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Budakeszi, Hungary

Bender B.,Institute of Animal Biotechnology | Bender B.,ImmunoGenes Kft | Ivett Hoffmann O.,Institute of Animal Biotechnology | Negre D.,Ecole Normale Superieure de Lyon | And 3 more authors.
BioTechniques | Year: 2013

Efficient production of transgenic animals using low-titer lentiviral constructs remains challenging. Here we demonstrate that microinjec-tion of simian immundeficiency virus-derived lentiviral constructs can produce transgenic mice and rats with high efficiency even when using low-titer virus preparations.

Szebenyi K.,Hungarian Academy of Sciences | Furedi A.,Hungarian Academy of Sciences | Kolacsek O.,Hungarian Academy of Sciences | Csohany R.,Semmelweis University | And 11 more authors.
Journal of the American Society of Nephrology | Year: 2015

Intrarenal changes in cytoplasmic calciumlevels have a key role in determining pathologic and pharmacologic responses inmajor kidney diseases. However, cell-specific delivery of calcium-sensitive probes in vivo remains problematic. We generated a transgenic rat stably expressing the green fluorescent protein-calmodulin- based genetically encoded calcium indicator (GCaMP2) predominantly in the kidney proximal tubules. The transposon-based method used allowed the generation of homozygous transgenic rats containing one copy of the transgene per allele with a defined insertion pattern, without genetic or phenotypic alterations. We applied in vitro confocal and in vivo two-photon microscopy to examine basal calcium levels and ligand- and drug-induced alterations in these levels in proximal tubular epithelial cells. Notably, renal ischemia induced a transient increase in cellular calcium, and reperfusion resulted in a secondary calcium load, which was significantly decreased by systemic administration of specific blockers of the angiotensin receptor and the Na-Ca exchanger. The parallel examination of in vivo cellular calcium dynamics and renal circulation by fluorescent probes opens new possibilities for physiologic and pharmacologic investigations. Copyright © 2015 by the American Society of Nephrology.

Cervenak J.,ImmunoGenes Kft | Doleschall M.,Hungarian Academy of Sciences | Bender B.,ImmunoGenes Kft | Mayer B.,Semmelweis University | And 8 more authors.
mAbs | Year: 2013

Among the many functions of the neonatal Fc receptor (FcRn) for IgG, it binds to IgG-opsonized antigen complexes and propagates their traffic into lysosomes where antigen processing occurs. We previously reported that transgenic (Tg) mice and rabbits that carry multiple copies and overexpress FcRn have augmented humoral immune responses. Nuclear factor-kappa B (NFκB) is a critical molecule in the signaling cascade in the immune response. NFκB induces human FcRn expression and our previous in silico analysis suggested NFκB binding sites in the promoter region of the bovine (b) FcRn α-chain gene (FCGRT). Here, we report the identification of three NFκB transcription factor binding sites in the promoter region of this gene using luciferase reporter gene technology, electromobility shift assay and supershift analysis. Stimulation of primary bovine endothelial cells with the Toll-like receptor-4 ligand lipopolysaccharide (LPS), which mediates its effect via NFκB, resulted in rapid upregulation of the bFcRn expression and a control gene, bovine E-selectin. This rapid bFcRn gene induction was also observed in the spleen of bFcRn Tg mice treated with intraperitoneally injected LPS, analyzed by northern blot analysis. Finally, NFκB-mediated bFcRn upregulation was confirmed at the protein level in macrophages isolated from the bFcRn Tg mice using flow cytometry with a newly developed FcRn specific monoclonal antibody that does not cross-react with the mouse FcRn. We conclude that NFκB regulates bFcRn expression and thus optimizes its functions, e.g., in the professional antigen presenting cells, and contributes to the much augmented humoral immune response in the bFcRn Tg mice. © 2013 Landes Bioscience.

Schneider Z.,Eotvos Lorand University | Schneider Z.,Texas A&M University | Jani P.K.,ImmunoGenes Kft | Szikora B.,Eotvos Lorand University | And 11 more authors.
Frontiers in Immunology | Year: 2015

The neonatal Fc receptor (FcRn) plays key roles in IgG and albumin homeostasis, maternal IgG transport and antigen presentation of IgG-opsonized antigens. Previously we reported that transgenic (Tg) mice that overexpress the bovine FcRn (bFcRn) have augmented T dependent humoral immune response with increased IgG protection, higher level of antigen specific antibodies, greater number of antigen-specific B cells and effective immune response even against of weakly immunogenic epitopes. In the current study we analyzed the localization of the bFcRn in secondary lymphoid organs, and focused to demonstrate the in vivo impact of its overexpression in the spleen on the course of antibody production. bFcRn was highly expressed by red pulp macrophages and marginal zone macrophages in the spleen and by subcapsular sinus macrophages and macrophage-like cells in the interfollicular areas in the lymph node cortex. We also demonstrated that splenic dendritic cells of Tg mice express bFcRn and intraperitoneal immunization of these mice with T-dependent antigens led to more than threefold increase in the number of many antigen-specific activated T helper cells with increased size and numbers of germinal centers compared to wild type (wt) controls. bFcRn expression in splenic B cells was also detected and that may also contribute to the enhanced B cell activation. Finally, we demonstrated that these Tg mice developed efficient immune response against very low-dose of antigen, reflecting another important practical benefit of these Tg mice. © 2015 Schneider, Jani, Bence, Végh, Kövesdi, Iliás, Cervenak, Balogh, Kurucz and Kacskovics.

Vegh A.,ImmunoGenes Kft | Farkas A.,Eotvos Lorand University | Kovesdi D.,Eotvos Lorand University | Papp K.,Eotvos Lorand University | And 9 more authors.
PLoS ONE | Year: 2012

Our previous studies have shown that overexpression of bovine FcRn (bFcRn) in transgenic (Tg) mice leads to an increase in the humoral immune response, characterized by larger numbers of Ag-specific B cells and other immune cells in secondary lymphoid organs and higher levels of circulating Ag-specific antibodies (Abs). To gain additional insights into the mechanisms underlying this increase in humoral immune response, we further characterized the bFcRn Tg mice. Our Western blot analysis showed strong expression of the bFcRn transgene in peritoneal macrophages and bone marrow derived dendritic cells; and a quantitative PCR analysis demonstrated that the expression ratios of the bFcRn to mFcRn were 2.6- and 10-fold in these cells, respectively. We also found that overexpression of bFcRn enhances the phagocytosis of Ag-IgG immune complexes (ICs) by both macrophages and dendritic cells and significantly improves Ag presentation by dendritic cells. Finally, we determined that immunized bFcRn mice produce a much greater diversity of Ag-specific IgM, whereas only the levels, but not the diversity, of IgG is increased by overexpression of bFcRn. We suggest that the increase in diversity of IgG in Tg mice is prevented by a selective bias towards immunodominant epitopes of ovalbumin, which was used in this study as a model antigen. These results are also in line with our previous reports describing a substantial increase in the levels of Ag-specific IgG in FcRn Tg mice immunized with Ags that are weakly immunogenic and, therefore, not affected by immunodominance. © 2012 Végh et al.

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