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Biomedical subjects

G Valentini

Publications and source records attributed to G Valentini.

At least 19 recordsLinked to original sources
PubMed1999

Decay of the GRB 990123 optical afterglow: implications for the fireball model

Broad-band (ultraviolet to near-infrared) observations of the intense gamma ray burst GRB 990123 started approximately 8.5 hours after the event and continued until 18 February 1999. When combined with other data, in particular from the Robotic Telescope and Transient Source Experiment (ROTSE) and the Hubble Space Telescope (HST), evidence emerges for a smoothly declining light curve, suggesting some color dependence that could be related to a cooling break passing the ultraviolet-optical band at about 1 day after the high-energy event. The steeper decline rate seen after 1.5 to 2 days may be evidence for a collimated jet pointing toward the observer.

Journal Article↗
PubMed1999

Rapid detection of ornithine transcarbamylase activity by micellar electrokinetic chromatography.

A quantitative ultraviolet detection method for determining ornithine transcarbamylase (OTC-ase) activity using micellar electrokinetic chromatography (MEKC) is described. The method is based on the direct determination of citrulline formed upon enzymatic reaction. Using a background electrolyte consisting of 35 mM sodium tetraborate, pH 9.3, containing 65 mM sodium dodecyl sulfate (SDS), the peak of citrulline in the electropherogram was easily identified and integrated. This allowed us to determine the rate of formation of the reaction product and to calculate the kinetic parameter Km of the OTC-ase investigated. The capillary electrophoretic method developed was applied to the determination of OTC-ase activity in plasma samples for citrulline in the nanogram range.

Buffers↗
PubMed1999

Inhibition of protein denaturation by fatty acids, bile salts and other natural substances: a new hypothesis for the mechanism of action of fish oil in rheumatic diseases.

Natural hydrophobic substances like bile salts (cholate, deoxycholate, chenodeoxycholate, lithocholate and their conjugates with glycine and taurine), fatty acids (caprylic, capric, lauric, myristic, palmitic, stearic, oleic, linoleic, arachidonic, eicosapentaenoic and docosahexaenoic acid) were much more active (EC50 approximately 10(-4)-10(-5) M) than selected amino acids (EC50 > 10(-2) M) and inorganic salts (EC50 approximately 10(-1) M) in inhibiting heat-induced denaturation of human serum albumin in vitro. Fish oil, rich in n-3-polyunsaturated acids such as eicosapentaenoic acid and docosahexaenoic acid, administered p.o. (1 ml/kg) in the rat, protected ex vivo (after 2 hr) serum against heat-induced denaturation more than bendazac, a known antidenaturant drug. Thus, we speculated that the antidenaturant activity of fish oil may be partly (in addition to the known effect on endogenous eicosanoid composition) responsible for its beneficial effects in rheumatoid arthritis and other rheumatic conditions. In this connection, it is of note that the in vitro antidenaturant activity of fish oil fatty acids was higher than that of known antidenaturant drugs such as bendazac and bindarit and nonsteroidal anti-inflammatory drugs like phenylbutazone and indomethacin which could exert beneficial effects in chronic inflammatory conditions by stabilizing endogenous proteins.

Administration, Oral↗
PubMed1998

Antitumor efficacy of the combination of photodynamic therapy and chemotherapy in murine tumors.

Photodynamic therapy (PDT) is based on the administration of tumor-localizing photosensitizers followed by light exposure of the tumor mass. The photocytotoxic effects are mainly caused by the generation of singlet oxygen. Recently, PDT has been proposed for use in combination with anticancer chemotherapy with a view to exploiting any additive antitumor effect. We investigated the effect of PDT with photoactivated aluminum disulfonated phthalocyanine (AlS2Pc) combined with the antiblastic drugs Adriamycin (ADR) and cisplatinum (CDDP) on murine tumors. Mice bearing L1210 leukemia and P388 lymphoma were treated with ADR or CDDP and subsequently treated with PDT. Low chemotherapy doses were ineffective, but the combination of antiblastic drugs + PDT had a significantly additive antitumor effect. In conclusion, with this combined therapy we were able to greatly reduce the effective doses of antiblastic drugs, thus lowering their toxic effects on normal host tissues.

Animals↗
PubMed1998

Changes of glycosylation of serum proteins in psoriatic arthritis, studied by enzyme-linked lectin assay (ELLA), using concanavalin A.

Changes of glycosylation of serum proteins of patients with psoriatic arthritis were detected by lectin blotting and a new enzyme-linked lectin assay (ELLA) using concanavalin A (Con A). A good linear correlation was found between the total Con A-reactivity of serum and the serum levels of C-reactive protein and interleukin-6, which is known to regulate the glycosylation pattern of proteins upon inflammation. A good linear correlation was also observed between the immunoreactivity of alpha 1-antitrypsin, measured by ELISA, using a monoclonal antibody sensitive to glycosylation changes, and the erythrocyte sedimentation rate and the serum concentrations of soluble interleukin-2 receptor, an index of lymphocyte activation which correlated with some inflammatory parameters of disease activity. These protein changes, which are described here for the first time, deserve to be studied in further detail in view of their possible clinical applications.

Acute-Phase Proteins↗
PubMed1998

Development of a turbidimetric assay to study the effect of urinary components on calcium oxalate precipitation.

The pharmacological treatment of calcium urinary stones, most of which are made of calcium oxalate (CaOx), is only prophylactic. The causes of nephrolithiasis are often unclear, and a number of patients were found to be deficient in physiological inhibitors, e.g. citrate, pyrophosphate, magnesium, and specific proteins. The identification and characterization of these inhibitors can be performed in vitro by a number of methods, most of which are complex and time-consuming. Thus, we developed a simple turbidimetric method based on the precipitation of CaOx from a supersaturated solution. Using this approach, we determined that ionic strengths > 0.2 and pH < 5 inhibited the precipitation of CaOx. The first observation is of interest if one considers that the osmolarity of urine varies in the range of 50-1,400 mmol/kg, while the effect of pH is not usually seen in vivo because of the influence of other phenomena, such as the precipitation of uric acid. The activity of sodium chloride, magnesium and citrate was displayed at concentrations not far from their normal urinary level. Among several mono-, di- and tricarboxylic acids, like acetic, ascorbic, citric, isocitric, formic, fumaric, gluconic, glutaric, alpha-ketoglutaric, maleic, malic, malonic, propionic, pyruvic, succinic, and tartaric acid, only isocitric acid was more potent than citric acid. Pyrophosphate was a potent inhibitor in vitro, but its urinary level may not be sufficient for a significant effect in vivo. Amino acids like Ala, Arg, Asp, Glu, Gly, and Ser which are known to bind calcium, showed little activity. Work is in progress to search for new compounds potentially useful in the treatment of nephrolithiasis.

Calcium Oxalate↗
PubMed1998

Development of an HPLC assay to study the effect of endogenous and exogenous substances on heat-induced aggregation of human serum albumin.

The mechanism of action of non-steroidal anti-inflammatory drugs which are used in high doses in chronic inflammatory conditions is not clearly understood. Their known protein-stabilizing properties could play a significant role. The inhibition of cyclooxygenase may not be essential for their anti-rheumatic activity, since compounds with strong anti-denaturant properties and devoid of anti-inflammatory activity were shown to be effective in an experimental model of rheumatoid arthritis. Hence, to develop new anti-rheumatic drugs it is essential that a simple in vitro method to evaluate the anti-denaturant activity of endogenous and exogenous compounds is available. We developed a new assay, using gel permeation high performance liquid chromatography, to study the effect of endogenous and exogenous compounds on heat-induced aggregation of human serum albumin in conditions in which protein precipitation does not occur. Non-steroidal anti-inflammatory drugs like diclofenac, ibuprofen and naproxen inhibited the aggregation of albumin at low concentrations (EC50 10(-4)-10(-5) mol/l) comparable to those active in a classical turbidimetric method, whereas the effect of weak stabilizers, like sodium cloride and formic, fumaric, maleic, malonic, and succinic acid (EC50 10(-1)-10(-2) mol/l in the Mizushima test) was not detectable. Furthermore, the HPLC assay allowed the examination of a number of coloured substances, including bilirubin, which appeared to be a strong stabilizer of its physiological carrier, albumin. These data could be clinically relevant, since the drugs examined are used at very high doses in rheumatoid arthritis and related conditions, with plasma levels that could cause significant stabilization of serum albumin and perhaps other proteins.

Acids, Acyclic↗
PubMed1998

Effect of selected substances on heat-induced aggregation of albumin, IgG and lysozyme.

Compounds capable of inhibiting protein aggregation may find pharmacological applications in the treatment of a number of diseases called protein condensation diseases [Benedek (1997)], which include cataract, biliary and urinary lithiasis and certain rheumatic diseases. We examined the effect of selected compounds on heat-induced aggregation human serum albumin (HSA), IgG and lysozyme. HSA (0.2% w/v in 0.066 M sodium phosphate pH 5.3 at 22 degrees C), IgG (0.5% w/v in 0.066 M Tris pH 8.0 at 22 degrees C), and L (0.2 % w/v in 0.066 M CAPS pH 11.0 at 22 degrees C) were heated for 30 min at 70 degrees C in the presence or absence of different concentrations of the substance under examination and heat-induced aggregation of 100 microl aliquots was evaluated by measuring the absorbance at 595 nm using an automatic microplate reader. In these conditions, inhibition of aggregation could be due to an anti-denaturant effect or to interferences with the aggregation of denatured molecules, as previously described [Saso, Casini et al. (1998)]. However, this distinction may not be pharmacologically relevant when the target of the therapy is the prevention of abnormal phenomena of protein aggregation. Inorganic salts like NaCl and CaCl2 were active on the three proteins (IgG > HSA > L) but many ligands of HSA such as tryptophan, N-acetyl-tryptophan, caprylic acid, capric acid, cholic acid, deoxycholic acid, chenodeoxycholic acid, lithocholic acid and bendazac were active on their carrier but not on IgG and L, indicating that the latter proteins are more difficult to protect and that specific anti-denaturant and/or anti-aggregant compounds should be developed.

Albumins↗
PubMed1997

A solid tissue phantom for photon migration studies.

A solid tissue phantom made of agar, Intralipid and black ink is described and characterized. The preparation procedure is fast and easily implemented with standard laboratory equipment. An instrumentation for time-resolved transmittance measurements was used to determine the optical properties of the phantom. The absorption and the reduced scattering coefficients are linear with the ink and Intralipid concentrations, respectively. A systematic decrease of the reduced scattering coefficient dependent on the agar content is observed, but can easily be managed. The phantom is highly homogeneous and shows good repeatability among different preparations. Moreover, agar inclusions can be easily embedded in either solid or liquid matrixes, and no artefacts are caused by the solid-solid or solid-liquid interfaces. This allows one to produce reliable and realistic inhomogeneous phantoms with known optical properties, particularly interesting for studies on optical imaging through turbid media.

Agar↗
PubMed1997

Cardiorespiratory changes during gynaecological laparoscopy by abdominal wall elevation: comparison with carbon dioxide pneumoperitoneum.

We have studied the cardiorespiratory changes produced by abdominal wall elevation (AWE) or carbon dioxide pneumoperitoneum (PN) in 20 women undergoing gynaecological laparoscopy. Arterial pressure, heart rate, lung/chest complicance and blood-gas tensions were measured 10 min after induction of general anaesthesia (T0), 10 min after abdominal distension in the supine position (T1) and 10 min after the Trendelenburg position was assumed (T2). Visual analogue scores for pain were recorded 1 and 6 h after the end of surgery. We found that lung/chest compliance was reduced significantly in group PN at T1 and T2 compared with both T0 and group AWE. Diastolic arterial pressure increased significantly in group PN at T1 and T2 compared with both T0 and group AWE, while it remained unchanged in group AWE. Arterial PCO2 increased significantly only in group PN after pneumoperitoneum, while oxygenation was almost unchanged in both groups. AWE patients had greater abdominal pain 1 h after surgery. Six hours after surgery pain was similar in the two groups. These data indicate that abdominal wall elevation reduced pulmonary compliance less than a pneumoperitoneum in patients undergoing gynaecological laparoscopy.

Abdominal Muscles↗
PubMed1997

Fluorescence lifetime imaging of experimental tumors in hematoporphyrin derivative-sensitized mice.

Tumor detection has been carried out in mice sensitized with hematoporphyrin derivative (HpD) by measuring the spatial distribution of the fluorescence lifetime of the exogenous compound. This result has been achieved using a time-gated video camera and a suitable mathematical processing that led to the so-called "lifetime images." Extensive experimental tests have been performed on mice bearing the MS-2 fibrosarcoma or the L1210 leukemia. Lifetime images of mice show that the fluorescence decay of HpD is appreciably slower in the tumor than in healthy tissues nearby, allowing a reliable detection of the neoplasia. The lengthening of the lifetime in tumors depends little on the drug dose, which in our experiments could be lowered down to 0.1 mg/kg body weight, still allowing a definite tumor detection. In order to ascertain the results achieved with the imaging apparatus, high-resolution spectroscopy, based on a time-correlated single photon counting system, has also been performed to measure the fluorescence lifetime of the drug inside the tumor and outside. The outcomes obtained with two techniques are in good agreement.

Animals↗
PubMed1997

Recombinant pyruvate kinase type I from Escherichia coli: overproduction and revised C-terminus of the polypeptide.

The gene encoding pyruvate kinase type I (PKI) of Escherichia coli was amplified by PCR, cloned and sequenced. The gene product was overexpressed in E. coli, using an inducible T7 RNA polymerase-based expression system. The transformed cells contained sixtyfold the enzyme activity of the reference cells and enabled the purification of 30 mg of highly active PKI from 1 liter of culture. The gene sequence was determined and found to be different from the one previously reported, i.e., the T nucleotide at position 1351 was missing. This resulted in a downstream shift of the stop codon, thus the deduced polypeptide was 470 amino acids long instead of 462. In addition the twelve C-terminal amino acids of the former sequence were changed.

Amino Acid Sequence↗
PubMed1996

An essential lysine in the substrate-binding site of ornithine carbamoyltransferase.

Treatment of ornithine carbamoyltransferase from dolphin Stenella with pyridoxal phosphate, followed by reduction with NaBH4 resulted in complete loss of enzyme activity. The phosphate alone or the substrate analogue 2-aminovaleric acid moderately decreased the extent of inactivation, while carbamoyl phosphate plus 2-aminovaleric acid provided complete protection from inactivation. The partially inactivated enzyme showed K(m) values for substrates equivalent to those of native enzyme and lowered Kcat values. Two lysyl residues were substantially modified in the absence of ligands but only one of them was responsible for the inactivation of catalytic activity. Modification of a single subunit was sufficient to completely abolish the catalytic activity of the trimeric enzyme. The lysine involved has been identified as lysine 56 on the known primary structure of homologous human liver enzyme.

Amino Acid Sequence↗
PubMed1996

The allosteric regulation of pyruvate kinase.

Crystallographic and mutagenesis studies have unravelled the general features of the allosteric transition mechanism in pyruvate kinase. The enzyme displays a dramatic conformational change in going from the T- to the R-state. All three domains forming each subunit of the tetrameric enzyme undergo simultaneous and concerted rotations, in such a way that all subunit and domain interfaces are modified. This mechanism is unprecedented since in all tetrameric allosteric enzymes, characterised at atomic resolution, at least one of the domain or subunit interfaces remains unchanged on the T- to R-state transition. The molecular mechanism of allosteric regulation here proposed provides a rationale for the effect of single site mutations observed in the human erythrocyte pyruvate kinase associated with a congenital anaemia.

Allosteric Regulation↗
PubMed1996

Study of photoablation of rabbit corneas by Er:YAG laser.

BACKGROUND AND OBJECTIVE: This work studied the ablation mechanisms of rabbit corneas by the Erbium:YAG laser. The occurrence of thermal and mechanical damages in the tissue as a function of the laser fluence was also investigated. STUDY DESIGN/MATERIALS AND METHODS: The experiments were performed both on enucleated eyes and in vivo. An ultrafast imaging technique was used to investigate the dynamic evolution of the ablation. The treated samples underwent histological and ultrastructural study. RESULTS: A single high fluence laser shot led to the complete removal of the epithelium by a photomechanical effect. In eyes whose epithelium was manually removed, high fluence pulses resulted in evident tears in the stroma, whereas low fluence pulses led to few microns deep incisions, characterized by limited mechanical and thermal damages. CONCLUSION: The photomechanical action plays a significant role in the ablation of the cornea by Erbium laser. Precise control of the fluence is required to avoid cracking phenomena in the stroma.

Aluminum Silicates↗
PubMed1996

In vivo absorption spectrum of disulphonated aluminium phthalocyanine in a murine tumour model.

The absorption spectrum of aluminum phthalocyanine with an average disulphonation of 2.1 (hereafter called disulphonated aluminum phthalocyanine, A1S2Pc) was measured in vivo in a murine tumour model by means of time-resolved reflectance. Mice bearing the L1210 leukaemia were administered 2.5 or 5 mg/kg body weight (b.w.) of A1S2Pc intraperitoneally. Reflectance measurements were performed in the 650-695 nm range before and 1, 4 and 7 h after the drug administration. Fitting of the data with the diffusion theory allowed us to assess the absorption coefficient in both conditions (i.e. before and after). As a difference between the latter and the former data, the in vivo absorption spectrum of A1S2Pc was evaluated. 1 h after the administration of 2.5 mg/kg b.w. A1S2Pc, the absorption peak was centred at 685 nm, red-shifted about 15 nm with respect to the spectrum in aqueous solution. For the lower dose, the absorption line shapes 4 and 7 h after the administration remained very similar. The red shift of the absorption spectrum is consistent with the therapeutic efficacy of the photodynamic therapy which was measured at 672, 685 and 695 nm, and proved to be maximum at 685 nm for both the L1210 leukaemia and the MS-2 fibrosarcoma. With the higher drug dose, the absorption spectra taken from different animals showed significant differences. In particular, in some mice the line shape was similar to that measured with 2.5 mg/kg b.w., while in other subjects it showed a broadening or a second peak at shorter wavelengths. Measurements on some animals were performed also 18 and 24 h after the injection of 5 mg/kg b.w., leading to no time evolution or to a progressive line shape narrowing.

Animals↗
PubMed1996

Experimental test of theoretical models for time-resolved reflectance.

Four different expressions, derived from the diffusion theory or the random walk model, were used to fit time-resolved reflectance data for the evaluation of tissue optical properties. The experimental reflectance curves were obtained from phantoms of known optical parameters (absorption and transport scattering coefficients) covering the range of typical values for biological tissues between 600 and 900 nm. The measurements were performed using an instrumentation for time-correlated single-photon counting. The potential of the four methods in the assessment of the absorption and transport scattering coefficients was evaluated in terms of absolute error, linearity error, and dispersion of data. Each method showed different performances depending on the optical properties of the sample and the experimental conditions. We propose some criteria for the optimal choice of the fitting method to be used in different applications.

Biophysical Phenomena↗