[Intervention of psychological and ethical professionals of human science in obstetrical morbidity and mortality conferences]. / L'intervention des sciences humaines en psychologie et en éthique dans les revues de morbi-mortalité en obstétrique.

OBJECTIVE: To identify the defence mechanisms manifested by medical staff which could disturb the decision making, revealed by professionals of human science (PHS) in morbidity and mortality conferences (MMC). MATERIALS AND METHODS: Application of two methods of psychological intervention in MMC, conducted between March 1st, 2009 and November 30, 2010, in 20 randomized maternity among five perinatal networks: the method of inter-active problem solving targeted at the functioning of the teams and the method for developing professional practice centred on individual. The data collection was realized during analyse of case in MMC, with note-taking by two pair PHS. The oral expressions of RMM' participant were secondarily re-written, analyzed and classed by theme. RESULTS: Fifty-four MMC were performed. The mechanisms of defence have been identified by PHS intervention in MMC: denial of situation, pact of denegation, rift and overprotection. They were be identified by two PHS intervention methods, this consolidates these results. This intervention began staff medical to transformation at different level, in particular to improve the capacity of cooperation. CONCLUSION: The identification of the mechanisms of defence in MMC enables staff medical to improve communication and quality relationship between healthcare professionals. This could constitute an actual factor of practices improvement. However, complementary studies must be performed to confirm this hypothesis.

[Spermiogenesis: histone acetylation triggers male genome reprogramming]. / Spermiogenèse: l'acétylation des histones déclenche la reprogrammation du génome mâle.

During their post-meiotic maturation, male germ cells undergo an extensive reorganization of their genome, during which histones become globally hyperacetylated, are then removed and progressively replaced by transition proteins and finally by protamines. The latter are known to tightly associate with DNA in the mature sperm cell. Although this is a highly conserved and fundamental biological process, which is a necessary prerequisite for the transmission of the male genome to the next generation, its molecular basis remains mostly unknown. We have identified several key factors involved in this process, and their detailed functional study has enabled us to propose the first model describing molecular mechanisms involved in post-meiotic male genome reprogramming. One of them, Bromodomain Testis Specific (BRDT), has been the focus of particular attention since it possesses the unique ability to specifically induce a dramatic compaction of acetylated chromatin. Interestingly, a mutation was found homozygous in infertile men which, according to our structural and functional studies, disrupts the function of the protein. A combination of molecular structural and genetic approaches has led to a comprehensive understanding of new major actors involved in the male genome reprogramming and transmission.

Use of nucleic acid testing for blood donor screening of human immunodeficiency virus and hepatitis C virus in the Saudi population.

OBJECTIVE: To determine the risk of transfusion associated infection for human immunodeficiency virus and Hepatitis C virus using nucleic acid testing. METHODS: During March 1998, 400 donor blood samples from the Saudi population that were negative by serology were further tested for human immunodeficiency virus 1 and 2 and Hepatitis C virus using nucleic acid testing. RESULTS: A total of 400 were tested by nucleic acid testing, 381 of these were negative, 4 were indeterminate but were found to be negative on repeat testing and one seronegative sample was found to be positive for Hepatitis C virus. CONCLUSION: Due to the low prevalence of human immuno-deficiency virus in the Kingdom of Saudi Arabia, nucleic acid testing of blood donors by serology is adequate for screening. But the higher prevalence of Hepatitis C virus and increased risk of transmission would indicate that nucleic acid testing may be warranted for Hepatitis C virus in the near future.

High resolution x-ray analysis of two mutants of a curaremimetic snake toxin.

A previous mutational analysis of erabutoxin a (Ea), a curaremimetic toxin from sea snake venom, showed that the substitutions S8G and S8T caused, respectively, 176-fold and 780-fold affinity decreases for the nicotinic acetylcholine receptor (AchR). In view of the fact that the side-chain of Ser8 is buried in the wild-type toxin, we wondered whether these affinity changes reflect a direct binding contribution of S8 to the receptor and/or conformational changes that could have occurred in Ea as a result of the introduced mutations. To approach this question, we solved X-ray structures of the two mutants S8G and S8T at high resolution (0.18 nm and 0.17 nm, with R factors of 18.0% and 17.9%, respectively). The data show that none of the mutations significantly modified the toxin structure. Even within the site where the toxin binds to the receptor the backbone conformation remained unchanged. Therefore, the low affinities of the mutants S8T and S8G cannot be explained by a large conformational change of the toxin structure. Although we cannot exclude the possibility that undetectable structural changes have occurred in the toxin mutants, our data support the view that, although buried between loop I and II, S8 is part of the functional epitope of the toxin.

Crystal structures of alpha-mercaptoacyldipeptides in the thermolysin active site: structural parameters for a Zn monodentation or bidentation in metalloendopeptidases.

Three alpha-mercaptoacyldipeptides differing essentially in the size of their C-terminal residues have been crystallized in the thermolysin active site. A new mode of binding was observed for 3 [HS-CH(CH(2)Ph)CO-Phe-Tyr] and 4 [HS-CH((CH(2))(4)CH(3))CO-Phe-Ala], in which the mercaptoacyl moieties act as bidentates with Zn-S and Zn-O distances of 2.3 and 2.4 A, respectively, the side chains fitting the S(1), S(1)', and S(2)' pockets. Moreover, a distance of 3.1 A between the sulfur atom and the OE1 of Glu(143) suggests that they are H-bonded and that one of these atoms is protonated. This H-bond network involving Glu(143), the mercaptoacyl group of the inhibitor, and the Zn ion could be considered a "modified" transition state mimic of the peptide bond hydrolysis. Due to the presence of the hindering (5-phenyl)proline, the inhibitor HS-CH(CH(2)Ph)CO-Gly-(5-Ph)Pro (2) interacts through the usual Zn monodentation via the thiol group and occupancy of S(1)' and S(2)' subsites by the aromatic moieties, the proline ring being outside the active site. The inhibitory potencies are consistent with these structural data, with higher affinities for 3 (4.2 x 10(-)(8) M) and 4 (4.8 x 10(-)(8) M) than for 2 (1.2 x 10(-)(6) M). The extension of the results, obtained with thermolysin being considered as the model of physiological zinc metallopeptidases, allows inhibitor-recognition modes for other peptidases, such as angiotensin converting enzyme and neutral endopeptidase, to be proposed and opens interesting possibilities for the design of new classes of inhibitors.

Structures of escherichia coli CMP kinase alone and in complex with CDP: a new fold of the nucleoside monophosphate binding domain and insights into cytosine nucleotide specificity.

BACKGROUND: . Nucleoside monophosphate kinases (NMP kinases) catalyze the reversible transfer of a phosphoryl group from a nucleoside triphosphate to a nucleoside monophosphate. Among them, cytidine monophosphate kinase from Escherichia coli has a striking particularity: it is specific for CMP, whereas in eukaryotes a unique UMP/CMP kinase phosphorylates both CMP and UMP with similar efficiency. RESULTS: . The crystal structure of the CMP kinase apoenzyme from E. coli was solved by single isomorphous replacement and refined at 1.75 A resolution. The structure of the enzyme in complex with CDP was determined at 2.0 A resolution. Like other NMP kinases, the protein contains a central parallel beta sheet, the strands of which are connected by alpha helices. The enzyme differs from other NMP kinases in the presence of a 40-residue insert situated in the NMP-binding (NMPbind) domain. This insert contains two domains: one comprising a three-stranded antiparallel beta sheet, the other comprising two alpha helices. CONCLUSIONS: . Two features of the CMP kinase from E. coli have no equivalent in other NMP kinases of known structure. Firstly, the large NMPbind insert undergoes a CDP-induced rearrangement: its beta-sheet domain moves away from the substrate, whereas its helical domain comes closer to it in a motion likely to improve the protection of the active site. Secondly, residues involved in CDP recognition are conserved in CMP kinases and have no counterpart in other NMP kinases. The structures presented here are the first of a new family of NMP kinases specific for CMP.

Solubility diagram of the Rhodobacter sphaeroides reaction center as a function of PEG concentration.

In order to quantify the effect of polyethylene glycol 4000 (PEG) on the solubility of an integral membrane protein, we have crystallized the photochemical reaction center from Rhodobacter sphaeroides Y by batch method on a large range of PEG. The measurement of the solubility diagram display a semi-logarithmic dependence of solubility versus PEG concentration. Comparison of our results with previously published ones [Odahara, T., Ataka, M. and Katsura, M. (1994) Acta Cryst. D50, 639-642] suggests a notable effect of additional 1,2,3-heptane-triol and/or temperature on photochemical reaction center solubility.

Structure of the photochemical reaction centre of a spheroidene-containing purple-bacterium, Rhodobacter sphaeroides Y, at 3 A resolution.

The crystal structure of the photochemical reaction centre from Rhodobacter sphaeroides Y, a carotenoid-containing wild-type purple bacterium, has been determined at 3 A resolution. This membrane complex consists of three subunits (281, 307 and 260 residues, respectively) and ten cofactors. It was crystallized in presence of beta-D-octylglucoside. The crystals are orthorhombic with unit-cell dimensions, a = 143.7, b = 139.8, c = 78.7 A, space group P2(1)2(1)2(1) with four molecules in the unit cell. Refinement of the structure by X-PLOR and manual reconstructions yielded an R value of 22.1% for 19630 reflections between 7 and 3 A. The secondary structure is highly homologous to those determined for Rhodopseudomonas viridis (Protein Data Bank entry 1PRC) and Rhodobacter sphaeroides R26 (Protein Data Bank entry 4RCR) reaction centres. In the latter two structures one Fe(2+) ion located between the two quinones is coordinated by four histidines and one glutamic acid. In the Rhodobacter sphaeroides Y structure, Mn(2+) occupies the same position with identical ligands and geometry. The carotenoid conformation which is a non-planar 15-15'-cis spheroidene molecule in our structure differs from the 13-14-cis 2,4-dihydroneusporene in the Rhodopseudomonas viridis structure.

Response to upright exercise after cardiac transplantation.

There is little information on the hemodynamic response to upright exercise in patients who have undergone cardiac transplantation. We compared the hemodynamic and metabolic response to upright bicycle exercise in 11 patients with heart transplants and 12 controls. Patients performed two tests--a steady-state test with a right heart catheter and a maximal incremental test. During steady-state exercise at 20% of their predicted maximum workload, patients with heart transplants had a higher (mean +/- SD, p < 0.05) heart rate (108 +/- 11 vs. 96 +/- 15 beats/min), mean systemic blood pressure (116 +/- 17 vs. 101 +/- 11 mmHg), mean pulmonary artery pressure (29 +/- 9 vs. 22 +/- 3 mmHg), mean pulmonary wedge pressure (14 +/- 6 vs. 9 +/- 2), pulmonary (302 +/- 101 vs. 220 +/- 50 d-sec-cm-5-m2) and systemic (2049 +/- 531 vs. 1459 +/- 520) resistance indices, and lactate concentration (3.4 +/- 1.7 vs. 1.7 +/- 0.4 mmol/l), and a lower stroke index (39 +/- 8 vs. 50 +/- 8 ml/m2) compared with controls. Cardiac index, right atrial pressure, and mixed venous oxygen saturation were similar. During the maximal exercise test, patients with heart transplants achieved a significantly lower percentage of predicted maximum heart rate (77 +/- 13 vs. 91 +/- 8%), workload (70 +/- 25 vs. 102 +/- 23%), oxygen consumption (63 +/- 11 vs. 108 +/- 19%), and ventilation (67 +/- 18 vs. 89 +/- 15%) compared with controls. Heart transplant patients also had a lower blood pressure and anaerobic threshold. We conclude that heart transplant patients have an altered hemodynamic and metabolic response to upright bicycle exercise.

Flow cytometry and sorting of amphibian bladder endocytic vesicles containing ADH-sensitive water channels.

The water permeability of ADH target epithelial cells is believed to be regulated by a cycle of exo-endocytosis of vesicles containing functional water channels. These vesicles were selectively labeled in intact frog urinary bladders with an impermeant fluorescent marker, 6-carboxyfluorescein. Vesicle suspensions containing the labeled endosomes were obtained by homogenization and differential centrifugation of bladder epithelial cells. The osmotic permeability of the endocytic vesicles was measured, using a stopped-flow fluorescence technique, in the absence or in the presence of HgCl2. This permeability was found very high (500 microns/sec) and inhibited by 1 mM HgCl2 (90%), thus confirming the presence of water channels. The labeled endosomes were then separated from the other membrane vesicles by flow cytometry and sorting. Their protein content was analyzed by electrophoresis on ultrathin polyacrylamide gels. Two double bands were found at 71 and 55 kDa as well as a small band at 43 kDa. They respectively correspond to 31, 38 and 10% of the total amount of silver-stained proteins present in the sorted endosomes, while they only represent 2, 4, and less than 1% of the proteins contained in the vesicle suspension, before sorting. These highly enriched proteins (or at least one of them) are likely to be involved in the mechanism of water transport. Associated to their partial purification by differential centrifugation, the sorting of the endosomes by flow cytometry seems a good way to further characterize the water channel.

Fluorescence response and sensitivity determination for ATC 3000 flow cytometer.

The fluorescence emitted by labeled particles after interaction with exciting light is conditioned by laser beam geometry and by the mode of fluorescence collection and filtration. A laser elliptic focusing mode is described, and the fluorescence characteristics of the sample cell flow are calculated. Fluorescence collection and detection through optical filters were analyzed, and efficiency was calculated for the ATC 3000 flow cytometer (Odam-Bruker, Wissembourg, France). A mathematical model is proposed for calculation of the fluorescence signal and its fluctuations. The background noise for the ATC 3000 was quantified experimentally using fluorescent microspheres of a known number of bound equivalent fluorescein isothiocyanate (FITC) molecules. These experimental measurements were found to fit the theoretical predictions, thus validating the proposed model.