TY - JOUR A1 - Radke, Lars A1 - Sandig, Grit A1 - Lubitz, Annika A1 - Schließer, Ulrike A1 - von Horsten, Hans Henning A1 - Blanchard, Véronique A1 - Keil, Karolin A1 - Sandig, Volker A1 - Giese, Christoph A1 - Hummel, Michael A1 - Hinderlich, Stephan A1 - Frohme, Marcus T1 - In Vitro Evaluation of Glycoengineered RSV-F in the Human Artificial Lymph Node Reactor JF - Bioengineering N2 - Subunit vaccines often require adjuvants to elicit sustained immune activity. Here, a method is described to evaluate the efficacy of single vaccine candidates in the preclinical stage based on cytokine and gene expression analysis. As a model, the recombinant human respiratory syncytial virus (RSV) fusion protein (RSV-F) was produced in CHO cells. For comparison, wild-type and glycoengineered, afucosylated RSV-F were established. Both glycoprotein vaccines were tested in a commercial Human Artificial Lymph Node in vitro model (HuALN®). The analysis of six key cytokines in cell culture supernatants showed well-balanced immune responses for the afucosylated RSV-F, while immune response of wild-type RSV-F was more Th1 accentuated. In particular, stronger and specific secretion of interleukin-4 after each round of re-stimulation underlined higher potency and efficacy of the afucosylated vaccine candidate. Comprehensive gene expression analysis by nCounter gene expression assay confirmed the stronger onset of the immunologic reaction in stimulation experiments with the afucosylated vaccine in comparison to wild-type RSV-F and particularly revealed prominent activation of Th17 related genes, innate immunity, and comprehensive activation of humoral immunity. We, therefore, show that our method is suited to distinguish the potency of two vaccine candidates with minor structural differences. KW - glycoengineering KW - fucosylation KW - RSV KW - F-protein KW - NanoString Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:526-opus4-9831 SN - 2306-5354 VL - 4 IS - 3 ER - TY - JOUR A1 - Bay, Daniyah H. A1 - Busch, Annika A1 - Lisdat, Fred A1 - Iida, Keisuke A1 - Ikebukuro, Kazunori A1 - Nagasawa, Kazuo A1 - Karube, Isao A1 - Yoshida, Wataru T1 - Identification of G-quadruplex structures that possess transcriptional regulating functions in the Dele and Cdc6 CpG islands JF - BMC Molecular Biology N2 - G-quadruplex is a DNA secondary structure that has been shown to play an important role in biological systems. In a previous study, we identified 1998 G-quadruplex-forming sequences using a mouse CpG islands DNA microarray with a fluorescent-labeled G-quadruplex ligand. Among these putative G-quadruplex-forming sequences, G-quadruplex formation was verified for 10 randomly selected sequences by CD spectroscopy and DMS footprinting analysis. In this study, the biological function of the 10 G-quadruplex-forming sequences in the transcriptional regulation has been analyzed using a reporter assay. KW - Cdc6 KW - Dele KW - G-quadruplex KW - transcriptional regulation KW - 7OTD Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:526-opus4-9811 SN - 1471-2199 VL - 18 IS - 17 ER - TY - JOUR A1 - Hornemann, Andrea A1 - Sinning, Denise A1 - Cortes, Sofia A1 - Campino, Lenea A1 - Emmer, Peggy A1 - Kuhls, Katrin A1 - Ulm, Gerhard A1 - Frohme, Marcus A1 - Beckhoff, Burkhard T1 - A pilot study on fingerprinting Leishmania species from the Old World using Fourier transform infrared spectroscopy JF - Analytical and Bioanalytical Chemistry N2 - Leishmania species are protozoan parasites and the causative agents of leishmaniasis, a vector borne disease that imposes a large health burden on individuals living mainly in tropical and subtropical regions. Different Leishmania species are responsible for the distinct clinical patterns, such as cutaneous, mucocutaneous, and visceral leishmaniasis, with the latter being potentially fatal if left untreated. For this reason, it is important to perform correct species identification and differentiation. Fourier transform infrared spectroscopy (FTIR) is an analytical spectroscopic technique increasingly being used as a potential tool for identification of microorganisms for diagnostic purposes. By employing mid-infrared (MIR) spectral data, it is not only possible to assess the chemical structures but also to achieve differentiation supported by multivariate statistic analysis. This work comprises a pilot study on differentiation of Leishmania species of the Old World (L. major, L. tropica, L. infantum, and L. donovani) as well as hybrids of distinct species by using vibrational spectroscopic fingerprints. Films of intact Leishmania parasites and their deoxyribonucleic acid (DNA) were characterized comparatively with respect to their biochemical nature and MIR spectral patterns. The strains’ hyperspectral datasets were multivariately examined by means of variance-based principal components analysis (PCA) and distance-based hierarchical cluster analysis (HCA). With the implementation of MIR spectral datasets we show that a phenotypic differentiation of Leishmania at species and intra-species level is feasible. Thus, FTIR spectroscopy can be further exploited for building up spectral databases of Leishmania parasites in view of high-throughput analysis of clinical specimens. KW - Fourier transform infrared spectroscopy KW - hierarchical cluster analysis (HCA) KW - principal components analysis (PCA) KW - Leishmania KW - DNA KW - multivariate differentiation Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:526-opus4-10080 SN - 1432-1130 ER - TY - JOUR A1 - Gottschalk, Josefine Johanna A1 - Stollfuß, Carsten A1 - Lietzau, Kai-Henning A1 - Foitzik, Andreas A1 - Richetta, Maria T1 - Conceptual study for long-term monitoring of chemotherapeutic induced cell reactions by ESPI JF - tm - Technisches Messen N2 - During the last years, various approaches on an individualized drug therapy for benign cells have been researched. However, due to the complex topic a universal approach has not been found up until this point. Commonly, the effect of cytotoxic drugs on benign cells is in most cases the same compared to regular cells while the actual effect on patient still can't be predicted. In order to reduce unwanted side effects or unspecific drug reactions a test system for patients which allows to analyse the interaction between cytotoxic agents and the targeted cells is needed. Furthermore, this should also include an adequate measurement system which is capable to work in a natural environment and without any additional preparation. In terms of this work, a first proof of concept with different benign cells and cytotoxic agents is presented while monitoring the obtained displacement using electronic speckle pattern interferometry (ESPI). KW - interferometry KW - ESPI KW - anticancer drug KW - cellular displacement Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:526-opus4-10676 SN - 2196-7113 VL - 85 IS - 2 SP - 111 EP - 118 ER -