PubMed Health⌕ Search

Biomedical subjects

A Fraschini

Publications and source records attributed to A Fraschini.

At least 19 recordsLinked to original sources
PubMed2006

Enzyme-assisted photosensitization with rose Bengal acetate induces structural and functional alteration of mitochondria in HeLa cells.

Rose Bengal acetate (RB-Ac) can be used as a fluorogenic substrate for photosensitization of cells both in vivo and in vitro: once inside the cells, RB-Ac is converted into photoactive rose Bengal (RB) molecules which redistribute dynamically in the cytoplasm and, upon irradiation by visible green light, can damage organelles such as the endoplasmic reticulum, the Golgi apparatus, and the cytoskeleton. Recently, evidence has been provided that mitochondria may also be affected. The aims of the present study were to describe RB-induced photodamage of mitochondria in single HeLa cells and to define, on a quantitative basis, the effects of photosensitization on their morphofunctional features. HeLa cell cultures were exposed to 10(-5) M RB-Ac for 60 min and then irradiated with a light emitting diode at 530 nm (total light dose, 1.6 J/cm2). After irradiation, the cells were transferred to a drug-free complete medium and allowed to grow for 24-72 h. Using conventional and confocal fluorescence microscopy, transmission electron microscopy, and flow cytometry, we demonstrate that, in photosensitized cells, mitochondria undergo structural and functional alterations which can lead cells to apoptosis. Interestingly, in our system some cells were able to survive 72 h post-treatment and to recover, exhibiting the same mitochondrial structure, distribution and inner membrane potential as those in untreated controls. Taking into account that the photoactive molecules redistribute dynamically inside the cell upon RB-Ac administration, it may be hypothesized that cells can be differently affected by irradiation, depending on the relative amount and organelle location of the photosensitizer.

Flow Cytometry↗
PubMed2005

Changes in extranucleolar transcription during actinomycin D-induced apoptosis.

Actinomycin D (AMD) inhibits DNA-dependent RNA polymerases and its selectivity depends on the concentration used; at very high concentrations it may also induce apoptosis. This study investigates the effects of different concentrations (0.01 to 1 microg/ml) of AMD on RNA transcription and maturation and on the organization of nuclear ribonucleoproteins (RNPs), and their relationship with apoptosis induction. Human HeLa cells were used as a model system. At the lowest concentration used, AMD induced the segregation of the nucleolar components and impaired r-RNA synthesis, as revealed by the decreased immunopositivity for bromo-uridine incorporation and for DNA/RNA hybrid molecules. The synthesis of pre-mRNAs, on the contrary, was active, while the immunolabeling of snRNP proteins and of the SC-35 splicing factor strongly decreased on perichromatin fibrils (where they are involved in co-transcriptional splicing). This suggests that the post-transcriptional maturation of extranucleolar RNAs was also affected. Moreover, still in the absence of typical late morphological or biochemical signs of apoptosis (i.e. chromatin condensation), these cells displayed the early apoptotic features, i.e. the externalization of phosphatidylserine residues on the plasma membrane and propidium iodide exclusion in vivo. At the highest concentrations of AMD used, apoptosis massively occurred, with the typical morphological events (progressive chromatin condensation, clustering of snRNPs and SC-35 splicing factor, cell blebbing). However, transcription of hnRNAs was maintained in the residual areas of diffuse chromatin up to advanced apoptotic stages. The inhibition of rRNA synthesis and the defective pre-mRNA maturation seem to be part of the apoptotic process induced by AMD.

Antibiotics, Antineoplastic↗
PubMed1999

Nuclear phospholipids in human lymphocytes activated by phytohemagglutinin.

Using a specific ultracytochemical technique, the labelling with phospholipase A2-gold complex, we have followed nuclear phospholipids (PL) along the G1 phase in human lymphocytes activated by PHA. Our data point out two main results relating nuclear PL to the transcriptional activity, characteristic of the G1 phase, during which many different molecules necessary both for progression through G1 and for the start of S phase are synthesized. PL quantitative changes parallel those of hnRNPs and snRNPs, which are markers of the levels of transcriptional activity and processing. We found that nuclei of G0 lymphocytes, with a very low transcription level, are poor of PL as well as of RNPs. The amount of PL increases in activated lymphocytes, along all G1, until the beginning of S phase. At the same time, hnRNPs and snRNPs strongly increase and maintain higher levels than in control cells, till the beginning of S phase. PL are localized on nuclear structures where also RNPs involved in transcription and splicing, are located, i. e. perichromatin fibrils, interchromatin granules and the dense fibrillar component of the nucleolus. Since it is known that during S phase nuclear PL decrease, while both the enzyme activities related to their breakdown and their hydrolysis products increase, PL seem to be involved in the generation of signal molecules triggering DNA replication. We suggest that PL in the nucleus can be involved in multiple functions, depending on the phase of the cell cycle.

Cell Cycle Proteins↗
PubMed1996

Facts and paradoxes in current notions of nuclear organization and function.

Invisible compartments, identified rather by their activities than by their morphology, seem to operate in the nucleus. These compartments interrelate somehow, including mediation by the nuclear matrix. As our knowledge about the nucleus increases, more paradoxes become evident. We here consider some of them: 1) the well-known C-paradox of Cavalier-Smith, concerning the disproportionate amount of nuclear DNA content in comparison with the amount of DNA potentially able to transcribe; 2) the DNA folding in the chromatin fibre and its superorganization within the nucleus, which seems to be in opposition with the transcribing and self-replicating activities; 3) the elusive role of the DNA sequences with different degrees of repetitivity; and 4) the compartmentalization in the nucleus and how it relates to transcription, processing and transport of transcripts, and to DNA reduplication. We conclude by introducing the concept of species specific, minimal, but essential genome components, i.e. the elusive few thousand DNA bases that, in our hypothesis, act as a functional bridge between the nuclear matrix and chromatin.

Animals↗
PubMed1995

Nuclear phospholipids during the adaptation of human EUE cells to hypertonic stress.

The phospholipid component of interphase nuclei was analysed in EUE cells (an established cell line from embryonic human epithelium) grown in an isotonic culture medium and during the adaptation process to a hypertonic medium, using a highly specific ultracytochemical procedure, viz. labelling with the phospholipase A2 gold-complex. Within the nucleus, the phospholipids were localized in domains involved in different steps of the synthesis and processing of the RNA. These localizations did not vary at the two key steps of the adaptation process to hypertonic medium: short-term treatment (6 h) representing critical shock condition, and long-term growth (5 days) representing the adapted cells under survival conditions. On the contrary, deep changes of the labelling intensity of phospholipids at these sites occurred at the different times of hypertonic treatment and followed the same course as those observed in the ultramorphological patterns of transcription: the chromatin condensation, as evaluated by image analysis, the permanent nucleolar components, the interchromatin and the perichromatin granules. These data endorse the hypothesis that nuclear phospholipids could be involved in different steps of the transcriptional activity. They are indicative of the deep changes occurring in the EUE cells submitted to hypertonic stress.

Adaptation, Physiological↗
PubMed1992

TEM cytochemical study of the localization of phospholipids in interphase chromatin in rat hepatocytes.

The electron microscopy cytochemical detection of phospholipids in well-defined areas in the interphase nuclei of hepatocytes has been obtained by the acid haematein test, modified for electron microscopy and by the phospholipase A2-colloidal gold method. The specificity of both methods were controlled by enzymatic digestion with phospholipase. The main intra-nuclear localization of phospholipids is at the border between the condensed and dispersed chromatin, where non-ribosomal RNA is also revealed by RNase-gold labelling. Phospholipids are detected, too, over the clusters of interchromatin granules and in the fibrillar component of the nucleolus.

Animals↗
PubMed1991

A 33 kDa protein band is enhanced during long-term adaptation of EUE cells to a hypertonic medium.

A cell line derived from human embryonic epithelium (EUE cells) shows an enhanced expression of a 33 kDa protein when adapted to grow in a hypertonic medium containing 0.246 M NaCl (1.8 x the isotonic concentration). The maximum amount of this protein, followed by SDS-PAGE electrophoresis, was found after 4 days of adaptation; thereafter, the protein band remained fairly constant up to 30 days. When the cells were transferred back to a medium containing 0.137 M NaCl (isotonic medium), the protein pattern reverted to that of control cells. This protein is mainly localized in the cytosol, although a small part is associated with the 150,000 g pellet and needs detergents to be extracted. The molecular weight and the cellular location suggest a possible analogy with the so-called amphitropic proteins, that are known to interact with both the epidermal growth factor receptor and hydrophobic structures, such as the membrane phospholipids and the cytoskeletal components.

Adaptation, Physiological↗
PubMed1990

Changes of gene expression during long term adaptation of human EUE cells to a hypertonic medium: electrophoretic protein patterns and DNase I digestion in situ.

The effects of long term (1 to 10 days) growth in a hypertonic medium have been studied in human EUE cells. Following polyacrylamide gel electrophoresis, a change in the protein pattern has been found, with the progressive enhancement, during adaptation, of a 33 kDa band. Experiments of DNA digestion in situ by DNase I showed that chromatin DNA of cells grown in a hypertonic medium is more available to the enzyme cleavage. These findings show that long term hypertonic stress is able to induce a change in gene expression in EUE cells.

Cells, Cultured↗
PubMed1989

Free perforation due to colonic adenocarcinoma in Crohn's disease.

Free perforation of the bowel due to cancer in Crohn's disease is very rare. One case found among 75 cases of Crohn's disease is reported. The perforation occurred in a mucinous adenocarcinoma of the ascending colon in a 54-year-old woman with a 17-year history of Crohn's disease, who had undergone two previous operations of small bowel and colonic resection. A right hemicolectomy was performed; the patient had an uneventful recovery and does not show recurrence of cancer at 31-month follow-up. This is in contrast with a recent review, in which a very poor prognosis was claimed for the perforated colonic cancer in Crohn's disease.

Adenocarcinoma, Mucinous↗
PubMed1988

[Arterial hypertension with hypokalemia].

After a careful review of the literature on physiopathological role of the renin-angiotensin-aldosterone system in cases of arterial hypertension with hypokalaemia, a simple diagnostic procedure is suggested to differentiate between this and secondary hypertensions which, though few, are often curable. Three clinical cases examined in the medical department of Casalpusterlengo Hospital are presented in which the study of the renin-angiotensin system proved decisive for a correct diagnosis of the hypertension.

Adenocarcinoma↗
PubMed1986

Sperm-chromatin maturation in the mouse. A cytochemical approach.

Cytochemical techniques were used to study chromatin during spermiogenesis and sperm maturation in the mouse, starting from the stages at which the substitution of somatic histones by testis-specific proteins occurs. It was possible to distinguish and analyze the different temporal incidence of two processes involved in sperm maturation, i.e. chromatin condensation (a tridimensional highly compacted arrangement) and chromatin stabilization (a tough structure, which protects the genome DNA). The first process, involving a reduction in the nuclear size and a decrease in the amount of sperm DNA accessible to specific cytochemical reactions and stainings, was found to reach its maximum in caput-epididymidis spermatozoa, in which electron microscopy revealed that the sheared chromatin was mainly organized into 120-A-thick knobby fibers. No further changes were found in sperm up to their appearance in the fallopian tubes. On the contrary, chromatin stabilization, the onset of which occurs in the testis (at the late spermatid stage) via the formation of -S-S- cross-links, is completed in the vas deferens, where chromatin has a superstructure consisting of thicker fibers, with diameters of 210 and 350 A. The reductive cleavage of disulfides in vas-deferens spermatozoa does not completely destroy the superstructure of sperm chromatin, which could indicate 'coiling' of the basic knobby fiber. In fact, when the ion concentration was increased, the chromatin of vas-deferens spermatozoa appeared to be organized into fibers with diameters similar to those of the caput epididymidis. This unique organization of mature sperm chromatin should have an essential role in the fast swelling of spermatozoa during fertilization.

Animals↗
PubMed1985

Ultrastructural analysis of mouse sperm chromatin.

Chromatin organization was studied during the maturation processes in the epididymis and vas deferens; these processes lead to a potentially reversible inactivation of the genome. There are progressive increases in resistance to detergent action and -S-S- bridge reduction in both head membranes and chromatin. In all the sites studied, there was a basic "knobby" chromatin fiber of 110 A diameter. In the caput epididymidis only, in addition to the knobby fibers, there were some smooth fibers, which can be considered as markers of a transient situation in which the stabilization of DNA/protamine interactions has not completely been achieved. In the vas deferens, the knobby fibers, the diameters of which are multiples of that of the basic one, can be converted to single units by increasing the ionic strength.

Animals↗
PubMed1981

The effect of different fixatives on chromatin: cytochemical and ultrastructural approaches.

This study explores the effects of two types of fixative on chromatin. The first type (acrolein, glutaraldehyde) engenders a high degree of ultrastructural preservation. The other type are fixatives that are widely used in cytochemistry and cytogenetics (acetic acid, 3:1 by vol. methanol-acetic acid, methanol alone, formaldehyde). Lymphocytes of adult rats so-fixed in vitro were prepared for electron microscopy or microdensitometric evaluations of smears. Assessments were made of variations in their total protein, nuclear basic protein and DNA contents. DNA was determined both as Feulgen-positive material and by its binding of intercalating dyes (Methyl Green, specific for double-stranded polynucleotides). Our results showed that some fixatives break up the chromatin organization by acting on particular components of chromatin fibres. They can thus be considered to be destructive agents in situ. In addition, a revaluation of some aldehyde fixatives is proposed for both ultrastructural and cytochemical research.

Acrolein↗
PubMed1981

Analysis of some cytotopochemical parameters for a definition of the chromatin status.

A study of the curves of Feulgen hydrolysis kinetics has been performed on different cell types of the same animal species and on the same cell type from different species at different taxonomical level. The curves were analyzed by a computer programme of least square fit to the Bateman's function, which gives a description of DNA hydrolysis kinetics by three parameters only: 1) amount potentially stainable groups; 2) depurination rate constant; 3) depolymerization rate constant. The chromatin homogeneity within the nucleus seems to affect these parameters, together with the degree of chromatin compaction. Furthermore the thermal denaturation (100 degrees C), also after fractionate histone extraction, shows a resistant DNA fraction, which does not appear to be affected by the degree of chromatin compaction only.

Animals↗
PubMed1981

Some parameters for a cell cycle cytotopochemical study.

With the Feulgen-Naphthol Yellow double reaction we dosed, simultaneously on a cell, [1] the DNA nuclear content (as Feulgen positive material), [2] the protein cell content and [3] the Feulgen chromatin area; the degree of condensation is expressed by the ratio [1]/[3]. The evolution of the lymphocytes culture at different times of PHA stimulation was in this way studied with the aim of defining the existence of cell subpopulations pertaining at different phases or subphases of the cycle.

Blood Proteins↗