Electrophysiological and metabolic effects of CHF5074 in the hippocampus: protection against in vitro ischemia.

CHF5074 is a non-steroidal anti-inflammatory derivative holding disease-modifying potential for the treatment of Alzheimer's disease. The aim of the present study was to characterize the electrophysiological and metabolic profile of CHF5074 in the hippocampus. Electrophysiological recordings show that CHF5074 inhibits in a dose-dependent manner the current-evoked repetitive firing discharge in CA1 pyramidal neurons. This result is paralleled by a dose-dependent reduction of field excitatory post-synaptic potentials with no effect on the paired-pulse ratio. The effects of CHF5074 were not mediated by AMPA or NMDA receptors, since the inward currents induced by local applications of AMPA and NMDA remained constant in the presence of this compound. We also suggest a possible activity of CHF5074 on ASIC1a receptor since ASIC1a-mediated current, evoked by application of a pH 5.5 solution, is reduced by pretreatment with this compound. Moreover, we demonstrate that CHF5074 treatment is able to counteract in hippocampal slices the OGD-induced increase in alanine, lactate and acetate levels. Finally, CHF5074 significantly reduced the apoptosis in hippocampal neurons exposed to OGD, as revealed by cleaved-caspase-3 immunoreactivity and TUNEL staining. Overall, the present work identifies novel mechanisms for CHF5074 in reducing metabolic acidosis, rendering this compound potentially useful also in conditions of brain ischemia.

Effect of betamethasone in combination with antibiotics on gram positive and gram negative bacteria.

Betamethasone is an anti-inflammatory steroid drug used in cases of anaphylactic and allergic reactions, of Alzheimer and Addison diseases and in soft tissue injuries. It modulates gene expression for anti-inflammatory activity suppressing the immune system response. This latter effect might decrease the effectiveness of immune system response against microbial infections. Corticosteroids, in fact, mask some symptoms of infection and during their use superimposed infections may occur. Thus, the use of glucocorticoids in patients with sepsis remains extremely controversial. In this study we analyzed the in vitro effect of a commercial formulation of betamethasone (Bentelan) on several Gram positive and Gram negative bacteria of clinical relevance. It was found to be an inhibitor of the growth of most of the strains examined. Also the effect of betamethasone in combination with some classes of antibiotics was evaluated. Antibiotic-steroid combination therapy is, in such cases, superior to antibiotic-alone treatment to impair bacterial growths. Such effect was essentially not at all observable on Staphylococcus aureus or Coagulase Negative Staphylococci (CoNS).

Saliva metabolomics by NMR for the evaluation of sport performance.

The paper reports preliminary results of a study in order to verify that saliva is a bio-fluid sensitive to metabolite variations due to stress and fatigue in soccer athletes, and possibly, to identify potential markers of test of performance. Saliva samples of fourteen professional soccer players were collected before and after the stressful physical activity of the level 1 Yo-Yo intermittent recovery test and, also, physiological parameters were evaluated. The NMR spectra of saliva offer a metabolites profiling which was analyzed by Principal Component Analysis as a blind test. The results of NMR pre and post test shows that it was possible to cluster the best and the worst performing athletes and that the role of the actual player may be diagnosed by a different cluster of metabolites profile. Thus saliva can be considered a biofluid metabolically sensitive to the induced physical stress and, in the future, deeper investigated to monitor the performances in athletes.

Comparison of the action of different proteases on virulence properties related to the staphylococcal surface.

AIMS: The purpose of this study was to evaluate the antimicrobial efficacy of five different proteases belonging to two different families on Staphylococcus aureus and Staphylococcus epidermidis strains. METHODS AND RESULTS: We used three serine proteases and two metalloproteases in single species biofilm formation assays and in human cell invasion processes. Following each protease incubation with bacterial cells, surface protein patterns were analysed by SDS-PAGE and zymography. Some differently expressed proteins were identified by mass spectrometry. CONCLUSIONS: The effect of tested proteases on biofilm formation was not related to the protease category but was strain-dependent and was related to the biofilm formation capacity of each staphylococcal strain. Some proteases showed a nonspecific and indiscriminate effect on surface proteins, while others induced a discrete and reproducible action on protein profiles. SIGNIFICANCE AND IMPACT OF THE STUDY: The inhibition of the surface-related virulence factors is a promising avenue to overcome persistent infections caused by bacterial biofilms. To this end, we show here that proteases, in particular the metalloprotease serratiopeptidase, can interfere with adhesion and invasion of eukaryotic cells and biofilm formation in staphylococci and their use could represent a viable treatment for the development of novel combination therapies.

Holo and apo-transferrins interfere with adherence to abiotic surfaces and with adhesion/invasion to HeLa cells in Staphylococcus spp.

Staphylococcus aureus and Staphylococcus epidermidis are the major cause of infections associated with implanted medical devices. Colonization on abiotic and biotic surfaces is often sustained by biofilm forming strains. Human natural defenses can interfere with this virulence factor. We investigated the effect of human apo-transferrin (apo-Tf, the iron-free form of transferrin, Tf) and holo-transferrin (holo-Tf, the iron-saturated form) on biofilm formation by CA-MRSA S. aureus USA300 type (ST8-IV) and S. epidermidis (a clinical isolate and ATCC 35984 strain). Furthermore S. aureus adhesion and invasion assays were performed in a eukaryotic cell line. A strong reduction in biofilm formation with both Tfs was obtained albeit at very different concentrations. In particular, the reduction in biofilm formation was higher with apo-Tf rather than obtained with holo-Tf. Furthermore, while S. aureus adhesion to eukaryotic cells was not appreciably affected, their invasion was highly inhibited in the presence of holo-Tf, and partially inhibited by the apo form. Our results suggest that Tfs could be used as antibacterial adjuvant therapy in infection sustained by staphylococci to strongly reduce their virulence related to adhesion and cellular invasion.

Bacterial biofilm formation inhibitory activity revealed for plant derived natural compounds.

Use of herbal plant remedies to treat infectious diseases is a common practice in many countries in traditional and alternative medicine. However to date there are only few antimicrobial agents derived from botanics. Based on microbiological screening tests of crude plant extracts we identified four compounds derived from Krameria, Aesculus hippocastanum and Chelidonium majus that showed a potentially interesting antimicrobial activity. In this work we present an in depth characterization of the inhibition activity of these pure compounds on the formation of biofilm of Staphylococcus aureus as well as of Staphylococcus epidermidis strains. We show that two of these compounds possess interesting potential to become active principles of new drugs.

Postexplant residual cavity hemostasis with a TachoSil patch.

Intracapsular nephrectomy as the standard explant technique for a unfunctional graft is associated with intra- and postoperative complications due mainly to hemorrhage, fluid collections, effusions from damaged tissues and the residual cavity. We have reported herein a positive experience with the use of a collagen medical sponge patch into the surgical site for hemostasis. Use of this device produced a reduction in postoperative bleeding and collections with shortened time of drainage, reduced infection risk, and earlier discharge.

Low limb and allograft rescue with iliofemoral graft for external iliac artery dissection: case report.

The technical difficulties in performing a transplantation can often be joined by an accident that demands an urgent solutions, putting the life of the patient in serious risk. In our case we were forced to perform an iliofemoral PTFE graft substitution to save the vascularization of the right lower limb. The cause was constituted by a dissection in the external iliac artery wall immediately after upstream declamping. Fortunately quick understanding of the complication made us stop the lesion downstream before it involved the femoral district. In addition the renal artery was already not declamped and the organ not perfused. Successively we anastomosed the renal artery to the PTFE graft with an end-to side anastomosis. Our first target was obviously to preserve the low limb vascularization but, with this solution we managed also to save the allograft. Actually renal function is conserved and with an optimal state of the vascular graft.

An indirect method to measure trimerization constants using surface plasmon resonance.

The ability of surface plasmon resonance to precisely measure kinetic binding constants was exploited here to indirectly evaluate the thermodynamic dissociation trimerization constant (K(d)) of a designed chimeric protein, IZN-23, derived from an isoleucine zipper and a portion of the N-terminal helix residues of HIV-1 gp41. The method uses two monoclonal antibodies (mAbs) that display different off-rates when binding the monomeric or trimeric IZN-23. A detailed description of the data analysis strategy employed to unravel the K(d) trimerization constant from the observed off-rate kinetic values is presented, and the potential exploitation of this technique in different fields is highlighted.

A biphasic daily pattern of slow wave activity during a two-day 90-minute sleep wake schedule.

Twenty-four hour sleep patterns were measured in six healthy male volunteers during a 90-minute short sleep-wake (SW 30:60) cycle protocol for 48 hours. Sleep pressure estimates (amount of Slow Wave Sleep [SWS], SWA, and Rate of Synchronization [RoS: the rate of SWA build-up at the beginning of the NREM period]) were compared with the 24-hour patterns of body temperature (Tb24) and sleep propensity. A moderate sleep debt was incurred over the 48 hour study as indicated by decreased levels of 24 hour sleep. On day 1, ultradian patterns of REM and SWS sleep were prominent; on day 2, more prominent were circadian patterns of REM sleep, SWS, Sleep Latency, TST and Tb24. Also on Day 2, biphasic patterns of SWA and RoS were expressed, with peaks occurring during the falling and rising limbs of Tb24. The biphasic peaks in SWA and RoS may be associated with phase-specific interactions of the circadian pacemaker with the sleep homeostat during conditions of moderate sleep pressure. Further research is needed to replicate the finding and to identify biological factors that may underlie the twelve hour pattern in SWA.

Is serum troponin T a useful marker of myocardial damage in newborn infants with perinatal asphyxia?

AIM: To assess the correlation of echocardiographic signs of myocardial damage to serum cardiac troponin T (cTnT) concentrations in newborn infants with perinatal asphyxia. METHODS: Electocardiograms (ECG) and echocardiograms (Echo) were obtained during the first 24 h of life from 29 asphyxiated and 30 control infants and correlated with cTnT concentrations. The echocardiographic parameters included systolic ventricular performance, preload, afterload, diastolic function, stroke volume (SV), left ventricular output (LVO), hyperechogenity of the papillary muscles and insufficiency of the atrioventricular valves. RESULTS: LVO and SV were lower but CTnT were significantly higher in asphyxiated than in control infants: 0.15 (010-0.23) vs. 0.05 (0.02-0.13), p < 0.001). Asphyxiated infants with signs of myocardial damage were associated with significantly higher cTnT than those without, 0.20 (0.11-0.28) and 0.11 (0.05-0.14 ug/L), p = 0.04. CONCLUSION: Cardiac troponin may prove to be valuable in evaluating myocardial damage in birth asphyxia. However, the degree of prematurity may complicate the assessment.

Correlated responses in body composition to divergent selection for exponential growth rate to 14 or 42 days of age in chickens.

Chicks divergently selected for 14-d (14H and 14L) or 42-d (42H and 42L) exponential growth rate (EGR) over five generations were used to determine correlated responses between growth at different ages and body composition. Regression coefficient estimates across five generations of selection were not significant for any line at either age for percentage total body water or protein. Genetic correlations between EGR from hatching to 14 d of age (EGR14) and 42-d percentage carcass fat were -0.18, -0.57, 0.63, and -0.79 among the 14H, 14L, 42H, and 42L lines, respectively. Genetic correlations between EGR from hatching to 42 d of age (EGR42) and 42-d percentage carcass fat were 0.09, -0.67,0.50, and -0.75 among the 14H, 14L, 42H, and 42L lines, respectively. During the short-term selection experiment, selection for fast EGR14 or EGR42 increased fat at the age of selection. However, selection for fast EGR42 increased body weight and percentage fat at 42 d of age (DOA), whereas selection for fast EGR14 increased body weight but not fat at 42 DOA. Therefore, it is possible to simultaneously select for high body weight at, or near, the inflection point of the growth curve without increasing fat deposition or obesity by taking advantage of the lack of a genetic correlation between EGR14 and body fat percentage at later ages.

2. the effect of feeding excess methionine on live performance, carcass traits, and ascitic mortality.

Appetite suppressant activity of dietary excess Met was investigated in a 7-wk trial for the potential to restrict feed intake and the early growth of broilers as a means of reducing the incidence of ascitic mortality. A basal starter diet meeting the minimum NRC (1994) requirements for broilers was compared with a diet providing Met above the requirement (1.86% of the diet). Excess dietary Met was fed to the broilers from 4 to 11 d. Grower and finisher diets were provided from 22 to 35 d and from 36 to 49 d. Birds in two rooms were exposed to cool temperatures to induce ascites. The temperature program for the cool rooms was as follows: 12 h at 24 C daily (0800 to 2000) and 12 h at 15 C nightly (2000 to 0800) from 14 to 49 d, whereas the other two rooms were maintained at 24 C (control rooms). Body weight gain was depressed by 20.4% when excess Met was fed from 4 to 11 d. Feed conversion was also negatively affected by the excess dietary Met. A brief recovery was achieved from 11 to 14 d upon refeeding the well-balanced basal starter diet. Full compensatory growth was achieved by 49 d when body weight was equal to that of birds fed the basal diet. Excess dietary Met did not affect the carcass traits with the exception of abdominal fat pad, which was reduced by the excess dietary Met in the control rooms, whereas the opposite was true in the cool rooms. Ascitic mortality was not altered by dietary Met, however, the total mortality was significantly reduced by excess Met in the control rooms.

1. Appetite suppressant activity of supplemental dietary amino acids and subsequent compensatory growth of broilers.

This study was conducted to take advantage of the appetite-suppressant effect of excessive dietary amino acids in reducing feed intake and, in turn, restricting the early rapid growth of broilers to minimize metabolic disorders. Dietary amino acids were supplemented to a basal diet to yield a total of 1.57, 2.57, and 3.57% His; 2.7, 4.3, and 5.9% Lys; 1.36, 2.16, and 2.96% Met; 2.8, 3.8, and 4.8% Thr; and 1.27, 2.27, and 3.27% Trp and were fed to 408 chicks from 4 to 11 d of age. Fifteen dietary treatments of His, Lys, Met, Thr, and Trp were compared to the basal diet. Feed consumption was measured daily. Body weight measurements were taken at 0, 4, 7, 11, 14, and 21 d. At 21 d, pectoralis major and minor muscles, liver, and abdominal fat pad were weighed. High levels of Met and His caused the greatest depression in appetite from 4 to 11 d, and Thr, Trp, and Lys were found to be less potent. The exponential growth rate (EGR) of birds from 4 to 11 d of age was significantly reduced by the intermediate and high levels of the amino acid supplementation. From 11 to 14 d, EGR was greatest with high levels of Met or Trp, indicating more potential compensatory growth realized with these treatments. The high level of His decreased the percentage of pectoralis minor muscle yield, whereas the high level of Lys and Met increased the percentage of liver compared to those fed the basal diet. These results indicate that it is possible to use excessive individual amino acids in diets to suppress the appetite and early rapid growth to alleviate or minimize metabolic disorders.

Measurement of homonuclear three-bond J(H(N)Halpha) coupling constants in unlabeled peptides complexed with labeled proteins: application to a decapeptide inhibitor bound to the proteinase domain of the NS3 protein of hepatitis C virus (HCV).

A new isotope-filtered experiment has been designed to measure homonuclear three-bond J(H(N)Halpha) coupling constants of unlabeled peptides complexed with labeled proteins. The new experiment is based on the 3D HNHA pulse scheme, and belongs to the 'quantitative J-correlation' type. It has been applied to a decapeptide inhibitor bound to the proteinase domain of the NS3 protein of human hepatitis C virus (HCV).

A fragment of prosaposin (SGP-1) from rooster sperm promotes sperm-egg binding and improves fertility in chickens.

A protein isolated from the supernatant of cryopreserved rooster sperm was found to increase the capability of cryopreserved rooster sperm to bind in vitro to the perivitelline membrane of a chicken egg and substantially raise fertility after artificial insemination (AI). That activity was partially purified and termed universal primary sperm-egg binding protein (UPSEBP). Insufficient protein remained from 6 x 10(11) sperm, despite retention of bioactivity, to allow sequencing. We deduced that the protein may be related to prosaposin (also termed SGP-1, for sulfated glycoprotein-1), and we used published amino acid sequences of prosaposin as a guide for synthesis of peptides. Certain peptides were found to increase in vitro sperm-egg binding and increase fertility of frozen-thawed or fresh rooster sperm, in a manner similar to semipurified UPSEBP. Active epitopes were in a 60 amino acid sequence, reflecting the intervening sequence between saposins A and B, plus short extensions into saposins A and B. Highest activity was found when this synthetic peptide was oxidized to form a disulfide bond between terminal cysteines. Antibody against a synthetic peptide consisting of 58 of these 60 amino acids bound to a 7-9 kilodalton protein in UPSEBP. Collectively, the data support the conclusion that UPSEBP is a fragment of prosaposin. Because prosaposin is in semen in humans and animals, these observations have broad implications for possible cause and therapy of one type of subfertility.

Improved sensitivity in indirect monitoring of chemical shifts of proton-heteronuclear spin pairs (1H-13C and 1H-15N) in 3D and 4D NMR spectroscopy.

In three-dimensional and four-dimensional experiments on doubly labelled proteins not only heteronuclear (13C or 15N) but also proton (1H) frequencies are often indirectly monitored, rather than being directly observed. In this communication we show how in these experiments by overlaying 1H and heteronuclear evolutions one can obtain decreased apparent relaxation rates of 1H signals, yielding improved sensitivity. The new method applies to spin pairs like 1H-15N, as in amide groups, or 1H-13C, as in methine groups of alpha or aromatic systems.