Interleukin-6 release from human skeletal muscle during exercise: relation to AMPK activity.

We tested the hypothesis that IL-6 release from muscle during exercise may be related to muscle activity of 5'-AMP-activated protein kinase (AMPK). Eight healthy, well-trained young men completed two 60-min trials on a bicycle ergometer at 70% of their peak oxygen uptake in either a glycogen-depleted or a glycogen-loaded state. IL-6 was released from the leg already after 10 min of exercise in the glycogen-depleted state, whereas no significant release was observed at any time in the loaded state. Nevertheless, plasma IL-6 increased similarly in the two trials from approximately 0.8 pg/ml at rest to approximately 4.5 pg/ml after 60 min of exercise. Activity of alpha1-AMPK (160%) and alpha2-AMPK (145%) was increased at rest in the glycogen-depleted compared with the loaded situation. During exercise, alpha1-AMPK activity did not change from resting levels in both trials, whereas alpha2-AMPK activity increased only in the glycogen-depleted state. After 60 min of exercise in the glycogen-depleted state, individual values of alpha2-AMPK activity correlated significantly (r = 0.87, P < 0.006) with individual values of IL-6 release as well as with average IL-6 release over the entire 60 min (r = 0.86, P < 0.006). The present data are compatible with a role for AMPK in IL-6 release during exercise or a role for IL-6 in activating AMPK. Alternatively, both AMPK and IL-6 are independent sensors of a low muscle glycogen concentration during exercise. In addition, leg release of IL-6 cannot alone explain the increase in plasma IL-6 during exercise.     

Effects of glucose, glucose plus branched-chain amino acids, or placebo on bike performance over 100km

Madsen, Klavs, Dave A. MacLean, Bente Kiens, and DirkChristensen. Effects of glucose, glucose plus branched-chain aminoacids, or placebo on bike performance over 100 km. J. Appl. Physiol. 81(6): 2644-2650, 1996.This studywas undertaken to determine the effects of ingesting either glucose(trial G) or glucose plusbranched-chain amino acids (BCAA; trialB), compared with placebo (trialP), during prolonged exercise. Nine well-trained cyclists with a maximal oxygen uptake of 63.1 ± 1.5 mlO₂ ^· min^{1 ·} kg¹performed three laboratory trials consisting of 100 km of cycling separated by 7 days between each trial. During these trials, the subjects were encouraged to complete the 100 km as fast as possible ontheir own bicycles connected to a magnetic brake. No differences inperformance times were observed between the three trials (160.1 ± 4.1, 157.2 ± 4.5, and 159.8 ± 3.7 min, respectively). Intrial B, plasma BCAA levels increased from339 ± 28 µM at rest to 1,026 ± 62 µM after exercise(P < 0.01). Plasma ammoniaconcentrations increased during the entire exercise period for allthree trials and were significantly higher intrial B compared withtrials G andP (P < 0.05). The respiratory exchange ratio was similar in the threetrials during the first 90 min of exercise; thereafter, it tended todrop more in trial P than intrials G andB. These data suggest that neitherglucose nor glucose plus BCAA ingestion during 100 km of cyclingenhance performance in well-trained cyclists.

     

Differential behavior of cytotoxic effector cells against HLA antigens in strong genetic linkage disequilibrium

Five sets of cytotoxic effector cells were generated, using haploidentical, first degree relatives in five different families, against the HLA-A3; B7 serological determinants combined with different DR antigens. When tested against a panel of cells bearing combinations of the HLA-A, -B and -DR antigens it was shown that the HLA-B7 antigen was as strong a CML target determinant alone as it was in the presence of HLA-A3. The strength of the HLA-A3 antigen as target determinant varied. With effector cells primed to the HLA-A3; B7; DR2 haplotype, the A3 antigen alone behaved as a weak target determinant. When the same target cells were tested with the effector cells generated against HLA-A3; B7 without DR2, the A3 antigen behaved as a strong target determinant. A number of target cells lacking the serologically detectable HLA determinants present on the sensitizing HLA haplotype were identified as being killed by specific effector cells. These data suggest either a number of new CML target determinants controlled by different loci or the presence of a single, new locus with multiple alleles controlling CML targets.     

Comparison of myosin isoenzymes present in skeletal and cardiac muscles of the Arctic charr Salvelinus alpinus (L.). Sequential expression of different myosin heavy chains during development of the fast white skeletal muscle.

The expression of myosin isoforms and their subunit composition in the white skeletal body musculature of Arctic charr (Salvelinus alpinus) of different ages (from 77-day embryos until about 5 years old) was studied at the protein level by means of electrophoretic techniques. Myosin from the white muscle displayed three types of light chain during all the developmental stages examined: two myosin light chains type 1 (LC1F) differing in both apparent molecular mass and pI, one myosin light chain type 2 (LC2F) and one myosin light chain type 3 (LC3F). The fastest-migrating form of LC1F seemed to be predominant during the embryonic and eleutheroembryonic periods. The slowest-migrating form of LC1F was predominant in the 5-year-old fish. Between 1 year and 4 years, both types of LC1F were present in similar amounts. Cardiac as well as red muscle myosin from 3-year-old fish had two types of light chain. The myosin light chains from atria and ventriculi were indistinguishable by two-dimensional electrophoresis, but were different from the myosin light chains from red muscle. Neither the light chains from cardiac nor red muscle were coexpressed with the myosin light chains of white muscle at any of the developmental stages examined. Two myosin heavy chain bands were resolved by SDS/glycerol/polyacrylamide gel electrophoresis of the extract from embryos. One of the bands was present in minor amounts. The other, and most abundant, band comigrated with the only band found in the extracts of white muscle myosin from older fish. One-dimensional Staphylococcus aureus V8 protease peptide mapping of these bands revealed some differences during development of the white muscle tentatively interpreted as follows. The myosin heavy chain band present in minor amounts in the embryos may represent an early embryonic form that is replaced by a late embryonic or foetal form in the eleutheroembryos. The foetal myosin heavy chain appears to be present until the resorption of the yolk sack and beginning of the free-swimming stage. A new form of myosin heavy chain, termed neonatal and probably expressed around hatching, is present until about 1 year of age.(ABSTRACT TRUNCATED AT 400 WORDS)     

Lewontin and Kojima Meet Fisher: Linkage in a Symmetric Model of Sex Determination

The effect of linkage and epistasis on the evolution of the sex-ratio is studied in a symmetric two-locus model of autosomal sex determination closely related to the symmetric viability model of R. C. Lewontin and K. Kojima. R. A. Fisher's expectation of an even sex ratio for autosomal sex determination by a single gene governs the dynamics when the loci are tightly linked. However, recombination may preclude optimization of the sex ratio just as occurs in viability selection models. Many of the evolutionary phenomena known for the symmetric viability model also occur here. In addition, we exhibit a series of new phenomena related to the presence of surfaces of even sex ratio.     

Attachment sites of primary binding proteins L1, L2 and L23 on 23 S ribosomal RNA of Escherichia coli

The attachment sites of the primary binding proteins L1, L2 and L23 on 23 S ribosomal RNA of Escherichia coli were examined by a chemical and ribonuclease footprinting method using several probes with different specificities. The results show that the sites are confined to localized RNA regions within the large ribonuclease-protected ribonucleoprotein fragments that were characterized earlier. They are as follows: (1) L1 recognizes a tertiary structural motif in domain V centred on two interacting internal loops; the main protein interaction sites occur at the internal loop/helix junctions. (2) The L2 site constitutes a single irregular stem/loop structure in the centre of domain IV where non-Watson-Crick pairing is likely to occur. (3) L23 recognizes a tertiary structural motif involving a single terminal loop structure and part of an adjacent internal loop at the centre of domain III. Each of the three primary binding proteins, whose presence is essential for ribosomal assembly, has been associated with important ribosomal functions: L1 lies in the E-site for deacylated tRNA binding while L2 and L23 have been implicated in the P and A substrate sites, respectively, of the peptidyl transferase centre. Moreover, each of the protein sites, but particularly those of L2 and L23, lies at the centre of RNA domains where they can maximally influence both the assembly of secondary binding proteins and the function of the RNA region.     

Physical and genetic mapping of the genomes of five Mycoplasma hominis strains by pulsed-field gel electrophoresis.

We present the complete maps of five Mycoplasma hominis genomes, including a detailed restriction map and the locations of a number of genetic loci. The restriction fragments were resolved by field inversion gel electrophoresis or by the contour-clamped homogeneous-electric-field system of pulsed-field gel electrophoresis. All the ApaI, SmaI, BamHI, XhoI, and SalI restriction sites (total of 21 to 33 sites in each strain) were placed on the physical map, yielding an average resolution of 26 kb. The maps were constructed using three different approaches: (i) size determination of DNA fragments partially or completely cleaved with one or two restriction enzymes, (ii) hybridization analysis with purified restriction fragments and specific probes, and (iii) use of linking clones. A genetic map was constructed by hybridization with gene-specific probes for rpoA, rpoC, rrn, tuf, gyrB, hup, ftsY, the unc operon, the genes for two M. hominis-specific antigenic membrane proteins, and one gene encoding a protein with some homology to Escherichia coli alanyl-tRNA synthetase. The positions of mapped loci were partially conserved in the five strains except in one strain in which a 300-kb fragment was inverted. The numbers and order of mapped restriction sites were only partly conserved, and this conservation was restricted to certain regions. The gene order was compared with the gene order established for other bacteria and was found to be identical to that of the phylogenetically related Clostridium perfringens. The genome size of the M. hominis strains varied from 704 to 825 kb.     

The human placental transferrin receptor: Reconstitution into liposomes and electron microscopy

Human transferrin receptor was isolated from Triton X-100 solubilized placental plasma membranes by a rapid one-step chromatographic procedure based on immunoadsorption of the receptortransferrin complex on anti-transferrin Sepharose and lectin-affinity on wheat germ agglutinin. Following exchange of Triton X-100 with CHAPS or n-octylglucoside, the purified receptor was incorporated into egg phosphatidylcholine liposomes upon, detergent removal by dialysis (lipid/protein ratio 15:1 to 45:1 (w/w) Reconstitution of the receptor was confirmed by trypsin cleavage to dissociate the large extracellular receptor domain from the liposomal membranes. Electron micrographs of the receptor-lipid recombinants negatively stained with sodium sillicotungstate, showed ographs of the receptor-lipid recombinants negatively stained with sodium sillicotungstate, showed that the receptor molecules distributed very inhomogeneously on the liposomes, most receptors being clustered. Single copies of the receptor were seen as elongate structures (5×10 nm) oriented with their long axis parallel to the liposome surface and separated from this by a 2–3 nm gap. This result provides evidence for a narrow connecting link between the globular extracellular receptor domain and the membrane spanning segment.Abbreviations CHAPS 3-[(3-cholamidopropyl)dimethylammonio]-1-propane sulfonate - PAGE polyacrylaminde gel electrophoresis - PC phosphatidylcholine - PMSF phenylmethylsulfonyl fluoride - SDS sodium dodecyl sulfate - WGA wheat germ agglutinin     

Chlamydia trachomatis contains a protein similar to the Legionella pneumophila mip gene product

A 27kDa Chlamydia trachomatis L2 protein was characterized by the use of monoclonal antibodies and by two-dimensional gel electrophoresis. The protein was shown to be located in the membrane of reticulate bodies as well as elementary bodies. Its synthesis could be detected from 10 hours post-infection. Cloning and sequence analysis of the distal part of the gene revealed an open reading frame of 175 amino acids. Comparison of the deduced amino acid sequence with the NBRF data base revealed significant homology between the 27 kDa chlamydial membrane protein and the product of the macrophage infectivity potentiator (mip) gene of Legionella pneumophila.     

Immunocytochemical demonstration of somatotropin-like and prolactin-like activity in the brain of Calamoichthys calabaricus (Actinopterygii)

Summary Cellular binding of anti-bSTH and anti-oPRL IgG is demonstrated in the brain and the pituitary gland of the African freshwater fish Calamoichthys calabaricus by means of the unlabeled antibody enzyme method at the light microscopic level. In the brain, somatotropin and prolactin are demonstrated in separate neurons in the preoptic area. The somatotropinergic and prolactinergic perikarya are distinct from those of the hypothalamic-hypophysial neurosecretory neurons, i.e., those stainable with aldehyde fuchsin presumed to be vasotocinergic and isotocinergic. The somatotropinergic and prolactinergic neuronal perikarya give rise to separate beaded axons which pass either ventroposteriorly into the infundibulum, terminating in the neurohypophysis, or ventro-laterally through the wall of the preoptic recess, terminating near the superficial capillary bed covering this part of the brain surface. Moreover, coarse dendrite-like processes of both kinds of immuno-reactive neurons extend towards, and end in, the third ventricle. Binding sites in the brain to antisera against hLH, hFSH, hTSH and anti-(1–24) ACTH IgG, all reactive in the pituitary, are not observed in the neurons confined to the preoptic area.Supported by the Danish Natural Sciences Research CouncilThe authors wish to thank Professor Dr K.G. Wingstrand, University of Copenhagen, Denmark, for placing two series of C. calabaricus at their disposal. They would also like to thank the National Institute of Arthritis, Metabolism and Digestive Diseases, Bethesda, USA, for the generous gift of antisera against the subunits of human LH, TSH and FSH, and likewise Dr L. Hummer, Glostrup Hospital, Copenhagen, Denmark, for the gift of the anti-(1-24)ACTH IgG