Chemotactic tripeptides incorporating at position 2 α-aminoacid residues with unsaturated side chains

New N-For-Met-Leu-Phe-OMe (fMLF-OMe) analogues incorporating three different gamma-delta-didehydro-alpha-aminoacid residues (namely: Alg = (S)-Allylglycine; Dag = Diallylglycine; Cpg = 1-Aminocyclopent-3-ene-1-carboxylic acid) replacing the native (S)-Leucine have been synthesized and their activity towards human neutrophils has been evaluated in comparison with that shown by the reference tripeptide fMLF-OMe. Chemotaxis, lysozyme release and superoxide anion production have been measured. (1)H NMR titration experiments and NOESY spectrum of the Cpg containing model 10 have been discussed in order to ascertain the preferred solution conformations. A fully extended (C(5)) conformation at position 2 and a folded conformation with two consecutive gamma-turns (C(7) structure) have been proposed for the Dag and Cpg containing tripeptides, respectively.     

Gamete cryobanks for laboratory research: Developing a rapid and easy-to-perform protocol for the cryopreservation of the sea urchin Paracentrotus lividus (Lmk, 1816) spermatozoa

Gamete cryopreservation is a biotechnology that can guarantee a continuous supply of gametes, regardless of the seasonal reproductive cycle. In this study we developed a protocol for the cryopreservation of the sea urchin Paracentrotuslividus spermatozoa, with a view to the creation of cryobanks of semen to be used as a model system in laboratory research and ecotoxicological tests. All the key phases of the procedure were separately considered and the effect on sperm motility was evaluated by means of computer assisted analysis. The best results were obtained using 7% dimethylsulfoxide in 1% NaCl plus 0.04 M trehalose as the extender, at a freezing rate of −20 °C/min. On thawing, in semen samples cryopreserved in accordance with this protocol the velocity parameters of the sub-population of rapid sperm (best performing spermatozoa) did not significantly differ from semen on collection; in addition also the fertilization ability was restored, and about 50% of normal developed plutei larvae were obtained by thawed semen. The developed protocol is rapid and easy-to-perform; moreover, the use of gametes from reared urchins makes it unnecessary to continuously collect specimens from natural populations, making this procedure a promising starting point for the creation of alternative and more sustainable methodologies in laboratory research on sea urchin gametes and embryos.     

Training load,recovery and game performance in semiprofessional male basketball: influence of individual characteristics and contextual factors

This study examined the effects of individual characteristics and contextual factors on training load, pre-game recovery and game performance in adult male semi-professional basketball. Fourteen players were monitored, across a whole competitive season, with the session-RPE method to calculate weekly training load, and the Total Quality Recovery Scale to obtain pre-game recovery scores. Additionally, game-related statistics were gathered during official games to calculate the Performance Index Rating (PIR). Individual characteristics and contextual factors were grouped using k-means cluster analyses. Separate mixed linear models for repeated measures were performed to evaluate the single and combined (interaction) effects of individual characteristics (playing experience; playing position; playing time) and contextual factors (season phase; recovery cycle; previous game outcome; previous and upcoming opponent level) on weekly training load, pre-game recovery and PIR. Weekly load was higher in guards and medium minute-per-game (MPG) players, and lower for medium-experienced players, before facing high-level opponents, during later season phases and short recovery cycles (all p < 0.05). Pre-game recovery was lower in centers and high-experience players (p < 0.05). Game performance was better in high-MPG players (p < 0.05) and when facing low and medium-level opponents (p < 0.001). Interestingly, players performed better in games when the previous week’s training load was low (p = 0.042). This study suggests that several individual characteristics and contextual factors need to be considered when monitoring training load (playing experience, playing position, playing time, recovery cycle, upcoming opponent level), recovery (playing experience, playing position) and game performance (opponent level, weekly training load, pre-game recovery) in basketball players during the competitive season.     

Synthesis and bioactivity of chemotactic tetrapeptides: fMLF-OMe analogues incorporating spacer aminoacids at the lateral positions

A small library of N-For and N-Boc tetrapeptidic analogues of the chemotactic tripeptide For-Met-Leu-Phe-OMe (fMLF-OMe), obtained by incorporating three different spacer aminoacids (Gly, βAla and Pro) between the native residues of Met and Leu (N-For- and N-Boc-Met-Xaa-Leu-Phe-OMe; Xaa² series) and Leu and Phe (N-For- and N-Boc-Met-Leu-Xaa-Phe-OMe; Xaa³ series), have been synthesized and examined for their biological activity as agonists and antagonists. Chemotaxis, lysozyme release and superoxide anion production have been measured. All the N-For analogues maintain good to moderate chemotactic activity with the βAla³ 15 model reaching the maximum value. All the N-Boc tetrapeptides are efficient chemotactic antagonists. Conversely, with the exception of the moderate antagonistic activity exhibited by the N-Boc Xaa² models against lysozyme release, all the other N-Boc analogues do not show significant activity against both superoxide anion and lysozyme release.     

Pyrazole[3,4-e][1,4]thiazepin-7-one derivatives as a novel class of Farnesoid X Receptor (FXR) agonists

A virtual screening procedure was applied to the discovery of structurally diverse non-steroidal Farnesoid X Receptor (FXR) agonists. From 117 compounds selected by virtual screening, a total of 47 compounds were found to be FXR agonists, with 34 of them showing activity below a concentration of 20 μM. 1H-Pyrazole[3,4-e][1,4]thiazepin-7-one-based hit compound 7 was chosen for hit-to-lead optimization. A large number of 1H-pyrazole[3,4-e][1,4]thiazepin-7-one derivatives was designed, synthesized, and evaluated by a cell-based luciferase transactivation assay for their agonistic activity against FXR. Most of them exhibited low micromolar range of potency and very high efficacy.     

An Anionic Porphyrin Binds <Emphasis Type="Italic">β</Emphasis>-Lactoglobulin A at a Superficial Site Rich in Lysine Residues

Binding of small ligands to globular proteins remains a major research topic in biophysics. We have studied the binding of several photoactive dyes to β-lactoglobulin (BLG), as a model to investigate the photoinduced effects of porphyrins on proteins. A combination of optical spectroscopies (fluorescence, circular dichroism) and molecular docking simulations were used to estimate the pH-dependence of the binding parameters and the docking location for meso-tetrakis(sulfonatophenyl)-porphyrin (TPPS). We have observed that the binding of TPPS is not modulated by the pH-mediated conformational transition of the protein (i.e., Tanford transition). Binding of TPPS appears to occur with some degree of negative cooperativity. Moreover, TPPS remains bound even upon partial denaturation of the protein. These results are consistent with a superficial binding site at a location removed from the aperture of the interior β-barrel. Binding occurs through electrostatic interactions between the negative SO₃ ⁻ groups of TPPS and positively charged Lys and Arg residues. This is the first study that explores the interaction of an anionic porphyrin with BLGA in a pH range that spans across the Tanford transition. Establishing the location of the binding site will enable us to explain the photoinduced conformational effects mediated by TPPS on BLG. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

A long-term genetic model for the evolution of sexual preference: the theories of Fisher and Zahavi re-examined

 Long-term co-evolution of male's sexual extravagance and female's preference for it is studied. Fisher's ``Sexy Son' principle is checked against Zahavi's Handicap Principle. It is shown that although both principles are equally likely to explain this sort of co-evolution in the short run, only the second one allows for a long-term evolutionarily stable females' preference for costly male's extravagance. It is shown, however, that Fisher's argument, although not sufficient on its own to explain long-term persistence of females' choice, may tacitly appear as an indispensable component for the application of Zahavi's theory to the important case of dense polygenous populations. Received: 7 March 2001 / Revised version: 15 November 2001 / Published online: 26 June 2002     

Effects of extender composition, cooling rate and freezing on the fertilisation viability of spermatozoa of the Pacific oyster (Crassostrea gigas)

The aim of this research was to optimise protocols for freezing spermatozoa of the Pacific oyster. All the phases of the cryopreservation procedure (choice of cryoprotectant, cooling, freezing, and thawing) were studied in relation to the species of spermatozoa to restore on thawing the morphological and physiological characteristics of fresh semen. The choice of type and concentration of cryoprotectant in which semen is incubated before freezing is fundamental for a successful cryopreservation: the cryoprotectants (dimethylsulfoxide--Me(2)SO, ethylene glycol--EG, propylene glycol-PG, and glycerol in concentrations between 5 and 15%) were tested for their toxicity on the semen exposed up to 30 min at +26 degrees C (room temperature) by evaluating its ability to fertilise and the embryo development to the regular D larval stage. The best cryoprotectants, Me(2)SO, EG, and PG 5, 10, and 15% respectively, were used for the pre-cooling (adaptation/cooling) tests. Two different adaptation/cooling procedures were tested: (A) from +26 degrees C to 0-2 degrees C (2.6 degrees C/min) and (B) at +26 degrees C for 15 min. Lastly, using the cryoprotectants and the adaptation procedure (B) that had given the best results in the preceding stages of the experiment, four cooling rates were tested: 6, 11, 16, and 21 degrees C/min. It was seen that the semen that was incubated with EG 10%, adapted at +26 degrees C for 15 min, and then cooled at a rate of 6 degrees C/min showed a percentage of regular D larvae on thawing comparable to that of fresh semen (p > 0.05).     

Storage of buffalo (Bubalus bubalis) semen

Characteristics of buffalo semen, diluents used for liquid storage, aspects involved in freezing and thawing of semen are reviewed, and fertility results after artificial insemination (AI) with frozen-thawed semen are given.