Biomedical subjects

G Donelli

Publications and source records attributed to G Donelli.

At least 37 records · Page 2Linked to original sources

PubMed1996

Antioxidant N-acetyl-cysteine increasing cell adhesion capability could facilitate the biocompatibility processes.

Cell adhesion plays an important role in several cell processes and functions, including differentiation, proliferation and death. An important role for cell attachment to medical devices in biocompatibility studies has also been hypothesized. In this paper we report that the use of the antioxidant drug N-acetyl-cysteine is capable of increasing the adhesion properties of epithelial cells in culture. This is associated with a modification of specific cytoskeletal element assembly, such as microfilament system molecules. In contrast, no quantitative alterations in the expression of certain surface receptors for extracellular matrix molecules, such as VLA2, VLA3 and VLA6, are found. These data seem to indicate that intracellular oxidative balance, in particular of thiol groups, could play a key role in the cell adhesion properties and that N-acetyl-cysteine treatment, acting as 'thiol supply', could be of importance in several circumstances, including biocompatibility of medical devices.

Acetylcysteine↗

PubMed1996

Bacteroides fragilis enterotoxin induces cytoskeletal changes and surface blebbing in HT-29 cells.

Certain strains of the anaerobic bacterium Bacteroides fragilis are known to produce an enterotoxin of about 20 kDa which is able to induce a fluid response in ligated intestinal loops and a cytotoxic response in HT-29 cells. It presents protease activity, belonging to a family of metalloproteases termed metzincins. In order to investigate the mode of action of the enterotoxin in cultured cells, we performed a study with HT-29 cells, using both fluoresence and electron microscopy. Treated cells underwent morphological changes, mainly consisting of the retraction of the cell body and the formation of numerous blebs on the cell surface. The microfilament system was reorganized, the F-actin being condensed as a ring at the cell periphery, whereas other cell organelles appeared to be unaffected. All these changes, clearly visible after 3 h of exposure to the toxin, were reversed within 24 h of treatment. By inhibiting the protease activity of the toxin with specific metal chelators, the cytoskeletal effects were also prevented. Thus, B. fragilis enterotoxin appears to act on cells by reversibly modifying the actin cytoskeleton, an effect probably dependent on its proteolytic activity.

Bacterial Toxins↗

PubMed1996

An epidemiological study on viral infantile diarrhoea in Tirana.

During the period May 1993-April 1994, an epidemiological survey was conducted on enteric viruses which cause gastroenteritis in infants and young children in Tirana, Albania. Specimens from 321 cases were screened by direct electron microscopy and by an enzyme-linked immunosorbent assay specific for rotavirus group A antigen. By ultrastructural analysis, rotaviruses were detected in 10.3% of cases and adenoviruses in 0.6%, whereas small round structured viruses and small round viruses were found in 2.8% and 2.2% of cases, respectively. Different percentages of rotavirus excretors were revealed by enzyme-linked immunosorbent assay (12.15%) and electron microscopy. Samples rotavirus-positive in at least one of these assays were also analyzed by agglutination of latex particles and electron microscopy results were confirmed. Analysis of electron microscopy-positive samples by rotaviral RNA polyacrylamide gel electrophoresis showed five different long electropherotypes of rotavirus among which a single, largely predominant electropherotype (65.5%) was observed.

Adenoviridae↗

PubMed1996

Intestinal microsporidiosis in Italian individuals with AIDS.

A survey on microsporidiosis in individuals with AIDS presenting chronic diarrhoea was carried out in Italy, over a four-year period (1992-1995). Three out of 72 (4.2%) individuals were found positive, on intestinal biopsies, for Enterocytozoon bieneusi by light microscopy and transmission electron microscopy (TEM). Sixteen individuals with AIDS, from a second group of subjects, were confirmed positive, by TEM, for intestinal microsporidiosis due to Enterocytozoon bieneusi. Of these 19 cases, 10 (52.6%) were homosexual men. Two of these individuals, under albendazole treatment, showed also spores with unusual features. The prevalence of intestinal microsporidiosis (12-50%) reported in European countries, Australia and North America, where homosexuality is the major HIV risk factor (63-77%), is higher than in Italy, where homosexual men represent only 16% of the total number of AIDS cases.

AIDS-Related Opportunistic Infections↗

PubMed1995

Characterization of SA-11 rotavirus receptorial structures on human colon carcinoma cell line HT-29.

The involvement of different cell membrane components in the receptor structures for SA-11 rotavirus was investigated. As experimental model, the human enterocyte-like HT-29 cell line, was used because of its closer resemblance to the in vivo viral cellular target as compared to other in vitro systems. Rotavirus was incubated with whole membranes or their separated protein and lipid fractions before infection. Either isolated cell membranes or lipid components were capable of binding to the virus and to prevent infection, whereas proteins did not show any inhibitory activity. Among lipids, the glycolipid fraction was shown to impede rotaviral antigen synthesis with a dose-dependent relationship, whereas phospholipids failed to prevent viral infection. To confirm these findings, membranes and target cells were subjected to different enzymatic treatments prior to infection. In addition, HT-29 cells were also incubated with different lectins before infection. The blocking activity of membranes was inhibited by treatment with ceramide glycanase, neuraminidase, and beta-galactosidase but not by treatment with proteases or heat (100 degrees C). Viral infection was prevented by preincubation of target cells with lectins specific for sialic acid and galactose or with ceramide glycanase, neuraminidase, and beta-galactosidase, whereas protease treatments were not active. The results of these experimental procedures indicate that glycolipids containing specific carbohydrate moieties, such as sialic acid and galactose, contribute to the SA-11 rotavirus receptor structure on HT-29 cells.

Adenocarcinoma↗

PubMed1995

Enhancement of rotavirus infectivity by saturated fatty acids.

The effect of different saturated fatty acids from 10 to 16 carbon atom chains and some derivatives on the infectivity of SA-11 rotavirus was examined. Both fatty acids and derivatives induced an increase of rotavirus infected LLC-MK2 cells when present during viral absorption to host cells. Capric acid and palmitic acid were the most effective with a dose-dependent relationship. These last lipids, in the same experimental conditions, failed to restore the susceptibility to infection of LLC-MK2 cells made resistant by neuraminidase treatment or to allow cell infection by non-infectious single-shelled viral particles. Results obtained suggest that the enhancing effect on viral infectivity by saturated fatty acids requires previous binding of rotaviral outer capsid proteins to sialic acid containing cell receptors.

Animals↗

PubMed1995

P-170 glycoprotein (P-170) is involved in the impairment of natural killer cell-mediated cytotoxicity in HIV+ patients.

In the present study we analyze peripheral blood lymphocytes (PBL) from patients with human immunodeficiency virus (HIV) infection for both phenotypic expression and function of P-glycoprotein (P-170). This transmembrane efflux pump is known to be one of the mechanisms responsible for the multidrug resistance (MDR) in cancer therapy and it is also constitutively expressed in normal PBL. P-170 function, evaluated as Rhodamine 123 (Rh123) efflux in flow cytometry, was found to be significantly reduced in CD16+ natural killer (NK) cells from patients with HIV infection. Interestingly, this reduced efflux significantly correlates with the decreased NK cytotoxicity observed in HIV+ patients, as evaluated against the NK-specific K562 target cell line. These results support a possible role of the P-170-related pump in specific immunological lymphocyte function such as NK cell-mediated cytotoxicity.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

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↗

PubMed1995

Transmembrane P-glycoprotein (P-gp/P-170) in HIV infection: analysis of lymphocyte surface expression and drug-unrelated function.

P-glycoprotein (P-gp/P-170), a transmembrane efflux pump known to be one of the mechanisms responsible for multidrug resistance in cancer therapy, is constitutively expressed in several solid human tissues as well as in normal peripheral blood lymphocytes and bone marrow cells. In particular, this molecule has been associated with the transport of perforin and other cytolysins in natural killer (NK) and T cytotoxic lymphocytes. In the present study, we analyzed peripheral blood lymphocytes (PBLs) from controls and HIV+ patients for phenotypic expression and function of the P-gp/P-170 molecule. We found that 90% of all PBL subsets (i.e., CD4+, CD8+, CD56+, and CD19+ cells) expressed surface P-gp/P-170 both in controls and HIV+ patients. However, a significant decrease in CD4+/P-170+ and CD19+/P-170+ cells was observed in HIV+ individuals with respect to controls. PHA and IL-2 stimulation of PBLs was unable to increase the expression of P-gp/P-170 both in controls and HIV+ patients, despite the increased detection of the CD25 molecule. On the other hand, stimulation with anti-CD3 determined a significant increase in lymphocyte P-gp/P-170. The function of P-gp/P-170, assessed by a flow cytometric assay for rhodamine-123 (Rh123) efflux, was significantly reduced in CD16+ NK cells and CD19+ B cells from HIV+ patients. The Rh123 efflux by NK cells correlated (p < 0.01) with the NK cytotoxicity against the 51Cr-labeled K562 cell line. Last, the effect of the antiretroviral drugs AZT, ddI, and ddC on P-gp expression and function was evaluated. The dideoxynucleoside compounds did not inhibit P-gp/P-170 function of normal mononuclear cells in vitro, and did not increase P-gp/P-170 expression in vivo, in patients undergoing antiretroviral therapy with AZT. These findings provide further evidence of a possible involvement of the P-gp/P-170 system in specific immunological lymphocyte functions, and especially in cytotoxic-type functions. In addition, it is possible to suggest, on the basis of our experimental data, that the dideoxynucleoside class of antiretroviral agents does not contribute to the phenotypic and functional alterations related to P-glycoprotein during HIV infection.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed1995

Escherichia coli cytotoxic necrotizing factor 1: evidence for induction of actin assembly by constitutive activation of the p21 Rho GTPase.

Cytotoxic necrotizing factor type 1 (CNF1) induces in HEp-2 cells an increase in F-actin structures, which was detectable by fluorescence-activated cell sorter analysis 24 h after addition of this factor to the culture medium. Increase in F-actin was correlated with the augmentation of both the cell volume and the total cell actin content. Actin assembly-disassembly is controlled by small GTP-binding proteins of the Rho family, which have been reported recently to be modified by CNF1 treatment. Clostridium difficile toxin B and Clostridium botulinum exoenzyme C3, both known to act on the Rho GTPase, were used as biological tools to study the effect of CNF1 on this protein. CNF1 incubated before, during, or after exposure to the chimeric toxin C3B (which is the product of a genetic fusion between the DNA coding for C3 and the one coding for the B fragment of diphtheria toxin) protected HEp-2 cells from the disruption of F-actin structures caused by inactivation of the Rho GTPase through its ADP-ribosylation. On the other hand, C. difficile toxin B cytopathic effect was not observed upon preincubation of cells with CNF1. Toxins acting through a Rho-independent mechanism, such as cytochalasin D and Clostridium spiroforme iota-like toxin, could not be modified in their cellular activities by CNF1 treatment. All of our results suggest that CNF1 modifies the Rho molecule, thus probably protecting this GTPase from further bacterial toxin modification.

ADP Ribose Transferases↗

PubMed1995

Zonula occludens toxin modulates tight junctions through protein kinase C-dependent actin reorganization, in vitro.

The intracellular signaling involved in the mechanism of action of zonula occludens toxin (ZOT) was studied using several in vitro and ex vivo models. ZOT showed a selective effect among various cell lines tested, suggesting that it may interact with a specific receptor, whose surface expression on various cells differs. When tested in IEC6 cell monolayers, ZOT-containing supernatants induced a redistribution of the F-actin cytoskeleton. Similar results were obtained with rabbit ileal mucosa, where the reorganization of F-actin paralleled the increase in tissue permeability. In endothelial cells, the cytoskeletal rearrangement involved a decrease of the soluble G-actin pool (-27%) and a reciprocal increase in the filamentous F-actin pool (+22%). This actin polymerization was time- and dose-dependent, and was reversible. Pretreatment with a specific protein kinase C inhibitor, CGP41251, completely abolished the ZOT effects on both tissue permeability and actin polymerization. In IEC6 cells ZOT induced a peak increment of the PKC-alpha isoform after 3 min incubation. Taken together, these results suggest that ZOT activates a complex intracellular cascade of events that regulate tight junction permeability, probably mimicking the effect of physiologic modulator(s) of epithelial barrier function.

Actins↗

PubMed1995

Electropherotypes of rotavirus strains causing gastroenteritis in infants and young children in Tirana, Albania, from 1988 to 1991.

During 1988-1991, an epidemiological survey was conducted in Tirana (Albania) on group A rotavirus strains which cause gastroenteritis in infants and young children. Rotaviruses were detected in 312 of 1,241 (25.1%) examined specimens from children with acute diarrhoea. Viruses were detected throughout the study period. Among the 72 rotavirus strains tested for double-stranded RNA (dsRNA) electrophoretic migration pattern, 9 different electropherotypes were recognized, 1 of those being more frequent than the others. At the beginning and at the end of the examined period (1988 and 1990-1991) two different long electropherotypes were predominant, whereas in 1989 (middle period) short electropherotypes were common indicating an involvement of virus strains with short electropherotypes in hospitalization-requiring diarrhoeas occurring in the area surveyed in that year.

Age Factors↗

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

Detection of enteroadherent Escherichia coli associated with diarrhoea in Italy.

One hundred and sixty-eight isolates of Escherichia coli obtained in Italy from 112 children with diarrhoea and 56 age-matched controls were examined by the HEp-2 cell adhesion assay. Sixteen strains showed localised adherence (LA), 29 showed diffuse adherence (DA) and eight strains showed aggregative adherence (AA). No adhesion pattern was significantly associated with disease. Strains that showed LA or AA were further characterised by serotyping, fluorescent actin staining (FAS) test and hybridisation with the EPEC adherence factor (EAF), E. coli attaching and effacing (eae) and enteroaggregative (EAgg) DNA probes. Strains that showed poor LA were FAS-negative, did not belong to EPEC serotypes and did not hybridise with EPEC probes. Conversely, the two strains that showed a good LA pattern belonged to serotype O128:H2, were FAS positive and hybridised with the eae probe. No isolate hybridised with the EAF probe. Only three of the eight strains with the AA pattern hybridised with the EAgg probe. Probe positivity correlated with the ability to produce clumps at the surface of the liquid culture and to agglutinate rat erythrocytes. In two of these EAgg probe-positive strains, electronmicroscopy revealed the presence of fibrillar bundles which seem to mediate bacterial aggregation.

Bacterial Adhesion↗

PubMed1994

Influence of particle size and chemical composition on efficiency of clearance mechanisms: electron microscopy studies on humans.

This article compares the presence of solid particles in lung parenchyma samples collected from accident victims and in bronchoalveolar lavage fluid taken from patients diagnosed with pulmonary carcinoma. Analysis by electron microscopy showed differences in particle size between the two groups, which could be attributable both to differences in original particle size and to their solubility in the biological environment.

Bronchoalveolar Lavage Fluid↗

PubMed1994

[Infections associated with intra- and extravascular catheters: factors involved in microorganism-biomaterial interactions].

Infections is one of the most common cause of catheter failure as well as the most difficult to manage, most often requiring catheter removal. Staphylococcus is the etiologic agent of such infections more frequently isolated, particularly Staphylococcus epidermidis. Several factors have been suggested to be involved in bacteria-biomaterial interactions such as catheter surface morphology, molecular biofilm and bacterial virulence features. Different strategies have been tried to avoid the development to catheter-associated infections: among them adsorption of antibiotic molecules to the catheter surface might represent a successful tool to improve catheter implant life.

Bacterial Adhesion↗

PubMed1994

Bacterial protein toxins acting on the cell cytoskeleton.

A number of bacterial protein toxins are known to exert their cytotoxic activity via a modification of cytoskeletal components. Some toxins induce the ADP-ribosylation of actin whereas others interact with the cytoskeleton by an unknown mechanism. Understanding the mode of action of such toxins at cellular level could provide useful information on their role in vivo as virulence factors.

Actin Cytoskeleton↗

PubMed1994

Rotavirus and poliovirus co-infection in HT-29 cells.

The effect of a mixed poliovirus-rotavirus infection in HT-29 cells, a gut tumour derived cell line highly susceptible to both viruses, has been analyzed. The obtained results showed an increase of poliovirus multiplication in cells super-infected or co-infected with rotavirus, whereas the pre-infection with poliovirus had an interfering effect on rotavirus replication.

Antigens, Viral↗