Institute of Clinical Chemistry and Pathobiochemistry
Institute of Clinical Chemistry and Pathobiochemistry
Barucker C.,Free University of Berlin |
Barucker C.,McGill University |
Harmeier A.,Free University of Berlin |
Harmeier A.,Hoffmann-La Roche |
And 13 more authors.
Journal of Biological Chemistry | Year: 2014
Although soluble species of the amyloid-β peptide Aβ42 correlate with disease symptoms in Alzheimer disease, little is known about the biological activities of amyloid-β (Aβ). Here, we show that Aβ peptides varying in lengths from 38 to 43 amino acids are internalized by cultured neuroblastoma cells and can be found in the nucleus. By three independent methods, we demonstrate direct detection of nuclear Aβ42 as follows: (i) biochemical analysis of nuclear fractions; (ii) detection of biotin-labeled Aβ in living cells by confocal laser scanning microscopy; and (iii) transmission electron microscopy of Aβ in cultured cells, as well as brain tissue of wild-type and transgenic APPPS1 mice (overexpression of amyloid precursor protein and presenilin 1 with Swedish and L166P mutations, respectively). Also, this study details a novel role for Aβ42 in nuclear signaling, distinct from the amyloid precursor protein intracellular domain. Chromatin immunoprecipitation showed that Aβ42 specifically interacts as a repressor of gene transcription with LRP1 and KAI1 promoters. By quantitative RT-PCR, we confirmed that mRNA levels of the examined candidate genes were exclusively decreased by the potentially neurotoxic Aβ42 wild-type peptide. Shorter peptides (Aβ38 or Aβ40) and other longer peptides (nontoxic Aβ42 G33A substitution or Aβ43) did not affect mRNA levels. Overall, our data indicate that the nuclear translocation of Aβ42 impacts gene regulation, and deleterious effects of Aβ42 in Alzheimer disease pathogenesis may be influenced by altering the expression profiles of disease-modifying genes. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.
PubMed | University of Liverpool, Institute of Clinical Chemistry and Pathobiochemistry, Heinrich Heine University Düsseldorf, University of Heidelberg and 2 more.
Type: Journal Article | Journal: Oncogenesis | Year: 2016
Hypoxia-inducible factor 1 (Hif1) is a key regulator of cellular adaptation and survival under hypoxic conditions. In pancreatic ductal adenocarcinoma (PDAC), it has been recently shown that genetic ablation of Hif1 accelerates tumour development by promoting tumour-supportive inflammation in mice, questioning its role as the key downstream target of many oncogenic signals of PDAC. Likely, Hif1 has a context-dependent role in pancreatic tumorigenesis. To further analyse this, murine PDAC cell lines with reduced Hif1 expression were generated using shRNA transfection. Cells were transplanted into wild-type mice through orthotopic or portal vein injection in order to test the in vivo function of Hif1 in two major tumour-associated biological scenarios: primary tumour growth and remote colonization/metastasis. Although Hif1 protects PDAC cells from stress-induced cell deaths in both scenarios-in line with the general function Hif1-its depletion leads to different oncogenic consequences. Hif1 depletion results in rapid tumour growth with marked hypoxia-induced cell death, which potentially leads to a persistent tumour-sustaining inflammatory response. However, it simultaneously reduces tumour colonization and hepatic metastases by increasing the susceptibility to anoikis induced by anchorage-independent conditions. Taken together, the role of Hif1 in pancreatic tumorigenesis is context-dependent. Clinical trials of Hif1 inhibitors need to take this into account, targeting the appropriate scenario, for example palliative vs adjuvant therapy.
Muller C.,Institute of Clinical Chemistry and Pathobiochemistry |
Gardemann A.,Institute of Clinical Chemistry and Pathobiochemistry |
Keilhoff G.,Institute of Biochemistry and Cell Biology |
Peter D.,Institute of Clinical Chemistry and Pathobiochemistry |
And 3 more authors.
Free Radical Research | Year: 2010
Excessive flux of free fatty acids (FFA) into the liver contributes to liver impairment in non-alcoholic fatty liver disease (NAFLD). It remains unclear how FFA contribute to impairment of hepatocytes. This study treated hepatocytes with linoleic acid and palmitate to investigate the early event triggering FFA-mediated impairment. It determined cell viability, content of nitrite/nitrate and triacylglycerides (TG), inducible nitric oxide synthase (iNOS) protein, oxidation of cardiolipin (CL) as well as formation of F2-isoprostanes in the presence of insulin and glucose. Linoleic acid caused significant decrease in cell viability. It is shown that palmitate caused induction of iNOS resulting in increased nitrite/nitrate concentration and slight increase in TG content. Linoleic acid led to a decrease in nitrite/nitrate concentration parallelled by massive TG accumulation in combination with increased oxidation of CL and increased F2- isoprostane levels. It is concluded that nitric oxide (NO) concentration regulates FFA-dependent TG accumulation and oxidative stress in rat hepatocytes. © 2010 Informa UK, Ltd.
PubMed | Otto Von Guericke University of Magdeburg, The University of Oklahoma Health Sciences Center, University of Health Sciences, Lahore, University of Greifswald and 4 more.
Type: Journal Article | Journal: Journal of the American Society of Nephrology : JASN | Year: 2015
Ischemia-reperfusion injury (IRI) is the leading cause of ARF. A pathophysiologic role of the coagulation system in renal IRI has been established, but the functional relevance of thrombomodulin (TM)-dependent activated protein C (aPC) generation and the intracellular targets of aPC remain undefined. Here, we investigated the role of TM-dependent aPC generation and therapeutic aPC application in a murine renal IRI model and in an in vitro hypoxia and reoxygenation (HR) model using proximal tubular cells. In renal IRI, endogenous aPC levels were reduced. Genetic or therapeutic reconstitution of aPC efficiently ameliorated renal IRI independently of its anticoagulant properties. In tubular cells, cytoprotective aPC signaling was mediated through protease activated receptor-1- and endothelial protein C receptor-dependent regulation of the cold-shock protein Y-box binding protein-1 (YB-1). The mature 50 kD form of YB-1 was required for the nephro- and cytoprotective effects of aPC in vivo and in vitro, respectively. Reduction of mature YB-1 and K48-linked ubiquitination of YB-1 was prevented by aPC after renal IRI or tubular HR injury. aPC preserved the interaction of YB-1 with the deubiquitinating enzyme otubain-1 and maintained expression of otubain-1, which was required to reduce K48-linked YB-1 ubiquitination and to stabilize the 50 kD form of YB-1 after renal IRI and tubular HR injury. These data link the cyto- and nephroprotective effects of aPC with the ubiquitin-proteasome system and identify YB-1 as a novel intracellular target of aPC. These insights may provide new impetus for translational efforts aiming to restrict renal IRI.