Skeletal muscle metabolism is unaffected by DCA infusion and hyperoxia after onset of intense aerobic exercise

This study investigated whether hyperoxic breathing (100% O(2)) or increasing oxidative substrate supply [dichloroacetate (DCA) infusion] would increase oxidative phosphorylation and reduce the reliance on substrate phosphorylation at the onset of high-intensity aerobic exercise. Eight male subjects cycled at 90% maximal O(2) uptake (VO(2 max)) for 90 s in three randomized conditions: 1) normoxic breathing and saline infusion over 1 h immediately before exercise (CON), 2) normoxic breathing and saline infusion with DCA (100 mg/kg body wt), and 3) hyperoxic breathing for 20 min at rest and during exercise and saline infusion (HYP). Muscle biopsies from the vastus lateralis were sampled at rest and after 30 and 90 s of exercise. DCA infusion increased pyruvate dehydrogenase (PDH) activation above CON and HYP (3.10 +/- 0.23, 0.56 +/- 0.08, 0.69 +/- 0.05 mmol x kg wet muscle(-1) x min(-1), respectively) and significantly increased both acetyl-CoA and acetylcarnitine (11.0 +/- 0.7, 2.0 +/- 0.5, 2.2 +/- 0.5 mmol/kg dry muscle, respectively) at rest. However, DCA and HYP did not alter phosphocreatine degradation and lactate accumulation and, therefore, the reliance on substrate phosphorylation during 30 s (CON, 51.2 +/- 5.4; DCA, 56.5 +/- 7.1; HYP, 69.5 +/- 6.3 mmol ATP/kg dry muscle) and 90 s of exercise (CON, 90.6 +/- 9.5; DCA, 107.2 +/- 13.0; HYP, 101.2 +/- 15.2 mmol ATP/kg dry muscle). These data suggest that the rate of oxidative phosphorylation at the onset of exercise at 90% VO(2 max) is not limited by oxygen availability to the active muscle or by substrate availability (metabolic inertia) at the level of PDH in aerobically trained subjects.     

Mollusc larval shell formation: amorphous calcium carbonate is a precursor phase for aragonite

The larval shells of the marine bivalves Mercenaria mercenaria and Crassostrea gigas are investigated by polarized light microscopy, infrared spectroscopy, Raman imaging spectroscopy, and scanning electron microscopy. Both species contain similar shell ultrastructures. We show that larval shells contain amorphous calcium carbonate (ACC), in addition to aragonite. The aragonite is much less crystalline than non-biogenic aragonite. We further show that the initially deposited mineral phase is predominantly ACC that subsequently partially transforms into aragonite. The postset juvenile shell, as well as the adult shell of Mercenaria also contains aragonite that is less crystalline than non-biogenic aragonite. We conclude that ACC fulfills an important function in mollusc larval shell formation. It is conceivable that ACC may also be involved in adult shell formation.     

Projection structure at 8 A resolution of the melibiose permease,an Na-sugar co-transporter from Escherichia coli

The ion-coupled sugar membrane symporter or co-transporter melibiose permease (MelB), responsible for alpha-galactoside accumulation in Escherichia coli, is a representative member of the glycoside-pentoside- hexuronide family of the vast class of electrochemical potential-driven porters. Pure solubilized preparations of a MelB recombinant protein were subjected to two-dimensional crystallization trials and several crystal forms were observed. Two of these appeared as large wide tubes suitable for analysis by electron crystallography. Flattened tubes on carbon support film, embedded in amorphous ice prior to electron cryomicroscopy, showed two-sided plane group symmetries P12(1) or P222(1), with unit cell dimensions a = 89.9 A, b = 51.6 A, gamma = 91.9 degrees and a = 188.9 A, b = 48.8 A, gamma = 90 degrees, respectively. The projection map from the P222(1 )crystals at 8 A resolution displayed an asymmetric protein unit consisting of two domains lining a central and curve-shaped cleft. Together, the MelB monomer could host the 12 predicted transmembrane alpha-helices. Overall, the MelB helix packing arrangement compared more favorably with that of the Na(+)/H(+) antiporter NhaA than that of the oxalate antiporter.     

New and bioactive compounds from Streptomyces strains residing in the wood of Celastraceae

Wood from three different plants of the Celastraceae growing in their natural habitats in Brazil (Maytenus aquifolia Mart.) and South Africa [Putterlickia retrospinosa van Wyk and Mostert, P. verrucosa (E. Meyer ex Sonder) Szyszyl.] was established as a source of endophytic bacteria using a medium selective for actinomycetes. Two isolates were identified as Streptomyces setonii and S. sampsonii whereas two others were not assignable to any of the known Streptomyces species. They were preliminarily named Streptomyces Q21 and Streptomyces MaB-QuH-8. The latter strain produces a new chloropyrrol and chlorinated anthracyclinone. The chloropyrrol showed high activity against a series of multiresistent bacteria and mycobacteria.     

Indole alkaloids from Rauvolfia bahiensis A.DC. (Apocynaceae)

Four indole alkaloids, 12-methoxy-N(a)-methyl-vellosimine, demethoxypurpeline, 12-methoxyaffinisine, and 12-methoxy-vellosimine, in addition to picrinine, vinorine, raucaffrinoline, normacusine B, norseredamine, seredamine, 10-methoxynormacusine B, norpurpeline and purpeline, were isolated from the bark or leaf extracts of Rauvolfia bahiensis.     

Fine mapping of a quantitative trait locus for twinning rate using combined linkage and linkage disequilibrium mapping

A novel and robust method for the fine-scale mapping of genes affecting complex traits, which combines linkage and linkage-disequilibrium information, is proposed. Linkage information refers to recombinations within the marker-genotyped generations and linkage disequilibrium to historical recombinations before genotyping started. The identity-by-descent (IBD) probabilities at the quantitative trait locus (QTL) between first generation haplotypes were obtained from the similarity of the marker alleles surrounding the QTL, whereas IBD probabilities at the QTL between later generation haplotypes were obtained by using the markers to trace the inheritance of the QTL. The variance explained by the QTL is estimated by residual maximum likelihood using the correlation structure defined by the IBD probabilities. Unlinked background genes were accounted for by fitting a polygenic variance component. The method was used to fine map a QTL for twinning rate in cattle, previously mapped on chromosome 5 by linkage analysis. The data consisted of large half-sib families, but the method could also handle more complex pedigrees. The likelihood of the putative QTL was very small along most of the chromosome, except for a sharp likelihood peak in the ninth marker bracket, which positioned the QTL within a region <1 cM in the middle part of bovine chromosome 5. The method was expected to be robust against multiple genes affecting the trait, multiple mutations at the QTL, and relatively low marker density.     

Zinc-mediated dimerization and its effect on activity and conformation of staphylococcal enterotoxin type C

Staphylococcal enterotoxins are superantigen exotoxins that mediate food poisoning and toxic shock syndrome in humans. Despite their structural and functional similarities, superantigens display subtle differences in biological properties and modes of receptor binding as a result of zinc atoms bound differently in their crystal structures. For example, the crystal structures of the staphylococcal enterotoxins in the type C serogroup (SECs) contain a zinc atom coordinated by one aspartate and two histidine residues from one molecule and another aspartate residue from the next molecule, thus forming a dimer. This type of zinc ligation and zinc-mediated dimerization occurs in several SECs, but not in most other staphylococcal enterotoxin serogroups. This prompted us to investigate the potential importance of zinc in SEC-mediated pathogenesis. Site-directed mutagenesis was used to replace SEC zinc binding ligands with alanine. SEC mutants unable to bind zinc did not have major conformational alterations although they failed to form dimers. Zinc binding was not essential for T cell stimulation, emesis, or lethality although in general the mutants were less pyrogenic. Thus the zinc atom in SECs might represent a non-functional heavy atom in an exotoxin group that has diverged from related bacterial toxins containing crucial zinc atoms.     

Patterns of infection with the nematodes Syphacia obvelata and Aspiculuris tetraptera in conventionally maintained laboratory mice

Data on the frequency, distribution and mean intensity of the helminth fauna recovered from outbred and inbred mice conventionally maintained in Brazilian animal houses, are reported. The oxyurid nematodes Syphacia obvelata and Aspiculuris tetraptera presented overall frequencies of 91.5% and 8.5%, respectively. The frequency of S. obvelata in animals of three groups out of the four investigated ranged from 9% to 74% and A. tetraptera from 17% to 83%, since animals of one of the groups were negative for helminths. Infections due to a single species were observed in 62% of the animals, compared to 16% related to associations. The frequency of single infections in each group varied from 58.6% to 100% whereas associations varied from 24.1% to 41.4%. The analysis of specific mean intensities showed that S. obvelata was represented by 13.35 to 66.58 specimens/host and A. tetraptera by 5.85 to 16.75 specimens/host.     

The effect of TGF-beta receptor binding peptides on smooth muscle cells

TGF-beta1 is a potent regulator of vascular smooth muscle cell (VSMC) proliferation, migration, and extracellular matrix (ECM) synthesis. In this study, we selected two peptides, IM-1 and IM-2, that bind to the TGF-beta type II receptor (TGF-beta RII) using phage display. IM-1 and IM-2 bind to the TGF-beta RII, with a K(d) of 1 microM. Like TGF-beta, IM-1 induced VSMC chemotaxis and PAI-1 mRNA expression, as determined using Boyden chambers and real time quantitative PCR. In contrast, IM-2 had no effect on VSMC chemotaxis or PAI-1 induction. Induction of ECM synthesis, involving proteins such as osteopontin and alpha-smooth muscle actin, was determined by ELISA. Osteopontin expression was inhibited by both peptides, but TGF-beta-induced alpha-smooth muscle actin expression could only be inhibited by IM-1. In conclusion, IM-1 activity on VSMC is agonistic with TGF-beta, except for ECM synthesis, whereas the IM-2 peptide is antagonistic for some examined TGF-beta functions.     

MDR1 in taste buds of rat vallate papilla: functional, immunohistochemical, and biochemical evidence

Multidrug resistance P-glycoprotein (MDR1) is a membrane proteinof 150-170 kDa that catalyzes the ATP-driven efflux of hydrophobic xenobiotics, including fluorescent dyes, from cells. Expressed in manyepithelial tissues and in the endothelia of the blood-brain barrier,the MDR1 protein provides major routes of detoxification. We found thattaste cells of the rat vallate papilla (VP; posterior tongue) had onlya slow increase in fluorescence due to uptake of the hydrophobic dyecalcein acetoxymethyl ester. However, the development of fluorescencewas accelerated two- to threefold by substrates and/orinhibitors of MDR1, such as verapamil, tamoxifen, and cyclosporin A,and by addition of the transport-blocking antibody to MDR1, UIC2.Western blots of vallate tissue rich in taste buds withthe MDR1-specific monoclonal antibodies C219 and C494 revealed animmunoreactive protein at ~170 kDa. In contrast, the lingual epithelium surrounding the VP showed a much weaker band with these antibodies. Furthermore, using the antibodies C494 and UIC2 with tissuesections, MDR1-like immunoreactivity was found in taste cells. Theseresults show that MDR1 is present and functional in vallate taste cellsof the rat. MDR1-related transport may achieve active elimination ofxenobiotics from the sensory cells and thereby protect the peripheraltaste organs from potentially harmful molecules contained in ananimal's food.