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Kraków, Poland

Wolkow P.P.,Jagiellonian University | Kosiniak-Kamysz W.,Jagiellonian University | Osmenda G.,Jagiellonian University | Wilk G.,Jagiellonian University | And 10 more authors.

Background: The genetic background of atherosclerosis in type 2 diabetes mellitus (T2DM) is complex and poorly understood. Studying genetic components of intermediate phenotypes, such as endothelial dysfunction and oxidative stress, may aid in identifying novel genetic components for atherosclerosis in diabetic patients. Methods: Five polymorphisms forming two haplotype blocks within the GTP cyclohydrolase 1 gene, encoding a rate limiting enzyme in tetrahydrobiopterin synthesis, were studied in the context of flow and nitroglycerin mediated dilation (FMD and NMD), intima-media thickness (IMT), and plasma concentrations of von Willebrand factor (VWF) and malondialdehyde (MDA). Results: Rs841 was associated with FMD (p = 0.01), while polymorphisms Rs10483639, Rs841, Rs3783641 (which form a single haplotype) were associated with both MDA (p = 0.012, p = 0.0015 and p = 0.003, respectively) and VWF concentrations (p = 0.016, p = 0.03 and p = 0.045, respectively). In addition, polymorphism Rs8007267 was also associated with MDA (p = 0.006). Haplotype analysis confirmed the association of both haplotypes with studied variables. Conclusions: Genetic variation of the GCH1 gene is associated with endothelial dysfunction and oxidative stress in T2DM patients. © 2014 Wolkow et al.This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Source

Cremer C.,University Hospita | Vierbuchen T.,University Hospita | Hein L.,University Hospita | Fischer R.,Fraunhofer Institute for Molecular Biology and Applied Ecology | And 3 more authors.
Journal of Immunotherapy

Human cytolytic fusion proteins (hCFPs) are therapeutically efficacious recombinant polypeptides comprising a target cell-specific binding component and a human effector domain that induces apoptosis. Compared with former generations of immunotoxins, which contain immunogenic cytotoxic domains derived from bacteria or plants, hCFPs contain solely human proteins that do not induce an immune response, thus avoiding the development of neutralizing antibodies. Here, we investigated the suitability of human angiogenin (Ang) mutants as effector domains. We engineered 3 different Ang variants that outperformed the wild-type enzyme by replacing amino acid residues with key roles in the protein's catalytic activity and its interaction with the ribonuclease inhibitor RNH1. The cytotoxic potential of these mutants was compared with wild-type Ang by fusing each to the CD64-specific single-chain variable fragment H22. All hCFPs were successfully expressed in HEK293T cells and purified from the cell culture supernatant by immobilized metal ion affinity chromatography. The Ang mutant-based hCFPs showed normal binding activity towards human interferon-g-stimulated CD64+ HL-60 cells and activated human macrophages isolated from peripheral blood mononuclear cells, but increased cytotoxicity based on reduced affinity towards RNH1 and higher ribonucleolytic activity. © 2015 Wolters Kluwer Health, Inc. All rights reserved. Source

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