31 research outputs found

    Maszkot viselve – mindig szerepben : Petrás Éva: Álarcok mögött. Nagy Töhötöm életei.

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    "The Sisters of the Redeemer in the Trauma of Dispersion" : The Sisters of the Divine Redeemer in the 1950s and 1960s in the Light of Recollections and State Security Reports

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    While researching the history of the Sisters of the Divine Redeemer, also referred to as the Sisters of the Redeemer, it became clear that the ordeals of the Second World War and the Communist dictatorship had a profound impact on the congregation, which was engaged in nursing and teaching. The sources allow us to reconstruct the horrors of the advancing battlefront and the sisters' flight, along with their determination to provide social assistance and their role in saving Jews. The Communist regime that emerged after the war forced the congregation into an increasingly impossible situation, depriving them of their teaching positions and nursing vocation. Their internment in 1950 and the revocation of the congregation's operating license seemed to have eliminated the community entirely. However, recollections of the events of the 1950s and 1960s, together with state security reports, attest that the congregation survived in the form of a “subterranean stream,” and that tiny communities of sisters continued to pursue their monastic vocation, often in a single apartment that functioned as a mini convent. The traumas they had experienced rarely crushed the sisters' inner sense of peace, and they strove to cope with the harassments inflicted by the party-state by adapting to the new situation

    Investigation of sphingosin-1-phosphate-triggered matriptase activation using a rat primary hepatocyte model

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    Sphingosine-1-phosphate (S1P) has been reported as a matriptase activator. The aim of this study was to reveal if S1P can influence hepcidin production. Furthermore, we investigated how S1P can affect the viability and the redox status of primary hepatocytes. Rat primary hepatocytes were cultivated for 72 h and were treated with 50, 200, 1000 ng/ml S1P. Cell-free supernatants were collected every 24 h. Cell viability was tested by a colorimetric method using tetrazolium compound (MTS). The hepcidin levels in the cell-free supernatants were examined with hepcidin sandwich ELISA to determine the effect of S1P on the hepcidin-modulating ability of matriptase. In order to estimate the extent of S1P-generated oxidative stress, extracellular H2O2 measurements were performed by the use of fluorescent dye. Based on the findings, S1P treatment did not cause cell death for 72 h at concentrations up to 1000 ng/ml. S1P did not influence the extracellular H2O2 production for 72 h. The hepcidin levels were significantly suppressed in hepatocytes exposed to S1P treatment. Further studies would be needed to explore the exact mechanism of action of S1P

    Serum MASP-1 in complex with MBL activates endothelial cells.

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    The complement system plays an important role in the induction of inflammation. In this study we demonstrate that the initiation complexes of the lectin pathway, consisting of mannose-binding lectin (MBL) and associated serine proteases (MASPs) elicit Ca2+ signaling in cultured endothelial cells (HUVECs). This is in agreement with our previous results showing that the recombinant catalytic fragment of MASP-1 activates endothelial cells by cleaving protease activated receptor 4. Two other proteases, MASP-2 and MASP-3 are also associated with MBL. Earlier we showed that recombinant catalytic fragment of MASP-2 cannot activate HUVECs, and in this study we demonstrate that the same fragment of MASP-3 has also no effect. We find the same to be the case if we use recombinant forms of the N-terminal parts of MASP-1 and MASP-2 which only contain non-enzymatic domains. Moreover, stable zymogen mutant form of MASP-1 was also ineffective to stimulate endothelial cells, which suggests that in vivo MASP-1 have the ability to activate endothelial cells directly as well as to activate the lectin pathway simultaneously. We show that among the components of the MBL-MASPs complexes only MASP-1 is able to trigger response in HUVECs and the proteolytic activity of MASP-1 is essential. Our results strengthen the view that MASP-1 plays a central role in the early innate immune response

    A versatile modular vector set for optimizing protein expression among bacterial, yeast, insect and mammalian hosts

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    We have developed a unified, versatile vector set for expression of recombinant proteins, fit for use in any bacterial, yeast, insect or mammalian cell host. The advantage of this system is its versatility at the vector level, achieved by the introduction of a novel expression cassette. This cassette contains a unified multi-cloning site, affinity tags, protease cleavable linkers, an optional secretion signal, and common restriction endonuclease sites at key positions. This way, genes of interest and all elements of the cassette can be switched freely among the vectors, using restriction digestion and ligation without the need of polymerase chain reaction (PCR). This vector set allows rapid protein expression screening of various hosts and affinity tags. The reason behind this approach was that it is difficult to predict which expression host and which affinity tag will lead to functional expression. The new system is based on four optimized and frequently used expression systems (Escherichia coli pET, the yeast Pichia pastoris, pVL and pIEx for Spodoptera frugiperda insect cells and pLEXm based mammalian systems), which were modified as described above. The resulting vector set was named pONE series. We have successfully applied the pONE vector set for expression of the following human proteins: the tumour suppressor RASSF1A and the protein kinases Aurora A and LIMK1. Finally, we used it to express the large multidomain protein, Rho-associated protein kinase 2 (ROCK2, 164 kDa) and demonstrated that the yeast Pichia pastoris reproducibly expresses the large ROCK2 kinase with identical activity to the insect cell produced counterpart. To our knowledge this is among the largest proteins ever expressed in yeast. This demonstrates that the cost-effective yeast system can match and replace the industry-standard insect cell expression system even for large and complex mammalian proteins. These experiments demonstrate the applicability of our pONE vector set

    Multiple roles of complement MASP-1 at the interface of innate immune response and coagulation

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    MASP-1 is a versatile serine protease that cleaves a number of substrates in human blood. In recent years it became evident that besides playing a crucial role in complement activation MASP-1 also triggers other cascade systems and even cells to mount a more powerful innate immune response. In this review we summarize the latest discoveries about the diverse functions of this multi-faceted protease. Recent studies revealed that among MBL-associated serine proteases, MASP-1 is the one responsible for triggering the lectin pathway via its ability to rapidly autoactivate then cleave MASP-2, and possibly MASP-3. The crystal structure of MASP-1 explains its more relaxed substrate specificity compared to the related complement enzymes. Due to the relaxed specificity, MASP-1 interacts with the coagulation cascade and the kinin generating system, and it can also activate endothelial cells eliciting pro-inflammatory signaling. © 2014 Elsevier Ltd. All rights reserved

    Cutting Edge: A New Player in the Alternative Complement Pathway, MASP-1 Is Essential for LPS-Induced, but Not for Zymosan-Induced, Alternative Pathway Activation

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    The complement system is a sophisticated network of proteases. In this article, we describe an unexpected link between two linear activation routes of the complement system: the lectin pathway (LP) and the alternative pathway (AP). Mannose-lectin binding-associated serine protease (MASP)-1 is known to be the initiator protease of the LP. Using a specific and potent inhibitor of MASP-1, SGMI-1, as well as other MASP-1 inhibitors with different mechanisms of action, we demonstrated that, in addition to its functions in the LP, MASP-1 is essential for bacterial LPS-induced AP activation, whereas it has little effect on zymosan-induced AP activation. We have shown that MASP-1 inhibition prevents AP activation, as well as attenuates the already initiated AP activity on the LPS surface. This newly recognized function of MASP-1 can be important for the defense against certain bacterial infections. Our results also emphasize that the mechanism of AP activation depends on the activator surface

    CR3 is the dominant phagocytotic complement receptor on human dendritic cells.

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    Dendritic cells (DCs) play a decisive role in immunity; they interact with various pathogens via several pattern recognition and different opsonophagocytotic receptors, including Fc- and complement-receptors. beta2-integrins, including complement receptors CR3 (CD11b/CD18) and CR4 (CD11c/CD18) participate in many immunological processes, especially those involving cell migration, adherence, and phagocytosis. Human monocyte derived dendritic cells (MDCs) are known to express CR3 as well as CR4, however possible differences regarding the role of these receptors has not been addressed so far. Our aim was to explore whether there is a difference between the binding and uptake of various complement-opsonized microorganisms, mediated by CR3 and CR4. Studying the expression of receptors during differentiation of MDCs we found that the appearance of CD11b decreased, whereas that of CD11c increased. Interestingly, both receptors were present in the cell membrane in an active conformation. Here we demonstrate that ligation of CD11b directs MDCs to enhanced phagocytosis, while the maturation of the cells and their inflammatory cytokine production are not affected. Blocking CD11c alone did not change the uptake of opsonized yeast or bacteria by MDCs. We confirmed these results using siRNA; namely downregulation of CD11b blocked the phagocytosis of microbes while silencing CD11c had no effect on their uptake. Our data clearly demonstrate that complement C3-dependent phagocytosis of MDCs is mediated mainly by CR3
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