Regulation of nonphosphorylated and phosphorylated forms of neurofilament proteins in the prefrontal cortex of human opioid addicts.

The neurofilament (NF) proteins (NF-H, NF-M, and NF-L for high, medium, and low molecular weights) play a crucial role in the organization of neuronal shape and function. In a preliminary study, the abundance of total NF-L was shown to be decreased in brains of opioid addicts. Because of the potential relevance of NF abnormalities in opioid addiction, we quantitated nonphosphorylated and phosphorylated NF in postmortem brains from 12 well-defined opioid abusers who had died of an opiate overdose (heroin or methadone). Levels of NF were assessed by immunoblotting techniques using phospho-independent and phospho-dependent antibodies, and the relative (% changes in immunoreactivity) and absolute (changes in ng NF/microg total protein) amounts of NF were calculated. Decreased levels of nonphosphorylated NF-H (42-32%), NF-M (14-9%) and NF-L (30-29%) were found in the prefrontal cortex of opioid addicts compared with sex, age, and postmortem delay-matched controls. In contrast, increased levels of phosphorylated NF-H (58-41%) and NF-M (56-28%) were found in the same brains of opioid addicts. The ratio of phosphorylated to nonphosphorylated NF-H in opioid addicts (3.4) was greater than that in control subjects (1.6). In the same brains of opioid addicts, the levels of protein phosphatase of the type 2A were found unchanged, which indicated that the hyperphosphorylation of NF-H is not the result of a reduced dephosphorylation process. The immunodensities of GFAP (the specific glial cytoskeletol protein), alpha-internexin (a neuronal filament related to NF-L) and synaptophysin (a synapse-specific protein) were found unchanged, suggesting a lack of gross changes in glial reaction, other intermediate filaments of the neuronal cytoskeletol, and synaptic density in the prefrontal cortex of opioid addicts. These marked reductions in total NF proteins and the aberrant hyperphosphorylation of NF-H in brains of opioid addicts may play a significant role in the cellular mechanisms of opioid addiction.

Statistical discrimination and employers' recruitment : Practices for low-skilled workers

This paper deals with the recruitment strategies of employers in the low-skilled segment of the labour market. We focus on low-skilled workers because they are overrepresented among jobless people and constitute the bulk of the clientele included in various activation and labour market programmes. A better understanding of the constraints and opportunities of interventions in this labour market segment may help improve their quality and effectiveness. On the basis of qualitative interviews with 41 employers in six European countries, we find that the traditional signals known to be used as statistical discrimination devices (old age, immigrant status and unemployment) play a somewhat reduced role, since these profiles are overrepresented among applicants for low skill positions. However, we find that other signals, mostly considered to be indicators of motivation, have a bigger impact in the selection process. These tend to concern the channel through which the contact with a prospective candidate is made. Unsolicited applications and recommendations from already employed workers emit a positive signal, whereas the fact of being referred by the public employment office is associated with the likelihood of lower motivation.

Acidez Potencial Estimada pelo Método do pH SMP em Solos do Estado da Paraíba

O método do pH SMP, em virtude de sua praticidade analítica, tem sido amplamente utilizado na estimativa da acidez potencial (H+Al), em substituição à solução de acetato de cálcio 0,5 mol L-1 a pH 7,0, em laboratórios de análise de solo no Brasil. No entanto, tornam-se imprescindíveis estudos regionalizados para estimar com segurança os valores da acidez potencial. Este trabalho objetivou ajustar uma equação de regressão com boa capacidade preditiva dos valores de H+Al, a partir da medida do pH(SMP), para solos do Estado da Paraíba, visando sua utilização em análises de rotina para estimar a acidez potencial. Foram utilizadas 90 amostras de solo coletadas nas várias regiões do Estado. Essas amostras foram analisadas para pH(H2O), pH(SMP), H+Al e pH do sobrenadante da solução de acetato de cálcio. Foram observadas estreitas correlações quadrática simples do pH de equilíbrio da suspensão de acetato de cálcio com a acidez potencial e linear simples com o pH(SMP), com elevada capacidade preditiva (R2 = 0,93 e R2 = 0,95, respectivamente). Os resultados permitiram concluir que a acidez potencial (H+Al), expressa em cmolc dm-3 pode ser estimada pelo método do pH(SMP), por meio da equação H+Al = 57,108 -13,338** SMP + 0,7637** SMP2 (R2 = 0,87).

Assessment of driving capability through the use of clinical and psychomotor tests in relation to blood cannabinoids levels following oral administration of 20 mg dronabinol or of a cannabis decoction made with 20 or 60 mg Delta9-THC.

Delta(9)-Tetrahydrocannabinol (THC) is frequently found in the blood of drivers suspected of driving under the influence of cannabis or involved in traffic crashes. The present study used a double-blind crossover design to compare the effects of medium (16.5 mg THC) and high doses (45.7 mg THC) of hemp milk decoctions or of a medium dose of dronabinol (20 mg synthetic THC, Marinol on several skills required for safe driving. Forensic interpretation of cannabinoids blood concentrations were attempted using the models proposed by Daldrup (cannabis influencing factor or CIF) and Huestis and coworkers. First, the time concentration-profiles of THC, 11-hydroxy-Delta(9)-tetrahydrocannabinol (11-OH-THC) (active metabolite of THC), and 11-nor-9-carboxy-Delta(9)-tetrahydrocannabinol (THCCOOH) in whole blood were determined by gas chromatography-mass spectrometry-negative ion chemical ionization. Compared to smoking studies, relatively low concentrations were measured in blood. The highest mean THC concentration (8.4 ng/mL) was achieved 1 h after ingestion of the strongest decoction. Mean maximum 11-OH-THC level (12.3 ng/mL) slightly exceeded that of THC. THCCOOH reached its highest mean concentration (66.2 ng/mL) 2.5-5.5 h after intake. Individual blood levels showed considerable intersubject variability. The willingness to drive was influenced by the importance of the requested task. Under significant cannabinoids influence, the participants refused to drive when they were asked whether they would agree to accomplish several unimportant tasks, (e.g., driving a friend to a party). Most of the participants reported a significant feeling of intoxication and did not appreciate the effects, notably those felt after drinking the strongest decoction. Road sign and tracking testing revealed obvious and statistically significant differences between placebo and treatments. A marked impairment was detected after ingestion of the strongest decoction. A CIF value, which relies on the molar ratio of main active to inactive cannabinoids, greater than 10 was found to correlate with a strong feeling of intoxication. It also matched with a significant decrease in the willingness to drive, and it matched also with a significant impairment in tracking performances. The mathematic model II proposed by Huestis et al. (1992) provided at best a rough estimate of the time of oral administration with 27% of actual values being out of range of the 95% confidence interval. The sum of THC and 11-OH-THC blood concentrations provided a better estimate of impairment than THC alone. This controlled clinical study points out the negative influence on fitness to drive after medium or high dose oral THC or dronabinol.

Proteomic analysis of membrane rafts of melanoma cells identifies protein patterns characteristic of the tumor progression stage.

The molecular mechanisms controlling the progression of melanoma from a localized tumor to an invasive and metastatic disease are poorly understood. In the attempt to start defining a functional protein profile of melanoma progression, we have analyzed by LC-MS/MS the proteins associated with detergent resistant membranes (DRMs), which are enriched in cholesterol/sphingolipids-containing membrane rafts, of melanoma cell lines derived from tumors at different stages of progression. Since membrane rafts are involved in several biological processes, including signal transduction and protein trafficking, we hypothesized that the association of proteins with rafts can be regulated during melanoma development and affect protein function and disease progression. We have identified a total of 177 proteins in the DRMs of the cell lines examined. Among these, we have found groups of proteins preferentially associated with DRMs of either less malignant radial growth phase/vertical growth phase (VGP) cells, or aggressive VGP and metastatic cells suggesting that melanoma cells with different degrees of malignancy have different DRM profiles. Moreover, some proteins were found in DRMs of only some cell lines despite being expressed at similar levels in all the cell lines examined, suggesting the existence of mechanisms controlling their association with DRMs. We expect that understanding the mechanisms regulating DRM targeting and the activity of the proteins differentially associated with DRMs in relation to cell malignancy will help identify new molecular determinants of melanoma progression.

Muscle protein waste in tumor-bearing rats is effectively antagonized by a beta 2-adrenergic agonist (clenbuterol). Role of the ATP-ubiquitin-dependent proteolytic pathway.

Tissue protein hypercatabolism (TPH) is a most important feature in cancer cachexia, particularly with regard to the skeletal muscle. The rat ascites hepatoma Yoshida AH-130 is a very suitable model system for studying the mechanisms involved in the processes that lead to tissue depletion, since it induces in the host a rapid and progressive muscle waste mainly due to TPH (Tessitore, L., G. Bonelli, and F. M. Baccino. 1987. Biochem. J. 241:153-159). Detectable plasma levels of tumor necrosis factor-alpha associated with marked perturbations in the hormonal homeostasis have been shown to concur in forcing metabolism into a catabolic setting (Tessitore, L., P. Costelli, and F. M. Baccino. 1993. Br. J. Cancer. 67:15-23). The present study was directed to investigate if beta 2-adrenergic agonists, which are known to favor skeletal muscle hypertrophy, could effectively antagonize the enhanced muscle protein breakdown in this cancer cachexia model. One such agent, i.e., clenbuterol, indeed largely prevented skeletal muscle waste in AH-130-bearing rats by restoring protein degradative rates close to control values. This normalization of protein breakdown rates was achieved through a decrease of the hyperactivation of the ATP-ubiquitin-dependent proteolytic pathway, as previously demonstrated in our laboratory (Llovera, M., C. García-Martínez, N. Agell, M. Marzábal, F. J. López-Soriano, and J. M. Argilés. 1994. FEBS (Fed. Eur. Biochem. Soc.) Lett. 338:311-318). By contrast, the drug did not exert any measurable effect on various parenchymal organs, nor did it modify the plasma level of corticosterone and insulin, which were increased and decreased, respectively, in the tumor hosts. The present data give new insights into the mechanisms by which clenbuterol exerts its preventive effect on muscle protein waste and seem to warrant the implementation of experimental protocols involving the use of clenbuterol or alike drugs in the treatment of pathological states involving TPH, particularly in skeletal muscle and heart, such as in the present model of cancer cachexia.

Alterations of Neuromuscular Function after the World's Most Challenging Mountain Ultra-Marathon

We investigated the physiological consequences of the most challenging mountain ultra-marathon (MUM) in the world: a 330-km trail run with 24000 m of positive and negative elevation change. Neuromuscular fatigue (NMF) was assessed before (Pre-), during (Mid-) and after (Post-) the MUM in experienced ultra-marathon runners (n = 15; finish time = 122.43 hours +/-17.21 hours) and in Pre- and Post- in a control group with a similar level of sleep deprivation (n = 8). Blood markers of muscle inflammation and damage were analyzed at Pre- and Post-. Mean +/- SD maximal voluntary contraction force declined significantly at Mid- (-13+/-17% and -10+/-16%, P<0.05 for knee extensor, KE, and plantar flexor muscles, PF, respectively), and further decreased at Post- (-24+/-13% and -26+/-19%, P<0.01) with alteration of the central activation ratio (-24+/-24% and -28+/-34% between Pre- and Post-, P<0.05) in runners whereas these parameters did not change in the control group. Peripheral NMF markers such as 100 Hz doublet (KE: -18+/-18% and PF: -20+/-15%, P<0.01) and peak twitch (KE: -33+/-12%, P<0.001 and PF: -19+/-14%, P<0.01) were also altered in runners but not in controls. Post-MUM blood concentrations of creatine kinase (3719+/-3045 Ul.1), lactate dehydrogenase (1145+/-511 UI.L-1), C-Reactive Protein (13.1+/-7.5 mg.L-1) and myoglobin (449.3+/-338.2 microg.L-1) were higher (P<0.001) than at Pre- in runners but not in controls. Our findings revealed less neuromuscular fatigue, muscle damage and inflammation than in shorter MUMs. In conclusion, paradoxically, such extreme exercise seems to induce a relative muscle preservation process due likely to a protective anticipatory pacing strategy during the first half of MUM and sleep deprivation in the second half.

Muscle protein waste in tumor-bearing rats is effectively antagonized by a beta 2-adrenergic agonist (clenbuterol). Role of the ATP-ubiquitin-dependent proteolytic pathway.

Tissue protein hypercatabolism (TPH) is a most important feature in cancer cachexia, particularly with regard to the skeletal muscle. The rat ascites hepatoma Yoshida AH-130 is a very suitable model system for studying the mechanisms involved in the processes that lead to tissue depletion, since it induces in the host a rapid and progressive muscle waste mainly due to TPH (Tessitore, L., G. Bonelli, and F. M. Baccino. 1987. Biochem. J. 241:153-159). Detectable plasma levels of tumor necrosis factor-alpha associated with marked perturbations in the hormonal homeostasis have been shown to concur in forcing metabolism into a catabolic setting (Tessitore, L., P. Costelli, and F. M. Baccino. 1993. Br. J. Cancer. 67:15-23). The present study was directed to investigate if beta 2-adrenergic agonists, which are known to favor skeletal muscle hypertrophy, could effectively antagonize the enhanced muscle protein breakdown in this cancer cachexia model. One such agent, i.e., clenbuterol, indeed largely prevented skeletal muscle waste in AH-130-bearing rats by restoring protein degradative rates close to control values. This normalization of protein breakdown rates was achieved through a decrease of the hyperactivation of the ATP-ubiquitin-dependent proteolytic pathway, as previously demonstrated in our laboratory (Llovera, M., C. García-Martínez, N. Agell, M. Marzábal, F. J. López-Soriano, and J. M. Argilés. 1994. FEBS (Fed. Eur. Biochem. Soc.) Lett. 338:311-318). By contrast, the drug did not exert any measurable effect on various parenchymal organs, nor did it modify the plasma level of corticosterone and insulin, which were increased and decreased, respectively, in the tumor hosts. The present data give new insights into the mechanisms by which clenbuterol exerts its preventive effect on muscle protein waste and seem to warrant the implementation of experimental protocols involving the use of clenbuterol or alike drugs in the treatment of pathological states involving TPH, particularly in skeletal muscle and heart, such as in the present model of cancer cachexia.

Iowa Department of Transportation Major Items - Preliminary Quantity Report, February 19, 2013; With Addendums

Weekly letting report.

Iowa Department of Transportation Major Items - Preliminary Quantity Report, May 21, 2013; With Addendums

Weekly letting report.

Deficiency in monocarboxylate transporter 1 (MCT1) in mice delays regeneration of peripheral nerves following sciatic nerve crush.

Peripheral nerve regeneration following injury occurs spontaneously, but many of the processes require metabolic energy. The mechanism of energy supply to axons has not previously been determined. In the central nervous system, monocarboxylate transporter 1 (MCT1), expressed in oligodendroglia, is critical for supplying lactate or other energy metabolites to axons. In the current study, MCT1 is shown to localize within the peripheral nervous system to perineurial cells, dorsal root ganglion neurons, and Schwann cells by MCT1 immunofluorescence in wild-type mice and tdTomato fluorescence in MCT1 BAC reporter mice. To investigate whether MCT1 is necessary for peripheral nerve regeneration, sciatic nerves of MCT1 heterozygous null mice are crushed and peripheral nerve regeneration was quantified electrophysiologically and anatomically. Compound muscle action potential (CMAP) recovery is delayed from a median of 21days in wild-type mice to greater than 38days in MCT1 heterozygote null mice. In fact, half of the MCT1 heterozygote null mice have no recovery of CMAP at 42days, while all of the wild-type mice recovered. In addition, muscle fibers remain 40% more atrophic and neuromuscular junctions 40% more denervated at 42days post-crush in the MCT1 heterozygote null mice than wild-type mice. The delay in nerve regeneration is not only in motor axons, as the number of regenerated axons in the sural sensory nerve of MCT1 heterozygote null mice at 4weeks and tibial mixed sensory and motor nerve at 3weeks is also significantly reduced compared to wild-type mice. This delay in regeneration may be partly due to failed Schwann cell function, as there is reduced early phagocytosis of myelin debris and remyelination of axon segments. These data for the first time demonstrate that MCT1 is critical for regeneration of both sensory and motor axons in mice following sciatic nerve crush.

Suunnitelmatalous Neuvosto-Karjalassa 1928-1941 : paikallistason rooli Neuvostoliiton teollistamisessa

Summary: Planned economy in Soviet Karelia 1928-1941. The role of the local level in the industrialisation of the Soviet Union

Regulation of P-selectin glycoprotein ligand 1 (PSGL-1) interactions with selectins : role of the decameric repeats and of the cytoplasmic domain

SUMMARY BACKGROUND: P-selectin glycoprotein ligand 1 (PSGL-1) is a major selectin ligand, mediating leukocyte rolling along inflamed vascular wall. It is a mucin-like homodimer composed of a N-terminal domain which binds selectins, followed by 14-16 decameric repeats (DR), a transmembrane domain and a cytoplasmic tail, which may be involved in regulating leukocyte rolling and in generating intracellular signals, through its binding to moesin and Syk. P- and L-selectin binding is dependent on core-2 O-glycosylation and tyrosine sulfation of PSGL-1 N-terminus. However, a minor part of E-selectin-mediated rolling is dependent on N-terminal O-glycans; additional binding sites may thus be involved. In this project, we studied whether (1) PSGL-1 DR and (2) PSGL-1 cytoplasmic residues which bind moesin, were also involved in the regulation of selectin-dependent rolling. METHODS: Several mutated cDNAs were obtained: (1) PSGL-1 DR were either deleted, or substituted by platelet GPlba macroglycopeptide, (2) Ser-336, -348, Lys-337 and Arg-338 were mutated to alanine; moreover, truncation mutants retaining only 6 or 2 cytoplasmic residues were also generated. Transfected CHO expressing mutant PSGL-1 were tested for their ability to bind soluble selectin chimeras and to support selectin-dependent rolling under flow conditions. RESULTS: (1) Deletion of the DR had a dramatic effect on P- and L-selectin-dependent cell recruitment and rolling stability, which could only partially be compensated for, by GPlba substitution. In addition, we observed that DR create a binding site for E-selectin and thus support PSGL-1-dependent rolling. (2) Flow assays revealed that the moesin-binding site, in particular Ser-336, plays a crucial role in regulating the recruitment, velocity and rolling stability of PSGL-1-expressing cells on P- and L-selectin. CONCLUSIONS: Data presented here highlight the structure -function relationship of PSGL-1 DR. Moreover, they reveal a crucial role for the moesin-binding residues in regulating P-and L-selectin-dependent rolling. RÉSUMÉ CONTEXTE: PSGL-1 (P-selectin glycoprotein ligand 1) est un ligand majeur des sélectines permettant le roulement des leucocytes le long de la paroi vasculaire enflammée. C'est un homodimère de type mucine, composé d'un domaine N-terminal liant les sélectines, suivi de 14-16 répétitions décamèriques (RD), d'un domaine transmembranaire et d'une queue cytoplasmique qui pourrait être impliquée dans la régulation du roulement leucocytaire et la génération de signaux intracellulaires, via sa liaison à la moésine et à Syk. La liaison à la Pet à la L-sélectine dépend de la présentation par le N-terminus de PSGL-1 de O-glycans sur des structures core-2 et de tyrosines sulfatées. Cependant, une fraction mineure du roulement médié par la E-sélectine dépend des O-glycans N-terminaux; des sites de liaisons supplémentaires pourraient donc être impliqués. Dans ce projet, nous avons étudié si (1) les RD de PSGL-1 ainsi que (2) les résidus cytoplasmiques liant la moésine, étaient impliqués dans la régulation du roulement dépendant des sélectines. MÉTHODES: Plusieurs ADN codant des formes mutées de PSGL-1 ont été obtenus: (1) Les RD de PSGL-1 ont été soit ôtées, soit remplacées par le macroglycopeptide de la GPlba plaquettaire, (2) les Ser-336, -348, la Lys-337 et l'Arg-338 ont été mutées en alanine; par ailleurs, des mutants tronqués ne retenant plus que 6 ou 2 résidus cytoplasmiques ont également été générés. Des CHO transfectées exprimant PSGL-1 muté ont été testées pour leur capacité à lier des sélectines chimériques solubles et à soutenir un roulement dépendant des sélectines dans des conditions de flux. RÉSULTATS: (1) La perte des RD a eu un effet dramatique sur le recrutement cellulaire et la stabilité de roulement dépendant des P- et L-sélectine, qui n'a pu être que partiellement compensé par la substitution par la GPlba. De plus, nous avons observé que les RD forment un site de liaison pour la E-sélectine et soutiennent ainsi le roulement dépendant de PSGL-1. (2) Les tests de flux ont révélé que le site de liaison à la moésine, notamment la Ser-336, joue un rôle crucial dans la régulation du recrutement, de la vitesse et de la stabilité du roulement des cellules exprimant PSGL-1 sur les P- et L-sélectine. CONCLUSIONS; Les données présentées ici ont permis d'éclaircir la relation structure -fonction des RD de PSGL-1. Par ailleurs, elles révèlent un rôle crucial pour les résidus liant la moésine dans le roulement dépendant des P- et L-sélectine. RÉSUMÉ DESTINÉ À UN LARGE PUBLIC Pour accomplir ses fonctions, le sang circule sur un réseau de 96'000 kilomètres; ainsi, il approvisionne les cellules de l'organisme en énergie, il transporte diverses substances, il assure la défense contre les pathogènes et il participe à la régulation de la température corporelle. Le sang contient plusieurs types de cellules: la grande majorité sont les globules rouges, auxquels il faut ajouter les plaquettes (dont le rôle est de colmater les lésions vasculaires) et les globules blancs (leucocytes) qui, bien que présents en très faible quantité (moins de 0.01 %), jouent un rôle crucial en cas d'infection ou d'inflammation. Une attaque par un pathogène provoque plusieurs changements (rougeur, chaleur, gonflement, douleur), qui sont des manifestations de l'inflammation. Pour atteindre l'agent infectieux, des globules blancs spécialisés (les granulocytes) doivent quitter la circulation sanguine. Afin de faciliter leur capture, les vaisseaux sanguins vont exprimer des protéines telles que les sélectines, qui sont reconnues par une protéine leucocytaire appelée PSGL-1 (P-selectin glycoprotein ligand 7). L'interaction des sélectines avec PSGL-1 soutient le roulement du globule blanc le long de la paroi vasculaire, à une vitesse très inférieure à celle du flux sanguin. Ce roulement conduit à l'activation du globule blanc par des molécules de l'inflammation, permettant son adhésion ferme, puis son arrêt. Finalement, le granulocyte va migrer à travers la paroi du vaisseau pour atteindre et éliminer les causes de l'inflammation. L'adhésion est un processus intéressant à caractériser, car outre l'inflammation, il est également impliqué dans l'artériosclérose, l'infarctus, la métastatisation et la thrombose. Dans ce travail, nous nous sommes intéressés à définir les rôles des différents domaines de PSGL-1 dans la régulation de son interaction avec les sélectines. En effet, en plus de son extrémité extracellulaire de haute affinité pour les sélectines, PSGL-1 est composé de plusieurs séquences répétées hautement glycosylées et d'une courte région intracellulaire, dont les fonctions n'avaient pas été étudiées auparavant. En créant des formes mutées de PSGL-1, nous avons pu montrer qu'un roulement efficace des leucocytes nécessite la présence des régions répétitives et du domaine intracellulaire au complet.