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1.
Acta Otorhinolaryngol Ital ; 34(2): 99-104, 2014 Apr.
Article in English | MEDLINE | ID: mdl-24843219

ABSTRACT

Head and neck squamous cell carcinoma of the (HNSCC) represents approximately 5% of malignant tumours in Italy. HNSCC are commonly treated with surgery or radiotherapy, or a combination of such therapies. The objectives of treatment are maximum cure rate balanced with organ preservation, restoration of form and function, reduction of morbidities and improvement or maintenance of the patient's quality of life. Immediate reconstructive surgery: local, regional or free flaps are now widely advised in the treatment of these patients. Microsurgical transfer requires expertise, is time and resource consuming, and as a whole requires substantial costs. These considerations introduce some concerns about the wide or indiscriminate use of free flap reconstructive surgery. When considering cost-benefit outcomes of such treatment, the main objective is undoubtedly, survival. This data is underreported in the current literature, whereas functional outcomes of free flaps have been largely diffused and accepted. This study collects data from 1178 patients treated with free flap reconstructive surgery following ablation of HNSCC in a group of Italian tertiary hospitals, all members of the Head & Neck Group affiliated with the Italian Society of Microsurgery. According to many authors, free flap surgery for HNSCC seems to be a beneficial option for treatment even in terms of survival.


Subject(s)
Carcinoma, Squamous Cell/mortality , Carcinoma, Squamous Cell/surgery , Free Tissue Flaps , Head and Neck Neoplasms/mortality , Head and Neck Neoplasms/surgery , Adolescent , Adult , Aged , Aged, 80 and over , Female , Humans , Male , Middle Aged , Plastic Surgery Procedures/methods , Retrospective Studies , Squamous Cell Carcinoma of Head and Neck , Survival Rate , Young Adult
2.
Acta Otorhinolaryngol Ital ; 32(5): 336-8, 2012 Oct.
Article in English | MEDLINE | ID: mdl-23326015

ABSTRACT

Extracranial meningiomas of the head and neck region are rare neoplasms, the majority being a secondary location of a primary intracranial tumour. We herewith report three rare cases of extracranial meningiomas, located in the temporal muscle, parotid gland and nasal cavity, together with complete pathological, immunohistochemical and ultrastructural studies. Prognosis of this tumour is generally excellent. Surgical excision is the treatment of choice, with no need for further treatment; nevertheless, differential diagnosis must consider other more common tumours of the head and neck and be based on histopathologic examination and relative techniques, including examination of frozen sections. This procedure is particularly useful assessing surgical treatment and should be performed whenever possible to exclude the malignant nature of the lesion and avoid over-treatment. All three patients underwent surgery and are alive and disease-free.


Subject(s)
Head and Neck Neoplasms/diagnosis , Meningioma/diagnosis , Adult , Aged , Female , Head and Neck Neoplasms/pathology , Humans , Immunohistochemistry , Meningioma/pathology
3.
Acta Otorhinolaryngol Ital ; 27(5): 227-32, 2007 Oct.
Article in English | MEDLINE | ID: mdl-18198751

ABSTRACT

To assess whether locking-screw titanium plates (UniLOCK) and pedicled pectoralis major myocutaneous flaps are a valid alternative to complex reconstruction with bony free flaps in poor prognosis or poor performance status oncological patients with mandibular defects, a retrospective evaluation has been made of outcomes in 27 consecutive cases. No patient died perioperatively. Mean operating time was 270 minutes. Post-operative course was uneventful in 14. Mean follow-up was 13 months with no loss to follow-up. Twelve patients are alive and well, 12 died from their malignancy, two from non-neoplastic causes, and one from second cancer. Plate exposure - the main problem with bridging plates - occurred in 6 (22%, 4 early, 2 late), 4 with symphyseal and 2 with postero-lateral defects: removal was necessary in 2; 2 died with the plate exposed, and 2 had successful re-coverage, increasing the final success rate from 78% to 85%. Most patients considered the aesthetic outcome acceptable, however all edentulous patients complained of unsatisfactory dental rehabilitation. From the acceptable success rate, it may be concluded that bridging plates represent a useful reconstruction method, provided they are well covered by viable muscular tissue. They should be offered to patients contraindicated for more invasive procedures or with limited functional needs, or poor prognosis.


Subject(s)
Head and Neck Neoplasms/surgery , Mandible/surgery , Pectoralis Muscles/transplantation , Plastic Surgery Procedures/methods , Skin Transplantation/methods , Surgical Flaps , Titanium/therapeutic use , Adult , Aged , Biocompatible Materials , Female , Humans , Male , Middle Aged , Neoplasm Staging
4.
Exp Brain Res ; 167(4): 641-8, 2005 Dec.
Article in English | MEDLINE | ID: mdl-16078027

ABSTRACT

Curcumin, an extract from the plant Curcuma longa with well-known antioxidant and anti-inflammatory activities, was tested as protective agent against excitotoxicity in rat retinal cultures. A 24 h-treatment with curcumin reduced N-methyl-D: -aspartate (NMDA)-mediated excitotoxic cell damage, estimated as decrease of cell viability and increase in apoptosis. The protection was associated with decrease of NMDA receptor-mediated Ca(2+) rise and reduction in the level of phosphorylated NR1 subunit of the NMDA receptor. These results enlighten a new pharmacological action of the plant extract, possibly mediated by a modulation of NMDA receptor activity.


Subject(s)
Anti-Inflammatory Agents, Non-Steroidal/pharmacology , Calcium/metabolism , Curcumin/pharmacology , N-Methylaspartate/toxicity , Neurons/drug effects , Retina/drug effects , Animals , Apoptosis/drug effects , Blotting, Western , Cell Survival/drug effects , Cells, Cultured , Immunohistochemistry , Neurons/metabolism , Phosphorylation , Rats , Rats, Wistar , Receptors, N-Methyl-D-Aspartate/drug effects , Retina/cytology , Retina/metabolism
5.
Environ Res ; 96(1): 62-71, 2004 Sep.
Article in English | MEDLINE | ID: mdl-15261785

ABSTRACT

The RAW 246.7 macrophage cell line was exposed in vitro to aged crystalline silica particles of respirable size for 24 h at a range of doses starting from 15 microg/2 x 10(6) cells, which is a realistic exposure level of macrophages in the airways of ambiently exposed individuals. The particle sample used for the experiments was prepared to mimic some aspects of ambient crystalline silica particles: size distribution, morphology, and surface reactivity. Our purpose was to determine whether a nontoxic quartz load comparable to that of ambient exposure would be able to induce macrophage activation and impairment of the phagocytic ability, factors altering the lung's capacity to deal with increased particle loads (as occurs during high-pollution episodes) or infections and affecting the local and systemic responses through the release of biologically active compounds (cytokines, reactive oxygen species, NO, isoprostanes). Exposure of RAW 264.7 cells to aged silica particles induced macrophage activation (evidenced by the morphological features observed with scanning electron microscopy and by the release of TNF-alpha and IL-6) and impairment of phagocytosis of test particles, even at noncytotoxic doses. The reduction of the phagocytic function of the cells after silica treatment was dose-dependent, as evidenced by an increase of the population of unphagocytic cells, paralleled by a decrease of the actively phagocytizing cell population. We evaluated the oxidative stress induced by aged silica particles, quantifying the peroxidation products (8-isoprostanes) in the culture media of treated cells, and found a strong release at low doses. Isoprostanes are a complex family of compounds which have been used as in vivo markers of lipid peroxidation in human disorders, but that, as far as we know, have never been evaluated in relation to airborne particulate matter exposure. Lipid peroxides are involved in various cellular events in the inflammatory response, and isoprostanes are also supposed to exert important biological actions on airway and pulmonary vascular smooth muscles and on platelets.


Subject(s)
Air Pollutants/adverse effects , Macrophages, Peritoneal/drug effects , Silicon Dioxide/adverse effects , Animals , Interleukin-6/metabolism , Macrophages, Peritoneal/metabolism , Macrophages, Peritoneal/ultrastructure , Mice , Phagocytosis , Tumor Necrosis Factor-alpha/metabolism
6.
J Biomed Mater Res B Appl Biomater ; 65(1): 133-6, 2003 Apr 15.
Article in English | MEDLINE | ID: mdl-12632382

ABSTRACT

The effects of perfluorohexyloctane (F6H8), recently investigated as a long-term artificial vitreous substitute, were studied in vitro, with the use of rat retinal cultures seeded on microporous inserts that allow the cell layer to be in contact with the material to be tested, on the apical side, and with the nutrient medium, on the basal side. After 72 h of treatment with F6H8, retinal cultures lost the characteristic two-layered organization with glial cells at the bottom and neuronal cells on top of them. They appeared to be composed of only one layer of polyhedrical, flattened, and disconnected cells. TUNEL assay revealed an evident increase in the percentage of apoptotic cells in F6H8-treated cultures (30.1 +/- 4.5), compared to control (10.3 +/- 2.6) and perfluoroctane-treated cultures (10.1 +/- 1.7). Immunolabeling of MAP-2, a protein of neuronal cytoskeleton, evidenced a marked loss of neurites. The results suggest that F6H8 is harmful to retinal cells in vitro and can therefore be potentially noxious to the retina as an artificial vitreous substitute.


Subject(s)
Apoptosis/drug effects , Cell Communication/drug effects , Fluorocarbons/adverse effects , Retina/cytology , Animals , Biocompatible Materials/adverse effects , Cells, Cultured , Embryo, Mammalian , Materials Testing , Microtubule-Associated Proteins/analysis , Neurites/drug effects , Rats , Rats, Wistar , Retina/drug effects
7.
J Physiol Paris ; 96(3-4): 313-6, 2002.
Article in English | MEDLINE | ID: mdl-12445911

ABSTRACT

A characteristic feature of neuritic plaques in Alzheimer's disease is represented by the presence of activated astrocytes, surrounding dystrophic neurons and beta-amyloid deposition. To explore the role of astrocytes in in vitro beta-amyloid neurotoxicity, we studied the effect of beta-amyloid treatment in hippocampal neurons in two different cell models: pure cultures, where neurons were grown in absence of astrocytes and mixed cultures, where neurons were seeded on a confluent layer of astrocytes. We evaluated two characteristic aspects of in vitro beta-amyloid neurotoxicity: reduction of cell viability and degeneration of the neuritic tree. We demonstrated that neurons growing on astrocytes were more prone to the detrimental effect of the amyloid peptide, with respect to neurons grown in absence of the glial component. Our results support the hypothesis that beta-amyloid-astrocyte interaction can adversely condition neurons and contribute to neuronal damage in Alzheimer's disease.


Subject(s)
Amyloid beta-Peptides/toxicity , Astrocytes/metabolism , Cell Communication/drug effects , Neurons/metabolism , Animals , Astrocytes/cytology , Cell Survival/drug effects , Cells, Cultured , Coculture Techniques , Hippocampus/cytology , Neurons/cytology , Rats , Rats, Wistar
8.
Glia ; 34(1): 68-72, 2001 Apr 01.
Article in English | MEDLINE | ID: mdl-11284021

ABSTRACT

Astrocytosis is a common feature of amyloid plaques, the hallmark of Alzheimer's disease (AD), along with activated microglia, neurofibrillary tangles, and beta-amyloid (beta A) deposition. However, the relationship between astrocytosis and neurodegeneration remains unclear. To assess whether beta A-stimulated astrocytes can damage neurons and contribute to beta A neurotoxicity, we studied the effects of beta A treatment in astrocytic/neuronal co-cultures, obtained from rat embryonic brain tissue. We found that in neuronal cultures conditioned by beta A-treated astrocytes, but not directly in contact with beta A, the number of apoptotic cells increased, doubling the values of controls. In astrocytes, beta A did not cause astrocytic cell death, nor did produce changes in nitric oxide or prostaglandin E(2) levels. In contrast, S-100 beta expression was remarkably increased. Our data show for the first time that beta A--astrocytic interaction produces a detrimental effect on neurons, which may contribute to neurodegeneration in AD.


Subject(s)
Amyloid beta-Peptides/pharmacology , Apoptosis/drug effects , Astrocytes/drug effects , Hippocampus/drug effects , Neurons/drug effects , Alzheimer Disease/physiopathology , Animals , Apoptosis/physiology , Astrocytes/physiology , Cells, Cultured , Cerebral Cortex , Embryo, Mammalian , Hippocampus/physiology , Neurons/physiology , Rats , Rats, Wistar
9.
Int J Cancer ; 88(5): 772-7, 2000 Dec 01.
Article in English | MEDLINE | ID: mdl-11072247

ABSTRACT

Pancreatic acinar cell carcinoma (PAC) is a rare pancreatic tumor for which no information about chromosomal and gene anomalies is available. We performed genome-wide allelotyping of 9 PACs using DNA from 5 frozen and 4 paraffin-embedded samples and 76 PCR-amplified, chromosome-specific microsatellite markers. High degrees of allelic loss were found, with a mean fractional allelic loss of 0.33. Chromosomes 1p, 4q and 17p showed loss of heterozygosity in >70% of cases and chromosomes 11q, 13q, 15q and 16q, in 60% to 70% of cases. Chromosomes 3q, 6q, 8q, 18q and 21q showed loss in 50% to 60% of cases. All of the remaining chromosomes showed no or few allelic losses. The resulting allelotype of PAC is markedly different from that of either ductal or endocrine tumors of the pancreas, and the involvement of chromosomes 4q and 16q appears to be characteristic of this tumor type. High-resolution mapping of the 12 frequently altered chromosomes in 5 cases with 222 markers permitted subchromosomal localization of regions of consensus loss on 5 chromosomes, including 1p36.31, 3p25.2, 4q26-31.1, 15q15-22.1 and 16q21-q22.1. Our findings suggest that PAC tumorigenesis involves molecular pathways different from those occurring in more common pancreatic tumor types.


Subject(s)
Carcinoma, Acinar Cell/genetics , Chromosome Deletion , Pancreatic Neoplasms/genetics , Adolescent , Adult , Aged , Alleles , Chromosome Mapping , Female , Humans , Immunohistochemistry , Male , Middle Aged
10.
Cytogenet Cell Genet ; 89(3-4): 199-203, 2000.
Article in English | MEDLINE | ID: mdl-10965122

ABSTRACT

We describe the complete sequence, genomic organization, and FISH chromosome mapping of the human VAMP2. We identified a 7-kb clone, pISSHG2b3A, containing the entire structure of VAMP2. Previous studies performed by others identified a 5-kb clone, pVPC5-2, containing the incomplete VAMP2. The pVPC5-2 clone was partially sequenced and mapped to the broad region 17pter-->p12 by somatic cell hybridization. Our clone overlaps the pVPC5-2 clone and extends approximately 2 kb at the 3' end. In this study, we mapped this gene more precisely on 17p12 by FISH and we found a new polymorphic microsatellite, (GT)(7)CC(GT)(5), in exon V. This microsatellite, revealing three alleles with frequencies of 0.778, 0.139, and 0.083, might be useful for future linkage studies. Finally, we localized three previously known markers, stSG12859, TIGR-A002F11, and WIAF-1699 (alias stSG4044), in the 3' untranslated region of the gene.


Subject(s)
Chromosomes, Human, Pair 17/genetics , Genes/genetics , Membrane Proteins/genetics , Base Sequence , Chromosome Mapping , DNA/chemistry , DNA/genetics , Exons , Humans , In Situ Hybridization, Fluorescence , Introns , Microsatellite Repeats , Molecular Sequence Data , Polymorphism, Genetic , R-SNARE Proteins , Sequence Analysis, DNA
11.
Gene ; 253(2): 161-70, 2000 Aug 08.
Article in English | MEDLINE | ID: mdl-10940553

ABSTRACT

Per genes encode components of the circadian clocks controlling metabolic and behavioural rhythms. The human Per1 cDNA, RIGUI, was previously isolated and mapped on chromosome 17p12 (Sun, Z.S., Albrecht, U., Zhuchenko, O., Bailey, J., Eichele, G., Lee, C.C., 1997. RIGUI, a putative mammalian orthologue of the Drosophila period gene. Cell 90, 1003-1011). We have now isolated the entire genomic locus containing the human Per1 gene, in a search for genes associated with CpG-rich sequences. The hPer1 gene spans 15kb of human genomic DNA and is composed of 23 exons, flanked by 5' and 3' regulatory regions. Comparison of the hPer1 genomic clone with the dbEST database revealed homologies with putative alternative transcripts. Functional mapping within the 5' CpG-rich regulatory region enabled us to locate the hPer1 promoter core in a 510bp-long sequence centred around a TATA box, which supports high levels of hPer1 transcription. A second regulatory region was formally identified in intron 1, which appears to exert a negative role in transcriptional control of hPer1. These regions may be differentially involved in tissue-specificity, and/or circadian regulation, of the human hPer1 gene transcription.


Subject(s)
Genes/genetics , Nuclear Proteins/genetics , Promoter Regions, Genetic/genetics , 3T3 Cells , Animals , Base Sequence , Cell Cycle Proteins , Chloramphenicol O-Acetyltransferase/genetics , Chloramphenicol O-Acetyltransferase/metabolism , Chromosome Mapping , Chromosomes, Human, Pair 17/genetics , Cloning, Molecular , DNA/chemistry , DNA/genetics , DNA/isolation & purification , Drosophila Proteins , Exons , Humans , In Situ Hybridization, Fluorescence , Introns , Mice , Molecular Sequence Data , Period Circadian Proteins , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolism , Sequence Analysis, DNA , Transcription, Genetic
12.
Genes Chromosomes Cancer ; 28(3): 294-9, 2000 Jul.
Article in English | MEDLINE | ID: mdl-10862035

ABSTRACT

Pancreatic acinar cell carcinoma (PAC) is a rare pancreatic tumor for which no information about chromosomal anomalies is available. We examined six primary PACs by comparative genomic hybridization (CGH). All cases showed chromosomal changes. A total of 106 gains and 48 losses was detected. Consensus regions of gain were identified on chromosomes 1, 12, and X: 1q21 in four cases, 1q42 in three cases, 12p11.2 in four cases, and Xq12-21 in three cases. Recurrent losses were found at 16p13.2-p13.1 in three cases and at 16q23 in three cases. To verify these chromosomal imbalances, microsatellite analysis of matched normal and tumor DNA was performed using PCR-amplified markers for chromosomes 1, 12, and 16 in the regions showing nonrandom gains or losses. This analysis showed allelic imbalances in tumor DNA consistent with the CGH profiles. Our CGH study suggests that PAC shows a characteristic pattern of chromosomal alterations, involving gain at 1q, 12p, and Xq and loss of sequences at 16p and 16q. This pattern appears unique among solid tumors and is markedly different from that detected in pancreatic ductal carcinomas by the same technique. This suggests that PAC tumorigenesis involves different molecular pathways than those involved in the more common pancreatic ductal tumors. Genes Chromosomes Cancer 28:294-299, 2000.


Subject(s)
Carcinoma, Acinar Cell/genetics , Chromosome Aberrations/genetics , Pancreatic Neoplasms/genetics , Adult , Aged , Alleles , Female , Genetic Markers , Humans , Male , Middle Aged , Nucleic Acid Hybridization
13.
Genes Chromosomes Cancer ; 26(3): 203-9, 1999 Nov.
Article in English | MEDLINE | ID: mdl-10502317

ABSTRACT

We used arbitrarily primed polymerase chain reaction (AP-PCR) fingerprinting to identify chromosomal imbalances in six primary mediastinal B-cell lymphomas (PMBLs). Seventy-four chromosomal imbalances were detected, consisting of 49 sequence gains and 25 losses. Amplifications on chromosome X were seen in five cases, four of which involved the same chromosomal locus. Nonrandom gains at the same locus were also identified on chromosomes 2 and 7 in four cases and on chromosomes 5, 9, and 12 in three cases. Five PMBLs were also analyzed by comparative genomic hybridization (CGH), which found chromosome arm 9p amplification as the only nonrandom imbalance. Our data demonstrate that chromosomal amplifications outnumber losses in PMBL. These mainly involve chromosomes 9 and X and may reflect more complex phenomena, such as translocations or other chromosomal rearrangements, as AP-PCR found coexistent gains and losses on these chromosomes. Comparison between AP-PCR and CGH suggests that anomalies affecting the same chromosomal regions may occur at much higher frequencies than expected by CGH, suggesting that genomic amplifications are usually confined to DNA segments smaller than the megabase long segments required for detection in CGH. Modest increases in genetic material may be as effective as higher-level amplifications when affecting sites where a proto-oncogene resides.


Subject(s)
Chromosome Aberrations , DNA Fingerprinting/methods , Lymphoma, B-Cell/genetics , Mediastinal Neoplasms/genetics , Polymerase Chain Reaction/methods , Chromosomes, Human, Pair 2 , Chromosomes, Human, Pair 5 , Chromosomes, Human, Pair 7 , Chromosomes, Human, Pair 9 , DNA Primers/genetics , DNA, Neoplasm/analysis , Female , Gene Amplification , Humans , Male , Nucleic Acid Hybridization , Proto-Oncogene Mas , Sequence Deletion , Translocation, Genetic , X Chromosome
15.
Biochim Biophys Acta ; 1379(1): 161-70, 1998 Jan 08.
Article in English | MEDLINE | ID: mdl-9468344

ABSTRACT

The possible cytotoxic effects of fusinite, a new charcoal-like electron paramagnetic resonance (EPR) oxygen probe, were evaluated in three cell types with very different characteristics and growth features: K562 (an erythroleukemic cell line which grows in suspension), A431 (an epidermal carcinoma cell line which grows in monolayer) and primary cultures of murine fibroblasts (which also grow in adhesion culture) utilizing morphological and functional studies as well as growth analyses. Scanning and transmission electron microscopy as well as fluorescence microscopy were used for the morphological analyses while conductometric relaxation studies in the radiowave frequency range, membrane resistance measurements and adenine nucleotide levels were utilized for the more subtle functional evaluation of cell parameters. The results show that the presence of fusinite particles, even after long internalization times, does not induce any cytotoxic effects in the cells studied. Thus, from these results, it can be deduced that fusinite is non-toxic as well as highly stable, inert and very sensitive to oxygen, and can be used with great success for cell studies where determination of oxygen concentration is important.


Subject(s)
Carbon/toxicity , Oxygen/analysis , Spin Labels , Adenine Nucleotides/analysis , Adenine Nucleotides/metabolism , Cell Division/drug effects , Cell Membrane/physiology , Cells, Cultured , Electric Conductivity , Electron Spin Resonance Spectroscopy , Microscopy, Electron , Microscopy, Fluorescence , Molecular Probes/toxicity , Phagocytosis
16.
Biomaterials ; 17(9): 921-8, 1996 May.
Article in English | MEDLINE | ID: mdl-8718938

ABSTRACT

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.


Subject(s)
Acetylcysteine/pharmacology , Antioxidants/pharmacology , Biocompatible Materials , Cell Adhesion/drug effects , Cell Line , Cytoskeletal Proteins/metabolism , Epithelial Cells , Epithelium/drug effects , Epithelium/metabolism , Extracellular Matrix Proteins/metabolism , Humans , Hyaluronan Receptors/metabolism , Integrin alpha3beta1 , Integrin alpha6beta1 , Integrins/metabolism , Intercellular Adhesion Molecule-1/metabolism , Materials Testing , Microscopy, Fluorescence , Receptors, Collagen
17.
Infect Immun ; 63(10): 3936-44, 1995 Oct.
Article in English | MEDLINE | ID: mdl-7558302

ABSTRACT

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.


Subject(s)
Actins/biosynthesis , Bacterial Toxins/toxicity , Botulinum Toxins , Cytotoxins/toxicity , Escherichia coli Proteins , GTP-Binding Proteins/metabolism , ADP Ribose Transferases/toxicity , Cells, Cultured , Cytochalasin D/pharmacology , rho GTP-Binding Proteins
18.
Biochim Biophys Acta ; 1243(1): 110-6, 1995 Jan 18.
Article in English | MEDLINE | ID: mdl-7827098

ABSTRACT

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.


Subject(s)
Carbon , Coal , Electron Spin Resonance Spectroscopy/methods , Molecular Probes , Oxygen/analysis , Cell Division , Microscopy, Electron , Oxygen Consumption
19.
J Photochem Photobiol B ; 26(3): 265-70, 1994 Dec.
Article in English | MEDLINE | ID: mdl-7853118

ABSTRACT

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.


Subject(s)
Cytoskeleton/radiation effects , Ultraviolet Rays , Actins/radiation effects , Actins/ultrastructure , Carcinoma, Squamous Cell , Cell Line , Cell Membrane/radiation effects , Cell Membrane/ultrastructure , Cytoskeleton/ultrastructure , Dose-Response Relationship, Radiation , Humans , Microscopy, Electron, Scanning , Microtubules/radiation effects , Microtubules/ultrastructure
20.
FEBS Lett ; 336(2): 335-9, 1993 Dec 27.
Article in English | MEDLINE | ID: mdl-8262257

ABSTRACT

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.


Subject(s)
Apoptosis , CD4 Antigens/metabolism , Apoptosis/drug effects , Cell Line , Cell Membrane/immunology , Cycloheximide/pharmacology , Down-Regulation , HIV/physiology , Humans , Tumor Necrosis Factor-alpha/pharmacology
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