Top PDF Cytomegalovirus replicon-based regulation of gene expression in vitro and in vivo.

Cytomegalovirus replicon-based regulation of gene expression in vitro and in vivo.

Cytomegalovirus replicon-based regulation of gene expression in vitro and in vivo.

At first glance, the presence of both a viral oriLyt and transgenes appears similar to an amplicon vector construct, as the amplicon also contains both elements and, in addition, a pac signal. However, the oriLyt within amplicon vectors serves only to amplify the vector DNA for efficient packaging of a high number of vector copies into helper virus capsids via the pac signal [14]. Interestingly, the phenomenon of transgene silencing of amplicon vectors by HDAC-dependent mecha- nisms has also been described [55]. Whether or not the context of the oriLyt itself affects the level of gene expression has, to our knowledge, not been investigated. Neither has the question of oriLyt-specific de-silencing of the vector by super-infection of cells carrying the vector been addressed. Taking the results of our study into account, we expect that expression of transgenes provided by amplicons derived from human pathogens would be affected by natural super-infection. This prediction is testable using gene therapy studies incorporating herpesvirus amplicon vectors. The strength of this effect would probably depend on details such as transcription orientation and distance from the oriLyt sequence. The replicon vector system for inducible gene expression should be applicable to all herpesvirus subfamilies. Moreover, most DNA viruses regulate late gene expression upon DNA replication, although it is achieved via different mechanisms. Such replicon vector systems may be applicable for polyomaviruses [56] and adenoviruses [57] with minor modifications. Notably, there are suggestions of a connection between the origins of replication and enhanced gene expression in higher eukaryotic genomes [58,59]. Trans-complementation of late proteins involved in herpesvirus morphogenesis is still a difficult task. Incorrect timing, aberrant intracellular distribution due to missing viral interaction partners and incorrect expression levels of the viral protein may explain poor complementation results. For instance, the isolated expres- sion of late herpesvirus promoters without providing an oriLyt in cis results in aberrant early expression of the transgene. Today, systems for inducible gene expression typically require the use of small chemical compounds such as tetracyclin or doxycyclin (as in the case of Tet-on/Tet-off systems) [60] or rapamycin (for FKBP12-based systems) [61]. In these systems, gene expression is activated synchronously and irrespective of the state of virus replication in all cells, whereas the oriLyt-based system uses viral DNA replication as the signal for induction. The expression of the gene increases in proportion with the amplification of the vector DNA and reflects the natural expression kinetics of late herpesvirus genes. As each cell is activated individually upon infection, the appropriate and correct timing of expression of the trans-complementing gene is determined by the infecting virus.
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Regulation and gene expression profiling of NKG2D positive human cytomegalovirus-primed CD4+ T-cells.

Regulation and gene expression profiling of NKG2D positive human cytomegalovirus-primed CD4+ T-cells.

The ligands for human NKG2D include MICA, MICB, ULBP1-3, RAET1E, RAET1G, and RAET1L [9,10]. NKG2D- ligands are either absent or expressed at low levels on healthy cells, but can be induced by cellular stress caused by tumor transfor- mation or infection [10,11]. The NKG2D/NKG2D-ligand system is frequently used by the immune system to recognize danger signals [10,11]. However, the system must be kept under strict control to avoid aberrant killing and tissue damage. Anomalous stimulation of the NKG2D/NKG2D-ligand system has been implicated in various autoimmune diseases, including crohn’s disease, celiac disease and rheumatoid arthritis [3,5,12]. In
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SOX9 governs differentiation stage-specific gene expression in growth plate chondrocytes via direct concomitant transactivation and repression.

SOX9 governs differentiation stage-specific gene expression in growth plate chondrocytes via direct concomitant transactivation and repression.

Cartilage and endochondral bone development require SOX9 activity to regulate chondrogenesis, chondrocyte proliferation, and transition to a non-mitotic hypertrophic state. The restricted and reciprocal expression of the collagen X gene, Col10a1, in hypertrophic chondrocytes and Sox9 in immature chondrocytes epitomise the precise spatiotemporal control of gene expression as chondrocytes progress through phases of differentiation, but how this is achieved is not clear. Here, we have identified a regulatory element upstream of Col10a1 that enhances its expression in hypertrophic chondrocytes in vivo. In immature chondrocytes, where Col10a1 is not expressed, SOX9 interacts with a conserved sequence within this element that is analogous to that within the intronic enhancer of the collagen II gene Col2a1, the known transactivation target of SOX9. By analysing a series of Col10a1 reporter genes in transgenic mice, we show that the SOX9 binding consensus in this element is required to repress expression of the transgene in non-hypertrophic chondrocytes. Forced ectopic Sox9 expression in hypertrophic chondrocytes in vitro and in mice resulted in down-regulation of Col10a1. Mutation of a binding consensus motif for GLI transcription factors, which are the effectors of Indian hedgehog signaling, close to the SOX9 site in the Col10a1 regulatory element, also derepressed transgene expression in non-hypertrophic chondrocytes. GLI2 and GLI3 bound to the Col10a1 regulatory element but not to the enhancer of Col2a1. In addition to Col10a1, paired SOX9–GLI binding motifs are present in the conserved non-coding regions of several genes that are preferentially expressed in hypertrophic chondrocytes and the occurrence of pairing is unlikely to be by chance. We propose a regulatory paradigm whereby direct concomitant positive and negative transcriptional control by SOX9 ensures differentiation phase-specific gene expression in chondrocytes. Discrimination between these opposing modes of transcriptional control by SOX9 may be mediated by cooperation with different partners such as GLI factors.
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Transcriptional and epigenetic regulation of KIAA1199 gene expression in human breast cancer.

Transcriptional and epigenetic regulation of KIAA1199 gene expression in human breast cancer.

To identify putative transcription factor-binding sites within the KIAA1199 promoter, we employed two programs (MatInspector and Alibaba2) to predict transcription factor binding sites based on DNA sequence. This analysis revealed transcriptional elements in the KIAA1199 promoter, including AP-1, NF-kB, and Twist-1, which are situated in the activator region (pro-1.4) (Fig. 4). AP-1 binding sequence (GAGT) that is located between 248 and 245 and its flanking regions on both sides are highly conserved (100%) between human and mouse. Although four putative NF-kB transcription binding sites were recognized, the first three putative NF-kB binding sites located in the proximal part of the promoter (2246/2234, 2324/2312, and 2704/2715), may not be involved in activation of the KIAA1199 promoter based on the finding that deletion mutants containing these three NF-kB binding sites did not affect transcriptional activity of the KIAA1199 promoter (Fig. 3B). In contrast, the deletion mutant encompassing the fourth putative NF-kB binding site (21345 and 21333) lost more than 50% transcriptional activity (Fig. 3B). In addition to AP-1 and NF-kB binding sites, two putative Twist-1 binding sites (2269/2249 and 2923/2903) were also identified. Deletion of Figure 1. Elevated expression of KIAA1199 in human cancers. A) By mining Oncomine and GEO databases, KIAA1199 expression pattern in more than 40 microarray data sets shows significant alteration (P,0.01). Representative data are presented. High KIAA1199 expression in various human cancers.1-breast cancer n:27, normal n:7, p = 6.97E-4 [41]; 2-colon cancer n:36, normal n:24, p = 1.7E-4[42]; 3-gastric cancer n:26, normal n:31, p = 3.69E-13[43]; 4-lung cancer n:45, normal n:65, p = 8.59E-9[44]; 5-head&neck cancer n:41, normal n:13, p = 2.35E-7[45]; 6-ovarian cancer n:6, normal n:4, p = 5.92E-4[46]; 7-pancreatic cancer n:11, normal n:11, p = 0.001 [47]. B) Expression of KIAA1199 in human cancer cell lines: Human prostate (LNCaP and Du145), and breast cancer (MCF-7 and MDA-MB-231) cell lines were examined by real time RT-PCR using KIAA1199 specific primers. The expression of KIAA1199 was normalized by house-keeping genes (HPRT-1 and GAPDH). The relative levels of genes were determined using the DDCt method. Each bar represents the mean 6 S.E (*,0.05).
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Overexpression of SMPX in adult skeletal muscle does not change skeletal muscle fiber type or size.

Overexpression of SMPX in adult skeletal muscle does not change skeletal muscle fiber type or size.

Transfected HEK293 cells were washed twice with ice cold PBS and lysed with lysis buffer A: (50 mM tris acetate, 0.27 M sucrose, 1 mM EDTA, 1 mM EGTA, 1 mM sodium orthovanadate, 10 mM a-glycerophosphate, 50 mM sodium fluoride, 5 mM sodium pyrophosphate, 1% (w/v) Triton X100, 0.2 mM PMSF, 1 mM benzamidine and 0.1% (v/v) b-mercaptoethanol). The lysates were then centrifuged at 4 uC and 13000G for 20 minutes and the supernatant saved. Protein concentration was then measured at 595 nm using a Bradford assay on Victor 2 (Wallac). 30 m g of protein was run on NuPAGE Novex Bis-Tris 4–12% Gels (Invitrogen), followed by Western blotting (NuPAGE Western Transfer Protocol, Invitrogen). Blots were developed using the ECL Western Blotting Detection kit (Amersham) according to the manufacturer’s instructions. A mouse anti-GFP antibody (Invitro- gen) was used to detect SMPX-EGFP and EGFP (Figure 1A). Expression of pCMS-EGFP-Smpx was verified with northern blotting on HEK-293 cells (data not shown)
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Gene expression dosage regulation in an allopolyploid fish.

Gene expression dosage regulation in an allopolyploid fish.

Interestingly, the results of the available studies on plants and invertebrates are not all coin- cident. Several showed that the majority of genes are expressed additively [48, 49], while other studies found higher levels of nonadditive expression [13, 44]. The causes for these apparent discrepancies are not yet clear [50]. Anyway, the considerable body of data gathered so far shows a possible positive correlation between size of the fraction of the nonadditively expressed genes and the magnitude of heterotic response. Also, increasing the number of diverse genome copies in an allopolyploid, usually leads to increasingly greater magnitudes of heterosis [51]. Nevertheless, there is no consensus about the amount or identity of the nonadditively express- ed genes [50, 52]. Concerning the S. alburnoides complex, the phenomenon of heterosis has been barely addressed, except for a few comparisons of growth and reproductive traits between diploid and triploid hybrids [53] and a comparative morphometric study [36], where the AA and PP parental genomotypes were not included. From these studies, mostly non-significant differences between diploid and triploid hybrids have been found, except for a marginal lon- gevity increase in triploids. Yet, the PAA genomotype is far more frequent in the natural popu- lations than PA. So, if we consider the number of non-additively expressed transcripts as an indicator of heterosis, PAA S. alburnoides are favored since the amount of additively expressed transcripts is higher than in PA genomotype. Also, according the Bateson-Dobzhansky-Muller Model a lower fitness in hybrids might result from a bad interaction between divergent ge- nomes due to the differential capacity of interaction between their proteins [54]. In this light, allopolyploid individuals have better chances to evade this weakness. Allopolyploids have more options to non-additively combine allele-specific regulated expressions and so, have higher chances to achieve an optimized and more functional expression pattern than one achieved merely additively.
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Enhancing T3 and cAMP responsive gene participation in the thermogenic regulation of fuel oxidation pathways

Enhancing T3 and cAMP responsive gene participation in the thermogenic regulation of fuel oxidation pathways

Objetivo: Identiicar enzimas das vias da glicólise, glicogenólise, lipólise, ciclo de Krebs, cadeia respiratória e fosforilação oxidativa possivelmente reguladas por T3 e cAMP. Materiais e méto- dos: Analisamos 56 genes metabólicos in silico mediante a identiicação dos elementos cis de regulação gênica responsivos ao T3 e cAMP (TREs, thyroid response elements e CREs, cAMP response elements), utilizando como referência o promotor da UCP1, SERCA2 e gliceraldeído 3-fosfato desidrogenase. Selecionamos somente os promotores com estudo funcional prévio in vitro. Resultados: 29/56 enzimas apresentaram TREs em suas regiões regulatórias, parte com escore ≥ 0,80 (melhor valor preditivo 1): citrato sintase, fosfoglucose isomerase, succinato de- sidrogenase A/C, UCP3, UCP2, UCP4, UCP5, fosfoglicerato mutase, gliceraldeído 3-P desidro- genase, glucoquinase, malato desidrogenase, acil-CoA transferase (tiolase), citocromo a3, e lactato desidrogenase; parte desconhecida como regulada por T3. Conclusão: Os resultados do presente estudo apontam para novos genes regulados por T3 e cAMP e, por conseguinte, sua contribuição na regulação da termogênese. Arq Bras Endocrinol Metab. 2010;54(4):381-9
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Braz. Dent. J.  vol.22 número5

Braz. Dent. J. vol.22 número5

O mixoma odontogênico (MO) é um tumor odontogênico benigno de origem mesenquimal caracterizado pela presença de células fusiformes ou estreladas dispostas em abundante matriz extracelular mucóide. A metilação do DNA é caracterizada pela adição de grupos metil em citosinas constituintes de ilhas CpG na região promotora do gene. A metilação pode diminuir a expressão de genes supressores de tumor e contribuir para o desenvolvimento de lesões neoplásicas. O objetivo deste trabalho foi avaliar o padrão de metilação nos genes P16 (CDKN2A), P21 (CDKN1A), P27 (CDKN1B), P53 (TP53), RB1 nos MO e na polpa dental. A metilação foi avaliada pela reação em cadeia da polimerase específica para a metilação. A transcrição dos genes foi estudada em alguns casos pela reação da transcriptase reversa (PCR quantitativa). Uma maior frequência de amostras não metiladas para os genes P27, P53 e RB1 foi observada nos MO quando comparados à polpa dental. Os MO expressaram RNAm de todos os genes avaliados. Considerando todas as amostras juntas, a expressão de Rb foi maior em amostras não metiladas comparadas as amostras parcialmente metiladas. Esta investigação mostrou a hipometilação dos genes P27, P53 e RB1 nos MO. Adicionalmente, a metilação nos genes supressores de tumor é um evento frequente em polpa dental normal.
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Modulation of the expression of the transcription factor Max in rat retinal ganglion cells by a recombinant adeno-associated viral vector

Modulation of the expression of the transcription factor Max in rat retinal ganglion cells by a recombinant adeno-associated viral vector

Adeno-associated viruses (AAVs) belong to the Parvoviridae family, small animal viruses with a linear single-stranded DNA genome, that replicate in the presence of a helper virus such as adenovirus (8). Gene transfer vectors based on AAV combine a number of advantages: AAV is naturally defective for replication and is considered nonpathogenic (9). It can transduce both proliferating and post-mitotic cells, transfer a gene to a number of cell types efficiently, and mediate long-term gene expression in the central nervous system, including the retina, as well as in the lungs, liver, or muscle cells, with minimal toxicity and cellular im- mune responses (10). A disadvantage of AAV is the limited packaging size of the virus, which cannot exceed 5,000 nucleotides. This,
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Investigating The Use Of Mobile Computing In Zimbabwe Polytechnics Case Of A Polytechnic In Zimbabwe

Investigating The Use Of Mobile Computing In Zimbabwe Polytechnics Case Of A Polytechnic In Zimbabwe

outside staff and lecture rooms, send and receive emails and communicate on social networks. This Polytechnic in particular, was one of the first institutions to install the wireless access points that accessed internet through the main fibre backbone. In 2009 it went on to procure laptops for staff members, starting with senior management, the Heads of departments and finally lectures. Students were then allowed to bring their own devices which could be configured to be able to access institutional WIFI (The Polytechnic ICT policy document, 2010). This was the beginning of mobile computing at the Polytechnic. Since then further strides were made in such areas as installation of applications that run on mobile devices through wireless connections, increasing internet bandwidth to improve speed as demand for internet went up, upgrading wireless access points to improve strength of connectivity, upgrading of servers to handle the demand and volumes and expansion of campus area network to cover the whole Polytechnic, procurement of more mobile devices. And the institution now boasts of such things as e-learning, m- learning, m-education, among other technologies that are giving it a competitive advantage over sister Polytechnics.
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Directly transforming PCR-amplified DNA fragments into plant cells is a versatile system that facilitates the transient expression assay.

Directly transforming PCR-amplified DNA fragments into plant cells is a versatile system that facilitates the transient expression assay.

Transient Expression of PCR-fragments in Epidermal Cells The components designed in the cassette of a PCR-fragment include a promoter, such as CaMV 35S promoter [16] or UBQ10 promoter [17], coding sequences of the gene of interest (CDS), and the terminator (NOS) (Figure 1A). Initially, PCR-fragments were generated through PCR amplification from the template plasmid DNA which contains the backbone of vector pUC18. The pair of universal primers PVU-F (located at 180 bp upstream of the promoter) and PVU-R (located at 100 bp downstream of NOS terminator) was used (Figure 1A). Amplified PCR-fragments were briefly purified with the extraction of phenol/chloroform and precipitated in ethanol [18]. In the first task, we prepared PCR- fragments of 35S-GFP-NOS from plasmid p35S-GFP. To remove residues of template plasmid DNA from the PCR-fragments, we purified PCR-fragments 35S-GFP-NOS through agarose gel extraction. Then, purified PCR-fragments 35S-GFP-NOS were transformed into epidermal cells of Onion peels with the biolistic bombardment delivery system. To compare transformation efficiency, plasmid DNA p35S-GFP was transformed in parallel. For a negative control, we transformed PCR-fragments generated from the same template plasmid but not containing GFP sequences. GFP signal was compared in transformations with PCR-fragments and with plasmid DNA. Results showed that GFP fluorescent signal was detected in the transformation with 35S- GFP-NOS or p35S-GFP, no GFP signal was detectable in the negative control (Figure S1). These results indicated that PCR- fragment based transient expression system (PCR-TES) could be used as an alternative way to assess gene expression in plant cells. Next, we compared the efficiency between PCR-fragments transformation and plasmids transformation. PCR-fragment 35S- GFP-NOS and plasmid p35S-Cherry were cotransformed into epidermal cells of Onion peels. Results showed that cells expressing Cherry fluorescent signal (red) were also displaying GFP fluores- cent signal (green) (Figure 1B), suggesting that transformation with PCR-fragments, instead of plasmid DNA, could be a substantial approach for transiently assessing a gene expression in plant cells. In attempted to examine the applicability of PCR-TES for analyzing subcellular localizations of genes, three cellular markers were tested in PCR-TES. PCR-fragments of YFP-mTn (for showing actin filaments) [19], NAG-YFP (for labeling golgi apparatus) [20] and ER-YFP (for showing endoplasmic reticulum) [21] were cotransformed with their correspondent plasmids (p35S- CFP-mTn, p35S-NAG-CFP and p35S-ER-CFP) into epidermal cells of Onion peels and Arabidopsis leaves, respectively. Results demonstrated that PCR-fragments (showing YFP fluorescence) of each cellular marker were expressed as efficient as their correspondent plasmids (showing CFP fluorescence) at their cellular locations (Figure 1C).
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Genes Regulating Maternal Recognition of Pregnancy in Domestic Animals: an Update

Genes Regulating Maternal Recognition of Pregnancy in Domestic Animals: an Update

cell adhesion and defence against invading microorganisms. Galectins do not possess a signal peptide or transmembrane spanning domain, and are secreted from the cells by a nonclassical pathway. Several forms of galectins: galectin-1 (LGALS1), galectin-3 (LGALS3), Galectin-5 (LGALS5), Galectin-9 (LGALS9) and Galectin-15 (LGALS15) have been discovered. Galectin-1 possess adhesive and anti-adhesive roles, immunomodulator in maternofetal tolerance as well as embryo implantation. However, galectin-1- null mouse embryos develop normally and do not produce any overt phenotypic abnormalities (Poirier and Robertson 32 ). Galectin-3 (LGALS3) is a galactose-specific lectin, expression of which increases significantly during the secretory phase of the menstrual cycle (von Wolff et al. 2005). Mice lack both galectins-1 and -3 implant normally but a further family member, galectin-5, is present and may compensate. Galectin-9 has been identified in mid- and late-secretory and decidual phases in human endometrium, with the expression in glandular and luminal epithelial but not stromal or immune cells (Smalley and Ley 2005). Recently, a new galectin family member, galectin-15, has been discovered in the endometrium of sheep, which has a prospective role in trophectoderm attachment (Farmer et al. 2008). Ovine endometrial Galectin-15 contains a conserved carbohydrate recognition domain (CRD) that binds β-galactosides, but the carbohydrate-binding specificity for each galectin appears to be different. In addition to a conserved CRD, it also contains predicted cell attachment sequences (LDV and RGD) that could mediate binding to integrins in the ECM proteins (Kimber and Spanswick 2000). Galectin-15 mRNA was detected only in the endometrial LE and superficial ductal GE (Gray et al. 2004) of ovine uterus and was not detected before day 10, appeared and then increased 13-fold between days 10 and 14, and then noticeably decreased between days 14 and 16 in cyclic, but not pregnant ewes. It was present at low levels on days 10 and 12, but was abundant on days 14 and 16 of pregnancy in uterine flushing. The proposed extracellular role of galectin-15 in the uterine lumen is functionally to bind and cross- link β-galactosides on glycoproteins, such as mucins, integrins, fibronectin, laminin and other glycoproteins and glycolipids, thereby allowing it to function as a heterophilic cell adhesion molecule bridging the blastocyst and the endometrial LE.
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Regulation of inflammatory gene expression in PBMCs by immunostimulatory botanicals.

Regulation of inflammatory gene expression in PBMCs by immunostimulatory botanicals.

induced (epiregulin, matrix metallopeptidase 14, podoplanin, VEGF, thrombospondin-1) are key players in angiogenesis. This agrees with recent reports which suggest that Astragalus promotes angiogenesis and induces VEGF expression [38,66]. Most of the above genes are not known to be responsive to LPS with the exception of VEGF and possibly sphingosine 1-phosphate phosphatase 2 which both contain NF-kB binding sites in their promoters [67,68]. Additionally, LPS is known to adversely impair wound healing [69]. This suggests that alternate components of Astragalus are likely mediating the wound healing and angiogenic effects and may be modulating the activities of LPS. Regarding blood pressure modulation, prostaglandin-endoperoxide synthase 2 (COX-2), adrenomedulin (ADM) and monoamine oxidase were all induced and are known to be involved in vasodilation. LPS induces both COX-2 and ADM and may be an active component associated with blood pressure modulation [65,70]. The vasode- pressor role of the COX-2 gene product has been shown to be regulated and dependent on the activity and ratio of both COX-1 and COX-2 [71]. Notably, treatment of PBMCs with the Astragalus extract led to a high induction of COX-2 transcription while slightly inhibiting COX-1 gene expression possibly supporting a vasodepressor effect. Similarly, several proteins were induced which can affect blood coagulation, including coagulation factor III, integrin b3, and thrombomodulin, and these genes have not been shown to be LPS-induced. Interestingly, thrombomodulin can prevent LPS induction of inflammatory molecules by the inhibition of NF-kB signaling [72]. Botanical components, as well as the natural presence of LPS in the Astragalus extracts may together be responsible for the induction of genes involved in angiogenesis, modulation of blood pressure, and blood clotting [73,74,75]. Further studies are required to determine the effects of other components present in the Astragalus extracts that may modulate the activities of LPS in putative physiological roles.
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Human Capital and the Recent Fall of Earnings Inequality in Brazil

Human Capital and the Recent Fall of Earnings Inequality in Brazil

This paper evaluates the factors that have contributed to the decline in earnings inequality in Brazil, for the first time in decades, by means of a flexible decomposition technique and counterfactual exercises. The variance of (log) earnings declined by about a quarter between 1995 and 2009. We find that, until the end of the 1990s, most of the fall happened within education and age groups, with very little role for our observable measures of skill. But, in the new century, the between-groups component also contributed significantly to the fall in inequality, mostly through the fall in the education wage differentials. Returns to experience have also declined, especially among the less skilled workers. We find that the education composition of the workforce also contributed
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Centralization And Decentralization In Planning

Centralization And Decentralization In Planning

Thecases in which they frequentlyusedecentralizationare: *The desire toincrease the participation ofthe populationandbecause of itsgreat importancein achieving thedecisions and policiesbe realisticandsome of the propertiesthat helpin the success ofdevelopment policiesto reachtheir goals Messahaalarge size ofpopulationandthe stateso that it cannotaddressthe centralpolicies ofeachsprawlingspaceissueswhich forces theupperbodies of theplanningmake room fordecentralizedmanagement styleandtakethe brunt ofthe most important inthe process of preparingdevelopment plans Multiplicity ofnationalities andracesas it isin such circumstancesbeuniting of interestsis not possible orfar-fetchedprocess, which requires giving
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In vivo and in vitro effect of imipramine

In vivo and in vitro effect of imipramine

For in vitro studies, synaptic plasma mem- branes from cerebral cortex of 75-day-old untreated rats were used as the membrane enzyme preparation. Imipramine or fluoxe- tine was dissolved in Tris-HCl buffer, pH 7.4, and added to the incubation medium to a final concentration ranging from 0.1 to 0.5 mM. Control assays did not contain the drugs in the incubation medium. The concentra- tion of the drugs used in the present investi- gation was chosen on the basis of previous studies using imipramine in vitro (12) and the solubility of fluoxetine in the buffer.
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Arq. Bras. Cardiol.  vol.98 número5

Arq. Bras. Cardiol. vol.98 número5

multiple benefits of physical training on survival and the harmful effects of a sedentary lifestyle made this therapeutic modality required to treat patients with heart disease, including those with systolic ventricular dysfunction, who have already shown a pathological increasing in ventricular volumes and masses. Its benefits regarding functional capacity can be explained by effects on endothelial function, peripheral vascular resistance and changes in the structure of skeletal muscle, although significant improvement in left ventricular ejection fraction have not been demonstrated.
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SOCIAL ECONOMY – A FORM OF INCLUSION AND OF ''REACTIVATING'' OF LABOR IN THE CONTEXT OF THE CURRENT CRISIS

SOCIAL ECONOMY – A FORM OF INCLUSION AND OF ''REACTIVATING'' OF LABOR IN THE CONTEXT OF THE CURRENT CRISIS

)n the context of the cohesion policy, solidarity must represent a support for development . For that purpose, solidarity can be seen as a help for self‐help and its success depends a great deal on the capacity and the training of the people to whom the support of making maximum profit out of these addresses to. This support does not mean exclusively financial support, although it is necessary and important but, of all things, it means an exchange of experiences and cooperation, the development of capacity through training, open discussions with the interested factors and last but not least a critic, but a constructive dialogue between the various levels of government: European, national, regional, local. )n other words, a functional labor market should represent a catalyst for the general objective of the European Union – social and economical cohesion – because it has in view the connections with the different markets of the services and of the goods and generates the necessary income for supporting the participation of the individuals, bringing them together, placing them in collaborations. )n this context, the starting points for promoting the inclusion through the activities of social economy have in view: adapting the institutional environment, developing the public‐private partnership, developing the social dialogue between players, investments in the human capital and supporting the exchange of good practices within the European Union.
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Electronic Government In Democratic Public Service In One Door Integrated Permit Handling Services Agency In The City Of Samarinda

Electronic Government In Democratic Public Service In One Door Integrated Permit Handling Services Agency In The City Of Samarinda

Development of information technologies and communication in a public sector have to pay more attention to the complexity in its implementation rather than focusing on best practice and strategies which are universal to prescribe how successfully applied Electronic Government program. As a result of advanced civilization for conducting residents of the state of being high-profile figures in the service in the era of democracy. Due to existence of interaction in the form of consultations will find a pattern, approaches such as what is appropriate to the needs of the community in exercising the functions of participation to the state. Based on the results of research on public services in the context of democratic consultation. Look about how the process of consultation with citizens. At the end result appears that the consultants who holds the project activities which implement electronic government that coupled with the community members who said that if the process of consultation having no democratic value. Certainly, it is negative in the development of democracy in the region licensing office, where community involvement not be used as reference to build an understanding together in achieving common interests in accordance with the concept of new public service. Criticism of the statement above , delivered by George (2002) that the failure of the process of the interaction between citizens with the government is located on the level of bureaucracy that is not acceptable to open direct communication. Given only communication with the use of symbols and an intermediary. Statement above supported by Robbins (2005) that stiff of a bureaucracy that has become part the past, of communications for interaction are the dominant choice for in the success of a form of service. All stiffness will perish along
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Ministro Joana TM 2011.pdf

Ministro Joana TM 2011.pdf

domain of blaR1. The overexpression of blaI caused no significant alterations in the resistance phenotype: a slightly large halo could be noticed but the same effect was observed in the control experiment with the empty plasmid vector. Nevertheless, contrary to what was observed with blaR1, the blaI overexpression did not originate a hyper-sensitive phenotype, suggesting that this latter gene is not toxic to the cell. It is reasonable to assume that the overexpression of blaI alone would not reproduce the phenotype observed with the introduction of the complete bla plasmid, since the inducer is not present. Concerning the overexpression of L3 domain of BlaR1 in COL-I, no significant effect on the phenotypic expression of resistance was observed as well. BlaR1 has two functional domains, the sensor domain that extends to the extracellular medium where β-lactams can bind, and the transducer domain that lies in the cytoplasm and has four transmembrane α-helices interconnected by three loops (Fig. 10). Importantly, the zinc metalloprotease domain, present in loop 3 (L3), is cleaved during the induction process and is thought to be the connective element between BlaR1 and BlaI (57). Although no toxic effect was observed in these experiments, the lack of effect on COL-I phenotype suggests that the overexpressed L3 domain is not functional and that other parts of BlaR1 are required for its activation.
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