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Biomedical subjects

A Bellelli

Publications and source records attributed to A Bellelli.

At least 73 records · Page 4Linked to original sources
PubMed1992

A ribosomal protein is specifically recognized by saporin, a plant toxin which inhibits protein synthesis.

Many plants express enzymes which specifically remove an adenine residue from the skeleton of the 28 S RNA in the major subunit of the eukaryotic ribosome (ribosome inactivating proteins, RIPs). The site of action of RIPs (A4324 in the rRNA from rat liver) is in a loop structure whose nucleotide sequence all around the target adenine is also conserved in those species which are completely or partially insensitive to RIPs. In this paper we identify a covalent complex between saporin (the RIP extracted from Saponaria officinalis) and ribosomal proteins from yeast (Saccharomyces cerevisiae), by means of chemical crosslinking and immunological or avidin-biotin detection. The main complex (mol. wt. congruent to 60 kDa) is formed only with a protein from the 60 S subunit of yeast ribosomes, and is not detected with ribosomes from E. coli, a resistant species. This observation supports the hypothesis for a molecular recognition mechanism involving one or more ribosomal proteins, which could provide a 'receptor' site for the toxin and favour optimal binding of the target adenine A4324 to the active site of the RIP.

Antineoplastic Agents, Phytogenic↗
PubMed1992

Cooperative cyanide dissociation from ferrous hemoglobin.

Rapid reduction of cyano-met hemoglobin (Hb+CN-) leads to the formation of an intermediate species, the cyanide derivative of ferrous hemoglobin, which dissociates to unliganded hemoglobin because of the extremely low affinity of the ligand for the ferrous heme iron. The properties of the intermediate were studied by transient spectroscopy in human hemoglobin and its isolated alpha and beta chains, in the presence and absence of CO. When mixing with dithionite, the time courses of reduction of the heme iron and dissociation of cyanide overlap considerably; addition to the reaction mixture of the redox indicator methyl viologen considerably increases the rate of reduction and allows unequivocal determination of the spectroscopic and kinetic properties of the intermediate. The results show that (i) the dissociation of cyanide from the isolated alpha and beta chains (as well as the (alpha CO)2(beta + CN-)2 hybrid) is a simple process; (ii) the two chains display similar rate parameters, but show spectroscopic inequivalence, both in the Soret and the visible regions; (iii) cooperative effects are shown to control the rate of dissociation of cyanide from hemoglobin, similarly to what happens for oxygen; and (iv) allosteric effectors (typically inositol hexaphosphate) increase the overall rate of dissociation by stabilization of the T state. We have, therefore, shown for the first time that the dissociation of cyanide from ferrous hemoglobin is controlled by the quaternary state, thereby adding one more ligand to the analysis of the structure-function relationships in hemoglobin.

Cyanides↗
PubMed1992

Biochemical and rheodynamic properties of red blood cells crosslinked with glutaraldehyde.

New data on the properties of red blood cells (RBC) cross-linked with glutaraldehyde are presented (see also Biochem. Biophys. Res. Commun., 1988, 156, 970-977). Equilibrium and kinetic data show that by carrying out the fixation procedure in the absence of oxygen but in the presence of allosteric effectors (e.g., stabilizing the low-affinity (T) quaternary state of hemoglobin), it is possible to maintain, at least in part, the biochemical functions of the crosslinked hemoglobin inside the cell. Moreover, we show that the oxygen affinity of fixed red blood cells (RBC) is still modulated, even though to a smaller degree, by the allosteric effector bezafibrate (BZF), which is able to cross the fixed RBC membrane. Membrane filtration experiments indicate that the higher rigidity of fixed RBC alters significantly their rheodynamic properties and show that in order to exploit "engineered" RBC as "blood substitutes," more flexible cross-linking reagents may offer significant advantages.

Bezafibrate↗
PubMed1992

[Cholesteatoma of the Prussak's space. Diagnosis with computerized tomography].

Prussak's space localization represents the early stage of the cholesteatoma arising from the pars flaccida of the tympanic membrane. From Prussak's space the mass spreads to the antrum and to mastoid air cells. Its demonstration is infrequent probably due to the limited use of CT in inflammatory diseases of the middle ear. On CT scans, cholesteatomas present as small soft-tissue masses, and their growth is responsible for scutum and ossicular chain erosion. The mass may also spread posteriorly to the aditus and antrum, and involve tendons and ligaments, which is a different behavior than that of fluid collections or granulation tissue. The authors report 9 cases of Prussak's cholesteatoma evaluated by means of HRCT.

Cholesteatoma↗
PubMed1991

On the mechanism and rate of substrate oxidation by amine oxidase from lentil seedlings.

The kinetics of reaction between lentil seedlings amine oxidase and two amine substrates, namely putrescine and dimethylaminomethylbenzylamine, have been studied by rapid mixing with diode array detection. In this way several wavelengths can be monitored at once allowing the simultaneous measurement of enzyme bleaching and formation of a yellow radical intermediate. The two substrates are oxidized at rates that differ by one order of magnitude in favor of putrescine. Of the individual five rate constants measured and/or calculated from the experimental ones, k2 alone, the monomolecular transformation of ES to EP accounts for this difference. The reoxidation step is instead not rate limiting and identical for the two substrates.

Amine Oxidase (Copper-Containing)↗
PubMed1991

Brief communication. on the problem of immunological detection of antigens in skeletal remains.

A detailed investigation with affinity-chromatography-purified fractions of antihemoglobin serum from rabbit shows that the hemoglobin content of human bones dating back 15 to 3,000 years may be very small. Some of the previous results (Ascenzi et al., 1985) indicating a high hemoglobin titer were +vitiated because of an unexpected cross-reactivity of bone extracts with the hemoglobin-unreactive fraction of the antiserum.

Animals↗
PubMed1991

[Mucocele of the paranasal sinus. CT assessment].

The authors report 7 cases of mucoceles of the paranasal sinuses evaluated with plain films and CT. All patients were submitted to surgery. Five patients were affected with frontal mucoceles and 4 with sphenoid ones. In 2 cases the lesion was double. Symptoms were correlated with both site and compressive changes due to expansile growth. All cases were evaluated by means of axial and coronal pre- and post-contrast scans: a satisfactory morphological depiction of the lesion was always achieved. CT values were also calculated, and the relationship of mucocele to the adjacent structures was demonstrated. The modalities of expansile growth, the lack of enhancement in post-contrast scans, and the possible "ring sign" in cases with intracranial spread allow a correct radiological diagnosis. CT is the diagnostic method of choice for best lesion demonstration: it makes a correct diagnosis possible and allows the thorough evaluation of the involved structures for surgical planning.

Adult↗
PubMed1990

Transient spectroscopy of the reaction of cyanide with ferrous myoglobin. Effect of distal side residues.

The reaction of cyanide metmyoglobin with dithionite conforms to a two-step sequential mechanism with formation of an unstable intermediate, identified as cyanide bound ferrous myoglobin. This reaction was investigated by stopped-flow time resolved spectroscopy using different myoglobins, i.e. those from horse heart, Aplysia limacina buccal muscle, and three recombinant derivatives of sperm whale skeletal muscle myoglobin (Mb) (the wild type and two mutants). The myoglobins from horse and sperm whale (wild type) have in the distal position (E7) a histidyl residue, which is missing in A. limacina Mb as well as the two sperm whale mutants (E7 His----Gly and E7 His----Val). All these proteins in the reduced form display an extremely low affinity for cyanide at pH less than 10. The differences in spectroscopy and kinetics of the ferrous cyanide complex of these myoglobins indicate a role of the distal pocket on the properties of the complex. The two mutants of sperm whale Mb are characterized by a rate constant for the decay of the unstable intermediate much faster than that of the wild type, at all pH values explored. Therefore, we envisage a specific role of the distal His (E7) in controlling the rate of cyanide dissociation and also find that this effect depends on the protonation of a single ionizable group, with pK = 7.2, attributed to the E7 imidazole ring. The results on A. limacina Mb, which displays the slowest rate of cyanide dissociation, suggests that a considerable stabilizing effect can be exerted by Arg E10 which, according to Bolognesi et al. (Bolognesi, M., Coda, A., Frigerio, F., Gatti, C., Ascenzi, P., and Brunori, M. (1990) J. Mol. Biol. 213, 621-625), interacts inside the pocket with fluoride bound to the ferric heme iron. A mechanism of control for the rate of dissociation of cyanide from ferrous myoglobin, involving protonation of the bound anion, is discussed.

Animals↗
PubMed1990

Effect of aromatic isothiocyanates on the functional properties of human hemoglobin. Role of the stereochemistry of the charged group.

The effect of chemical modification of hemoglobin with six derivatives of benzene isothiocyanate has been studied. The negatively charged reagents (isothiocyanates of benzoic and benzenesulfonic acids) markedly inhibit the interaction of hemoglobin with allosteric effectors such as H+, Cl- and organic phosphates; the affinity for heme ligands in the absence of effectors is reduced but cooperativity is maintained, making these modified hemoglobins suitable models for a possible 'blood substitute'. The only uncharged reagent tested (isothiocyanate of benzenesulfonamide) increases the oxygen affinity of hemoglobin and affects only slightly the interaction with heterotropic ligands; its potential use as an antisickling drug is under study.

Benzene Derivatives↗
PubMed1990

Ligand binding to a hemoprotein lacking the distal histidine. The myoglobin from aplysia limacina (Val(E7)).

The time course of ligand recombination to the myoglobin from Aplysia limacina, which has Val(E7), was measured following photolysis by flashes of 35 ps to 300 ns with a time resolution of 10 ps or 1 ns. CO shows only biomolecular recombination. O2 has a small geminate reaction with a half-time of tens of picoseconds, but no nanosecond geminate reaction. NO has two picosecond relaxations with half-times of 70 ps (15%) and 1 ns (80%) and one nanosecond relaxation with a half-time of 4.6 ns. The biomolecular rates for O2 and NO are the same: 2 x 10(7) M-1 s-1. Methyl and ethyl isonitriles have a geminate reaction with a half-time of 35 ps. Ethyl isonitrile has, in addition, a nanosecond relaxation (25%) with a half-time of 100 ns. t-Butyl isonitrile has four geminate relaxations (10 ps, 35 ps, 1 ns, and 1 microseconds). Analysis of the results suggests much easier movement of ligand between the heme pocket and the exterior than in sperm whale myoglobin (His(E7]. The reactivity of the heme is little different, placing the effect of the differences from sperm whale myoglobin on the distal side of the heme.

Animals↗
PubMed1990

Cooperative ligand binding of crosslinked hemoglobins at very high temperatures.

Human hemoglobin was reacted with the bifunctional reagent bis(3,5-dibromosalicyl) fumarate to yield a derivative (Hb alpha alpha) crosslinked between the two alpha-chains; when the reaction was carried out with HbA already crosslinked between the two beta-chains by 2-nor-2-formylpyridoxal 5'-phosphate, a doubly crosslinked derivative (Hb alpha alpha beta beta) was obtained. We have observed that both modified hemoglobins are extremely stable up to temperatures of at least 85 degrees C. The carbon monoxide binding kinetics of both crosslinked hemoglobins, studied at temperatures between 15 and 85 degrees C, by means of stopped flow and flash photolysis techniques, show that the ligand-linked allosteric transition is maintained even at the highest temperatures. These results are also relevant to the mechanism of thermal unfolding of human hemoglobin, since they show that dissociation into alpha beta dimers (and exposure of the relatively hydrophobic dimer-dimer interfaces) is an obligatory step in the irreversible denaturation of deoxy and carbon monoxy hemoglobin.

Aspirin↗
PubMed1990

Binding and internalization of ricin labelled with fluorescein isothiocyanate.

The toxic lectin ricin has been covalently labelled with fluorescein isothiocyanate on the enzymatically active A chain. The fluorescein reacted toxin maintains its biological activity. The lateral diffusion coefficient of cell surface bound ricin, studied in two cell lines by fluorescence photobleaching recovery, is D = 1 - 2 x 10(-10) cm2/s. Fluorescence microscopy provides preliminary evidence for secondary endosomes in the cytoplasm.

Animals↗
PubMed1990

Inhibition of tumor growth, invasion and metastasis in papain-immunized mice.

Papain-immunized mice possess serum antibodies which cross-react with cathepsin-B- and cathepsin-H-like endopeptidases isolated from B16 melanoma cells. The growth rate, invasion and metastasis of both the B16 melanoma and the Lewis lung carcinoma were inhibited in mice immunized with papain. These animals presented an increased mean survival time as compared to the tumor-bearing nonimmunized controls. Quantitative microscopy suggested that vasodilation and edema, associated with tumor invasion, are, at least partially, sustained by proteolytic enzymes, being strongly reduced when tumor cells were inoculated in papain-immunized mice.

Abdominal Muscles↗
PubMed1990

[Fibrous dysplasia of the facial bones. A CT study].

The authors report 7 cases of fibrous dysplasia of the facial bones which were evaluated with CT. The involvement of the facial bones by fibrous dysplasia is an uncommon event, which causes different syndromes according to the extent of bone invasion. In all the cases occurred to our observation CT allowed an exact spatial evaluation of the dysplasia, and therefore a correct surgical planning. Moreover, CT densitometric values and CT appearance of fibrous dysplasia often suggested the correct diagnosis.

Adolescent↗
PubMed1990

The amino acid sequence of a ribosome-inactivating protein from Saponaria officinalis seeds.

The complete primary structure of saporin SO-6, a ribosome-inactivating protein extracted from Saponaria officinalis seeds, has been determined. The sequence was reconstructed following purification and analysis of peptides obtained after digestion of the protein with different proteolytic agents. The protein is composed of 253 amino acids, corresponding to a molecular weight of 28,621 Da. Comparison of the primary structure of SO-6 with the sequence deduced from cDNA, shows amino acid substitutions in 11 positions, suggesting a tissue-related genetic variability. When the sequence of saporin is compared to those of two related proteins, ricin A chain and trichosanthin, a low degree of similarity (12%) is found; nevertheless some considerations about structure-function relationships and evolution of RIPs are possible.

Amino Acid Sequence↗
PubMed1989

Clinical response to auranofin in patients with psoriatic arthritis.

Fifty-two patients with psoriatic arthritis (PA), treated with auranofin (AF), were entered into a one year prospective, open study. The total group showed a significant increase in frequency of HLA antigens A1 and B38, and a reduction of B5 when compared to healthy controls. There was a remission or an important improvement of disease in the 51% of 45 patients who completed the study. The rate of withdrawal due to side effects was low (8.8%) and the toxicity was mild in nature (diarrhoea and mucocutaneous rash). We prospectively sought predictors of response using HLA antigens, and clinical and laboratory parameters at the beginning of therapy. The only 3 factors found to be related to outcome were duration of psoriasis, physician and patient assessment of disease activity. No laboratory data or HLA specificities could be associated with substantial response to AF therapy.

Arthritis↗
PubMed1989

Microspectroscopy of red blood cells.

Spectroscopic techniques have been widely employed to analyze properties of macromolecules and dynamics of intracellular events on bulk preparations of cells. The development of computer controlled microspectrophotometers has made possible the study of the same events in single cells, often providing significant and unexpected results. This paper briefly reviews experimental works carried out in our laboratories on single red blood cells. Microspectrophotometric techniques were applied which make use of the fact that ligand binding to intracellular haemoglobin is associated with optical changes. Information on the relative abundance of different haemoglobin components inside single erythrocytes of trout blood was obtained from spectra of air equilibrated samples, taking advantage of the extreme pH sensitivity of one of the four haemoglobin components. The kinetics of oxygen and carbon monoxide binding to haemoglobin has been followed and demonstrated to correspond to a zero order process, with a rate much slower than that characteristic for haemoglobin in solution. These results demonstrate that the process is diffusion limited; computer simulations suggest that ligand uptake is limited by the time required for the diffusion from the extracellular space of enough ligand molecules for total saturation of intraerythrocytic haemoglobin. Finally, oxygen dissociation curves in single red blood cells can be obtained by means of particular flow cell, with promising results for the study of physiological and pathological processes (namely red cell sickling in drepanocytosis).

Animals↗