No description available.

Bone tissue is continuously remodeled by bone-forming osteoblasts and bone-resorbing osteoclasts, in processes tightly regulated by hormones, cytokines and growth factors. CD47, a ubiquitously expressed protein, and one of its ligands, signal-regulatory protein alpha (SIRPα), are two cell-surface proteins belonging to the immunoglobulin (Ig)-superfamily. The interaction between CD47 and SIRPα is important for, amongst other processes, the fusion of macrophages into giant cells, which are closely related to osteoclasts. The aim of the present study was to gain knowledge about the role of CD47–SIRPα interaction and resultant downstream signaling pathways in bone cell differentiation, formation and function. The addition of antibodies against CD47 or SIRPα inhibited the formation of multinucleated osteoclasts from bone marrow monocytes (BMMs) in culture. Moreover, a significant decrease in the number of osteoclasts was detected in CD47-/- BMM cultures compared to CD47+/+ cultures. In line with these in vitro results, we found fewer osteoclasts in vivo in the trabecular bone of CD47-/- mice, as compared to CD47+/+ bone. Interestingly, an extended analysis of the trabecular bone of CD47-/- mice revealed that the bone volume, mineralizing surface, mineral apposition rate, bone formation rate and osteoblast number were also significantly reduced compared with CD47+/+ mice, indicating the importance of CD47 in osteoblast differentiation. In vitro studies of bone marrow stromal (BMS) cells from CD47-/- mice or SIRPα-mutant mice (mice lacking the signaling domain of SIRPa) showed a blunted expression of osteoblast-associated genes. Moreover, these altered genotypes were associated with reduced activity of the bone mineralization-associated enzyme alkaline phosphatase as well as a reduced ability to form mineral. To reveal the molecular mechanisms by which CD47 activation of SIRPα is important for BMS cell differentiation, we studied signaling downstream of SIRPα in the absence of CD47. In BMS cells lacking CD47, a considerable reduction in the levels of tyrosine phosphorylated SIRPα was detected, and the subsequent recruitment of the Src-homology-2 (SH2) domain-containing protein tyrosine phosphatase (SHP-2)–phosphoinositide 3-kinase (PI3K)–Akt2 signaling module was nearly abolished. In conclusion, the interaction between CD47 and SIRPα results in the activation of the SHP-2–PI3K–Akt2 pathway, which is necessary for normal osteoblast differentiation. In CD47-/- mice and SIRPα-mutant mice, this interaction is perturbed, which prevents normal osteoblast differentiation and subsequent mineral formation. In addition, the altered BMS cell phenotype results in an impaired ability to stimulate osteoclast differentiation.

The present thesis deals with the description of the current state on the course of Morava river from small hydro Kromeriz after a small water plant Hodonin. Subsequently, I deal with the description of search of weir plants and weirs for the construction of small hydropower (especially their heads, structures and, where appropriate, possible adaptation). Then I deal with completion new hydroelectric plants on this founded weir plants and weirs which are in section between the small hydroelectric power Kromeriz and Lanžhot city on the Czech Republic side (Brodské on the Slovak Republic side). In this work is mainly focus on the calculation of installed performance in individual locations (and also the calculation of achievable performance in these locations), machinery design (for the use of selected sets from Hydrohrom company) and the way the configuration of the power switch. If it is in those places, řeším even more possibilities of this construction is splavnění view of the river, especially in the field of Hodonin - Lanžhot. In addition, deals with the design of line side cubicle of each switch to local distribution networks (draft routes). In the last phase deals with the size of the production in each generation, calculations of costs of constructing these plants, approximate calculation of the production (of the flows of 2007) and time of return on your investment in the construction.

No / Summary. Background: Endothelial nitric oxide synthase (eNOS) activity in endothelial cells is regulated by post-translational phosphorylation of critical serine, threonine and tyrosine residues in response to a variety of stimuli. However, the post-translational regulation of eNOS in platelets is poorly defined. Objectives: We investigated the role of tyrosine phosphorylation in the regulation of platelet eNOS activity. Methods: Tyrosine phosphorylation of eNOS and interaction with the tyrosine phosphatase SHP-1 were investigated by coimmunoprecipitation and immunoblotting. An in vitro immunoassay was used to determine eNOS activity together with the contribution of protein tyrosine phosphorylation. Results: We found platelet eNOS was tyrosine phosphorylated under basal conditions. Thrombin induced a dose- and time-dependent increase in eNOS activity without altering overall level of tyrosine phosphorylation, although we did observe evidence of minor tyrosine dephosphorylation. In vitro tyrosine dephosphorylation of platelet eNOS using a recombinant protein tyrosine phosphatase enhanced thrombin-induced activity compared to thrombin alone, but had no effect on endothelial eNOS activity either at basal or after stimulation with bradykinin. Having shown that dephosphorylation could modulate platelet eNOS activity we examined the role of potential protein phosphatases important for platelet eNOS activity. We found SHP-1 protein tyrosine phosphatase, co-associated with platelet eNOS in resting platelets, but does not associate with eNOS in endothelial cells. Stimulation of platelets with thrombin increased SHP-1 association with eNOS, while inhibition of SHP-1 abolished the ability of thrombin to induce elevated eNOS activity. Conclusions: Our data suggest a novel role for tyrosine dephosphorylation in platelet eNOS activation, which may be mediated by SHP-1.

No description available.

Résumé : La néphropathie diabétique (ND) est la principale cause d’insuffisance rénale de stade terminal en Amérique du Nord. Les podocytes, cellules épithéliales hautement spécialisées du glomérule, supportent et maintiennent les mécanismes de filtration glomérulaire. Des biopsies de reins de patients diabétiques ont montré que le nombre de podocytes est significativement réduit chez les patients avec un diabète récent. Néphrine est une protéine transmembranaire qui a été démontrée comme ayant un rôle majeur dans le maintien de l’intégrité de ces cellules. Une diminution de l’expression de néphrine est observée chez les personnes atteintes de la ND. Des études ont démontré que la phosphorylation en tyrosine de néphrine était impliquée dans la régulation de l’inhibition des voies de l’apoptose et le remodelage du cytosquelette d’actine. Notre laboratoire a publié que l’expression de la tyrosine phosphatase SHP-1 était augmentée dans les podocytes exposés à des concentrations élevées de glucose (HG). Les résidus tyrosines de néphrine sont contenus dans des séquences pouvant être reconnues par SHP-1. Notre hypothèse est que SHP-1 interagit avec néphrine, et que l’augmentation de l’expression de SHP-1 en condition d’hyperglycémie et de diabète viendrait déréguler les voies de signalisation de néphrine, contribuant aux dommages des podocytes dans la maladie. Des coimmunoprécipitations dans des podocytes montrent une interaction entre SHP-1 et néphrine, qui est augmentée en condition HG. Cette augmentation en HG était associée à une baisse des niveaux de phosphorylation de néphrine. La surexpression de la forme inactive de SHP-1 dans les podocyte rétablie les niveaux de phosphorylation de néphrine en condition HG. Dans un modèle de surexpression avec des cellules HEK, la surexpression de SHP-1 diminue les niveaux de phosphorylation des tyrosines 1176/1193 et 1217, qui sont associées au remodelage de l’actine. Des coimmunoprécipitations avec des mutants de néphrine montrent que les tyrosines 1114 et 1138 sont essentielles pour l’interaction de SHP-1 avec néphrine. Dans un modèle murin de diabète de type 1, une diminution de l’expression et de la phosphorylation de néphrine sont observée comparativement aux souris de type sauvage. Ces diminutions sont associées avec une augmentation de l’expression de SHP-1. En conclusion, l’augmentation de l’expression de SHP-1 en condition d’hyperglycémie réduit les niveaux de phosphorylation en tyrosine de néphrine et vient potentiellement inhiber ses voies de signalisation dans le diabète, contribuant à la dysfonction podocytaire et à la néphropathie diabétique. / Abstract : Diabetic nephropathy (DN) is the leading cause of end-stage renal disease in North America. Podocytes are highly specialized epithelial cells involved in the glomerular filtration process. Morphometric observation from kidney biopsies of diabetic patients showed a significant reduction in the number of podocytes in patients with short duration of diabetes before the apparition of microalbuminuria. Nephrin, a transmembrane protein found in the slit diaphragm, has been found to play a key role in the integrity of the podocytes. Clinical observations indicated that nephrin expression was reduced in kidney biopsy of diabetes patients. Recent studies have shown that phosphorylation of tyrosine residues of nephrin participate in intracellular pathways regulating actin dynamics and podocyte survival. Our laboratory has recently published that the expression of the tyrosine phosphatase SHP-1 is elevated in podocytes exposed to high glucose concentrations (HG). Nephrin contains sequences that are known to be potential target for SHP-1. Our hypothesis is that SHP-1 can interact with nephrin, and the increase of SHP-1 expression in diabetic nephropathy deregulates nephrin-mediated pathways, contributing to podocyte’s damage in the disease. Coimmunoprecipitation experiments show an interaction between SHP-1 and nephrin which is increased in podocytes exposed to HG. Overexpression of the inactive form of SHP-1 in podocytes exposed to HG restores nephrin phosphorylation. In HEK cells, overexpression of SHP-1 reduces nephrin phosphorylation specifically on tyrosine 1176/1193 and 1217, which regulates actin dynamics. Coimmunoprecipitation experiments with nephrin mutants show that tyrosine 1114 and 1138 are essentials to the interaction between SHP-1 and nephrin. In a type 1 diabetic murine model, a reduction of the expression and phosphorylation levels of nephrin are observed. Both reductions are associated with an increase in SHP-1 expression. In conclusion, diabetes triggered SHP-1 expression in podocytes which reduces nephrin tyrosine phosphorylation and potentially inhibits nephrin signaling in diabetes, contributing to podocytes dysfunction in diabetic nephropathy.

<p>Predmet istraživanja ove doktorske disertacije<br />je bio formiranje i proučavanje osobina i<br />pona&scaron;anja organskih aerosola nastalih<br />atomiziranjem stabilnih emulzija dominantno<br />pod dejstvom centrifugalne sile. Kori&scaron;ćene su<br />vodene emulzije komercijalnih sredstava za<br />hlađenje i podmazivanje (SHP), koja se koriste<br />pri obradi metalnih odlivaka. Cilj istraživanja je<br />bio ispitati i objasniti uticaj koncentracije SHP<br />emulzije, brzine strujanja vazduha, prirode kako<br />ulja, tako i osobina emulzija, kao &scaron;to su gustina,<br />viskoznost, povr&scaron;inski napon, kao i udaljenost<br />od mesta atomiziranja na masenu koncentraciju,<br />broj i veličinu kapi aerosola u kontrolisanim<br />laboratorijskim uslovima.<br />Uslovi eksperimenta isključili su uticaj toplote<br />nastale kao posledica trenja alata i delova koji<br />se obrađuju. Pored toga isključeno je prisustvo<br />čvrstih mikronskih čestica koje potiču od delova<br />koji se obrađuju, kao i prisutvo plivajućeg<br />hidrauličnog ulja i mikroorganizama koji uvek<br />prate realan industrijski fluid.<br />Eksperimentalni program je realizovan na tri<br />komercijalna SHP sredstva različitog porekla.<br />Proučavani su aerosoli formirani atomiziranjem<br />emulzije tri različite koncentracije uljne faze: 1,<br />6 i 10 %. Pri jednakim eksperimentalnim<br />uslovima ispitivan je i aerosol formiran, od<br />vode kori&scaron;ćene za pripremu emulzija. Određene<br />su osobine ulja i emulzija koje su od značaja za<br />atomiziranje tečnosti. Proučavan je i uticaj<br />brzine ventilacionog vazduha na osobine<br />aerosola. Eksperimenti su realizovani pri<br />brzinama vazduha od 1, 3, 6, 8 m/s.<br />Uzorkovanje aerosola je realizovano u komori,<br />na ulazu u ventilacini vod i duž ventilacione<br />cevi dužine 8m na pet mernih tačaka koje se<br />nalaze na 0,5, 1, 2, 4, 6, 8 m u odnosu na<br />komoru.<br />Značajan doprinos ove doktorske disertacije je<br />postavljanje teorije da atomiziranjem emulzije<br />kao heterogene tečnosti, nastaje heterogen<br />aerosol, pri čemu mogu nastati kapi različite<br />prirode, između ostalog i kapi čistog ulja. Na taj<br />način je obja&scaron;njen fenomen da porastom<br />koncentracije emulzije raste veličina kapi<br />nastalog aerosola. Aerosoli nastali od<br />koncentrovanijih emulzija pokazuju uređenije<br />strujanje kroz ventilacionu cev, &scaron;to je od<br />značaja za uspe&scaron;nije projektovanje ventilacionih<br />sistema i filtara za njihovu separaciju u cilju<br />za&scaron;tite zdravlja radnika i za&scaron;tite okoline.</p> / <p> This PhD Thesis presents an experimental<br /> study of organic aerosols formed from stable<br /> water emulsions, predominantly by centrifugal<br /> force under laboratory conditions. Emulsions of<br /> metalworking fluids (MWF) were used in the<br /> experiments. The aim of this investigation was<br /> to explain the influence of MWF emulsion<br /> concentration, oil and emulsion properties<br /> (density, viscosity, and surface tension), air<br /> velocity, as well as distance from atomization<br /> generator on aerosol behavior and properties as<br /> size distribution, mass and number<br /> concentration. The experimental conditions<br /> excluded the influence of the heat arising from<br /> the friction between the tool and the<br /> workpieces. In this way the mechanism of<br /> aerosol formation by the<br /> evaporation/condensation is minimized.<br /> Besides, the presence of solid micrometer sized<br /> particulates originated from the work piece is<br /> excluded, as well as the presence of tramp oil<br /> and microorganisms, always accompanying a<br /> real industrial fluid.<br /> Three different commercial MWFs were<br /> investigated at three different oil-in-water<br /> emulsion concentrations, 1, 6, and 10 %<br /> respectively. Water aerosol was investigated<br /> under same experimental conditions. Properties<br /> of MWF oils and emulsion, which are important<br /> for liquid atomization, were determined.<br /> Influence of ventilation air velocity on aerosol<br /> properties was investigated as well.<br /> Experiments were realized under four different<br /> air velocities: 1, 3, 6, and 8 m/s. Samples were<br /> taken at three different points: at the camber, at<br /> the entrance of ventilation pipe, and at five<br /> sampling points along the pipe. Ventilation pipe<br /> was 8 m long, and sampling point were at 0.5, 1,<br /> 2, 4, 6, 8 m from the aerosol camber.<br /> The main contribution of this dissertation is<br /> the new theory that atomization of emulsion as a<br /> heterogeneous fluid leads to the formation of<br /> heterogeneous aerosol, containing droplets of a<br /> different nature, including droplets of pure oil.<br /> This theory explains the phenomenon that the<br /> aerosol droplet sizes increase with the increase<br /> of the emulsion concentration. Obtained results<br /> show that aerosols formed from the emulsions<br /> of higher concentration (6 and 10 %) have less<br /> chaotic flow through the ventilation pipe. This<br /> observation is important for better design of<br /> ventilation systems and filters for mist<br /> separation in order to protect human health and<br /> the environment.&nbsp;</p>

SHP-1 is a protein tyrosine phosphatase that often targets the phosphatidylinositol 3'-kinase (PI3K)/Akt signalling pathway. PI3K/Akt signalling regulates cell growth and survival, proliferation and differentiation. Growth factor-stimulated PI3K phospholipid production at the plasma membrane helps to recruit 3'-phosphoinositide-dependent protein kinase-1 (PDK1) and Akt, where PDK1 phosphorylates and activates the pro-survival kinase Akt.<p>Tyrosine phosphorylation of PDK1 may regulate its function and, perhaps more importantly, its nuclear localization. Yet, it is unclear how PDK1 is imported into the nucleus as it does not contain a nuclear localization signal (NLS), although it does contain a nuclear export signal (NES). Interestingly, several tyrosines in PDK1 are targets for Src kinase and are putative target motifs for SHP-1, which does have an NLS.<p>Hypothesis: SHP-1 and PDK1 form a tyrosine-dependent, nucleo-cytoplasmic shuttling complex. <p>Removal of serum from C6 glioma cell cultures induces a platelet-derived growth factor receptor (PDGFR)-sensitive redistribution of PI3K lipid kinase activity to the nucleus. PDK1 tyrosine phosphorylation and its association with SHP-1 are also increased, as is the accumulation of both SHP-1 and PDK1 in the nucleus. Site-directed mutagenesis of tyrosine residues in PDK1 reveals that tyrosine 9 (Tyr9) and Tyr376 are important for the interaction of PDK1 with SHP1, whereas Tyr333 and Tyr 373 are not. Using pharmacological and genetic manipulations, it was demonstrated that SHP-1 and PDK1 shuttle between the nucleus and cytoplasm, and that the C-terminal-expressed NLS of SHP-1 facilitates shuttling, while dephosphorylation of PDK1 Tyr9 and Tyr376 regulates the rate of PDK1 (and by virtue of association, SHP-1) export from the nucleus. The SHP-1/PDK1 complex, which is constitutive in most cell lines, is functionally relevant as indicated by its requirement for NGF-induced differentiation of preneuronal cells to a neuronal phenotype.

SHP-1 is a protein tyrosine phosphatase that often targets the phosphatidylinositol 3'-kinase (PI3K)/Akt signalling pathway. PI3K/Akt signalling regulates cell growth and survival, proliferation and differentiation. Growth factor-stimulated PI3K phospholipid production at the plasma membrane helps to recruit 3'-phosphoinositide-dependent protein kinase-1 (PDK1) and Akt, where PDK1 phosphorylates and activates the pro-survival kinase Akt.<p>Tyrosine phosphorylation of PDK1 may regulate its function and, perhaps more importantly, its nuclear localization. Yet, it is unclear how PDK1 is imported into the nucleus as it does not contain a nuclear localization signal (NLS), although it does contain a nuclear export signal (NES). Interestingly, several tyrosines in PDK1 are targets for Src kinase and are putative target motifs for SHP-1, which does have an NLS.<p>Hypothesis: SHP-1 and PDK1 form a tyrosine-dependent, nucleo-cytoplasmic shuttling complex. <p>Removal of serum from C6 glioma cell cultures induces a platelet-derived growth factor receptor (PDGFR)-sensitive redistribution of PI3K lipid kinase activity to the nucleus. PDK1 tyrosine phosphorylation and its association with SHP-1 are also increased, as is the accumulation of both SHP-1 and PDK1 in the nucleus. Site-directed mutagenesis of tyrosine residues in PDK1 reveals that tyrosine 9 (Tyr9) and Tyr376 are important for the interaction of PDK1 with SHP1, whereas Tyr333 and Tyr 373 are not. Using pharmacological and genetic manipulations, it was demonstrated that SHP-1 and PDK1 shuttle between the nucleus and cytoplasm, and that the C-terminal-expressed NLS of SHP-1 facilitates shuttling, while dephosphorylation of PDK1 Tyr9 and Tyr376 regulates the rate of PDK1 (and by virtue of association, SHP-1) export from the nucleus. The SHP-1/PDK1 complex, which is constitutive in most cell lines, is functionally relevant as indicated by its requirement for NGF-induced differentiation of preneuronal cells to a neuronal phenotype.

No description available.