Biomedical subjects

N Franceschini

Publications and source records attributed to N Franceschini.

At least 55 records · Page 3Linked to original sources

PubMed1993

Dye-induced photolesion in the mammalian retina: glial and neuronal reactions.

Irradiation in the presence of a dye applied to the extracellular space is known to produce degenerative features in irradiated neurones and fibers. In the present study, we confirmed the potential use of this procedure as a lesion technique by showing the removal of degenerating elements as part of the glial reaction to the lesion. The dye Rose Bengal was applied to the vitreous body of a rat eye and a T-shaped irradiation pattern was projected onto the retina within the absorption band of the dye. Degenerative features were restricted to the irradiated area, which could be readily identified from its shape. Retinae examined after various survival times showed that macrophages invaded the damaged area within 1 day, and that mitotic activity of reactive glial cells subsequently occurred in the vicinity of the wound. Both cell types were identified by their structural features. Macrophages were also revealed by a staining technique using the dye Nile Red, whereas reactive glial cells were immunolabeled with an antibody directed against the glial fibrillary acidic protein. Reactive glial cells helped the macrophages to gradually remove injured cells and damaged processes. Their main task, however, appeared to be in scar formation, since their processes seemed to suture the lips of the wound together and restore the limiting membrane at the inner retina. After 2 months' survival time, the parent ganglion cells of most disrupted axon bundles had retrogradely degenerated, but regenerating ganglion cell axons were also observed. These results provide some new data about healing processes in the retina. They demonstrate that the dye-induced photolesion technique can be used to either remove or axotomize selected neurones in neural networks which have been made optically accessible.

Animals↗

PubMed1993

A class-A beta-lactamase from Pseudomonas stutzeri that is highly active against monobactams and cefotaxime.

A beta-lactamase produced by Pseudomonas stutzeri was purified to protein homogeneity, and its physicochemical and catalytic properties were determined. Its profile was unusual since, in addition to penicillins, the enzyme hydrolysed second- and third-generation 'beta-lactamase-stable' cephalosporins and monobactams with similar efficiencies. On the basis of the characteristics of the interaction with beta-iodopenicillanic acid, the enzyme could be classified as a class-A beta-lactamase. However, when compared with most class-A beta-lactamases, it exhibited significantly lower kcat./Km values for the compounds usually considered to be the best substrates of these enzymes.

Cefotaxime↗

PubMed1993

In vitro activity of cefodizime (HR-221) in combination with beta-lactamase inhibitors.

Cefodizime (formerly HR221) was tested either for in vitro microbiological activity or for its stability to beta-lactamases in the presence of two beta-lactamase inhibitors (clavulanic acid, tazobactam). Cefodizime was a poor substrate of class C enzymes but hyperproducer strains were generally resistant with or without a beta-lactamase inhibitor used in combination. On the contrary, class A enzymes were able to hydrolyze cefodizime. However, strains expressing class A beta-lactamase were susceptible to cefodizime in combination with clavulanic acid.

Anti-Bacterial Agents↗

PubMed1993

Pigment granule migration and phototransduction are triggered by separate pathways in fly photoreceptor cells.

We studied the mechanism of pigment granule migration in fly photoreceptors. We found that these granules migrate transiently with red light stimulation. This suggests that excitation of metarhodopsin triggers granule migration. We also found that cyclic GMP is one of the activators of pigment granule migration. These findings suggest that pigment granule migration is triggered separately from phototransduction, which is initiated by the excitation of rhodopsin.

Animals↗

PubMed1993

S-adenosylhomocysteine hydrolase from Acinetobacter calcoaceticus: purification and partial characterization.

S-adenosylhomocysteine hydrolase (SAHase) was purified to homogeneity from the Gram negative strain Acinetobacter calcoaceticus 501. The molecular weight of the native enzyme, estimated by gel permeation, was about 288 KDa, while sodium dodecyl sulfate polyacrylamide gel electrophoresis yielded a relative molecular mass of 48 KDa. The determination of the coenzyme content gave 4 mol of NAD+ and 2 mol of NADH per mol of enzyme. The isoelectric point of native SAHase was at pH 5.1. When assayed in the hydrolytic direction, the Km for S-adenosylhomocysteine and the Vmax of the enzyme for this substrate were 84 microM and 357 mumol/min/mg, respectively; in the synthetic direction, instead, the Km for adenosine and the corresponding Vmax value were 1.6 microM and 37 mumol/min/mg. Substrate analogs were tested for their ability to act as inhibitors and inactivators of the enzyme. Among these compounds, 9-beta-arabinofuranosyl adenine (Ara A) appeared as the most powerful competitive inhibitor (Ki = 18 microM) as well as the strongest time-dependent inactivator. The common feature of all the assayed analogs was the presence of the adenine ring in their molecular structure. It can thus concluded that the presence of the adenine moiety is an essential element in substrate and/or inhibitor interaction with this bacterial enzyme.

Acinetobacter calcoaceticus↗

PubMed1992

Resistance to beta-lactams in Mycobacterium fortuitum.

It is widely assumed that the high level of intrinsic resistance to beta-lactam antibiotics exhibited by mycobacteria results from the combination of factors including permeability to the drugs, beta-lactamase production, and affinity for penicillin-binding proteins (PBPs). We conducted an evaluation of the second and third factors by isolating nitrosoguanidine-induced mutants from the beta-lactamase-producing strain Mycobacterium fortuitum ATCC 19542 that displayed either elevated or reduced resistance to various beta-lactam antibiotics. The mutants studied included D1 (a beta-lactamase producer with high penicillin resistance), gamma 27 (a low-level beta-lactamase producer with low penicillin resistance), and D316 (a high-level beta-lactamase producer with high penicillin resistance). In all strains examined, four major PBPs, named 1, 2a, 2b, and 3, with apparent molecular weights of 102,000, 90,000, 87,000, and 50,000, respectively, were found. The MICs of various beta-lactams toward ATCC 19542 and its mutants were considered in the context of beta-lactamase production, the quantity of PBPs synthesized, and their affinities for beta-lactam antibiotics. The data obtained show that beta-lactamase production is likely to be an important factor in the expression of resistance by clinical isolates and that PBP alterations can contribute to resistance at least in laboratory-derived mutants.

Anti-Bacterial Agents↗

PubMed1991

Cloning and nucleotide sequencing of the gene encoding the beta-lactamase from Citrobacter diversus.

The gene coding for the class A beta-lactamase of Citrobacter diversus has been cloned and sequenced. It contains the information for a 294-amino-acid precursor protein, including a 27-residue N-terminal signal peptide. The deduced sequence of the N-terminal portion of the mature protein is in excellent agreement with that determined by microsequencing of the protein and readily explains the pI differences observed between the naturally occurring forms I and II of the enzyme. The sequence of the mature protein exhibits a very high degree of similarity with that of the Klebsiella oxytoca class A beta-lactamase.

Amino Acid Sequence↗

PubMed1991

Proteolytic interconversion and N-terminal sequences of the Citrobacter diversus major beta-lactamases.

The N-terminal sequences of the two major beta-lactamases produced by Citrobacter diversus differed only by the absence of the first residue in form II and the loss of five amino acid residues at the C-terminal end. Limited proteolysis of the homogeneous form I protein yielded a variety of enzymatically active products. In the major product obtained after the action of papain, the first three N-terminal residues of form I had been cleaved, whereas at the C-terminal end the treated enzyme lacked five residues. However, this cannot explain the different behaviours of form I, form II and papain digestion product upon chromatofocusing. Form I, which was sequenced up to position 56, exhibited a very high degree of similarity with a Klebsiella oxytoca beta-lactamase. The determined sequence, which contained the active serine residue, demonstrated that the chromosome-encoded beta-lactamase of Citrobacter diversus belong to class A.

Amino Acid Sequence↗

PubMed1991

Some molecular properties of Citrobacter diversus beta-lactamases.

Citrobacter diversus ULA-27, a clinical isolate showing a broad resistance pattern towards both penicillins and cephalosporins, produces chromosome encoded beta-lactamases. However, the strain remains susceptible to some cephamycins, imipenem, ceftazidime and tetracyclines. Crude bacterial extracts analyzed by isoelectric focusing on polyacrylamide gels, revealed the presence of two main isoforms and some "satellite" bands focusing in the pH range 5.7-7.2. The isoform showing the pIs 6.8 and 6.2 were characterized as class "A" beta-lactamases (according to Ambler's classification) based on the rate of interaction of beta-iodopenicillanate and the amino acid sequence around the active site serine. The substrate specificity of the Citrobacter diversus beta-lactamases explains the resistance phenomenon of this bacterium to penicillins and cephalosporins.

Cephamycins↗

PubMed1991

Beta-lactamase induction antagonizes beta-lactam susceptibilities in Citrobacter diversus and Enterobacter cloacae clinical isolates.

Inducible beta-lactamases were obtained after exposure to several beta-lactams in clinical isolates of Enterobacter cloacae and Citrobacter diversus. Enzyme production was related to the inducer and medium composition. beta-lactamase is able to inactivate only labile compounds, thus generating minimum inhibitory concentrations higher than in the absence of the inducer; imipenem susceptibilities usually were not changed.

Citrobacter↗

PubMed1991

Beta-lactamase of Mycobacterium fortuitum: kinetics of production and relationship with resistance to beta-lactam antibiotics.

The kinetics of both intracellular and extracellular beta-lactamase production and the relationship between extracellular enzyme and in vitro susceptibility of Mycobacterium fortuitum to beta-lactam antibiotics have been studied. To this end we used a panel of stable nitrosoguanidine-induced mutants of M. fortuitum derived from the parental strain ATCC 19542 and differing in beta-lactamase production from 0.0001 to 278 U/liter in Mueller-Hinton broth. For overproducers of beta-lactamase (mutants A188, B180, C207, D316, and E31), MICs of benzylpenicillin, amoxicillin, ampicillin, and cephaloridine progressively increased with the amount of enzyme released into the medium, whereas MICs of imipenem and cefoxitin did not. The resistance of the mutants to amoxicillin was reduced up to 32-fold by clavulanic acid, whereas that to ampicillin was reduced 8-fold by sulbactam. These data suggest that the enzyme participated in the mechanisms of resistance to the beta-lactam antibiotics. However, for a mutant of M. fortuitum (gamma 27) with virtually nonexistent beta-lactamase production, the antibiotics still had relatively high MICs (for instance, benzylpenicillin and cephaloridine had MICs of 64 and 32 micrograms/ml, respectively). This suggests that, aside from beta-lactamase production, other mechanisms such as cell wall permeability and/or affinity for penicillin-binding proteins could coexist in M. fortuitum and explain its natural resistance to beta-lactam antibiotics.

Anti-Bacterial Agents↗

PubMed1991

Some molecular properties of human seminal transferrin (HSmT) in comparison with human serum transferrin (HSrT).

In this paper we report some structural features of human seminal transferrin (HSmT) in comparison with the homologous protein purified from human serum (HSrT). In particular, the sequence of the first 13 N-terminal amino acids of HSmT shows 12/13 of identity with the first 13 N-terminal amino acids of HSrT, the ninth residue of the former protein being not definitely determined. Moreover, HSrT and HSmT analysed under the same conditions, by means of reversed phase HPLC, thiol groups determination and second derivative spectroscopy, show a different content of amino acids. In particular, HSmT exhibits mainly: i) a lower Asx/Glx ratio; ii) a reduction of about 50% in Cys residues; iii) a decrease of Tyr and Trp residues. Eventually oligosaccharide parallel analyses of HSmT and HSrT show the same glycosidic bond and almost the same sugar content (around 5.5% w/w); conversely, HSmT lacks of sialic acid residues and probably it contains fucose. These results, taken all together, could be sound of interest to a better understanding of the possible physiological roles of HSmT.

Amino Acid Sequence↗

PubMed1990

Characterization of a beta-lactamase produced in Mycobacterium fortuitum D316.

A beta-lactamase from Mycobacterium fortuitum D316 was purified and some physico-chemical properties and substrate profile determined. On the basis of its N-terminal sequence and of its sensitivity to beta-iodopenicillanate inactivation, the enzyme appeared to be a class A beta-lactamase, but its substrate profile was quite unexpected, since nine cephalosporins were among the eleven best substrates. The enzyme also hydrolysed ureidopenicillins and some so-called 'beta-lactamase-stable' cephalosporins.

Amino Acid Sequence↗

PubMed1990

Dye-induced 'photo-degeneration' and 'photo-permeabilization' of mammalian neurons in vivo.

Dyes are known to induce neuronal 'photo-degeneration' and 'photo-permeabilization' in fly photoreceptor cells in vivo. In the present study, we attempted to generalize this photodynamic damage to vertebrate neurons, using the rat retina, a brain part which is optically accessible in vivo. After intravitreal injection of the photosensitizing dye Rose Bengal (RB), irradiation of the retina of a living rat with a T-shaped microbeam was found to induce striking 'optograms' which could be observed on the excised retina. The T-shaped pattern which was to be seen in the translucent retina under transmitted light was attributed to neuronal degeneration of the neurons irradiated in the presence of RB, as attested by classical degenerative features such as a cytoplasmic darkening or a drastic swelling. The T-shaped pattern could also be observed on adding the dye Lucifer yellow to the extracellular space of the retina either in vitro or in vivo, showing that the cells irradiated in the presence of RB became permeable. These structural reactions were observed in the cells in the inner nuclear layer (INL) and ganglion cell layer (GCL), in the processes in both plexiform layers, and in the ganglion cell axons crossing this area, whereas the photoreceptors in the outer retina appeared to be undamaged. From these reactions, due to photo-degeneration and photo-permeabilization, it was possible to identify the photodynamic damage to the nervous system histologically at the macroscopic, cellular and ultrastructural levels. In view of its accuracy and reproducibility, the photo-lesion technique holds great potential as a tool for investigating various nervous systems.

Animals↗

PubMed1990

Dye-induced photopermeabilization and photodegeneration: a lesion technique useful for neuronal tracing.

Using as a neural system the fly retina, which is visually accessible in vivo, we describe a lesion technique that takes advantage of the photodynamic damage produced by extrinsic dyes. Contrary to the photo-inactivation technique described by Miller and Selverston (1979), this technique does not involve intracellular injection, since the dye is applied to the extracellular space of the tissue. This treatment was found to trigger neuronal degeneration and cell permeabilization in fly photoreceptor neurones. We coined the names 'photodegeneration' and 'photopermeabilization' for these two phenomena. While the technique can be used to delete given neurones from the neural circuit after several days' survival time, it was found to produce adequate cytoplasmic labelling for anatomical studies with both light and electron microscopy. Since the area occupied by the degenerating cells is restricted to the light spot imaged onto the nervous tissue, the resolution with this lesion technique can range from single cells to whole neuronal populations. The remarkable precision of the 'photolesions' produced in this way makes this technique a powerful tool for physiological and anatomical investigations on real neural networks, whenever these can be made optically accessible in vivo or in situ.

Animals↗

PubMed1990

Broad spectrum beta-lactamases of Citrobacter diversus.

Citrobacter diversus NF85 produced a chromosomal beta-lactamase that was induced by a variety of beta-lactam antibiotics. Two major forms of the enzyme, with isoelectric points (pI's) of 5.7 and 6.2, were found in crude cell extracts. Derepressed mutants of NF85, generated by nitrosoguanidine treatment, displayed different levels of beta-lactamase expression to the parent strain and had different patterns of resistance to a range of beta-lactam antibiotics. Those mutants of NF85 that were totally derepressed, expressing high, constitutive levels of enzyme, were found to have an additional beta-lactamase activity with a pI of 6.8.

Anti-Bacterial Agents↗

PubMed1990

CTX and its desacetyl derivative (des-CTX): interaction with some representative beta-lactamases and their related pattern of resistance to newer selected clinical isolates.

In this study the kinetic features of cefotaxime (CTX) and desacetyl-cefotaxime towards several representative beta-lactamases were investigated. Desacetyl-CTX was more stable to hydrolysis in comparison with cefotaxime for all the investigated enzymes. However, a cephalosporinase produced in Acinetobacter was progressively inactivated by both CTX and des-CTX. After prolonged incubation, dialysis partially restored the enzyme activity. Finally, both compounds were tested against selected resistant strains. It is concluded that des-CTX, because of either poor hydrolysis or prolonged half-life in body fluids, could contribute in vivo to the good antimicrobial properties of cefotaxime.

Anti-Bacterial Agents↗

PubMed1989

Selective illumination of single photoreceptors in the house fly retina: local membrane turnover and uptake of extracellular horseradish peroxidase (HRP) and lucifer yellow.

Single photoreceptor cells in the compound eye of the housefly Musca domestica were selectively illuminated and subsequently compared electron-microscopically with the unilluminated photoreceptors in the immediate surroundings. The rhabdomeres of the illuminated cells remain largely unaffected, but the cells show an increase in the number of coated pits, various types of vesicles, and degradative organelles; some of the latter organelles are described for the first time in fly photoreceptors. Coated pits are found not only at the bases of the microvilli, but also in other parts of the plasma membrane. Degradative organelles, endoplasmic reticulum (ER) and mitochondria aggregate in the perinuclear region. The rough ER and smooth ER are more elaborate, the number of Golgi stacks, free ribosomes and polysomes is increased, and the shape and distribution of heterochromatin within the nuclei are altered. Illuminated photoreceptors also interdigitate extensively with their neighbouring secondary pigment cells. These structural changes in illuminated fly photoreceptor cells indicate an increase in membrane turnover and cellular metabolism. When applied to the eye, Lucifer Yellow spreads into the extracellular space and is taken up only by the illuminated photoreceptor cells. These cells show the same structural modifications as above. Horseradish peroxidase applied in the same way is observed in pinocytotic vesicles and degradative organelles of the illuminated cells. Hence, the light-induced uptake of extracellular compounds takes place in vivo at least partially as a result of an increase in pinocytosis.

Animals↗