Thermally Excited Resonating Membrane Mass Flow Sensor

A mass flow sensor based on the frequency shift of a resonating microstructure is being developed, using a measurement principle of the thermoanemometry type. The sensor is to be applied for mass flows up to 10 standard cubic centimeters per minute (sccm; 10sccm = 0.17 mg s-1), with a high sensitivity, a high resolution and a fast response. Here we report on the first prototype consisting of a 2 μm thick membrane: the temperature elevation of the thermally excited vibrating membrane affects its resonance frequency. The three-dimemsional heat transfer within the membrane and the mass flow is modeled, and expressions are derived for the resonance frequencies of initially curved and stressed membranes. Experiments have been carried out for nitrogen flows of up to 500 sccm passing over thermally excited membranes. Predicted and measured values for the shift of the resonance frequency agree well. The sensitivity of the average temperature elevation to the mass flow is quite small: at 10 sccm the cooling effect of the mass flow is only 0.2% of the heat loss by conduction to the substrate. At a resonance frequency of 5.0 kHz, and an average temperature elevation of the mebrane of 8°C, this still leads to a frequency change of 13 Hz in the mass flow range from zero to 10 sccm. Suggestions are presented for increasing the sensitivity of the sensor

Subgruppen-spezifische in vivo Transduktion von CD4+ und CD8+ T-Zellen

T cell-based immunotherapy is a promising approach in treating haematological and solid malignancies. The specificity of a T cell is determined by its T cell receptor (TCR). The TCR recognizes epitopes within peptides that are bound to major histocompatibility complex (MHC) molecules. Current T cell-based therapies focus on CD8+ T cells, disregarding the CD4+ T cell population. The inclusion of CD4+ T cells could lead to a more complete T cell response. Current manufacturing protocols utilise ex vivo transduction, a protracted and costly process unsuited for generating mixed T cell products. The aim of this thesis was to show subset-specific gene transfer targeting T cell subsets in vitro and in vivo with a gammaretroviral (gRV) vector system pseudotyped with a Measles virus (MV) envelope. This approach simplifies the manufacturing process of mixed therapeutic T cell products while bypassing ex vivo culture. In the scope of this thesis, TCRs targeting a model antigen, Influenza A/PR/8/34 H1N1 hemagglutinin (HA), were selected, sequenced, optimised, and integrated into MV pseudotyped gRV specific for either CD4 (MVm4) or CD8 (MVm8). The TCRs used were two MHC class I H 2Kd-restricted TCRs called Clone 1 and Clone 4 as well as the MHC class II H2 IEd restricted SFE TCR. Two cancer cell lines capable of tumorigenesis in immunocompetent BALB/c mice were chosen for in vitro and in vivo evaluation, and derivate cell lines expressing the target antigen were generated. The MV pseudotyped gRV vectors MVm4 and MVm8 were used to transduce unsorted splenocytes in vitro, delivering a transgene encoding for the model TCRs to their corresponding subsets. CD8+ T cells transduced with either Clone 1 or Clone 4 TCR exhibited secretion of IFNγ relative to the dose of antigenic stimulus upon co-culture with peptide loaded splenocytes and antigen-expressing cancer cell lines. Correspondingly, CD4+ T cells transduced with the SFE TCR showed antigen dependent secretion of IL-2 in co culture experiments with dendritic cells loaded with peptide or cancer cell lysate. MVm4 and MVm8 were tested in an in vivo model - they were applied systemically to BALB/c mice to redirect their T cell response against the antigenic stimulus of HA-expressing cancer cells. Homing, expansion, and contraction of T cells was monitored via live imaging for a timeframe of 73 days by luciferases co-expressed with the TCR. In conclusion, the MV pseudotyped gRV system was successfully adopted to BALB/c mice, confirming targeted in vivo transduction of CD8+ T cells and providing the first evidence of targeted in vivo transduction of CD4+ T cells leading to a functional MHC class II restricted T cell response.T-Zell-basierte Immuntherapie ist ein vielversprechender Fortschritt in der Behandlung von hämatologischen und soliden malignen Erkrankungen. Der T-Zell-Rezeptor (TCR) bestimmt die Spezifität von T-Zellen. Mittels TCR erkennen T-Zellen Epitope innerhalb von Peptiden, die durch Haupthistokompatibilitätskomplex-Moleküle (MHC) präsentiert werden. Derzeitige Immuntherapien fokussieren sich auf CD8+ T-Zellen und vernachlässigen die Rolle von CD4+ T Zellen. Der Einschluss von CD4+ T-Zellen würde eine umfassendere T-Zell-Antwort induzieren. Aktuelle Herstellungsprotokolle verwenden langwierige und kostspielige ex vivo Transduktion, ein Prozess, der nicht für die Herstellung gemischter T-Zell-Produkte geeignet ist. Das Ziel dieser Dissertation war der T-Zell Subgruppen-spezifische in vitro und in vivo Gentransfer mittels Masernvirus (MV) Hüllprotein-pseudotypisierter gammaretroviraler Vektoren (gRV). Diese Methode vereinfacht den Herstellungsprozess T-Zell-basierter Immuntherapien und umgeht die ex vivo T-Zell-Kultur. Im Rahmen dieser Dissertation wurden TCRs ausgewählt, die ein Modellantigen, Influenza A/PR/8/1934 H1N1 Hämagglutinin (HA), erkennen. Diese wurden sequenziert, optimiert und in MV Hüllprotein pseudotypisierte gRV, spezifisch entweder für CD4 (MVm4) oder CD8 (MVm8), integriert. Die verwendeten TCRs waren die zwei MHC Klasse I H 2Kd restringierten TCR Clone 1 und Clone 4, sowie der MHC Klasse II H2-IEd restringierte SFE TCR. Es wurden zwei Krebszelllinien für in vitro und in vivo Versuche ausgewählt, die eine Tumorerkrankung in immunkompetenten BALB/c Mäusen etablieren und Antigen exprimierende Derivate dieser Zelllinien generiert. Mit den MV Hüllprotein-pseudotypisierten gRV MVm4 und MVm8 wurden unsortierte Splenozyten in vitro transduziert und ein Transgen, kodierend für den passenden TCR für die jeweilige Subgruppe, übertragen. Transduzierte CD8+ T-Zellen, die entweder Clone 1 oder Clone 4 TCR exprimierten, zeigten eine dosisabhängige IFNγ-Sekretion in Ko Kultur Experimenten nach Antigenstimulus, sowohl durch Peptid-beladene Splenozyten als auch durch Antigen-exprimierende Zelllinien. Demgemäß sekretierten SFE TCR transduzierte CD4+ T-Zellen dosisabhängig IL-2 in Ko-Kultur-Experimenten mit Peptid- oder Krebszelllysat-beladenen dendritischen Zellen. MVm4 and MVm8 wurden in einem in vivo Modell erprobt, indem sie systemisch BALB/c Mäusen appliziert wurden, um deren T Zellantwort gegen HA-exprimierende Krebszelllinien zu richten. Homing, Expansion und Kontraktion der T-Zellen wurde über einen Zeitraum von 73 Tagen im Live-Imaging mittels, neben dem TCR koexprimierter, Luziferase dargestellt. Zusammenfassend lässt sich sagen, dass das MV Hüllprotein pseudotypisierte gRV-System die spezifische in vivo Transduktion von CD8+ T-Zellen in BALB/c Mäusen zeigen konnte, weiterführend wurden erstmalig die zielgerichtete in vivo-Transduktion von CD4+ T-Zellen und die resultierende MHC Klasse II restringierte T-Zellantwort gezeigt

el caso de un centro de idiomas: consideraciones acerca de una captación de estudiantes más eficaz

Incluye bibliografía y anexosEl presente trabajo analiza los diferentes elementos que tienen relevancia para las estrategias de captación de estudiantes de un Centro de Idiomas (CI) que opera en el marco de una universidad privada en Montevideo, Uruguay. La investigación partió de una demanda institucional, expresada por el centro en la fase inicial del estudio. Esta se basó en la necesidad de conocer las motivaciones de los estudiantes para la elección de cursos de lenguas extranjeras, con el fin de orientar la toma de decisiones respecto a la oferta y los niveles de los cursos, así como para definir la comunicación y el marketing de las actividades del centro. La demanda referida apunta a un aumento de estudiantes matriculados en los cursos de lenguas extranjeras del CI. El estudio se realizó mediante el método de casos que integró técnicas cualitativas y cuantitativas de investigación. Se realizaron entrevistas con dos estudiantes del CI y un estudiante de la universidad, además de la entrevista exploratoria con la coordinadora del CI al inicio del estudio. También se analizaron documentos proporcionados por el centro. La combinación de métodos se hizo con el objetivo de favorecer la triangulación de datos. En el trabajo se abordaron las motivaciones para el estudio de lenguas extranjeras, el marketing educativo en la era digital, el trabajo en equipo, así como los procesos de desarrollo institucional; todo en pos de una mejor estrategia de captación de estudiantes. El liderazgo y el trabajo en equipo son presentados como promotores de la mejora proyectada. A partir de los resultados de la investigación se elaboró un plan de mejora, en acuerdo con el centro estudiado, con el fin de atender la problemática señalada y proponer posibles vías de solución. A modo de conclusión se propone un modelo que reúne los tres principales factores que inciden en la captación de estudiantes del centro de idiomas: el marketing educativo, el apoyo institucional y el contexto coyuntural

"Designer cytokines" targeting the tumor vasculature - think global and act local

Tumor necrosis factor (TNF) was discovered in 1975 as a lipopolysaccharide-induced serum factor that causes necrosis of tumors (Carswell et al, 1975). It was later found that TNF and cachectin, a factor causing wasting disease, were one and the same molecule (Beutler et al, 1985). Studies on the inflammatory activity of TNF have been translated into clinical success, namely blocking antibodies used to suppress autoimmune diseases. Research on TNF anti-tumor activity, in contrast, has not yet resulted in a therapeutic breakthrough. This may change, based on a study by Huyghe et al (2020) describing novel "designer cytokines" (TNF and interferon-γ) that increase local activity by targeting the CD13-positive tumor vasculature, while simultaneously lowering the binding affinity to the respective cytokine receptor, thereby reducing off-target effects on normal cells

Stress hormones or general well-being are not altered in immune-deficient mice lacking either T- and B- lymphocytes or Interferon gamma signaling if kept under specific pathogen free housing conditions

It is controversially discussed whether immune-deficient mice experience severity in the absence of infection. Because a comprehensive analysis of the well-being of immune-deficient mice under specific pathogen free conditions is missing, we used a multi-parametric test analyzing, corticosterone, weight, nest building and facial expression over a period of 9 month to determine the well-being of two immune-deficient mouse lines (recombination activating gene 2- and interferon gamma receptor-deficient mice). We do not find evidence for severity when comparing immune-deficient mice to their heterozygous immune-competent littermates. Our data challenge the assumption that immune-deficiency per se regardless of housing conditions causes severity. Based on our study we propose to use objective non-invasive parameters determined by laboratory animal science for decisions concerning severity of immune-deficient mice