Extracellular vesicles from human cardiac progenitor cells inhibit cardiomyocyte apoptosis and improve cardiac function after myocardial infarction.

AIMS: Recent evidence suggests that cardiac progenitor cells (CPCs) may improve cardiac function after injury. The underlying mechanisms are indirect, but their mediators remain unidentified. Exosomes and other secreted membrane vesicles, hereafter collectively referred to as extracellular vesicles (EVs), act as paracrine signalling mediators. Here, we report that EVs secreted by human CPCs are crucial cardioprotective agents. METHODS AND RESULTS: CPCs were derived from atrial appendage explants from patients who underwent heart valve surgery. CPC-conditioned medium (CM) inhibited apoptosis in mouse HL-1 cardiomyocytic cells, while enhancing tube formation in human umbilical vein endothelial cells. These effects were abrogated by depleting CM of EVs. They were reproduced by EVs secreted by CPCs, but not by those secreted by human dermal fibroblasts. Transmission electron microscopy and nanoparticle tracking analysis showed most EVs to be 30-90 nm in diameter, the size of exosomes, although smaller and larger vesicles were also present. MicroRNAs most highly enriched in EVs secreted by CPCs compared with fibroblasts included miR-210, miR-132, and miR-146a-3p. miR-210 down-regulated its known targets, ephrin A3 and PTP1b, inhibiting apoptosis in cardiomyocytic cells. miR-132 down-regulated its target, RasGAP-p120, enhancing tube formation in endothelial cells. Infarcted hearts injected with EVs from CPCs, but not from fibroblasts, exhibited less cardiomyocyte apoptosis, enhanced angiogenesis, and improved LV ejection fraction (0.8 ± 6.8 vs. -21.3 ± 4.5%; P < 0.05) compared with those injected with control medium. CONCLUSION: EVs are the active component of the paracrine secretion by human CPCs. As a cell-free approach, EVs could circumvent many of the limitations of cell transplantation.

Ultrastructural evidence of exosome secretion by progenitor cells in adult mouse myocardium and adult human cardiospheres.

The demonstration of beneficial effects of cell therapy despite the persistence of only few transplanted cells in vivo suggests secreted factors may be the active component of this treatment. This so-called paracrine hypothesis is supported by observations that culture media conditioned by progenitor cells contain growth factors that mediate proangiogenic and cytoprotective effects. Cardiac progenitor cells in semi-suspension culture form spherical clusters (cardiospheres) that deliver paracrine signals to neighboring cells. A key component of paracrine secretion is exosomes, membrane vesicles that are stored intracellularly in endosomal compartments and are secreted when these structures fuse with the cell plasma membrane. Exosomes have been identified as the active component of proangiogenic effects of bone marrow CD34(+) stem cells in mice and the regenerative effects of embryonic mesenchymal stem cells in infarcted hearts in pigs and mice. Here, we provide electron microscopic evidence of exosome secretion by progenitor cells in mouse myocardium and human cardiospheres. Exosomes are emerging as an attractive vector of paracrine signals delivered by progenitor cells. They can be stored as an "off-the-shelf" product. As such, exosomes have the potential for circumventing many of the limitations of viable cells for therapeutic applications in regenerative medicine.

Análisis de convergencia de las regiones de la zona euro (1990-2010)

Since the birth of Economic and Monetary Union (EMU), policies have been implemented in the Member States of the European Union (EU) to lead them towards economic convergence. This article analyses the convergence of the 174 regions that exist in the 17 euro-zone countries in the years from 1990 to 2010. The article specifies a space-time econometric model using the hypotheses of conditioned beta-convergence and sigmaconvergence. The dependent variables of the model are per capita GDP and productivity and the explanatory variables are real economic variables. Beta-convergence is found to exist between the countries of the euro-zone in terms of per capita GDP, but there is divergence in terms of productivity, though only at country level. In other words, the hypothesis is confirmed that it is guidelines at country level that lead to hypothetical convergence and that the unfavourable performance of productivity is due, without doubt, to differences in behaviour between the active populations of the different euro zone countries

Can a Halo CME from the limb be geoeffective?

The probability for a halo coronal mass ejection (CME) to be geoeffective is assumed to be higher the closer the CME launch site is located to the solar central meridian. However, events far from the central meridian may produce severe geomagnetic storms, like the case in April 2000. In this work, we study the possible geoeffectiveness of full halo CMEs with the source region situated at solar limb. For this task, we select all limb full halo (LFH) CMEs that occurred during solar cycle 23, and we search for signatures of geoeffectiveness between 1 and 5 days after the first appearance of each CME in the LASCO C2 field of view. When signatures of geomagnetic activity are observed in the selected time window, interplanetary data are carefully analyzed in order to look for the cause of the geomagnetic disturbance. Finally, a possible association between geoeffective interplanetary signatures and every LFH CME in solar cycle 23 is checked in order to decide on the CME's geoeffectiveness. After a detailed analysis of solar, interplanetary, and geomagnetic data, we conclude that of the 25 investigated events, there are only four geoeffective LFH CMEs, all coming from the west limb. The geoeffectiveness of these events seems to be moderate, turning to intense in two of them as a result of cumulative effects from previous mass ejections. We conclude that ejections from solar locations close to the west limb should be considered in space weather, at least as sources of moderate disturbances.

Nicotine-induced antinociception, rewarding effects and physical dependence are decreased in mice lacking the preproenkephalin gene

It has been shown previously that the endogenous opioid system may be involved in the behavioral effects of nicotine. In the present study, the participation of endogenous enkephalins on nicotine responses has been investigated by using preproenkephalin knock-out mice. Acute nicotine-induced hypolocomotion remained unaffected in these mice. In contrast, antinociception elicited in the tail-immersion and hot-plate tests by acute nicotine administration was reduced in mutant animals. The rewarding properties of nicotine were then investigated using the place-conditioning paradigm. Nicotine induced a conditioned place preference in wild-type animals, but this effect was absent in knock-out mice. Accordingly, in vivo microdialysis studies revealed that the enhancement in dopamine extracellular levels in the nucleus accumbens induced by nicotine was also reduced in preproenkephalin-deficient mice. Finally, the somatic expression of the nicotine withdrawal syndrome precipitated in nicotine-dependent mice by mecamylamine was significantly attenuated in mutant animals. In summary, the present results indicate that endogenous opioid peptides derived from preproenkephalin are involved in the antinociceptive and rewarding properties of nicotine and participate in the expression of physical nicotine dependence.

Long-term changes in synaptic transmission of the lateral amygdala induced by strong "in vitro" activation

Résumé : L'amygdale latérale (AL) joue un .rôle essentiel dans la plasticité synaptique à la base du conditionnement de la peur. Malgré le faite que la majorité des cellules de l'AL reçoivent les afférentes nécessaires, une potentialisation dans seulement une partie d'entre elles est obligatoire afin que l'apprentissage de la peur ait lieu. Il a été montré que ces cellules expriment la forme active de CREB, et celui-ci a été associé aux cellules dites de type 'nonaccomrnodating' (nAC). Très récemment, une étude a impliqué les circuits récurrents de l'AL dans le conditionnement de la peur. Un lien entre ces deux observations n'a toutefois jamais été établi. t Nous avons utilisé un protocole in vitro de forte activation de l'AL, résultant dans l'induction de 'bursts' provenant de l'hippocampe et se propageant jusqu'à l'AL. Dans l'AL ces 'bursts' atteignent toutes les cellules et se propagent à travers plusieurs chemins. Utilisant ce protocole, nous avons, pour la première fois pu associer dans l'AL, des cellules connectées de manière récurrente avec des cellules de type nAC. Aussi bien dans ces dernières que dans les cellules de type 'accommodating' (AC), une diminution dans la transmission inhibitrice, à la fois exprimée de manière pré synaptique mais également indépendant de la synthèse de protéine a pu être observé. Au contraire, une potentialisation induite et exprimée au niveau pré synaptique ainsi que dépendante de la synthèse de protéine a pu être trouvé uniquement dans les cellules de type nAC. De plus, une hyperexcitabilité, dépendante des récepteurs NMDA a pu être observé, avec une sélection préférentielle des cellules du type nAC dans la génération de bursts. Nous avons également pu démontrer que la transformation d'un certain nombre de cellules de type AC en cellules dites nAC accompagnait cette augmentation générale de l'excitabilité de l'AL. Du faite da la grande quantité d'indices suggérant une connexion entre le système noradrénergique et les états de peur/d'anxiété, les effets d'une forte activation de l'AL sur ce dernier ont été investigués et ont révélés une perte de sa capacité de modulation du 'spiking pattern'. Finalement, des changements au niveau de l'expression d'un certain nombre de gènes, incluant celui codant pour le BDNF, a pu être trouvé à la suite d'une forte activation de l'AL. En raison du lien récemment décrit entre l'expression de la forme active de CREB et des cellules de type nAC ainsi que celui de l'implication des cellules de l'AL connectés de manière récurrente dans l'apprentissage de la peur, nos résultats nous permettent de suggérer un modèle expliquant comment la potentialisation des connections récurrentes entre cellules de type nAC pourrait être à la base de leur recrutement sélectif pendant le conditionnement de la peur. De plus, ils peuvent offrir des indices par rapport aux mécanismes à travers lesquels une sous population de neurones peut être réactivée par une stimulation externe précédemment inefficace, et induire ainsi un signal suffisamment fort pour qu'il soit transmit aux structures efférentes de l'AL. Abstract : The lateral nucleus of the amygdala (LA) is critically involved in the plasticity underlying fear-conditioned learning (Sah et al., 2008). Even though the majority of cells in the LA receive the necessary sensory inputs, potentiation in only a subset is required for fear learning to occur (Repa et al., 2001; Rumpel et al., 2005). These cells express active CREB (CAMP-responsive element-binding protein) (Han et al., 200, and this was related to the non-accommodating (nAC) spiking phenotype (Viosca et al., 2009; Zhou et al., 2009). In addition, a very recent study implicated recurrently connected cells of the LA in fear conditioned learning (Johnson et al., 2008). A link between the two observations has however never been made. In rats, we used an in vitro protocol of strong activation of the LA, resulting in bursting activity, which spread from the hippocampus to the LA. Within the LA, this activity reached all cells and spread via a multitude of pathways. Using this model, we were able to link, for the first time, recurrently connected cells in the LA with cells of the nAC phenotype. While we found a presynaptically expressed, protein synthesis independent decrease in inhibitory synaptic transmission in both nAC and accommodating (AC) cells, only nAC cells underwent a presynaptically induced and expressed, protein synthesis dependent potentiation. Moreover we observed an NMDA dependent hyperexcitability of the LA, with a preferential selection of nAC cells into burst generation. The transformation of a subset of AC cells into nAC cells accompanied this general increase in LA excitability. Given the considerable evidence suggesting a relationship between the central noradrenergic (NA) system and fear/anxiety states (Itoi, 2008), the effects of strong activation of the LA on the noradrenergic system were investigated, which revealed a loss of its modulatory actions on cell spiking patterns. Finally, we found changes in the expression levels of a number of genes; among which the one coding for $DNF, to be induced by strong activation of the LA. In view of the recently described link between nAC cells and expression of pCREB (phosphorylated cAMP-responsive element-binding protein) as well as the involvement of recurrently connected cells of the LA in fear-conditioned learning, our findings may provide a model of how potentiation of recurrent connections between nAC neurons underlies their recruitment into the fear memory trace. Additionally, they may offer clues as to the mechanisms through which a selected subset of neurons can be reactivated by smaller, previously ineffective external stimulations to respond with a sufficiently strong signal, which can be transmitted to downstream targets of the LA.

Gene expression in cultured endometrium from women with different outcomes following IVF.

Estradiol and progesterone are crucial for the acquisition of receptivity and the change in transcriptional activity of target genes in the implantation window. The aim of this study was to differentiate the regulation of genes in the endometrium of patients with recurrent implantation failure (IF) versus those who became pregnant after in vitro fertilization (IVF) treatment. Moreover, the effect of embryo-derived factors on endometrial transcriptional activity was studied. Nine women with known IVF outcome (IF, M, miscarriage, OP, ongoing pregnancy) and undergoing hysteroscopy with endometrial biopsy were enrolled. Biopsies were taken during the midluteal phase. After culture in the presence of embryo-conditioned IVF media, total RNA was extracted and submitted to reverse transcription, target cDNA synthesis, biotin labelling, fragmentation and hybridization using the Affymetrix Human Genome U133A 2.0 Chip. Differential expression of selected genes was re-analysed by quantitative PCR, in which the results were calculated as threshold cycle differences between the groups and normalized to Glyceraldehyde phosphate dehydrogenase and beta-actin. Differences were seen for several genes from endometrial tissue between the IF and the pregnancy groups, and when comparing OP with M, 1875 up- and 1807 down-regulated genes were returned. Real-time PCR analysis confirmed up-regulation for somatostatin, PLAP-2, mucin 4 and CD163, and down-regulation of glycodelin, IL-24, CD69, leukaemia inhibitory factor and prolactin receptor between Op and M. When the different embryo-conditioned media were compared, no significant differential regulation could be demonstrated. Although microarray profiling may currently not be sensitive enough for studying the effects of embryo-derived factors on the endometrium, the observed differences in gene expression between M and OP suggest that it will become an interesting tool for the identification of fertility-relevant markers produced by the endometrium.

Differentiation towards regulatory DC is maintained during RSV infection in airway epithelial cells

Airway epithelial cells have been shown to drive differentiation of monocytes into dendritic cells (DC) with suppressive phenotype. In this study we investigated the impact of virus-induced inflammatory mediator production on DC development. Monocyte differentiation into functional DC, as reflected by the expression of CD11c, CD123, BDCA-4 and DC-SIGN and the capacity to activate T cells, was similar for respiratory syncytial virus (RSV)- and mock-infected BEAS-2B and A549 cells. RSV-conditioned culture media resulted in a partially mature DC phenotype, but failed to upregulate CD80, CD83, CD86 and CCR7 and failed to release pro-inflammatory mediators upon TLR triggering. Nevertheless, these DCs were able to maintain an antiviral response by the release of type I IFN. Collectively, these data indicate that the airway epithelium maintains an important suppressive DC phenotype under inflammatory conditions induced by RSV infection.

Prevention of cutaneous melanoma: an epidemiological evaluation of the Swiss campaign.

A national information program, focusing on the main recognized risk factors (primary prevention) and on the potential benefits of early detection (secondary prevention) of cutaneous malignant melanoma, was launched in Switzerland in May 1988. The first campaign, based on a pilot study conducted in 1986 in the canton of Basel, was followed by a recall campaign in July 1989. This report describes the organization of this program and presents an assessment of its initial impact. The number of newly diagnosed cases increased more than twofold (+ 116%) in the two months following the launch of the first campaign (May to June 1988). This trend was accompanied by a statistically significant shift of case distribution towards younger ages (< 60 years; p = 0.003), and a non-significant shift was observed towards less advanced lesions (thickness < or = 1.5 mm). The incidence decreased quickly, though in the twelve month period between the two campaigns it remained 21% higher than before the inception of the program. No appreciable effects were detected from the recall campaign and no difference was seen among regions or between sexes.

Extracellular microvesicles from astrocytes contain functional glutamate transporters: regulation by protein kinase C and cell activation.

Glutamate transport through astrocytic excitatory amino-acid transporters (EAAT)-1 and EAAT-2 is paramount for neural homeostasis. EAAT-1 has been reported in secreted extracellular microvesicles (eMV, such as exosomes) and because the protein kinase C (PKC) family controls the sub-cellular distribution of EAATs, we have explored whether PKCs drive EAATs into eMV. Using rat primary astrocytes, confocal immunofluorescence and ultracentrifugation on sucrose gradient we here report that PKC activation by phorbol myristate acetate (PMA) reorganizes EAAT-1 distribution and reduces functional [(3)H]-aspartate reuptake. Western-blots show that EAAT-1 is present in eMV from astrocyte conditioned medium, together with NaK ATPase and glutamine synthetase all being further increased after PMA treatment. However, nanoparticle tracking analysis reveals that PKC activation did not change particle concentration. Functional analysis indicates that eMV have the capacity to reuptake [(3)H]-aspartate. In vivo, we demonstrate that spinal astrocytic reaction induced by peripheral nerve lesion (spared nerve injury, SNI) is associated with a phosphorylation of PKC δ together with a shift of EAAT distribution ipsilaterally. Ex vivo, spinal explants from SNI rats release eMV with an increased content of NaK ATPase, EAAT-1 and EAAT-2. These data indicate PKC and cell activation as important regulators of EAAT-1 incorporation in eMV, and raise the possibility that microvesicular EAAT-1 may exert extracellular functions. Beyond a putative role in neuropathic pain, this phenomenon may be important for understanding neural homeostasis and a wide range of neurological diseases associated with astrocytic reaction as well as non-neurological diseases linked to eMV release.

Lysine and arginine requirements of Salminus brasiliensis

The objective of this work was to determine the dietary lysine (DL) and dietary arginine (DA) requirements of dourado (Salminus brasiliensis), through dose-response trials using the amino acid profiles of whole carcasses as a reference. Two experiments were carried out in a completely randomized design (n=4). In the first experiment, groups of 12 feed-conditioned dourado juveniles (11.4±0.2 g) were stocked in 60 L cages placed in 300 L plastic indoor tanks in a closed circulation system. Fish were fed for 60 days on diets containing 1.0, 1.5, 2.0, 2.5, 3.0, or 3.5 % dietary lysine. In the second experiment, dourado juveniles (27.0±0.8 g) were fed for 60 days on semipurified diets containing arginine at 1.0, 1.5, 2.0, 2.5 or 3.0%, in similar conditions to those of the first experiment. Optimal DL requirements, as determined by broken-line analysis method for final weight, weight gain and specific growth rate, were 2.15% DL or 5% lysine in dietary protein, and 1.48% DA or 3.43% arginine in dietary protein. The best feed conversion ratio is attained with 2.5% DL or 5.8% lysine in dietary protein and 1.4% DA or 3.25% arginine in dietary protein.

System identification using a linear combination of cumulant slices

In this paper we develop a new linear approach to identify the parameters of a moving average (MA) model from the statistics of the output. First, we show that, under some constraints, the impulse response of the system can be expressed as a linear combination of cumulant slices. Then, thisresult is used to obtain a new well-conditioned linear methodto estimate the MA parameters of a non-Gaussian process. Theproposed method presents several important differences withexisting linear approaches. The linear combination of slices usedto compute the MA parameters can be constructed from dif-ferent sets of cumulants of different orders, providing a generalframework where all the statistics can be combined. Further-more, it is not necessary to use second-order statistics (the autocorrelation slice), and therefore the proposed algorithm stillprovides consistent estimates in the presence of colored Gaussian noise. Another advantage of the method is that while mostlinear methods developed so far give totally erroneous estimates if the order is overestimated, the proposed approach doesnot require a previous estimation of the filter order. The simulation results confirm the good numerical conditioning of thealgorithm and the improvement in performance with respect to existing methods.

Market entry order and competitive advantage of the firm

A firm that wishes to launch a new product to the market is faced with a difficult task of deciding what the best moment for the launch is. Timing may also be critical when a firm plans to adopt new processes or intends to head for new markets. The critical question the firm needs to tackle is whether it will try to reach the so-called first-mover advantage by acting earlier than its rivals. The first-mover position may reward the entrant with various opportunities to gain competitive advantage over later movers. However, there are also great risks involved in the early market entry, and sometimes the very first entrant fails even before the followers enter the market. The follower, on the other hand, may be able to free-ride on the earlier entrants' investments and gain from the languished uncertainties that characterize the new markets. According to the current understanding the occurrence of entry order advantages depends not only on the mechanism and attributes in the firm's environment that provide the initial opportunities but also on the firm's ability to capitalize on these advantage opportunities. This study contributes to this discussion by analyzing the linkages between the asset base of the firm, characteristics of the operating environment and the firm's entry timing orientation. To shed light on the relationship between the entry timing strategy and competitive advantage, this study utilizes the concept of entry timing orientation. The rationale for choosing this type of approach arises from the inability of previously employed research tools to reach the underlying factors that result in entry timing advantage. The work consists of an introductory theoretical discussion on entry timing advantages and of four research publication. The empirical findings support the understanding that entry timing advantage is related to the characteristics of the firm's operating environment but may also be related to firm-specific factors. This in turn suggests that some of the traditional ways of detecting and measuring first-mover advantage - which to some extent ignore these dimensions - may be outdated.

Els suburbia al Mediterrani de la Hispania Citerior, o els termòmetres de la salut de l’Imperi

Reflexión sobre los suburbios costeros de las principales ciudades de la provincia Hispania Citerior en base a criterios de dinamismo económico, concebidos como la causa principal de la aparición de las áreas periurbanas. Los procesos expansivos o regresivos se interrelacionan con la vitalidad de las actividades portuarias, determinantes en el crecimiento periurbano, o en la génesis de núcleos alejados de las ciudades ubicadas a escasa distancia de la costa. Este documento intenta aportar los rasgos comunes propios de la uniformidad urbanística del período altoimperial, y que desaparecieron en la antigüedad tardía como consecuencia de los procesos específicos de cada núcleo urbano, condicionado por la viabilidad de su modelo económico y su rol geoestratégico. De este modo, el análisis de las zonas suburbiales se manifiesta como el principal índice de prosperidad económica y urbanística. No en vano, estas mantuvieron su funcionalidad en época tardía, mientras la ciudad altoimperial fue desestructurándose.

Paperikoneen käyttöönoton onnistumisen riippuvuus valmistelutyöstä

Paperikoneinvestointi toteutetaan projekteina, joista muodostuvaa ketjua kutsutaan projektien elinkaareksi. Toteutusketjun viimeinen lenkki on käynnistykseen huipentuva käyttöönotto. Hyvä käyttöönotto palvelee investointia. Käyttöönotoissa koetaan erilaisia ongelmatilanteita, joista osa on satunnaisesti tai usein toistuvia. Investoinnin onnistumisen ja aikataulun asettama paine luo tilanteiden selvittämiseen erityishaasteita. Aina ei selvitä ilman takaiskuja. Tutkimustavoitteena oli kartoittaa käyttöönoton yleiset ongelmat ja niiden yhteydet projektihistoriaan, sekä ne projektinhallinnan osa-alueet, joita parantamalla varmistetaan käyttöönoton ja investoinnin onnistuminen. Tutkimus pohjautuu erilaisistainvestointiprojekteista ja käyttöönotoista saatuihin kokemuksiin, haastatteluihin (3 kpl) ja kyselypalautteeseen (42 kpl). Investoinnin onnistumisedellytykset luodaan projektihistoriassa, käyttöönotossa ne viimeistellään. Onnistuminen mitataan tuotto-odotusten saavuttamisena aikataulussa, johon vaikuttavia tekijöitä ovat tuotannon hallinta, käyttövarmuus ja markkinat. Käyttöönoton onnistumista tulee arvioida käyttöönottotehtävistäsuoriutumisen pohjalta, ei pelkästään investoinnille asetettujen aikataulu- ja tuotantotavoitteiden (laatu, määrä, hallinta) saavuttamisena, kuten usein tapahtuu. Tulosten perusteella käyttöönoton merkittävimmät ongelmat ovat tiedonkulun puutteet, ohjelmallisten korjausten suuri määrä ja palautumisajan riittämättömyys. Tärkeimmät painotukset ovat laiterikkojen estäminen, ohjelmavirheiden korjaaminen ja henkilöstön osaamisen varmentaminen. Konelinjan vaikeimmin hallittava osa on radan päänvienti. Osapuolten poikkeavat näkökulmat sekä työn laadun merkitys testauksissa, koulutuksissa ja kenttätoiminnoissa nousee tuloksista myös vahvasti esiin. Tutkimus on selvittänyt sille asetetut tavoitteet. Käyttöönoton merkittävimmät ongelmat syntyvät sen lähihistoriassa: testauksissa, koulutuksessa ja kenttätoiminnoissa. Käyttöönoton onnistumiseksi tulee painotusprojektinhallinnassa keskittää käynnistysvaiheen suunnitteluun sekä kenttätoimintojen hallintaan.