HDLs activate ADAM17-dependent shedding

The tumor necrosis factor-alpha (TNF) converting enzyme (ADAM17) is a metalloprotease that cleaves several transmembrane proteins, including TNF and its receptors (TNFR1 and TNFR2). We recently showed that the shedding activity of ADAM17 is sequestered in lipid rafts and that cholesterol depletion increased the shedding of ADAM17 substrates. These data suggested that ADAM17 activity could be regulated by cholesterol movements in the cell membrane. We investigated if the membrane cholesterol efflux induced by high-density lipoproteins (HDLs) was able to modify the shedding of ADAM17 substrates. HDLs added to different cell types, increased the ectodomain shedding of TNFR2, TNFR1, and TNF, an effect reduced by inhibitors active on ADAM17. The HDLs-stimulated TNF release occurred also on cell-free isolated plasma membranes. Purified apoA1 increased the shedding of TNF in an ABCA1-dependent manner, suggesting a role for the cholesterol efflux in this phenomenon. HDLs reduced the cholesterol and proteins (including ADAM17) content of lipid rafts and triggered the ADAM17-dependent cleavage of TNF in the non-raft region of the membrane. In conclusion, these data demonstrate that HDLs alter the lipid raft structure, which in turn activates the ADAM17-dependent processing of transmembrane substrates.     

Genetic variability in residual feed intake in rainbow trout clones and testing of indirect selection criteria (Open Access publication)

Little is known about the genetic basis of residual feed intake (RFI) variation in fish, since this trait is highly sensitive to environmental influences, and feed intake of individuals is difficult to measure accurately. The purpose of this work was (i) to assess the genetic variability of RFI estimated by an X-ray technique and (ii) to develop predictive criteria for RFI. Two predictive criteria were tested: loss of body weight during feed deprivation and compensatory growth during re-feeding. Ten heterozygous rainbow trout clones were used. Individual intake and body weight were measured three times at threeweek intervals. Then, individual body weight was recorded after two cycles of a three-week feed deprivation followed by a three-week re-feeding. The ratio of the genetic variance to the phenotypic variance was found high to moderate for growth, feed intake, and RFI (VG/VP = 0.63 ± 0.11, 0.29 ± 0.11, 0.29 ± 0.09, respectively). The index that integrates performances achieved during deprivation and re-feeding periods explained 59% of RFI variations. These results provide a basis for further studies on the origin of RFI differences and show that indirect criteria are good candidates for future selective breeding programs.     

TRP-2 specifically decreases WM35 cell sensitivity to oxidative stress

TRP-2 (dopachrome tautomerase) is a melanogenic enzyme whose expression was recently reported to modulate melanocyte response to different cytotoxic events. Here we studied a possible role of TRP-2 in the oxidative stress response in the amelanotic WM35 melanoma cell line. Cell viability assays showed that TRP-2 overexpression in WM35 cells reduced their sensitivity to oxidative stress. Comet assays linked TRP-2 expression to DNA damage protection, and high-performance liquid chromotography-tandem mass spectrometry experiments showed an increase in intracellular glutathione in TRP-2-overexpressing cells. These effects were specifically reversed when TRP-2 was silenced by RNA interference. Nevertheless, these properties appeared to depend on a particular cell environment because expression of TRP-2 failed to rescue HEK epithelial cells exposed to similar treatments.     

DYC-1, a protein functionally linked to dystrophin in Caenorhabditis elegans is associated with the dense body, where it interacts with the muscle LIM domain protein ZYX-1

In Caenorhabditis elegans, mutations of the dystrophin homologue, dys-1, produce a peculiar behavioral phenotype (hyperactivity and a tendency to hypercontract). In a sensitized genetic background, dys-1 mutations also lead to muscle necrosis. The dyc-1 gene was previously identified in a genetic screen because its mutation leads to the same phenotype as dys-1, suggesting that the two genes are functionally linked. Here, we report the detailed characterization of the dyc-1 gene. dyc-1 encodes two isoforms, which are expressed in neurons and muscles. Isoform-specific RNAi experiments show that the absence of the muscle isoform, and not that of the neuronal isoform, is responsible for the dyc-1 mutant phenotype. In the sarcomere, the DYC-1 protein is localized at the edges of the dense body, the nematode muscle adhesion structure where actin filaments are anchored and linked to the sarcolemma. In yeast two-hybrid assays, DYC-1 interacts with ZYX-1, the homologue of the vertebrate focal adhesion LIM domain protein zyxin. ZYX-1 localizes at dense bodies and M-lines as well as in the nucleus of C. elegans striated muscles. The DYC-1 protein possesses a highly conserved 19 amino acid sequence, which is involved in the interaction with ZYX-1 and which is sufficient for addressing DYC-1 to the dense body. Altogether our findings indicate that DYC-1 may be involved in dense body function and stability. This, taken together with the functional link between the C. elegans DYC-1 and DYS-1 proteins, furthermore suggests a requirement of dystrophin function at this structure. As the dense body shares functional similarity with both the vertebrate Z-disk and the costamere, we therefore postulate that disruption of muscle cell adhesion structures might be the primary event of muscle degeneration occurring in the absence of dystrophin, in C. elegans as well as vertebrates.     

Oligocarrageenans and tissue-dependant oxidative burst in Solieria chordalis (Rhodophyceae, Gigartinales)

The release of hydrogen peroxide by thallus fragments of the rhodophycean Solieria chordalis (C. Agardh) J. Agardh has been documented both in the presence and in the absence of oligosaccharides. Within 1 h, ramuli were able to release large amounts of peroxide in the absence of any chemical stress. Among potential elicitors tested, only degree of polymerization 1 (DP1) and DP7‐8 oligo‐iota‐carrageenans stimulated defense mechanisms in both axes and ramuli as shown by the occurrence of an oxidative burst. Chopping of the tissues had no effect on the intensity of the burst, therefore suggesting that mainly cortical cell layers were involved in the process. After 5 min incubation, a dose of 125 μg mL^?1 of an oligomeric mixture containing a large proportion of DP1 units proved to be sufficient to obtain a maximal response. The intensity of the burst was significantly higher with isolated ramuli than with pieces of the axis, with outer peroxide accumulations reaching 200 nmol g^?1 fresh weight of treated tissue. Altogether, our results show that S. chordalis is able to react to a simulated pathogen attack by an oxidative burst and that the capacity to carry out an oxidative burst is stronger in ramuli than in axes.     

An alternative HPLC method for the quantification of floridoside in salt-stressed cultures of the red alga Grateloupia doryphora

The floridoside content of the marine red alga Grateloupiadoryphora was measured using a new, alternative High PerformanceLiquid Chromatography method suitable for the analysis of a large number ofsamples. This assay was to study the quantitative changes in floridosideconcentration in G. doryphora cultivated in differentsalinities for 10 days. Marked changes in floridoside concentration occurredrapidly observed in reponse to osmotic stress, providing evidence of itsosmoregulatory function. Interestingly, after a rapid decrease at low salinityand a strong increase at high salinity, the floridoside content then slowlyreturned to its initial level over the last 3 days of culture. This observationconfirms the high physiological capacity of G. doryphoratowithstand environmental variations and probably contributes to its actualproliferation along French coasts.     

Characterization and quality control of recombinant adenovirus vectors for gene therapy

Highly purified recombinant adenovirus undergoes routine quality controls for identity, potency and purity prior to its use as a gene therapy vector. Quantitative characterization of infectivity is measurable by the expression of the DNA binding protein, an early adenoviral protein, in an immunofluorescence bioassay on permissive cells as a potency determinant. The specific particle count, a key quality indicator, is the total number of intact particles present compared to the number of infectious units. Electron microscopic analysis using negative staining gives a qualitative biophysical analysis of the particles eluted from anion-exchange HPLC. One purity assessment is accomplished via the documented presence and relative ratios of component adenoviral proteins as well as potential contaminants by reversed-phase HPLC of the intact virus followed by protein peak identification using MALDI-TOF mass spectrometry and subsequent data mining. Verification of the viral genome is performed and expression of the transgene is evaluated in in vitro systems for identity. Production lots are also evaluated for replication-competent adenovirus prior to human use. For adenovirus carrying the human IL-2 transgene, quantitative IL-2 expression is demonstrated by ELISA and cytokine potency by cytotoxic T lymphocyte assay following infection of permissive cells. Both quantitative and qualitative analyses show good batch to batch reproducibility under routine test conditions using validated methods.     

Effects of Sprint versus High-Intensity Aerobic Interval Training on Cross-Country Mountain Biking Performance: A Randomized Controlled Trial

Objectives

The current study compared the effects of high-intensity aerobic training (HIT) and sprint interval training (SIT) on mountain biking (MTB) race simulation performance and physiological variables, including peak power output (PPO), lactate threshold (LT) and onset of blood lactate accumulation (OBLA).

Methods

Sixteen mountain bikers (mean ± SD: age 32.1 ± 6.4 yr, body mass 69.2 ± 5.3 kg and VO_2max 63.4 ± 4.5 mL∙kg^-1∙min^-1) completed graded exercise and MTB performance tests before and after six weeks of training. The HIT (7–10 x [4–6 min—highest sustainable intensity / 4–6 min—CR100 10–15]) and SIT (8–12 x [30 s—all-out intensity / 4 min—CR100 10–15]) protocols were included in the participants’ regular training programs three times per week.

Results

Post-training analysis showed no significant differences between training modalities (HIT vs. SIT) in body mass, PPO, LT or OBLA (p = 0.30 to 0.94). The Cohen’s d effect size (ES) showed trivial to small effects on group factor (p = 0.00 to 0.56). The interaction between MTB race time and training modality was almost significant (p = 0.08), with a smaller ES in HIT vs. SIT training (ES = -0.43). A time main effect (pre- vs. post-phases) was observed in MTB race performance and in several physiological variables (p = 0.001 to 0.046). Co-variance analysis revealed that the HIT (p = 0.043) group had significantly better MTB race performance measures than the SIT group. Furthermore, magnitude-based inferences showed HIT to be of likely greater benefit (83.5%) with a lower probability of harmful effects (0.8%) compared to SIT.

Conclusion

The results of the current study suggest that six weeks of either HIT or SIT may be effective at increasing MTB race performance; however, HIT may be a preferable strategy.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT01944865","term_id":"NCT01944865"}}NCT01944865     

The PhysioFlow Thoracic Impedancemeter Is Not Valid for the Measurements of Cardiac Hemodynamic Parameters in Chronic Anemic Patients

The aim of the present study was to test the validity of the transthoracic electrical bioimpedance method PhysioFlow® to measure stroke volume in patients with chronic anemia. Stroke volume index (SVI), as well as cardiac index (CI) obtained by transthoracic electrical bioimpedance method and doppler echocardiography were compared in healthy subjects (n = 25) and patients with chronic anemia (i.e. mainly with sickle cell anemia; n = 32), at rest. While doppler echocardiography was able to detect difference in SVI between the two populations, the Physioflow® failed to detect any difference. Bland & Altman analyses have demonstrated no interchangeability between the two methods to assess CI and SVI in anemic patients and healthy subjects. While doppler echocardiography displayed a good concordance for SVI results with those obtained in the literature for anemic patients, the Physioflow® did not. Finally, in contrast to doppler echocardiography: 1) the CI obtained with the Physioflow® was not correlated with the hemoglobin level and 2) the stroke volume determined by the Physioflow® was highly influenced by body surface area. In conclusion, our findings indicate that the Physioflow® device is inaccurate for the measurement of SVI and CI in patients with chronic anemia and has a poor accuracy for the measurement of these parameters in African healthy subjects.