Exercise training in normobaric hypoxia in endurance runners. II. Improvement of mitochondrial properties in skeletal muscle.

This study investigates whether adaptations of mitochondrial function accompany the improvement of endurance performance capacity observed in well-trained athletes after an intermittent hypoxic training program. Fifteen endurance-trained athletes performed two weekly training sessions on treadmill at the velocity associated with the second ventilatory threshold (VT2) with inspired O2 fraction = 14.5% [hypoxic group (Hyp), n = 8] or with inspired O2 fraction = 21% [normoxic group (Nor), n = 7], integrated into their usual training, for 6 wk. Before and after training, oxygen uptake (VO2) and speed at VT2, maximal VO2 (VO2 max), and time to exhaustion at velocity of VO2 max (minimal speed associated with VO2 max) were measured, and muscle biopsies of vastus lateralis were harvested. Muscle oxidative capacities and sensitivity of mitochondrial respiration to ADP (Km) were evaluated on permeabilized muscle fibers. Time to exhaustion, VO2 at VT2, and VO2 max were significantly improved in Hyp (+42, +8, and +5%, respectively) but not in Nor. No increase in muscle oxidative capacity was obtained with either training protocol. However, mitochondrial regulation shifted to a more oxidative profile in Hyp only as shown by the increased Km for ADP (Nor: before 476 +/- 63, after 524 +/- 62 microM, not significant; Hyp: before 441 +/- 59, after 694 +/- 51 microM, P < 0.05). Thus including hypoxia sessions into the usual training of athletes qualitatively ameliorates mitochondrial function by increasing the respiratory control by creatine, providing a tighter integration between ATP demand and supply.     

Adapting light trap to catch household insects in central Cameroon: a pilot study

Various trapping methods and techniques developed for entomological studies are inefficient in attracting all arthropod groups. A new simple light trap, the basin light trap (BLT), to collect arthropod fauna around household in central Cameroon was thus designed. Its performance is evaluated by comparing collected data with those obtained by the Centre for Disease Control light trap (CDC) commonly used in mosquito studies. BLT were activated between 6 and 10 pm and CDC between 5 pm and 8 am. A total of 36,804 arthropods specimens were collected and sorted into 115 families within 20 orders and three phyla (Myriapoda, Arachnida and Hexapoda). BLT attracted more individuals (22,250 vs 14,554), families (106 vs 65) and significantly more families per night (33.0 ± 15.9 vs 21.0 ± 8.5) than CDC. Some families were collected by only one specific trap type while others overlapped between the two traps. Prosopistomatidae (Ephemeroptera) was the most abundant family collected by BLT. In contrast, Cecidomyiidae (Diptera) was the most abundant taxon trapped by CDC. These results indicate that collecting methods such as BLT complement available trapping methods in evaluating arthropod fauna in and around households.     

Chronic intake of red wine polyphenols by young rats prevents aging-induced endothelial dysfunction and decline in physical performance: role of NADPH oxidase

Aging is associated with oxidative stress-mediated endothelial dysfunction and decline in physical performance, which promote cardiovascular diseases. This study examined whether chronic intake of red wine polyphenols (RWPs), a rich source of natural antioxidants, prevents aging-related impairment of vascular function and physical exercise capacity. Vascular reactivity from 12, 20 and 40 week-old rats was assessed in organ chambers. Rats received from week 16 to 40 either solvent, RWPs or the antioxidant and NADPH oxidase inhibitor, apocynin. Aging was associated with blunted endothelium-dependent relaxations, oxidative stress (dihydroethidine staining), and an upregulation of eNOS, arginase I, NADPH oxidase p22phox and nox1 subunits, and AT1 and AT2 receptors (assessed by immunohistochemistry) in the mesenteric artery. RWPs and apocynin improved the endothelial dysfunction, normalized oxidative stress and the expression of the different proteins. RWPs also improved aging-related decline in physical exercise. Thus, intake of RWPs protects against aging-induced endothelial dysfunction and decline in physical performance. These effects likely involve the ability of RWPs to normalize oxidative stress and the expression of proteins involved in the formation of NO and the angiotensin II pathway.     

Mutations in cytoplasmic dynein lead to a Huntington's disease-like defect in energy metabolism of brown and white adipose tissues

The molecular motor dynein is regulated by the huntingtin protein, and Huntington's disease (HD) mutations of huntingtin disrupt dynein motor activity. Besides abnormalities in the central nervous system, HD animal models develop prominent peripheral pathology, with defective brown tissue thermogenesis and dysfunctional white adipocytes, but whether this peripheral phenotype is recapitulated by dynein dysfunction is unknown. Here, we observed prominently increased adiposity in mice harboring the legs at odd angles (Loa/+) or the Cramping mutations (Cra/+) in the dynein heavy chain gene. In Cra/+ mice, hyperadiposity occurred in the absence of energy imbalance and was the result of impaired norepinephrine-stimulated lipolysis. A similar phenotype was observed in 3T3L1 adipocytes upon chemical inhibition of dynein showing that loss of functional dynein leads to impairment of lipolysis. Ex vivo, dynein mutant adipose tissue displayed increased reactive oxygen species production that was, at least partially, responsible for the decreased cellular responses to norepinephrine and subsequent defect in stimulated lipolysis. Dynein mutation also affected norepinephrine efficacy to elicit a thermogenic response and led to morphological abnormalities in brown adipose tissue and cold intolerance in dynein mutant mice. Interestingly, protein levels of huntingtin were decreased in dynein mutant adipose tissue. Collectively, our results provide genetic evidence that dynein plays a key role in lipid metabolism and thermogenesis through a modulation of oxidative stress elicited by norepinephrine. This peripheral phenotype of dynein mutant mice is similar to that observed in various animal models of HD, lending further support for a functional link between huntingtin and dynein.     

Proteomic Analysis of C2C12 Myoblast and Myotube Exosome-Like Vesicles: A New Paradigm for Myoblast-Myotube Cross Talk?

Exosomes are nanometer-sized microvesicles formed in multivesicular bodies (MVBs) during endosome maturation. Exosomes are released from cells into the microenvironment following fusion of MVBs with the plasma membrane. During the last decade, skeletal muscle-secreted proteins have been identified with important roles in intercellular communications. To investigate whether muscle-derived exosomes participate in this molecular dialog, we determined and compared the protein contents of the exosome-like vesicles (ELVs) released from C2C12 murine myoblasts during proliferation (ELV-MB), and after differentiation into myotubes (ELV-MT). Using a proteomic approach combined with electron microscopy, western-blot and bioinformatic analyses, we compared the protein repertoires within ELV-MB and ELV-MT. We found that these vesicles displayed the classical properties of exosomes isolated from other cell types containing components of the ESCRT machinery of the MVBs, as well as numerous tetraspanins. Specific muscle proteins were also identified confirming that ELV composition also reflects their muscle origin. Furthermore quantitative analysis revealed stage-preferred expression of 31 and 78 proteins in ELV-MB and ELV-MT respectively. We found that myotube-secreted ELVs, but not ELV-MB, reduced myoblast proliferation and induced differentiation, through, respectively, the down-regulation of Cyclin D1 and the up-regulation of myogenin. We also present evidence that proteins from ELV-MT can be incorporated into myoblasts by using the GFP protein as cargo within ELV-MT. Taken together, our data provide a useful database of proteins from C2C12-released ELVs throughout myogenesis and reveals the importance of exosome-like vesicles in skeletal muscle biology.     

Internalization and down-regulation of the ALK receptor in neuroblastoma cell lines upon monoclonal antibodies treatment

Recently, activating mutations of the full length ALK receptor, with two hot spots at positions F1174 and R1275, have been characterized in sporadic cases of neuroblastoma. Here, we report similar basal patterns of ALK phosphorylation between the neuroblastoma IMR-32 cell line, which expresses only the wild-type receptor (ALK(WT)), and the SH-SY5Y cell line, which exhibits a heterozygous ALK F1174L mutation and expresses both ALK(WT) and ALK(F1174L) receptors. We demonstrate that this lack of detectable increased phosphorylation in SH-SY5Y cells is a result of intracellular retention and proteasomal degradation of the mutated receptor. As a consequence, in SH-SY5Y cells, plasma membrane appears strongly enriched for ALK(WT) whereas both ALK(WT) and ALK(F1174L) were present in intracellular compartments. We further explored ALK receptor trafficking by investigating the effect of agonist and antagonist mAb (monoclonal antibodies) on ALK internalization and down-regulation, either in SH-SY5Y cells or in cells expressing only ALK(WT). We observe that treatment with agonist mAbs resulted in ALK internalization and lysosomal targeting for receptor degradation. In contrast, antagonist mAb induced ALK internalization and recycling to the plasma membrane. Importantly, we correlate this differential trafficking of ALK in response to mAb with the recruitment of the ubiquitin ligase Cbl and ALK ubiquitylation only after agonist stimulation. This study provides novel insights into the mechanisms regulating ALK trafficking and degradation, showing that various ALK receptor pools are regulated by proteasome or lysosome pathways according to their intracellular localization.     

Adaptative evolution of the Vkorc1 gene in Mus musculus domesticus is influenced by the selective pressure of anticoagulant rodenticides

Anticoagulant rodenticides are commonly used to control rodent pests worldwide. They specifically inhibit the vitamin K epoxide reductase (VKORC1), which is an enzyme encoded by the Vkorc1 gene, involved in the recycling of vitamin K. Therefore, they prevent blood clotting. Numerous mutations of Vkorc1 gene were reported in rodents, and some are involved in the resistant to rodenticides phenotype. Two hundred and sixty‐six mice tails were received from 65 different locations in France. Coding sequences of Vkorc1 gene were sequenced in order to detect mutations. Consequences of the observed mutations were evaluated by the use of recombinant VKORC1. More than 70% of mice presented Vkorc1 mutations. Among these mice, 80% were homozygous. Contrary to brown rats for which only one predominant Vkorc1 genotype was found in France, nine missense single mutations and four double mutations were observed in house mice. The single mutations lead to resistance to first‐generation antivitamin K (AVKs) only and are certainly associated with the use of these first‐generation molecules by nonprofessionals for the control of mice populations. The double mutations, probably obtained by genetic recombination, lead to in vitro resistance to all AVKs. They must be regarded as an adaptive evolution to the current use of second‐generation AVKs. The intensive use of first‐generation anticoagulants probably allowed the selection of a high diversity of mutations, which makes possible the genetic recombination and consequently provokes the emergence of the more resistant mutated Vkorc1 described to date.