Lactate removal ability and graded exercise in humans

Venous lactate concentrations of nine athletes were recorded every 5 s before, during, and after graded exercise beginning at a work rate of 0 W with an increase of 50 W every 4th min. The continuous model proposed by Hughson et al. (J. Appl. Physiol. 62: 1975-1981, 1987) was well fitted with the individual blood lactate concentration vs. work rate curves obtained during exercise. Time courses of lactate concentrations during recovery were accurately described by a sum of two exponential functions. Significant direct linear relationships were found between the velocity constant (gamma 2 nu) of the slowly decreasing exponential term of the recovery curves and the times into the exercise when a lactate concentration of 2.5 mmol/l was reached. There was a significant inverse correlation between gamma 2 nu and the rate of lactate increase during the last step of the exercise. In terms of the functional meaning given to gamma 2 nu, these relationships indicate that the shift to higher work rates of the increase of the blood lactate concentration during graded exercise in fit or trained athletes, when compared with less fit or untrained ones, is associated with a higher ability to remove lactate during the recovery. The results suggest that the lactate removal ability plays an important role in the evolution pattern of blood lactate concentrations during graded exercise.     

Comparison of the glycolipid-binding specificities of cholera toxin and porcineEscherichia coli heat-labile enterotoxin: identification of a receptor-active non-ganglioside glycolipid for the heat-labile toxin in infant rabbit small intestine

The binding specificities of cholera toxin andEscherichia coli heat-labile enterotoxin were investigated by binding of¹²⁵I-labelled toxins to reference glycosphingolipids separated on thin-layer chromatograms and coated in microtitre wells. The binding of cholera toxin was restricted to the GM1 ganglioside. The heat-labile toxin showed the highest affinity for GM1 but also bound, though less strongly, to the GM2, GD2 and GD1b gangliosides and to the non-acid glycosphingolipids gangliotetraosylceramide and lactoneotetraosylceramide. The infant rabbit small intestine, a model system for diarrhoea induced by the toxins, was shown to contain two receptor-active glycosphingolipids for the heat-labile toxin, GM1 ganglioside and lactoneotetraosylceramide, whereas only the GM1 ganglioside was receptor-active for cholera toxin. Preliminary evidence was obtained, indicating that epithelial cells of human small intestine also contain lactoneotetraosylceramide and similar sequences. By computer-based molecular modelling, lactoneotetraosylceramide was docked into the active site of the heat-labile toxin, using the known crystal structure of the toxin in complex with lactose. Interactions which may explain the relatively high toxin affinity for this receptor were found.Abbreviations CT cholera toxin - CT-B B-subunits of cholera toxin - LT Escherichia coli heat-labile enterotoxin - hLT humanEscherichia coli heat-labile enterotoxin - pLT porcineEscherichia coli heat-labile enterotoxin - EI electron ionization     

Transforming growth factor-β in the early mouse embryo: Implications for the regulation of muscle formation and implantation

In a search for functions of transforming growth factor-β during early embryonic development we used two different experimental approaches. In the first we made use of embryonic stem (ES) cells. ES cells in culture differentiate to derivatives of all three germ layers and mimic some aspects of organogenesis when grown as aggregates in suspension to form embryoid bodies. Differentiation procedes further when the embryold bodies attach to suitable substrates. Muscle and neuronal cells are among the most readily identified cell types then formed. We examined the effect of all-trans retinoic acid (RA) and members of the transforming growth factor-β family(TGF-βl, TGF-β2) under these conditions in an assay where single aggregates formed in hanging microdrops in medium supplemented with serum depleted of lipophilic substances which would include retinoids. Endoderm-like cells formed under all conditions tested. RA at concentrations of 10⁸ M and 10⁷ M induced the formation of neurons but in the absence of RA or at concentrations up to 10?⁹ M, neurons were not observed. Instead, beating muscle formed in about one-third of the plated aggregates; this was greatly reduced when RA concentrations increased above 10?⁹ M. Immunofluorescent staining for muscle specific myosin showed that two muscle cell types could be distinguished: elongated, non-contractile myoblasts and mononucleate flat cells. The mononucleate flat cells appeared to correspond with rhythmically contracting muscle. The number of non-contractile myoblasts increased 3-fold over controls in the presence of 10?⁹ M RA. TGF-βs increased the number of contractile and non-contractile muscle cells by a factor 3 to 7 over controls, depending on the TGF-β isoform added and the muscle cell type formed. TGF-β2 also invariably increased the rate at which contracting muscle cells were first observed in replated aggregates. The stimulatory effect of TGF-βs on the formation of mononucleate flat cells was completely abrogated by RA at 10?⁹ M while the number of myoblasts under similar conditions was unchanged. These data suggest that a complex interplay between retinoids and TGF-β isoforms may be involved in regulation of differentiation in early myogenesis. In the second approach, neutralizing polyclonal rabbit antibodies specific for TGF-β2 were injected into the cavity of mouse blastocysts 3.5 days post coitum (pc). After 1 day in culture, embryos were transferred to pseudopregnant females. The number of decidua, embryos and resorptions were counted at day 8.5–9.5 pc. Control antibody injected embryos implanted with high efficiency (87%) compared with anti-TGF-β2 injected embryos which implanted with an efficiency of only 43%. If empty decidua (resorptions) were included, the overall recovery was 71% and 32% for control and experimental embryos, respectively. Embryos that were recovered showed no overt macroscopic abnormalities. These results together impiy functions for TGF-βs in implantation as well as in later development of the embryo. © 1993Wiley-Liss, Inc.     

Ornibactins—a new family of siderophores from Pseudomonas

Novel linear hydroxamate/hydroxycarboxylate siderophores from strains of Pseudomonas cepacia were isolated and named ornibactins. The ornibactins represent modified tetrapeptide siderophores, possessing the sequence l-Orn¹(N -OH, N -acyl)-d-threo-Asp(-OH)-l-Ser-l-Orn⁴(N -OH, N -formyl)-1,4-diaminobutane. The N -acyl groups of Orn¹(N -OH, N -acyl) may vary and represent the three acids 3-hydroxybutanoic acid, 3-hydroxyhexanoic acid and 3-hydroxyoctanoic acid, leading to a mixture of three different ornibactins, designated according to their acyl chain length as ornibactin-C4, ornibactin-C6 and ornibactin-C8. Each of the siderophores is accompanied by a small amount of a more hydrophilic component with a 16 a.m.u. higher mass. The structure elucidation was based on results from gas chromatography amino acid analysis, electrospray mass spectrometry, and one- and two-dimensional nuclear magnetic resonance techniques.     

Motor learning

Bilateral damage of the medial temporal lobe system prevents the formation of new declarative memories but leaves intact knowledge that was acquired before damage. For motor learning, no structure has been identified that plays a comparable role for the consolidation of motor memories. The deficits of motor learning are focal and show a similar allocation to the various of motor learning are focal and show a similar allocation to the various sensorimotor subsystems, as do the corresponding non-mnemonic functions. The involvement of sensorimotor circuitries changes during motor learning so that association areas are preferentially activated in the early stages, and cerebello- and striato-motor-cortical loops are preferentially activated in the late stages of motor learning. Recent neuroanatomical and neurophysiological findings on the effects of brain lesions in human and non-human primates are discussed.     

Prediction of osteoporotic fractures by postural instability and bone density.

OBJECTIVE--To investigate the utility of risk factors such as bone mineral density, lifestyle, and postural stability in the prediction of osteoporotic fractures. DESIGN--Longitudinal, epidemiological, and population based survey. SETTING--City of Dubbo, New South Wales. SUBJECTS--All residents of Dubbo aged > or = 60 on 1 January 1989. MAIN OUTCOME MEASURE--Incidence of fracture for individual subjects. RESULTS--The overall incidence of atraumatic fractures in men and women was 1.9% and 3.1% per annum respectively. The predominant sites of fracture were hip (18.9%), distal radius (18.5%), ribs and humerus (11.9% in each case), and ankle and foot (9.1% and 6.6% respectively). Major predictors of fractures in men and women were femoral neck bone mineral density, body sway, and quadriceps strength. Age, years since menopause, height, weight, and lifestyle factors were also correlated with bone mineral density and body sway and hence were indirect risk factors for fracture. Discriminant function analysis correctly identified 96% and 93% (sensitivities 88% and 81%) of men and women, respectively, who subsequently developed atraumatic fractures. Predictions based on this model indicated that a woman with a bone mineral density in the lowest quartile in the hip together with high body sway had a 8.4% probability of fracture per annum. This represented an almost 14-fold increase in risk of fracture compared with a woman in the highest bone mineral density quartile with low postural sway. An individual with all three predictors in the "highest risk" quartile had a 13.1% risk of fracture per annum. CONCLUSIONS--Bone mineral density, body sway, and muscle strength are independent and powerful synergistic predictors of fracture incidence.     

Genetic and structural analysis of a virulence determinant in polyomavirus VP1.

The LID strain of polyomavirus differs from other laboratory strains in causing a rapidly lethal infection of newborn C3H/Bi mice. This virulent behavior of LID was attenuated by dilution, yet at sublethal doses LID was able to induce tumors at a high frequency, like its parent virus PTA. By constructing and assaying LID-PTA recombinant viruses and by DNA sequencing, the determinant of virulence in LID was mapped to the major viral capsid protein, VP1. The VP1s of LID and PTA differed at two positions: at 185, LID has phenylalanine and PTA has tyrosine, and at 296, LID has alanine and PTA has valine. Results obtained with viruses constructed by site-directed mutagenesis showed that alanine at position 296 is sufficient to confer a fully virulent phenotype regardless of which amino acid is at position 185. However, with valine at position 296, an effect of phenylalanine at position 185 is apparent, as this virus possesses an intermediate level of virulence. A crystal structure of polyomavirus complexed with 3'-sialyl lactose previously indicated van der Waals contacts between the side chain of valine 296 and the sialic acid ring (T. Stehle, Y. Yan, T. L. Benjamin, and S. C. Harrison, Nature [London] 369:160-163, 1994). When this interaction was modeled with alanine, these contacts were greatly reduced. Direct confirmation that the substitutions in VP1 affected receptor binding was obtained by studying virus hemagglutination behavior. The ensemble of results are discussed in terms of the idea that a lower affinity of the virus for its receptor can result in more rapid spread and increased pathogenicity.     

Structural basis for differential receptor binding of cholera and Escherichia coli heat-labile toxins: influence of heterologous amino acid substitutions in the cholera B-subunit

The closely related B-subunits of cholera toxin (CTB) and Escherichia coli heat-labile enterotoxin (LTB) both bind strongly to GM1 ganglioside receptors but LTB can also bind to additional glycolipids and glycoproteins. A number of mutant CT B-subunits were generated by substituting CTB amino acids with those at the corresponding positions in LTB. These were used to investigate the influence of specific residues on receptor-binding specificity. A mutated CTB protein containing the first 25 residues of LTB in combination with LTB residues at positions 94 and 95, bound to the same extent as native LTB to both delipidized rabbit intestinal cell membranes, complex glycosphingolipids (polyglycosylceramides) and neolactotetraosylceramide, but not to non-GM1 intestinal glycosphingolipids. In contrast, when LTB amino acid substitutions in the 1–25 region were combined with those in the 75–83 region, a binding as strong as that of LTB to intestinal glycosphingolipids was observed. In addition, a mutant LTB with a single Gly-33→Asp substitution that completely lacked affinity for both GM1 and non-GM1 glycosphingolipids could still bind to receptors in the intestinal cell membranes and to polyglycosylceramides. We conclude that the extra, non-GM1 receptors for LTB consist of both sialylated and non-sialylated glycoconjugates, and that the binding to either class of receptors is influenced by different amino acid residues within the protein.     

Mapping and ordering of fragments of BK virus DNA produced by restriction endonucleases.

A total of 51 restriction sites were recognized within the BK virus genome by the combination of 10 different restriction endonucleases. These sites were mapped and oriented relative to one another as well as to the five fragments generated by the digestion of BK virus DNA with HindIII and EcoRI. The result was a comprehensive physical map suitable for in-depth characterization of the functions of BK virus at the molecular level.