Biomedical subjects

L Dini

Publications and source records attributed to L Dini.

At least 55 records · Page 3Linked to original sources

PubMed1995

Immunocytochemical study of binding and internalization of carrier-free Cu, Zn Superoxide dismutase by cultured rat hepatocytes.

Internalization of superoxide dismutase-gold complex by isolated liver cells has been shown to occur via receptor-mediated endocytosis. As colloidal gold may act as a carrier in this process, carrier-free human Cu, Zn superoxide dismutase was incubated with cultured rat hepatocytes. Light and electron microscopy immunocytochemistry revealed the binding and internalization of free native human Cu, Zn superoxide dismutase (hSOD) by cultured rat hepatocytes. Immunocytochemical demonstration of binding to the cell surface (hepatocytes were incubated with hSOD for 15 min. at 4 degrees C) and internalization (hepatocytes were incubated with hSOD for 15, 30 and 60 min. at 37 degrees C) of the carrier-free superoxide dismutase was achieved by using a monoclonal antibody selectively reacting with the human protein. The results obtained indicate that carrier-free superoxide dismutase is bound and internalized by rat hepatocytes in primary cultures and that the enzyme can enter the cells via a receptor-mediated endocytosis pathway. We followed the binding and the internalization process of hSOD thus validating the use of the native enzyme in the therapy of free radical-related diseases.

Animals↗

PubMed1994

Macrophage colony-stimulating factor enhances the susceptibility of macrophages to infection by human immunodeficiency virus and reduces the activity of compounds that inhibit virus binding.

The effects of macrophage colony-stimulating factor (M-CSF) on CD4 receptor expression, susceptibility to human immunodeficiency virus type 1 (HIV) infection, and anti-HIV activity of dextran sulfate and soluble-CD4 were studied in cultured, human primary macrophages. M-CSF stimulated macrophage cells to express the CD4 receptor, and this resulted in an increase of both the number of CD4+ cells and the density of the receptor on the cell surface. M-CSF also significantly enhanced the susceptibility of macrophage cells to HIV infection. Interestingly, the anti-HIV activity of dextran sulfate and soluble-CD4 (two compounds that interfere with HIV-CD4 binding with different mechanisms) was reduced 100-fold and fivefold, respectively, in M-CSF-treated macrophages. Human blood concentrations of M-CSF are reported to be similar to those used in this work (1,000 U/mL); thus, it is conceivable that also in vivo this cytokine may modify the susceptibility of macrophages to HIV and the ability of dextran sulfate and soluble CD4 to inhibit HIV replication. These results suggest that the in vitro study in M-CSF-treated macrophages of promising drugs inhibitors of HIV-CD4 binding could provide further insights into the potential efficacy of these compounds in patients.

Antigens, CD↗

PubMed1994

Atypical acrofacial dysostosis syndrome.

We describe a male fetus with a combination of defects, including severe mandibulofacial dysostosis, holoprosencephaly, upper limb deficiency, and microgastria. These abnormalities indicate a severe defect of gastrulation/blastogenesis predominantly affecting cephalad structures. This combination of anomalies has to our knowledge not previously been described. We propose that the anomaly pattern represents either a blastogenesis-related association, or a microgastria-limb deficiency polytopic field defect.

Abnormalities, Multiple↗

PubMed1994

Regulation of asialoglycoprotein receptor expression in rat hepatocytes cultured under proliferative conditions.

The expression of many glycoproteins on the surface of hepatocytes has been related to the state of differentiation of the liver. In particular, the asialoglycoprotein receptor (ASGP-R) is modulated in many physiological and pathological conditions in which hepatocyte proliferative activity is modified. We studied ASGP-R expression in primary monolayer cultures of rat hepatocytes stimulated to proliferate by the addition of epidermal growth factor and insulin. In proliferating hepatocytes the receptor concentration on the cell surface was lowered, with no modifications in its distribution; similarly, both the binding activity and the amount of specific transcripts were decreased. The decreases were related to the peak of cellular growth, as judged by [3H]thymidine incorporation into DNA. The results suggest the presence of a cell cycle-dependent ASGP-R expression also in in vitro systems; this control could be exerted by a decreased availability of receptor-specific mRNA molecules or on the stability of transcripts.

Animals↗

PubMed1994

Cystamine potently suppresses in vitro HIV replication in acutely and chronically infected human cells.

We have investigated the effects of cystamine on the replication of human immunodeficiency virus (HIV) in human lymphocytes and macrophages, the natural targets of HIV in vivo. Treatment of chronically infected macrophages with cystamine, at a concentration (500 microM) that did not show any cytotoxic or cytostatic effects, strongly decreased (> 80%) HIV-p24 antigen production and completely abolished the production of infectious viral particles. Cystamine does not affect viral transcription, translation or protein processing; indeed, all HIV proteins are present in a pattern similar to that of nontreated cells. Instead, cystamine interferes with the orderly assembly of HIV virions, as shown by electron microscopy analysis, that reveals only defective viral particles in treated cells. Moreover, suppression of HIV replication, due to the inhibition of proviral DNA formation was observed in acutely infected lymphocytes and macrophages pretreated with cystamine. These results show that cystamine potently suppresses HIV replication in human cells by contemporaneously blocking at least two independent steps of the viral life cycle, without affecting cell viability, suggesting that this compound may represent a new possibility towards the treatment of HIV-1 infection.

Antiviral Agents↗

PubMed1993

Modulation of the expression of galactose-specific receptors on Kupffer cells after partial hepatectomy and zymosan stimulation.

Partial hepatectomy (PH) and a single intravenous injection of zymosan were used to provoke expansion of the Kupffer cell population in rat liver. The number of Kupffer cells per unit of area of liver increased after both stimulations. During these growth phases the number of binding sites for galactose-exposing ligands was studied on Kupffer cell plasma membranes. Both partial hepatectomy and zymosan stimulation affected the binding of lactosylated bovine serum albumin conjugated to gold particles (LacBSA-Au5). The reduction observed depended on the time after the PH or zymosan injection studied. Low levels of binding sites were found at 24 h from PH and 5 days after zymosan stimulation and represented only 1/4 or 1/10 of the binding sites expressed on the cell surface of Kupffer cells from normal adult rats. In addition, in both experimental situations we observed that the clustered distribution of gold particles typical of adult Kupffer cell was changed.

Animals↗

PubMed1993

Galactose-specific receptor modulation related to the onset of apoptosis in rat liver.

The expression of the asialoglycoprotein receptor of hepatocytes and the galactose-specific receptors of non-parenchymal liver cells during the onset of apoptosis in liver of rats treated with lead nitrate was studied. During the involution of lead nitrate-induced hyperplasia in rat liver (occurring at 5 days after the injection) a significant increase of asialoglycoprotein receptor (ASGP-R) expression on hepatocytes coincided with the massive death by apoptosis of the same cells. The increase in the receptor expression was sustained by a large increase in the level of its specific mRNA. As a consequence of lead nitrate injection, we also detected a drastic change of the galactose-specific receptor expression and distribution on the surface of rat liver sinusoidal cells. However, the modulation of the receptor expression on the Kupffer cells did not parallel that observed for the ASGP-R: the peak of surface expression measured on hepatocytes always followed the one observed on Kupffer cells. Our data show a first evidence of a receptor modulation during the process of apoptosis. In fact, the entire carbohydrate recognition system of the liver is modulated during the onset of apoptosis induced by lead nitrate injection, but the pattern of modulation depends on the cellular types. We suggest that a physiological role for the hepatic carbohydrate recognition systems is related to the apoptosis of liver.

Animals↗

PubMed1992

Insulin effect on isolated rat hepatocytes: diacylglycerol-phosphatidic acid interrelationship.

It is widely accepted that insulin action does not involve inositol phospholipid hydrolysis through the stimulation of a phosphatidylinositol-specific phospholipase C (PI-PLC). This consideration prompted us to investigate the insulin effect on the mechanism leading to the accumulation of diacylglycerol (DAG) and phosphatidic acid (PA) in rat hepatocytes. Basically, insulin induces: (i) a significant increase of both [3H]glycerol and fatty acid labelling of DAG; (ii) a significant increase of PA labelling preceding DAG labelling and paralleled by a decrease of phosphatidylcholine (PC) labelling. These observations, which suggest an insulin-dependent involvement of a phospholipase D, are strengthened by the increase of PC-derived phosphatidylethanol in presence of ethanol. Finally, the observation that the PA levels do not return to basal suggests that other mechanisms different from PC hydrolysis, such as the stimulation of direct synthesis of PA, may be activated.

Animals↗

PubMed1992

Interaction of DNA with cationic liposomes: ability of transfecting lentil protoplasts.

The vesicle made of dipalmitoylphosphatidylcholine and stearylamine (9:1) were multilamellar and rather homogeneous in shape as seen by transmission electron microscopy. Upon addition of circular DNA plasmids of different lengths to the liposomes, the formation of vesicle clusters around the DNA filament was observed, with dimensions dictated by the ratio DNA/lipid. These liposomes were able to transfect lentil (Lens culinaris) protoplasts inside the cells two different reporter genes, chloramphenicol-acetyltransferase and beta-glucuronidase. The activity of these two enzymes could be found in the cell lysates after 24 h from the incubation of protoplasts with the lipid-DNA complexes.

Chloramphenicol O-Acetyltransferase↗

PubMed1992

In vivo and in vitro induction of 'tissue' transglutaminase in rat hepatocytes by retinoic acid.

Tissue transglutaminase (tTG) expression was found to be induced in rat liver following in vivo retinoic acid (RA) treatment (Piacentini et al. (1988) Biochem. J. 253, 33-38). Here we show that the increased enzyme expression in rat liver is at least partially the result of the action of RA in parenchymal cells. In fact, (a) when hepatocytes are isolated from RA-treated animals their transglutaminase protein content is much higher than in similarly isolated control cells; (b) higher tTG protein level is also found by immunoelectronmicroscopy in the hepatocytes of the RA-treated rats as compared with the very low amount detected in the controls; (c) RA induces tTG in hepatocytes under culture conditions as well. One of the functions of tTG is to form a protein polymer in dying apoptotic cells by epsilon(gamma-glutamyl)lysine and, specifically gamma-glutamylpolyamine cross-links (Fesus et al. (1989) FEBS Lett. 245, 150-154). Noteworthy, after in vivo and in vitro RA-treatment we could not determine any increase (there was even a slight decrease) in the number of the cross-linked apoptotic envelopes. In keeping with this is the significant reduction of protein bound gamma-glutamylpolyamine detected in hepatocytes exposed to RA in culture. These findings suggest that the RA-induced tTG in parenchimal cells is an inactive form.

Animals↗

PubMed1992

The clearance of apoptotic cells in the liver is mediated by the asialoglycoprotein receptor.

Apoptosing cells are actively phagocytosed in parenchymal tissues, thus preventing the inflammatory reaction which could derive from their slow uncontrolled degradation. The molecular mechanisms by which an apoptotic cell is recognized and taken up are largely unknown. We propose that the recognition of apoptotic hepatocytes is mediated by the sugar recognition systems of the liver, particularly the asialoglycoprotein receptor (ASGP-R). The results presented here demonstrated the participation of ASGP-R in the removal of apoptotic parenchymal cells, and indicate a new perspective for the understanding of its physiological role.

Acetylgalactosamine↗

PubMed1992

The simultaneous exposition of galactose and mannose-specific receptors on rat liver macrophages is developmentally regulated.

We studied the simultaneous binding of galactose and mannose-exposing ligands in sinusoidal rat liver cells during development and aging. The galactose-specific receptors were visualized using 17 nm diameter colloidal gold particles coupled with Lactosylated bovine serum albumine (LacBSA), while mannose-specific receptors were localized by means of 5 nm diameter particles adsorbed with mannan. We observed the presence of four different classes of Kupffer cells in relation to the ligands bound. The percentage of each group of Kupffer cells varied in relation to the age of the subject from which the sample was taken. There were few double-labelled cells in the livers from newborn rats, with numbers increasing with age to adulthood, and decreasing again in the older animals. Cells without labelling were in the majority after birth, but they decreased in number up to adulthood and increased again during subsequent aging. The numbers of single-labelled cells did not change significantly during liver maturation. We hypothesize that the exposition of galactose and mannose-specific receptorial systems is regulated by developmental conditions.

Aging↗

PubMed1992

Receptor-mediated endocytosis of galactose and mannose exposing ligands: an electron microscopic study on adult and neonatal cultured rat hepatocytes.

We compared the receptor-mediated endocytosis for galactose and mannose exposing ligands in primary cultures of hepatocytes from newborn and adult rats. The endocytic pathway was revealed ultrastructurally using colloidal gold particles coupled to lactosylated bovine serum albumin and invertase. The binding activity on the cell surfaces is observed by keeping the cells at 4 degrees C. For both ligands used, the binding capacity for hepatocytes from adult rats was greater than for neonatal cultured cells. Increasing the temperature to 37 degrees C, we observed that the protein-gold complexes entered the intracellular endocytic organelles. Within 5-15 min, the marker was confined in vesicles close to the cell surface and in the endosome, while after 60 min, the marker is found in lysosome-like compartments. We found that the process of endocytosis is similar for galactose and mannose exposing ligands. The organelles involved in the process of endocytosis in newborn cultured hepatocytes are not different in shape from those of cultured cells of adult rats, but the process of internalization is slower.

Aging↗

PubMed1992

[Critical analysis of the diagnostic imaging of focal hepatic lesions].

The aim of this paper is to evaluate the number and type of consecutive initial diagnoses of focal hepatic lesions obtained by abdominal US and CT. The diagnoses were coded according to the Index for Radiological Diagnoses (ACR). From January 1990 through April 1991, US and CT diagnosed focal hepatic lesion in 16.4% and 9.1% of cases, respectively. A lower number of focal hepatic lesions was diagnosed by CT, which however was more accurate as to the nature of the lesion itself. This is due to the fact that CT is often used to stage hepatic neoplasms already confirmed by US-guided fine-needle biopsy (FBN). The correlation between the initial diagnosis and actual clinical status demonstrates a high rate of occasional findings, especially with US. The rate of questionable diagnoses relative to primary or secondary malignancies was very high. This could be explained by caution in making a severe diagnosis, by awareness of the limits of macroscopic diagnostic techniques and by the immediate availability of US-guided FNB. In conclusion, the coding of consecutive initial diagnoses, by US and CT, could contribute to a practical evaluation of diagnostic imaging techniques, especially when correlated with the respective anamnestic and clinical data. Further studies are necessary to confirm this hypothesis.

Humans↗

PubMed1991

Cultured hepatocytes bind and internalize bovine serum amine oxidase-gold complexes.

Cultured hepatocytes express binding sites for bovine serum amine oxidase on their membrane surfaces as evaluated at the electron microscopic level by using enzyme-gold complexes. Hepatocytes show binding sites as small clusters of gold granules, not bound in a specialized region of the plasma membrane. The binding competition of enzyme-gold ligand to cells was achieved by preincubation with uncoupled bovine serum amine oxidase. In addition, N-acetyl-galactosamine, N-acetyl-glucosamine and Mannose, at the final concentration of 80 mM, partially inhibit the binding.

Amine Oxidase (Copper-Containing)↗

PubMed1991

Size and stability of dipalmitoylphosphatidylcholine/cholesterol unilamellar vesicles are affected by interaction with proteins.

The effect of entrapping the enzyme ascorbate oxidase into dipalmitoylphosphatidylcholine/cholesterol vesicles, was studied by conventional transmission electron microscopy and freeze-fracture. The freeze-fracture technique has definitely demonstrated the unilamellar nature of empty and enzyme-loaded vesicles. Images of freeze-fractured and label-fractured liposomes also indicate that the observed reduction of vesicles volume could be related to the localization of ascorbate oxidase across the membrane. The membrane localization of ascorbate oxidase may explain the oxidation of externally added ascorbate by intact enzyme-loaded liposomes. Finally, the ageing of liposomes appears to be accelerated in the presence of proteins.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed1991

"Tissue" transglutaminase is specifically expressed in neonatal rat liver cells undergoing apoptosis upon epidermal growth factor-stimulation.

We recently reported that activation of "tissue" transglutaminase (EC 2.3.2.13; tTG) in liver cells undergoing apoptosis determines extensive cross-linking of cellular proteins resulting in the formation of SDS-insoluble shells in the so-called "apoptotic bodies". In attempt to obtain further insight into the role played by tTG in apoptosis of liver cells, we investigated its expression in primary cultures of neonatal rat liver cells stimulated with epidermal growth factor (EGF). EGF-treatment of neonatal rat liver cells induces first hyperplasia of hepatocytes, followed by involution characterized by a high incidence of apoptosis. The proliferative phase of hepatocytes is paralleled by a 10-fold increase in tTG mRNA level, which is followed, during the phase of involution, by sequential increases in enzyme activity and levels of SDS-insoluble apoptotic bodies. tTG immunostaining at both the light- and electron-microscopic levels shows that the most intensive reaction is present in globular structures showing the typical morphological appearance of mature apoptotic bodies. In early apoptotic stages, tTG protein is localized in the perinuclear region of the cell. Intense immunostaining is also found in the apoptotic bodies present inside phagosomes within the cytoplasm of neighboring cells. This evidence confirms and extends our previous findings, indicating that tTG induction and activation specifically takes place in cells undergoing apoptosis, suggesting a key role for the enzyme in the apoptotic program.

Animals↗

PubMed1991

Chick hepatic lectins: an electron microscopic study on isolated hepatocytes during development.

We studied the carbohydrate recognition systems of hepatocytes isolated from 16-day-old embryos, 19-day-old embryos and chicks within 24 h of hatching. We localized and quantified at the ultrastructural level the binding sites for glycoproteins exposing terminal N-acetylglucosamine (GlcNAc), mannose and N-acetylgalactosamine (GalNAc) residues by means of protein-gold complexes. Binding sites specific for GlcNAc and mannose residues are present on hepatocytes from embryos and chicks. On the contrary GalNAc specific binding sites are exclusively observed on cells from 16-day-old embryos. The number and distribution of gold particles on hepatocyte cell surfaces depend on the binding sites and the age considered. We describe a modulation in the number of GlcNAc, and mannose specific receptors present on the cell surface between the embryonal stage and neonatal life.

Animals↗