytes, this enzyme is present in at least three different molecular forms, Ia, Ia* and Ib, sub-types of hexokinase type I, which show different intracellular distribution. Hexokinase Ia and Ib are soluble, whereas hexokinase Ia* is almost entirely bound to the mitochondria. Anion exchange chromatography of hexokinase from intact reticulocytes exposed to Fe(II)/ascorbate revealed a selective inactivation of forms Ia and Ib, whereas the form Ia* did not show any decay. Binding to the mitochondrial membrane seems to be responsible for the observed resistance of the form Ia* to the inactivation elicited by Fe(II)/ascorbate. Indeed, by using a cell-free system in which hexokinase Ia* was solubilized using Triton X-100, the decay in hexokinase activity induced by iron/ascorbate involved all three enzymatic forms.

Animals↗
PubMed1995

Rabbit brain hexokinase: evidence for the presence of two distinct molecular forms.

In mammalian brain tissue, most hexokinase is bound to the mitochondria and only a small amount of the enzyme is present in soluble form. In this study we report that, in rabbit brain, hexokinase is present in two distinct molecular forms, which we designated HKH and HKL, both of which are separable using hydrophobic interaction or anion-exchange chromatography. These two molecular forms can be detected when hexokinase is prepared at pH 7.4, whereas at pH 10.0 only the more hydrophobic form, HKH, is present. The two subtypes of hexokinase do not show significant differences in Km values for glucose and ATP, in Ki values for glucose-6-phosphate or in their molecular weights. HKH is able to rebind mitochondrial membranes, while HKL has lost this ability, suggesting that the hydrophobic peptide at the N-terminal has been removed. The susceptibility of the N-terminal peptide to proteolysis is completely inhibited by using antiproteolytic compounds, such as leupeptin or E-64. The results reported in this paper suggest that a cysteine protease, probably belonging to a the class of cathepsins, may be involved in the processing of bindable hexokinase to the non-bindable form in rabbit brain, and that the activity of this protease is pH-dependent.

Animals↗
PubMed1995

The effect of 50 Hz sinusoidal electric and/or magnetic fields on the rate of repair of DNA single/double strand breaks in oxidatively injured cells.

Exposure of cultured mammalian cells to 50 Hz electric (0.2-20 kV/m), magnetic (0.002-2 G), or combined electric and magnetic fields did not affect the rate of repair of DNA single strand breaks (SSB) induced by hydrogen peroxide. The same lack of effect was observed on the repair of both DNA SSBs and DNA double strand breaks (DSBs) in cells treated with the cocktail hydrogen peroxide/L-Histidine. These results indicate that exposure to electric and/or magnetic fields does not affect the machinery involved in the repair of DNA lesions in oxidatively injured cells thus suggesting that it is unlikely that field exposure might induce changes in the response of the cells to the tumor promoting or carcinogenic effects elicited by reactive oxygen species.

Animals↗
PubMed1994

Simultaneous high-performance capillary electrophoretic determination of reduced and oxidized glutathione in red blood cells in the femtomole range.

This paper describes a high-performance capillary electrophoretic (HPCE) method which allows a quick, simultaneous and quantitative determination of reduced (GSH) and oxidized (GSSG) glutathione in mammalian red blood cells using a Supelco-bonded hydrophilic phase capillary CElect-P150. The extraction procedure of GSH and GSSG from erythrocytes using Microcon-10 membranes is very simple and allows a correct evaluation of these compounds present in the red blood cells. Furthermore, the HPCE method does not require removal of the excess N-ethylmaleimide used to block the glutathione in its reduced state, making the simultaneous evaluation of GSH and GSSG possible in a very short time (ca. 4 min), with a sensitivity at femtomole level.

Animals↗
PubMed1994

Inactivation of rabbit red blood cell hexokinase activity promoted in vitro by an oxygen-radical-generating system.

Rabbit red blood cell hexokinase (EC 2.7.1.1) has been shown to be inactivated in vitro by incubating intact erythrocytes in the presence of an oxygen-radical-generating system represented by ascorbate and iron. It was interesting to note that among the glycolytic enzymes, only hexokinase was found to be susceptible to the action of oxygen radicals, suggesting that the loss of activity of this enzyme may be one of the first signals of cellular damage in rabbit red blood cells. This statement is supported by the fact that, under the experimental conditions used, we did not observe any significant plasma membrane lipid peroxidation nor intracellular proteolysis. Furthermore, mature erythrocytes are unable to synthesize hexokinase as well as other proteins de novo; therefore, the inactivation of this enzyme, which is both the first and one of the rate-limiting enzymes of the glycolytic pathway, could play an important role in determining metabolic impairment of red blood cells, with possible physiological implications. We also investigated the effect of various radical scavengers and antioxidants (glucose, vitamin E, dithiothreitol, flavonoids) which are able to influence the inactivation of hexokinase activity to different extents. Finally, under the experimental conditions used (90 min of incubation at 37 degrees C), we did not observe any difference in the hemolysis of rabbit red blood cells incubated in the presence or absence of ascorbate and iron (hemolysis was about 1% after 90 min of incubation), suggesting that the system used was able to furnish information about the cellular damage produced by oxygen radicals without provoking cell lysis.

Adenosine Triphosphatases↗
PubMed1994

Cell density-dependent regulation of ATP levels during the growth cycle of cultured Chinese hamster ovary cells.

Chinese hamster ovary (CHO) cells show a cell density-dependent modification of ATP levels during the growth cycle. Cells were seeded at a density of 500,000 cells/75 cm2 flask in 10 ml of growth medium and at various time intervals, samples were taken and assayed for cell number, for adenine (ATP, ADP, AMP) and pyridine (NAD+, NADP+) nucleotide levels and for the activity of some glycolytic enzymes. Glucose consumption was also evaluated. Experimental results indicated that the rate of cell growth was exponential for up to the 4th day of culture after which the cell number remained pratically unchanged up to the 9th day. Under these experimental conditions we found that, whereas the intracellular levels of NAD+ and some glycolytic enzymes were not significantly affected, a drop in ATP content was apparent after 48 hr of culture. The decline in ATP levels progressively increased, reaching a maximum after 4 days of culture, and then remained unchanged. In order to evaluate whether this effect on ATP was determined by a reduced availability of nutritional factors or was really a function of cell density, we also performed experiments similar to those reported above, with the exception that the cells were grown in 40 ml of culture medium. Under these experimental conditions, the exponential growth was longer (in comparison with the cell growth in 10 ml of medium) and a plateau was reached after 6 days of culture.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenine Nucleotides↗
PubMed1992

Induction/repair of strand breakage in mature and nascent DNA of cultured Chinese hamster ovary cells exposed to hydrogen peroxide.

Treatment of cultured mammalian cells with hydrogen peroxide results in the production of extensive DNA damage. Strand breakage was produced at the level of either nascent or mature DNA and the former target appeared slightly more resistant than the latter. Although inhibitors of the enzyme poly(ADP-ribose) polymerase similarly retarded the repair of such lesions, removal of DNA strand breaks was much slower for the newly synthesized DNA as compared to mature DNA.

Animals↗
PubMed1992

Laminar, columnar and topographic aspects of ocular dominance in the primary visual cortex of Cebus monkeys.

The representation of the two eyes in striate cortex (V1) of Cebus monkeys was studied by electrophysiological single-unit recordings in normal animals and by morphometric analysis of the pattern of ocular dominance (OD) stripes, as revealed by cytochrome oxidase histochemistry in V1 flat-mounts of enucleated animals. Single-unit recordings revealed that the large majority of V1 neurons respond to the stimulation of either eye but are more strongly activated by one of them. As in other species of monkey, neurons with preference for the stimulation of the same eye are grouped in columns 300-400 microns wide, spanning all cortical layers. Monocular neurons are clustered in layer IVc, specially in its deeper half (IVc-beta), and constitute less than 10% of the population of other layers. Neurons with equal responses to each eye are more commonly found in layer V than elsewhere in V1. In the supragranular layers and in granular layer IVc-alpha neurons strongly dominated by one of the eyes tend to be broadly tuned for orientation, while binocularly balanced neurons tend to be sharply tuned for this parameter. No such correlation was detected in the infragranular layers, and most neurons in layer IVc-beta responded regardless of stimulus orientation. Ocular dominance stripes are present throughout most of V1 as long, parallel or bifurcating bands alternately dominated by the ipsi- or the contralateral eye. They are absent from the cortical representations of the blind spot and the monocular crescent. The domains of each eye occupy nearly equal portions of the surface of binocular V1, except for the representation of the periphery, where the contralateral eye has a larger domain, and a narrow strip along the border of V1 with V2, where either eye may predominate. The orderliness of the pattern of stripes and the relationship between stripe arrangement and the representation of the visual meridians vary with eccentricity and polar angle but follow the same rules in different animals. These results demonstrate that the laminar, columnar and topographic distribution of neurons with different degrees of OD in V1 is qualitatively similar in New- and Old World monkeys of similar sizes and suggest that common ancestry, rather than parallel evolution, may account for the OD phenotypes of contemporaneous simians.

Animals↗
PubMed1992

Electric and/or magnetic field effects on DNA structure and function in cultured human cells.

Exposure of cultured K562 cells to 50 Hz electric (0.2-20 kV/m), magnetic (0.002-2 G), or combined electric and magnetic fields for up to 24 h did not result in the production of detectable DNA lesions, as assayed by the filter elution technique. The rate of cell growth was also unaffected as well as the intracellular ATP and NAD+ levels. These results indicate that, under the experimental conditions utilized in this study, 50 Hz electric, magnetic and electromagnetic fields are not geno- and cyto-toxic in cultured mammalian cells.

Adenosine Triphosphate↗
PubMed1990

Inhibition of Chinese hamster ovary cell DNA synthesis by hydrogen peroxide.

The DNA synthesis inhibitory effect of hydrogen peroxide has been examined under a number of experimental conditions. Results have indicated that the effect of the oxidant is more pronounced when the treatment is performed at 37 degrees C than at 4 degrees C and in low density as compared to high density cultures. In addition, similar levels of inhibition were achieved by measuring the incorporation of radiolabelled thymidine in the presence of, or following treatment with, the oxidant. Although early events seem to be responsible for the decreased rate of DNA synthesis, it would appear that hydrogen peroxide does not alter thymidine extracellularly and/or decrease the transport of the nucleoside across the plasma membrane, which may actually be slightly augmented. Thus, the previously illustrated results may represent an underestimate of the actual capacity of the oxidant to reduce DNA synthesis. This inference is further supported by the fact that the effect of hydrogen peroxide appears markedly enhanced in cells preloaded with the radiolabelled precursor. A temporal relationship seems to exist between the steady state level of DNA single strand breaks and the extent of DNA synthesis inhibition by hydrogen peroxide. The oxidant has no effect on DNA chain elongation. In conclusion, data presented in this paper suggest that early events, involving selective effects on replicon initiation, mediate the DNA synthesis inhibitory effect of hydrogen peroxide.

Animals↗
PubMed1990

[Transitory myocardial ischemia appearing after cranial trauma. Description of a case].

The authors describe a case of ECG alterations, indicating myocardial ischaemia, in a patient affected by mild cranial trauma. Both cycloergometric test and angiography of coronary vessels were normal. The authors conclude that every patient affected by brain injury should be monitored by the Cardiological Department, because of the high frequency of myocardial damage and lethal arrhythmias in these patients.

Coronary Disease↗
PubMed1989

Visual area MT in the Cebus monkey: location, visuotopic organization, and variability.

The representation of the visual field in the dorsal portion of the superior temporal sulcus (ST) was studied by multiunit recordings in eight Cebus apella, anesthetized with N2O and immobilized with pancuronium bromide, in repeated recording sessions. On the basis of visuotopic organization, myeloarchitecture, and receptive field size, area MT was distinguished from its neighboring areas. MT is an oval area of about 70 mm2 located mainly in the posterior bank of the superior temporal sulcus. It contains a visuotopically organized representation of at least the binocular visual field. The representation of the vertical meridian forms the dorsolateral, lateral, and ventrolateral borders of MT and that of the horizontal meridian runs across the posterior bank of ST. The fovea is represented at the lateralmost portion of MT, while the retinal periphery is represented medially. The representation of the central visual field is magnified relative to that of the periphery in MT. The cortical magnification factor in MT decreases with increasing eccentricity following a negative power function. Receptive field size increases with increasing eccentricity. A method to evaluate the scatter of receptive field position in multiunit recordings based on the inverse of the magnification factor is described. In MT, multiunit receptive field scatter increases with increasing eccentricity. As shown by the Heidenhain-Woelcke method, MT is coextensive with two myeloarchitectonically distinct zones: one heavily myelinated, located in the posterior bank of ST, and another, less myelinated, located at the junction of the posterior bank with the anterior bank of ST. At least three additional visual zones surround MT: DZ, MST, and FST. The areas of the dorsal portion of the superior temporal sulcus in the diurnal New World monkey Cebus are comparable to those described for the diurnal Old World monkey, Macaca. This observation suggests that these areas are ancestral characters of the simian lineage and that the differences observed in the owl monkey may be secondary adaptations to a nocturnal ecological niche.

Animals↗
PubMed1989

Morphological changes in Escherichia coli cells exposed to low or high concentrations of hydrogen peroxide.

Escherichia coli cells challenged with low or high concentrations of hydrogen peroxide are killed via two different mechanisms and respond with morphological changes which are also dependent on the extracellular concentration of the oxidant. Treatment with low concentrations (less than 2.5 mM) of H2O2 is followed by an extensive cell filamentation which is dependent on the level of H2O2 or the time of exposure. In particular, addition of 1.75 mM H2O2 results in a growth lag of approximately 90 min followed by partial increase in optical density, which was mainly due to the onset of the filamentous response. In fact, microscopic analysis of the samples obtained from cultures incubated with the oxidant for various time intervals has revealed that this change in morphology becomes apparent after 90 min of exposure to H2O2 and that the length of the filaments gradually increases following longer time intervals. Analysis of the ability of these cells to form colonies has indicated a loss in viability in the first 90 min of exposure followed by a gradual recovery in the number of cells capable of forming colonies. Measurement of lactate dehydrogenase in culture medium (as a marker for membrane damage) has revealed that a small amount of this enzyme was released from the cells at early times (less than 150 min) but not after longer incubation periods (300 min). Cells exposed to high concentrations of H2O2 (greater than 10 mM) do not filament and their loss of viability is associated with a marked reduction in cell volume. In fact, treatment with 17.5 mM H2O2 resulted in a time-dependent decrease of the optical density, clonogenicity, and cellular volume. In addition, these effects were paralleled by a significant release in the culture medium of lactate dehydrogenase thus suggesting that the reduced cell volume may be dependent on membrane damage followed by loss of intracellular material. This hypothesis is supported by preliminary results obtained in electron microscopy studies. In conclusion, this study further demonstrates that the response of E. coli to hydrogen peroxide is highly dependent on the concentration of H2O2 and further stresses the point that low or high concentrations of the oxidant result in the production of different species leading to cell death via two different mechanisms and/or capable of specifically affecting the cell shape.

Dose-Response Relationship, Drug↗
PubMed1988

Hypertensive emergencies in old age: effects of angiotensin converting enzyme inhibition.

Hypertensive emergencies, and to a certain extent their treatment, contribute to morbidity and mortality in elderly patients. We studied 22 hospitalized patients, aged 70-90 years, all of whom had moderate essential hypertensive. During acute hypertension, mean systolic and diastolic blood pressure rose to 230 +/- 24 and 120 +/- 22 mmHg, respectively. Symptoms of reduced tissue perfusion/oxygenation and/or organ failure occurred, forcing us to begin antihypertensive therapy. We administered 50 mg of the angiotensin converting enzyme (ACE) inhibitor captopril sublingually, and within 15 min, systolic blood pressure decreased by an average 60 +/- 16 mmHg and diastolic blood pressure by an average 25 +/- 14 mmHg. There was no significant change in the heart rate. In addition, we treated 22 comparable patients with 10 mg nifedipine sublingually and observed, in four cases, a greater fall in blood pressure (up to 90 mmHg) together with tachycardia. These results show the beneficial effects of captopril in the treatment of hypertensive emergencies in elderly patients. The absence of dangerous side effects indicates that ACE inhibitors can be used as first-choice drugs for the treatment of acute hypertensive crises, even in old age.

Administration, Sublingual↗