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Biomedical subjects

M Falconi

Publications and source records attributed to M Falconi.

At least 163 records · Page 9Linked to original sources
PubMed1990

Mapping the copper ligands of Cu,Zn superoxide dismutase by nuclear Overhauser enhancement of the isotropically shifted 1H-NMR lines of the Cu,Co derivative.

Transient nuclear Overhauser effect (NOE) experiments were made with bovine Cu,Co superoxide dismutase on the hyperfine shifted resonances of protons of the imidazole groups bound to the Cu,Zn cluster of the native enzyme. Assignments of lines based on the observed magnetization transfers were satisfactorily obtained only by altering the arrangement of the ligands around the catalytically active copper shown by X-ray crystallography.

Animals↗
PubMed1990

High-resolution detection of newly synthesized DNA by anti-bromodeoxyuridine antibodies identifies specific chromatin domains.

We analyzed the incorporation of bromodeoxyuridine (BrdUrd) into DNA in exponentially growing murine erythroleukemia cells (FLC-745), using fluorescent anti-BrdUrd antibodies with light microscopy and flow cytometry. The fine localization of the DNA replicating sites was investigated at the ultrastructural level by using a second antibody conjugated with colloidal gold. The latter approach, which does not require acidic denaturation of the DNA, enables preservation of good morphology and obtains a better resolution power than that of electron microscopic autoradiography, the percentage of labeled cells obtained with the two techniques being comparable. After short BrdUrd pulses, characteristic distribution of the labeling can be identified in the heterochromatin, in interchromatin domains, or at the boundary between the dispersed and the condensed chromatin. Similar patterns are also observable in the nuclear structures which condense after acid denaturation, suggesting that DNA replication takes place at fixed sites associated with the nuclear matrix.

Animals↗
PubMed1990

Retroperitoneal and peritoneal drainage and lavage in the treatment of severe necrotizing pancreatitis.

In severe necrotizing pancreatitis, the retroperitoneum is the main site both of autodigestion and of the production of toxins. With the aim of removing necrotic tissues and active enzymes from the retroperitoneum, we developed a surgical approach based on a wide exposure of the pancreas and on the insertion of multiple drainages with postoperative irrigations with hypertonic solutions and antiproteases. We treated 191 patients, and our results correlated with the timing of the operation. The operative mortality rate ranges from 8.1 in patients undergoing our procedure within 48 hours from the onset of the disease to 28.4 per cent when the operation was delayed for more than 96 hours. Our technique is detailed and the indications as to the timing of surgical treatment in instances of pancreatitis are discussed.

Acute Disease↗
PubMed1989

Conservation of local electric fields in the evolution of Cu,Zn superoxide dismutase.

The trend of the electric field and the value of the electric field flux, sensed by the superoxide substrate in the proximity of the active site, were found to be constant in three highly homologous Cu,Zn superoxide dismutases from ox, pig and sheep, which display large differences in net protein charge and distribution of electrically charged surface residues but very similar catalytic rate constants. The spatial relationship of charges on the protein surface apparently has been conserved during the evolution of this enzyme to create electrostatic facilitation of catalysis.

Amino Acid Sequence↗
PubMed1989

Crystallographic characterization and three-dimensional model of yeast Cu,Zn superoxide dismutase.

The Cu,Zn superoxide dismutase from yeast was crystallized in the orthorhombic space group P21212 with unit cell dimension a = 105.1 A,b = 142.2 A, c = 62.1 A. The crystals grow in 25 mM citrate, 10 mM phosphate buffer pH 6.5, and 6% (W/V) polyethylene glycol, with a Vm of 3,4 A3/dalton, for two dimers/asymmetric unit. The crystals were unstable in the mother liquor, but were stabilized by transfer to a 35% polyethylene glycol solution. This crystalline form diffracts at high resolution and is suitable for determination of the atomic structure. The three dimensional structure of the yeast enzyme could be model-built by computer graphics techniques using the bovine enzyme atomic coordinates as template. The proposed model requires removal of some salt bridges and non equivalence of the metal-binding sites in the subunits, in line with reported functional properties of the yeast enzyme.

Amino Acid Sequence↗
PubMed1989

Characterization of the structural genes for the DNA-binding protein H-NS in Enterobacteriaceae.

The promoter region of Escherichia coli hns, the structural gene for the DNA-binding protein H-NS, has been identified by use of a promoter search vector and the in vivo transcriptional start point by primer extension analysis. The homologous hns genes of two other Enterobacteriaceae, Proteus vulgaris and Serratia marcescens, were identified by heterologous hybridization with a DNA probe derived from E. coli hns, cloned and sequenced. Taking into account only the invariant nucleotides and amino acids, the homology of H-NS among the three organisms was found to be greater than 70% at the DNA level and greater than 75% at the protein level. The three hns genes were also found to have nearly identical transcriptional and translational signals.

Amino Acid Sequence↗
PubMed1989

Continuous peritoneal dialysis in acute experimental pancreatitis in dogs. Effect of aprotinin in the dialysate medium.

In 27 beagle dogs, acute necrotizing pancreatitis was induced by retrograde injection of autologous bile and trypsin into the main pancreatic duct. Animals were randomly assigned to the following treatments: group 1--(9 dogs) aprotinin 600,000 KIU/d by i.v. route; group 2--(9 dogs) peritoneal dialysis for 6 d plus 500,000 KIU/L of aprotinin in the dialysate fluid; group 3--(9 dogs) peritoneal dialysis without aprotinin in the dialysate fluid. All dogs of the group 1 died within 16 h following the induction of pancreatitis and extensive necrotizing and hemorrhagic changes were seen in the pancreatic and peripancreatic areas. Six dogs of the group 2 survived and no necrotizing changes were observed 30 or 50 d after the induction of pancreatitis. Three dogs of the group 3 survived, but slight necrotizing lesions were found at the autopsy. The survival rate was higher in dogs with peritoneal lavage (p = 0.0129) or with peritoneal lavage plus aprotinin (p less than 0.0001) than in those receiving i.v. aprotinin, indicating that the latter treatment has no beneficiary effect on the course of acute pancreatitis.

Acute Disease↗
PubMed1989

Pancreaticoduodenal graft in the rat: an original microsurgical technique.

The authors describe an original surgical technique for pancreatic transplantation carried out on an experimental rat model which duplicates as closely as possible the anatomic-functional situation of a pancreatic graft in man (one arterovenous vascular peduncle with one functioning kidney only). The technique entails a microanastomosis between the mesenteric artery and the portal vein of the donor organ with the left renal artery and vein of the recipient. This method appears to be technically encouraging and particularly useful in studying the early phases of ischemic graft injuries, a field in which the authors are particularly interested.

Animals↗
PubMed1988

Gradient fractionation of cycling and resting cells monitored by BrdUrd incorporation.

A number of techniques are currently employed for the fractionation of heterogeneous cell populations or for the separation of cells in different phases of their cycle. With the development of osmotically inert colloidal silica particles media, density gradient centrifugation became an established method for the separation and purification of cells and subcellular particles. We have applied this technique to the separation of cycling from resting Friend erythroleukemia cells, to obtain purified populations for further biological assays. The flow cytometric analysis of DNA content of the different fractions obtained by the gradient and stained with Propidium Iodide (PI), showed the S compartment highly concentrated in the 1.073/77 g/ml interface, while the upper levels of the gradient were highly enriched of cells in G1 phase. Moreover, the dual parameter analysis of DNA content by means of Bromodeoxyuridine (BrdUrd) incorporation and PI staining, showed that part of the cells in the 1.067/73 fraction represented the early S phase even if their DNA level, measured on the basis of PI fluorescence was within the diploid cell cluster. This method seems to be suitable to obtain pure cell fractions even when dealing with numerically large populations.

Animals↗
PubMed1988

Is the activity-linked electrostatic gradient of bovine Cu, Zn superoxide dismutases conserved in homologous enzymes irrespective of the number and distribution of charges?

Electrostatic potential calculations have been performed on three different Cu, Zn superoxide dismutases (superoxide: superoxide oxidoreductase, EC 1.15. 1.1), in order to evaluate the degree of conservation of the pattern of electrostatic interactions between O2- and the active site recently pointed out in bovine Cu Zn SOD. The three Cu, Zn SODs that have been selected for this study, namely the bovine, ovine, and porcine enzymes, are highly homologous as to reasonably assume identical three-dimensional structure but display large differences in their net charge, as shown by their pI's, which span over a wide pH range: 8.0 (sheep), 6.5 (pig), 5.2 (ox). Despite such a large difference in the net protein charge and in the spatial arrangement of electrostatic charges, electrostatic potential calculations show that the electrostatic channel directing the negatively charged substrate toward the positive catalytic site is strictly preserved with the same features for the three proteins. This suggests that the electrostatic funnel for conducting small anions into the active site is a highly conservative property in the evolution of Cu, Zn SOD.

Animals↗
PubMed1988

Proteins from the prokaryotic nucleoid: primary and quaternary structure of the 15-kD Escherichia coli DNA binding protein H-NS.

The primary sequence of H-NS (136 amino acid residues, Mr = 15,402), an abundant Escherichia coli DNA-binding protein, has been elucidated and its quaternary structure has been investigated by protein-protein cross-linking reactions. It was found that H-NS exists predominantly as a dimer, even at very low concentrations, but may form tetramers at higher concentrations and that the protein-protein interaction responsible for the dimerization is chiefly hydrophobic.

Amino Acid Sequence↗
PubMed1987

Ciprofloxacin penetration in pancreatic juice.

The penetration of ciprofloxacin in pancreatic juice was investigated in 5 patients with pancreatic fistula. The drug was administered as a single oral dose of 500 mg after which serial samples of pancreatic juice and serum were collected for ciprofloxacin assay. The following pharmacokinetic parameters (mean +/- SD) were estimated from the serum level versus time curves: clearance 11.51 +/- 2.85 (ml/min/kg); Vd area 3.08 +/- 1.20 ml/kg; terminal half-life 3.10 +/- 0.92 h; mean residence time 5.64 +/- 1.40 h. Ciprofloxacin serum levels declined rapidly after the third hour, whereas concentrations in pancreatic juice remained elevated (above 1 mg/1) for nearly 12 h. The pancreatic juice/serum ciprofloxacin concentration ratio increased gradually fom 0.63 +/- 0.45 after 0.5 h to 6.18 +/- 4.59 after 12 h (mean +/- SD). Our data indicate that while the drug elimination half-life from the serum is short, the time-course of ciprofloxacin levels in the pancreatic juice conforms to a much slower disappearance rate. In particular, the ciprofloxacin levels achieved in pancreatic juice are constantly greater than the MICs of the bacteria generally responsible for pancreatic infections.

Adult↗
PubMed1986

Conservative treatment of external pancreatic fistulas with parenteral nutrition alone or in combination with continuous intravenous infusion of somatostatin, glucagon or calcitonin.

Treatment with total parenteral nutrition (TPN) alone or combined with continuous intravenous infusion of either somatostatin or calcitonin or glucagon have been carried out upon 45 patients with a high output external pancreatic fistula. No significant difference among these treatment schedules was observed in the percentage of closure of fistulas (85 to 100 per cent of the patients), but patients treated with TPN plus somatostatin had the fistulas close within a significantly (p = 0.000028) shorter period of time. Moreover, this treatment was associated with the strongest inhibition of the output from the fistula (minus 82.3 per cent). Since treatment with somatostatin was not accompanied by any side effect, was followed by a quite rapid closure of the fistulas and allowed an estimated economic savings of about $2,100.00 dollars per patient, it seems to be advisable in the conservative treatment of external pancreatic fistulas.

Calcitonin↗
PubMed1986

[Epithelial splenic cysts].

The authors describe a case of an epithelial cyst of the spleen treated successfully by splenectomy. Diagnostic techniques and the principles adopted in treatment this rare disease are discussed.

Adult↗
PubMedDate not supplied

Membrane-perturbing activity of Viperidae myotoxins: an electrostatic surface potential approach to a puzzling problem.

Phospholipase-like myotoxins are a class of proteins present in Viperidae venom. Despite the high level of amino acid and structural homology with soluble phospholipases A(2), myotoxins are devoid of enzymatic activity and share cytolytic activity by means of a totally unknown mechanism involving the lipid bilayer perturbation. The distribution of electrostatic surface potentials of four myotoxins and seven phospholipases A(2) has been compared. The charge distribution is similar in all active non-cytolytic phospholipases with a strongly positive side corresponding to the domain interacting with the micellar substrate and with the opposite side negatively charged. In contrast, all myotoxins examined are positively charged on both sides. Myotoxin III, the only known example of a myotoxin sharing enzymatic activity, displays the same electrostatic surface potential as other related toxins. Using liposomes made with non-hydrolysable phospholipids, we demonstrate that myotoxin III perturbs the lipid bilayer like other myotoxins. Based on these results, a molecular model for myotoxin-membrane perturbing activity is proposed. In this model, potential double-face binding of myotoxic phospholipases A(2) to lipid surfaces could trigger a lipid bilayer destabilization and could generate a stable fusion pore, probably because of the presence of hydrophobic moieties that flank the cationic sites.

Cell Membrane Permeability↗