Amnion: a versatile tissue and cell source in tissue repair and regeneration

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Human amniotic membrane as newly identifed source of amniotic fuid pulmonary surfactant

Pulmonary surfactant (PS) reduces surface tension at the air-liquid interface in the alveolar epithelium of the lung, which is required for breathing and for the pulmonary maturity of the developing foetus. However, the origin of PS had never been thoroughly investigated, although it was assumed to be secreted from the foetal developing lung. Human amniotic membrane (hAM), particularly its epithelial cell layer, composes the amniotic sac enclosing the amniotic fuid. In this study, we therefore aimed to investigate a potential contribution of the cellular components of the hAM to pulmonary surfactant found in amniotic fuid. We identifed that cells within the native membrane contain lamellar bodies and express all four surfactant proteins as well as ABCA3. Lipidomic profling by nanoESI – MS/MS revealed the presence of the essential lipid species as found in PS. Also, the biophysical activity of conditioned cell culture supernatant obtained from hAM was tested with captive bubble surfactometry. hAM supernatant showed the ability to reduce surface tension, similar to human PS obtained from bronchoalveolar lavage. This means that hAM produces the essential PS-associated components and can therefore contribute as second potential source of PS in amniotic fuid aside from the foetal lung

Evidence for Regulated Interleukin-4 Expression in Chondrocyte-Scaffolds under In Vitro Inflammatory Conditions

OBJECTIVE: To elucidate the anti-inflammatory and anabolic effects of regulated expression of IL-4 in chondrocyte-scaffolds under in vitro inflammatory conditions. METHODS: Mature articular chondrocytes from dogs (n = 3) were conditioned through transient transfection using pcDNA3.1.cIL-4 (constitutive) or pCOX-2.cIL-4 (cytokine-responsive) plasmids. Conditioned cells were seeded in alginate microspheres and rat-tail collagen type I matrix (CaReS®) to generate two types of tissue-engineered 3-dimensional scaffolds. Inflammatory arthritis was simulated in the packed chondrocytes through exogenous addition of recombinant canine (rc) IL-1β (100 ng/ml) plus rcTNFα (50 ng/ml) in culture media for 96 hours. Harvested cells and culture media were analyzed by various assays to monitor the anti-inflammatory and regenerative (anabolic) properties of cIL-4. RESULTS: cIL-4 was expressed from COX-2 promoter exclusively on the addition of rcIL-1β and rcTNFα while its expression from CMV promoter was constitutive. The expressed cIL-4 downregulated the mRNA expression of IL-1β, TNFα, IL-6, iNOS and COX-2 in the cells and inhibited the production of NO and PGE(2) in culture media. At the same time, it up-regulated the expression of IGF-1, IL-1ra, COL2a1 and aggrecan in conditioned chondrocytes in both scaffolds along with a diminished release of total collagen and sGAG into the culture media. An increased amount of cIL-4 protein was detected both in chondrocyte cell lysate and in concentrated culture media. Neutralizing anti-cIL-4 antibody assay confirmed that the anti-inflammatory and regenerative effects seen are exclusively driven by cIL-4. There was a restricted expression of IL-4 under COX-2 promoter possibly due to negative feedback loop while it was over-expressed under CMV promoter (undesirable). Furthermore, the anti-inflammatory /anabolic outcomes from both scaffolds were reproducible and the therapeutic effects of cIL-4 were both scaffold- and promoter-independent. CONCLUSIONS: Regulated expression of therapeutic candidate gene(s) coupled with suitable scaffold(s) could potentially serve as a useful tissue-engineering tool to devise future treatment strategies for osteoarthritis

Proliferation and survival of human amniotic epithelial cells during their hepatic differentiation

Stem cells derived from placental tissues are an attractive source of cells for regenerative medicine. Amniotic epithelial cells isolated from human amnion (hAECs) have desirable and competitive characteristics that make them stand out between other stem cells. They have the ability to differentiate toward all three germ layers, they are not tumorigenic and they have immunosuppressive properties. Although liver transplantation is the best way to treat acute and chronic hepatic failure patients, there are several obstacles. Recently, stem cells have been spotlighted as alternative source of hepatocytes because of their potential for hepatogenic differentiation. In this work, we aimed to study the proliferation and survival of the hAECs during their hepatic differentiation. We have also analyzed the changes in pluripotency and hepatic markers. We differentiated amniotic cells applying a specific hepatic differentiation (HD) protocol. We determined by qRT-PCR that hAECs express significant levels of SOX-2, OCT-4 and NANOG during at least 15 days in culture and these pluripotent markers diminish during HD. SSEA-4 expression was reduced during HD, measured by immunofluorescence. Morphological characteristics became more similar to hepatic ones in differentiated cells and representative hepatic markers significantly augmented their expression, measured by qRT-PCR and Western blot. Cells achieved a differentiation efficiency of 75%. We observed that HD induced proliferation and promoted survival of hAECs, during 30 days in culture, evaluated by 3H-thymidine incorporation and MTT assay. HD also promoted changes in hAECs cell cycle. Cyclin D1 expression increased, while p21 and p53 levels were reduced. Immunofluorescence analysis showed that Ki-67 expression was upregulated during HD. Finally, ERK 1/2 phosphorylation, which is intimately linked to proliferation and cell survival, augmented during all HD process and the inhibition of this signaling pathway affected not only proliferation but also differentiation. Our results suggest that HD promotes proliferation and survival of hAECs, providing important evidence about the mechanisms governing their hepatic differentiation. We bring new knowledge concerning some of the optimal transplantation conditions for these hepatic like cells.Fil: Maymo, Julieta Lorena. Universidad de Buenos Aires; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales; ArgentinaFil: Riedel, Rodrigo Nicolas. Universidad de Buenos Aires; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales; ArgentinaFil: Pérez Alcázar, Germán Antonio. Hospital Universitario Virgen Macarena;Fil: Magatti, Marta. Istituto Ospedaliero;Fil: Maskin, Bernardo. Hospital Nacional Professor Dr. Alejandro Posadas; ArgentinaFil: Dueñas, José Luis. Hospital Universitario Virgen Macarena;Fil: Parolini, Ornella. Istituto Ospedaliero;Fil: Sánchez-Margalet, Víctor. Hospital Universitario Virgen Macarena;Fil: Varone, Cecilia Laura. Universidad de Buenos Aires; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales; Argentin

IL-1 and iNOS gene expression and NO synthesis in the superior region of meniscal explants are dependent on the magnitude of compressive strains

OBJECTIVE: Partial meniscectomy is known to cause osteoarthritis (OA) of the underlying cartilage as well as alter the load on the remaining meniscus. Removal of 30-60% of the medial meniscus increases compressive strains from a maximum of approximately 10% to almost 20%. The goal of this study is to determine if meniscal cells produce catabolic molecules in response to the altered loading that results from a partial meniscectomy. METHOD: Relative changes in gene expression of interleukin-1 (IL-1), inducible nitric oxide synthase (iNOS) and subsequent changes in the concentration of nitric oxide (NO) released by meniscal tissue in response to compression were measured. Porcine meniscal explants were dynamically compressed for 2 h at 1 Hz to simulate physiological stimulation at either 10% strain or 0.05 MPa stress. Additional explants were pathologically stimulated to either 0% strain, 20% strain or, 0.1 MPa stress. RESULTS: iNOS and IL-1 gene expression and NO release into the surrounding media were increased at 20% compressive strain compared to other conditions. Pathological unloading (0% compressive strain) of meniscal explants did not significantly change expression of IL-1 or iNOS genes, but did result in an increased amount of NO released compared to physiological strain of 10%. CONCLUSION: These data suggest that meniscectomies which reduce the surface area of the meniscus by 30-60% will increase the catabolic activity of the meniscus which may contribute to the progression of OA

Lehrer:innengesundheit und Stressresistenz und deren Kohärenz mit der persönlichen Einstellung

Die vorliegende Masterarbeit widmet sich der eingehenden Untersuchung der subjektiven Einstellung von Lehrkräften zur individuellen Stressresistenz - mit Fokus auf zwei zentrale Fragen: - Wie beeinflussen die individuelle Haltung und eine ausgewogene Work-Life-Balance die Stressresistenz von Lehrkräften? - Welche Eigenschaften und Bewältigungsstrategien fördern das Wohlbefinden und die Stressresistenz im Lehrberuf? Bestehende Forschung zeigt, dass Lehrkräfte in einem anspruchsvollen Umfeld mit physischen und psychischen Belastungen arbeiten. Diese Arbeit schließt eine Forschungslücke, indem sie die Beziehung zwischen persönlicher Kohärenz von Lehrkräften und deren Fähigkeit zur Stressbewältigung vertieft beleuchtet. Die notwendigen Ressourcen und Fähigkeiten für aktive Bewältigung werden ebenfalls erörtert. Die methodische Herangehensweise umfasst einerseits eine theoretische Auseinandersetzung mit den vielfältigen Stressoren, Stressreaktionen und Bewältigungsmechanismen im Lehrerberuf und andererseits die Durchführung qualitativer Interviews mit Lehrpersonal, um individuelle Sichtweisen zu erfassen. Dies ermöglicht eine tiefgreifende Erforschung individueller Perspektiven und praxiserprobter Bewältigungsstrategien. Abschließend wird auf mangelnde externe Hilfsmaßnahmen und individuelle Bewältigungsstrategien der Lehrerinnen und Lehrer eingegangen. Die Arbeit bietet einen tiefen Einblick in komplexe Zusammenhänge zwischen persönlicher Einstellung und Stressresistenz von Lehrkräften und legt den Grundstein für weitere Forschungen zur Förderung der Lehrer:innengesundheit in einem sich ständig wandelnden schulischen Umfeld.This master's thesis delves into the in-depth examination of teachers' subjective attitudes toward individual stress resilience, focusing on two central questions: • How do individual attitudes and a balanced work-life balance influence the stress resilience of teachers? • What specific qualities and coping strategies promote well-being and stress resilience in the teaching profession? Existing research suggests that teachers operate in a demanding environment with both physical and psychological stressors. This study addresses a research gap by thoroughly investigating the relationship between teachers' personal coherence and their ability to cope with stress. The necessary resources and skills for active coping are also discussed. The methodological approach involves a theoretical exploration of diverse stressors, stress reactions, and coping mechanisms in the teaching profession. Additionally, qualitative interviews with teaching staff are conducted to capture individual perspectives and practical coping strategies. The analysis sheds light on the complex interplay between personal attitudes and stress resilience among teachers, laying the groundwork for further research to enhance the health and well-being of educators in an ever-evolving school environment

Digitale Wirtschaft - digitale Betriebsstätte? : eine explorative Literaturanalyse zur Identifikation möglicher Probleme einer digitalen Betriebsstätte

Die Entwicklung von Informations- und Kommunikationstechnologien ermöglicht es Unternehmen grenzüberschreitend online Geschäfte zu tätigen. An den digital erwirtschafteten Gewinnen hat jedoch der Quellenstaat kein Besteuerungsrecht dieses liegt nur bei Bestehen einer Betriebsstätte iSd Art 5 OECD-Musterabkommen vor. Der Tatbestand einer Betriebsstätte knüpft allerdings an eine physische Präsenz an, welche im digitalen Zeitalter nicht mehr notwendig ist. Eine mögliche Lösung um digitale Geschäfte adäquat zu besteuern, wäre die Einführung einer digitalen Betriebsstätte. Im Rahmen dieser Masterarbeit wurde zunächst anhand der fallstudienhaften Analyse digitaler Innovationen untersucht, worin die Herausforderungen für die Besteuerung liegen und ob die digitale Betriebsstätte geeignet ist, um einen steuerlichen Anknüpfungspunkt im Quellenstaat zu schaffen. Dabei wurde ersichtlich, dass das Konzept der digitalen Betriebsstätte noch nicht ausgereift ist. Die Analyse der möglichen Probleme der digitalen Betriebsstätte zeigte, dass die Kriterien, die zum Bestehen einer digitalen Betriebsstätte führen, noch konkretisiert werden müssen. Weiters ist eine globale Lösung erforderlich, um Rechtsunsicherheiten und Doppelbesteuerung zu vermeiden. Die Hauptproblematik besteht jedoch darin, der digitalen Betriebsstätte tatsächlich auch Gewinne zuzuweisen - eine Adaptierung bzgl. der Verrechnungspreisbestimmung ist daher notwendig. Auch wenn bei digitalen Geschäftsmodellen durch die digitale Betriebsstätte eine Besteuerung theoretisch möglich wäre, kann die Einführung dieses Konzepts nicht empfohlen werden. Dafür gibt es aus heutiger Sicht noch zu viele Herausforderungen bei der Implementierung und die Gewinnzuteilung ist zwar möglich, jedoch sehr komplex. Zudem sollte von unilateralen Maßnahmen und Zwischenlösungen abgesehen werden, denn diese würden einen globalen Konsens zusätzlich erschweren.The transformation of the economy due to information and communication technologies enables multinational enterprises to do business online all over the world. Usually the source state does not have any right to tax the amount of profit, that was generated within the country through digital businesses. This is because of tax regulations currently valid, which declare that the source state only has a taxation right, if there is a permanent establishment. The problem is, that the concept of a permanent establishment is built on a physical presence of the company. However, in the digital age this is not necessary anymore. Therefore, the concept of a digital permanent establishment is being proposed by the OECD as one way to also tax digital businesses in the source state. In the course of this thesis analysis of the technological innovations of the digital world shows the challenges for their appropriate taxation and evaluates, if the concept of a digital permanent establishment could help to find a way of an adequate taxation. The results show that although there is a possibility to tax profits generated through digital businesses, the application of the digital permanent establishment will lead to several problems. Before the implementation can start, it would be necessary to have clear definitions of the criteria that result in the presence of a digital permanent establishment. Furthermore, to avoid legal uncertainty and cases of double taxation, there is the need for a global solution. The main challenge will be to find a suitable way of profit distribution among the business units of a corporate group. In consideration of these challenges, it is not possible to recommend the introduction of the digital permanent establishment. Furthermore, tax authorities should not implement any unilateral measures for taxing the digital economy, as this would make a global solution even more difficult.Elisabeth Hennerbichler, BScZusammenfassungen in Deutsch und EnglischAbweichender Titel laut Übersetzung des Verfassers/der VerfasserinKarl-Franzens-Universität Graz, Masterarbeit, 2018(VLID)346278