Reproductive cycle and trophic ecology in deep versus shallow populations of the Mediterranean gorgonian Eunicella singularis (Cap de Creus,northwestern Mediterranean Sea)

Coral Reefs - The annual gonad development of a shallow (20 m depth) population of the Mediterranean gorgonian Eunicella singularis was found to be closely synchronized with that of a deep...     

Loss of the preconditioning effect of rosuvastatin during sustained therapy: a human in vivo study

Studies have demonstrated that the acute administration of 3-hydroxy-3 methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors has protective effects in the setting of ischemia-reperfusion (IR). Previously, we demonstrated that a single dose of rosuvastatin prevented IR-induced endothelial dysfunction in humans through a cyclooxygenase-2-dependent mechanism. Whether the chronic administration of HMG-CoA reductase inhibitors provides similar protection remains controversial and is unknown in humans. Eighteen male volunteers were randomized to receive a single dose of rosuvastatin (20 mg) or placebo. Twenty-four hours later, endothelium-dependent, radial artery flow-mediated dilation (FMD) was measured before and after IR (15 min of upper arm ischemia followed by 15 min of reperfusion). In a separate protocol, 30 healthy volunteers were randomized in a double-blind fashion to receive oral rosuvastatin (20 mg/day) and placebo, rosuvastatin, and celecoxib (100 mg bid) or placebo alone, all for 21 days. Twenty-four hours after the final administration of study medication, FMD was measured before and after IR. Pre-IR FMD was similar between groups in both protocols. In the acute administration protocol, rosuvastatin significantly prevented the blunting of FMD associated with IR (FMD pre-IR: 8.4 ± 1.3%; post-IR: 6.2 ± 1.3%; P = 0.01 ANOVA, treatment group interaction). In the daily administration protocol, IR significantly blunted FMD in the placebo group (FMD pre-IR: 7.5 ± 0.9%; post-IR: 3.3 ± 0.7%; P < 0.001). Chronic treatment with rosuvastatin did not modify this ischemic injury (FMD pre-IR: 6.9 ± 0.4%; post-IR: 1.6 ± 1.0%; P < 0.001; P = NS ANOVA, treatment group interaction). Similarly, FMD responses post-IR in volunteers receiving rosuvastatin and celecoxib did not significantly differ from placebo (FMD pre-IR: 8.3 ± 0.9%; post-IR: 2.1 ± 0.8%; P < 0.001; P = NS ANOVA, treatment group interaction). In contrast to acute administration, chronic rosuvastatin does not prevent the development of IR-induced endothelial dysfunction in normal humans.     

Wdr5 is essential for osteoblast differentiation

Wdr5 is developmentally expressed in osteoblasts and accelerates osteoblast differentiation in vitro and in vivo. To address whether Wdr5 is essential for osteoblast differentiation, plasmid-based small interfering RNAs were used to stably suppress endogenous Wdr5 protein levels in MC3T3-E1 cells. Reduction of endogenous Wdr5 levels markedly inhibited osteoblast differentiation, evidenced by a significant decrease in alkaline phosphatase activity, Runx-2 and osteocalcin mRNAs, and absence of mineralized matrix formation. Wdr5 suppression also resulted in a reduction of histone H3 lysine 4 trimethylation, confirming its critical role in this modification. Because Wdr5 overexpression enhances canonical Wnt signaling in osteoblasts in vivo, the effects of Wdr5 silencing on this pathway were examined. The expression of the canonical Wnt target gene, c-myc, was decreased, whereas that of sfrp2, which is repressed by Wnt signaling, was increased with Wdr5 knockdown. Although only a minimal increase in apoptosis was observed, the antiapoptotic effect of Wnt signaling was also impaired with Wdr5 silencing. The expression of canonical Wnts was significantly decreased with Wdr5 knockdown, resulting in a decrease in nuclear beta-catenin protein levels. Activation of the canonical Wnt signaling pathway did not overcome the effects of Wdr5 knockdown on the expression of Wnt target genes. Chromatin immunoprecipitation demonstrated that Wdr5 is present on the Wnt1 promoter and on canonical Wnt response elements of the c-myc and Runx-2 promoters. These studies demonstrate that Wdr5 suppression interferes with the canonical Wnt signaling pathway at multiple stages and that optimal Wdr5 levels are required for induction of the osteoblast phenotype.     

Catalytic mechanisms and reaction intermediates along the hydrolytic pathway of a plant beta-D-glucan glucohydrolase.

BACKGROUND: Barley beta-D-glucan glucohydrolases represent family 3 glycoside hydrolases that catalyze the hydrolytic removal of nonreducing glucosyl residues from beta-D-glucans and beta-D-glucooligosaccharides. After hydrolysis is completed, glucose remains bound in the active site. RESULTS: When conduritol B epoxide and 2', 4'-dinitrophenyl 2-deoxy-2-fluoro-beta-D-glucopyranoside are diffused into enzyme crystals, they displace the bound glucose and form covalent glycosyl-enzyme complexes through the Odelta1 of D285, which is thereby identified as the catalytic nucleophile. A nonhydrolyzable S-glycosyl analog, 4(I), 4(III), 4(V)-S-trithiocellohexaose, also diffuses into the active site, and a S-cellobioside moiety positions itself at the -1 and +1 subsites. The glycosidic S atom of the S-cellobioside moiety forms a short contact (2.75 A) with the Oepsilon2 of E491, which is likely to be the catalytic acid/base. The glucopyranosyl residues of the S-cellobioside moiety are not distorted from the low-energy 4C(1) conformation, but the glucopyranosyl ring at the +1 subsite is rotated and translated about the linkage. CONCLUSIONS: X-ray crystallography is used to define the three key intermediates during catalysis by beta-D-glucan glucohydrolase. Before a new hydrolytic event begins, the bound product (glucose) from the previous catalytic reaction is displaced by the incoming substrate, and a new enzyme-substrate complex is formed. The second stage of the hydrolytic pathway involves glycosidic bond cleavage, which proceeds through a double-displacement reaction mechanism. The crystallographic analysis of the S-cellobioside-enzyme complex with quantum mechanical modeling suggests that the complex might mimic the oxonium intermediate rather than the enzyme-substrate complex.     

Metabolic heterogeneity of the muscle tissue: transmural distribution of glucose metabolizing enzymes across the left ventricular wall of control and hypertrophic rat heart

Mammalian cardiac muscle is a remarkably heterogeneous tissue, as judged from enzyme analysis of tissue from different myocardial layers of the left ventricle free-wall. Its diversity results from a spectrum of fibres with different metabolic properties and different location across the wall, which may be especially suited to the local range of functional demands.     

Pseudocontact shifts as constraints for energy minimization and molecular dynamics calculations on solution structures of paramagnetic metalloproteins

The pseudocontact shifts of NMR signals, which arise from the magnetic susceptibility anisotropy of paramagnetic molecules, have been used as structural constraints under the form of a pseudopotential in the SANDER module of the AMBER 4.1 molecular dynamics software package. With this procedure, restrained energy minimization (REM) and restrained molecular dynamics (RMD) calculations can be performed on structural models by using pseudocontact shifts. The structure of the cyanide adduct of the Met80Ala mutant of the yeast iso-1-cytochrome c has been used for successfully testing the calculations. For this protein, a family of structures is available, which was obtained by using NOE and pseudocontact shifts as constraints in a distance geometry program. The structures obtained by REM and RMD calculations with the inclusion of pseudocontact shifts are analyzed. Proteins 29:68–76, 1997. © 1997 Wiley-Liss, Inc.     

Occurrence and Identification of Yeast Species in Fermented Liquid Feed for Piglets

The major objective of the present study was to investigate the occurrence and identity of yeast species in fermented liquid feed (FLF) used for feeding piglets. In total, 40 different Danish farms were included in the analysis. The preparation and composition of FLF was found to be very heterogeneous with high variations in both yeast counts and yeast species composition. The yeast population varied between 6.0?×?10³ and 4.2?×?10⁷?cfug^?1 with an average yeast count of 8.7?×?10⁶?±?1.1?×?10⁷?cfug^?1. A total of 766 yeasts were isolated and identified by conventional and/or molecular typing techniques. The predominant yeast species in the FLF samples were found to be Candida milleri (58.4%), Kazachstania exigua (17.5%), Candida pararugosa (6.40%) and Kazachstania bulderi (5.09%). No clear separation between isolates of C. milleri and Candida humilis could be obtained based on sequencing of the D1/D2 region of the 26S rRNA gene. The combined use of ITS-RFLP analysis and phenotypic criteria did meanwhile suggest a closer relationship with C. milleri than C. humilis.