HALA!'s White Paper. Towards Higher Levels of Automation in ATM

• Objectives of HALA! • Main Activities • HALA! Magement Team • Participants • Intended Audience • Heritage in ATM and Automation • The new paradigm shift in Automation in ATM • Overall system performance as main driver for ATM Automation • The three interdependent dimensions for the paradigm change. • New roles assignment based on : • “best time” • “decision place” • “best player” • HALA! main research areas

A common mechanism of defective channel trafficking underlying DFNA2 hearing loss result in different cell surface expression levels of KCNQ4 mutants

KCNQ4 mutations underlie DFNA2, a subtype of autosomal dominant hearing loss. We had previously identified the pore-region p.G296S mutation that impaired channel activity in two manners: it greatly reduced surface expression and abolished channel function. Moreover, G296S mutant exerted a strong dominant-negative effect on potassium currents by reducing the channel expression at the cell surface representing the first study to identify a trafficking-dependent dominant mechanism for the loss of KCNQ4 channel function in DFNA2. Here, we have investigated the pathogenic mechanism associated with all the described KCNQ4 mutations (F182L, W242X, E260K, D262V, L274H, W276S, L281S, G285C, G285S and G321S) that are located in different domains of the channel protein. F182L mutant showed a wild type-like cell-surface distribution in transiently transfected NIH3T3 fibroblasts and the recorded currents in Xenopus oocytes resembled those of the wild-type. The remaining KCNQ4 mutants abolished potassium currents, but displayed distinct levels of defective cell-surface expression in NIH3T3 as quantified by flow citometry. Co-localization studies revealed these mutants were retained in the ER, unless W242X, which showed a clear co-localization with Golgi apparatus. Interestingly, this mutation results in a truncated KCNQ4 protein at the S5 transmembrane domain, before the pore region, that escapes the protein quality control in the ER but does not reach the cell surface at normal levels. Currently we are investigating the trafficking behaviour and electrophysiological properties of several KCNQ4 truncated proteins artificially generated in order to identify specific motifs involved in channel retention/exportation. Altogether, our results indicate that a defect in KCNQ4 trafficking is the common mechanism underlying DFNA2

Ionization levels of doped copper indium sulfide chalcopyrites

The electronic structure of modified chalcopyrite CuInS2 has been analyzed from first principles within the density functional theory. The host chalcopyrite has been modified by introducing atomic impurities M at substitutional sites in the lattice host with M = C, Si, Ge, Sn, Ti, V, Cr, Fe, Co, Ni, Rh, and Ir. Both substitutions M for In and M for Cu have been analyzed. The gap and ionization energies are obtained as a function of the M-S displacements. It is interesting for both spintronic and optoelectronic applications because it can provide significant information with respect to the pressure effect and the nonradiative recombination.

Higher levels of physical fitness are associated with a reduced risk of suffering sarcopenic obesity and better perceived health among elderly. The EXERNET multi-center study

Evaluar la asociación entre los niveles físicos de la aptitud, de calidad relacionada con la salud de la vida (CVRS) y la obesidad sarcopénica

Nutritional intake and body composition of adolescent soccer player of two different levels of performance: a comparative study

Soccer participation worldwide is increasing and every club try to discover new talents. It is well know that there is an important correlation between body composition (BC) and talent detection (TD) and when coaches and selectors choose players, they tend to choose them with optimum BC.

Effects of nutrition on digestión efficicency and gaseous emissions from slurry in growing-finishing pigs. I. Influence of the inclusión of two levels of orange pulp and carb meal in isofibrous diets

An experiment was conducted to investigate the effects of increasing the level of two sources of fibrous by-products, orange pulp (OP) and carob meal (CM), in iso-NDF growing-finishing pig diets on nutrient balance, slurry composition and potential ammonia (NH3) and methane (CH4) emissions. Thirty pigs (85.4 ± 12.3 kg) were fed five iso-nutritive diets: a commercial control wheat/barley (C) and four experimental diets including two sources of fibrous by-products (OP and CM) and two dietary levels (75 and 150 g/kg) in a 2 × 2 factorial arrangement. After a 14-day adaptation period, faeces and urine were collected separately for 7 days to measure nutrient digestibility and the excretory patterns of N from pigs (6 replicates per diet) housed individually in metabolic pens. For each animal, the derived NH3 and CH4 emissions were measured in samples of slurry over an 11- and 100-day storage periods, respectively. Source and level of the fibrous by-products affected digestion efficiency in a different way as the coefficients of total tract apparent digestibility (CTTAD) for dry matter (DM), organic matter (OM), fibre fractions and gross energy increased with OP but decreased with CM (P < 0.05). Crude protein CTTAD decreased with the inclusion of both sources of fibre, being lower at the highest dietary level. Faecal concentration of fibre fractions increased (P < 0.05) with the level of inclusion of CM but decreased with that of OP (P < 0.01). High dietary level for both sources of fibre increased (P < 0.02) CP faecal content but urine N content decreased (from 205 to 168 g/kg DM, P < 0.05) in all the fibre-supplemented compared to C diet. Additionally, the proportions of undigested dietary, water soluble, and bacterial and endogenous debris of faecal N excretion were not affected by treatments. The initial slurry characteristics did not differ among different fibre sources and dietary levels, except pH, which decreased at the highest by-product inclusion levels. Ammonia emission per kg of slurry was lower in all the fibre-supplemented diets than in C diet (from 2.44 to 1.81 g, P < 0.05). Additionally, slurries from the highest dietary level of by-products tended (P < 0.06) to emit less NH3 per kg of initial total Kjeldahl N and showed a lower B0, independently of the fibre source. Thus, the fibre sources and their dietary levels affected pig nutrient digestion and composition of urine and faeces, showing potential to decrease NH3 and CH4 emissions at high levels of inclusion, independently of type of fibre.

Cardioprotection from ischemia by a brief exposure to physiological levels of ethanol: Role of epsilon protein kinase C

Recent epidemiological studies indicate beneficial effects of moderate ethanol consumption in ischemic heart disease. Most studies, however, focus on the effect of long-term consumption of ethanol. In this study, we determined whether brief exposure to ethanol immediately before ischemia also produces cardioprotection. In addition, because protein kinase C (PKC) has been shown to mediate protection of the heart from ischemia, we determined the role of specific PKC isozymes in ethanol-induced protection. We demonstrated that (i) brief exposure of isolated adult rat cardiac myocytes to 10–50 mM ethanol protected against damage induced by prolonged ischemia; (ii) an isozyme-selective ɛPKC inhibitor developed in our laboratory inhibited the cardioprotective effect of acute ethanol exposure; (iii) protection of isolated intact adult rat heart also occurred after incubation with 10 mM ethanol 20 min before global ischemia; and (iv) ethanol-induced cardioprotection depended on PKC activation because it was blocked by chelerythrine and GF109203X, two PKC inhibitors. Consumption of 1–2 alcoholic beverages in humans leads to blood alcohol levels of ≈10 mM. Therefore, our work demonstrates that exposure to physiologically attainable ethanol levels minutes before ischemia provides cardioprotection that is mediated by direct activation of ɛPKC in the cardiac myocytes. The potential clinical implications of our findings are discussed.

A single dose of kainic acid elevates the levels of enkephalins and activator protein-1 transcription factors in the hippocampus for up to 1 year

Neuronal plasticity plays a very important role in brain adaptations to environmental stimuli, disease, and aging processes. The kainic acid model of temporal lobe epilepsy was used to study the long-term anatomical and biochemical changes in the hippocampus after seizures. Using Northern blot analysis, immunocytochemistry, and Western blot analysis, we have found a long-term elevation of the proconvulsive opioid peptide, enkephalin, in the rat hippocampus. We have also demonstrated that an activator protein-1 transcription factor, the 35-kDa fos-related antigen, can be induced and elevated for at least 1 year after kainate treatment. This study demonstrated that a single systemic injection of kainate produces almost permanent increases in the enkephalin and an activator protein-1 transcription factor, the 35-kDa fos-related antigen, in the rat hippocampus, and it is likely that these two events are closely associated with the molecular mechanisms of induction of long-lasting enhanced seizure susceptibility in the kainate-induced seizure model. The long-term expression of the proenkephalin mRNA and its peptides in the kainate-treated rat hippocampus also suggests an important role in the recurrent seizures of temporal lobe epilepsy.

Immediate and simultaneous sensory reorganization at cortical and subcortical levels of the somatosensory system

The occurrence of cortical plasticity during adulthood has been demonstrated using many experimental paradigms. Whether this phenomenon is generated exclusively by changes in intrinsic cortical circuitry, or whether it involves concomitant cortical and subcortical reorganization, remains controversial. Here, we addressed this issue by simultaneously recording the extracellular activity of up to 135 neurons in the primary somatosensory cortex, ventral posterior medial nucleus of the thalamus, and trigeminal brainstem complex of adult rats, before and after a reversible sensory deactivation was produced by subcutaneous injections of lidocaine. Following the onset of the deactivation, immediate and simultaneous sensory reorganization was observed at all levels of the somatosensory system. No statistical difference was observed when the overall spatial extent of the cortical (9.1 ± 1.2 whiskers, mean ± SE) and the thalamic (6.1 ± 1.6 whiskers) reorganization was compared. Likewise, no significant difference was found in the percentage of cortical (71.1 ± 5.2%) and thalamic (66.4 ± 10.7%) neurons exhibiting unmasked sensory responses. Although unmasked cortical responses occurred at significantly higher latencies (19.6 ± 0.3 ms, mean ± SE) than thalamic responses (13.1 ± 0.6 ms), variations in neuronal latency induced by the sensory deafferentation occurred as often in the thalamus as in the cortex. These data clearly demonstrate that peripheral sensory deafferentation triggers a system-wide reorganization, and strongly suggest that the spatiotemporal attributes of cortical plasticity are paralleled by subcortical reorganization.

CytA enables CryIV endotoxins of Bacillus thuringiensis to overcome high levels of CryIV resistance in the mosquito, Culex quinquefasciatus

Cry proteins produced by Bacillus thuringiensis are selective biodegradable insecticides used increasingly in bacterial insecticides and transgenic plants as alternatives to synthetic chemical insecticides. However, the potential for development of resistance and cross-resistance in target insect populations to Cry proteins used alone or in combination threatens the more widespread use of this novel pest control technology. Here we show that high levels of resistance to CryIV proteins in larvae of the mosquito, Culex quinquefasciatus, can be suppressed or reduced markedly by combining these proteins with sublethal quantities of CytA, a cytolytic endotoxin of B. thuringiensis. Resistance at the LC95 level of 127-fold for a combination of three CryIV toxins (CryIVA, B, and D), resulting from 60 generations of continuous selection, was completely suppressed by combining sporulated powders of CytA in a 1:3 ratio with sporulated powders of a CryIVA, CryIVB, and CryIVD strain. Combining the CytA strain with a CryIVA and CryIVB strain also completely suppressed mosquito resistance of 217-fold to the latter toxins at the LC95 level, whereas combination of CytA with CryIVD reduced resistance in a CryIVD-selected mosquito strain from greater than 1,000-fold to less than 8-fold. The CytA/CryIV model provides a potential molecular genetic strategy for engineering resistance management for Cry proteins directly into bacterial insecticides and transgenic plants.

A yeast sterol auxotroph (erg25) is rescued by addition of azole antifungals and reduced levels of heme

Genetic disruption of the Saccharomyces cerevisiae C-4 sterol methyl oxidase ERG25 gene leads to sterol auxotrophy. We have characterized a suppression system that requires two mutations to restore viability to this disrupted strain. One suppressor mutation is erg11, which is blocked in 14α-demethylation of lanosterol and is itself an auxotroph. The second suppressor mutation required is either slu1 or slu2 (suppressor of lanosterol utilization). These mutations are leaky versions of HEM2 and HEM4, respectively; addition of exogenous hemin reverses the suppressing effects of slu1 and slu2. Suppression of erg25 by erg11 slu1 (or erg11 slu2) results in a slow-growing strain in which lanosterol, the first sterol in the pathway, accumulates. This result indicates that endogenously synthesized lanosterol can substitute for ergosterol and support growth. In the triple mutants, all but 1 (ERG6) of the 13 subsequent reactions of the ergosterol pathway are inactive. Azole antibiotics (clotrimazole, ketoconazole, and itraconazole) widely used to combat fungal infections are known to do so by inhibiting the ERG11 gene product, the 14α-demethylase. In this investigation, we demonstrate that treatment of the sterol auxotrophs erg25 slu1 or erg25 slu2 with azole antibiotics paradoxically restores viability to these strains in the absence of sterol supplementation via the suppression system we have described.

Reduction in levels of the cyclin-dependent kinase inhibitor p27kip-1 coupled with transforming growth factor β neutralization induces cell-cycle entry and increases retroviral transduction of primitive human hematopoietic cells

Successful gene therapy depends on stable transduction of hematopoietic stem cells. Target cells must cycle to allow integration of Moloney-based retroviral vectors, yet hematopoietic stem cells are quiescent. Cells can be held in quiescence by intracellular cyclin-dependent kinase inhibitors. The cyclin-dependent kinase inhibitor p15INK4B blocks association of cyclin-dependent kinase (CDK)4/cyclin D and p27kip-1 blocks activity of CDK2/cyclin A and CDK2/cyclin E, complexes that are mandatory for cell-cycle progression. Antibody neutralization of β transforming growth factor (TGFβ) in serum-free medium decreased levels of p15INK4B and increased colony formation and retroviral-mediated transduction of primary human CD34+ cells. Although TGFβ neutralization increased colony formation from more primitive, noncycling hematopoietic progenitors, no increase in M-phase-dependent, retroviral-mediated transduction was observed. Transduction of the primitive cells was augmented by culture in the presence of antisense oligonucleotides to p27kip-1 coupled with TGFβ-neutralizing antibodies. The transduced cells engrafted immune-deficient mice with no alteration in human hematopoietic lineage development. We conclude that neutralization of TGFβ, plus reduction in levels of the cyclin-dependent kinase inhibitor p27, allows transduction of primitive and quiescent hematopoietic progenitor populations.

Increased levels of plasma lysosomal enzymes in patients with Lowe syndrome

Lowe syndrome is an X-linked disorder that has a complex phenotype that includes progressive renal failure and blindness. The disease is caused by mutations in an inositol polyphosphate 5-phosphatase designated OCRL. It has been shown that the OCRL protein is found on the surface of lysosomes and that a renal tubular cell line deficient in OCRL accumulated substrate phosphatidylinositol 4,5-bisphosphate. Because this lipid is required for vesicle trafficking from lysosomes, we postulate that there is a defect in lysosomal enzyme trafficking in patients with Lowe syndrome that leads to increased extracellular lysosomal enzymes and might lead to tissue damage and contribute to the pathogenesis of the disease. We have measured seven lysosomal enzymes in the plasma of 15 patients with Lowe syndrome and 15 age-matched male controls. We find a 1.6- to 2.0-fold increase in all of the enzymes measured. When the data was analyzed by quintiles of activity for all of the enzymes, we found that 95% of values in the lowest quintile come from normal subjects whereas in the highest quintile 85% of the values are from patients with Lowe syndrome. The increased enzyme levels are not attributable to renal insufficiency because there was no difference in enzyme activity in the four patients with the highest creatinine levels compared with the six patients with the lowest creatinine values.

Defects in embryonic hindbrain development and fetal resorption resulting from vitamin A deficiency in the rat are prevented by feeding pharmacological levels of all-trans-retinoic acid

Vitamin A is required for reproduction and normal embryonic development. We have determined that all-trans-retinoic acid (atRA) can support development of the mammalian embryo to parturition in vitamin A-deficient (VAD) rats. At embryonic day (E) 0.5, VAD dams were fed purified diets containing either 12 μg of atRA per g of diet (230 μg per rat per day) or 250 μg of atRA per g of diet (4.5 mg per rat per day) or were fed the purified diet supplemented with a source of retinol (100 units of retinyl palmitate per day). An additional group was fed both 250 μg of atRA per g of diet in combination with retinyl palmitate. Embryonic survival to E12.5 was similar for all groups. However, embryonic development in the group fed 12 μg of atRA per g of diet was grossly abnormal. The most notable defects were in the region of the hindbrain, which included a loss of posterior cranial nerves (IX, X, XI, and XII) and postotic pharyngeal arches as well as the presence of ectopic otic vesicles and a swollen anterior cardinal vein. All embryonic abnormalities at E12.5 were prevented by feeding pharmacological amounts of atRA (250 μg/g diet) or by supplementation with retinyl palmitate. Embryos from VAD dams receiving 12 μg of atRA per g of diet were resorbed by E18.5, whereas those in the group fed 250 μg of atRA per g of diet survived to parturition but died shortly thereafter. Equivalent results were obtained by using commercial grade atRA or atRA that had been purified to eliminate any potential contamination by neutral retinoids, such as retinol. Thus, 250 μg of atRA per g of diet fed to VAD dams (≈4.5 mg per rat per day) can prevent the death of embryos at midgestation and prevents the early embryonic abnormalities that arise when VAD dams are fed insufficient amounts of atRA.

A retro-inverso peptide corresponding to the GH loop of foot-and-mouth disease virus elicits high levels of long-lasting protective neutralizing antibodies

Peptides corresponding to the immunodominant loop located at residues 135–158 on capsid protein VP1 of foot-and-mouth disease virus (FMDV) generally elicit high levels of anti-peptide and virus-neutralizing antibodies. In some instances, however, the level of neutralizing antibodies is low or even negligible, even though the level of anti-peptide antibodies is high. We have shown previously that the antigenic activity of peptide 141–159 of VP1 of a variant of serotype A can be mimicked by a retro-inverso (all-d retro or retroenantio) peptide analogue. This retro-inverso analogue induced greater and longer-lasting antibody titers than did the corresponding l-peptide. We now show that a single inoculation of the retro-inverso analogue elicits high levels of neutralizing antibodies that persist longer than those induced against the corresponding l-peptide and confer substantial protection in guinea pigs challenged with the cognate virus. In view of the high stability to proteases of retro-inverso peptide analogues and their enhanced immunogenicity, these results have practical relevance in designing potential peptide vaccines.