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NF-κB in the Mammalian Cellular Response to Exposure with Ionising Radiation

Ruscher, Roland (2008) NF-κB in the Mammalian Cellular Response to Exposure with Ionising Radiation. Diploma, Universität Köln.

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Abstract

During space missions, astronauts are exposed not only to greater amounts of radiation than on Earth but also to different radiation qualities, which can result in immediate and long-term risks. Activation of the Nuclear Factor κB (NF-κB) pathway resulting in increased survival of irradiated cells might contribute to cancer risk after space radiation exposure. In this work, the effect of ionising radiation and/or of inhibition or knock out of several pathway components on the NF-κB signalling was investigated. Activation of NF-κB dependent gene expression was detected by means of stably transfected human embryonic kidney cells (HEK/293) which express the reporter gene destabilized Enhanced Green Fluorescent Protein (d2EGFP) under control of a NF-κB responsive promoter (HEK-pNF-κB-d2EGFP/Neo L2). d2EGFP fluorescence was determined by means of FACS analysis after diffuse or cell nucleus-targeted irradiation with α-particles performed at the PTB, Braunschweig. Both irradiation conditions were capable of activating the NF-κB pathway with a peak at 5x10<sup>6</sup> particles per cm<sup>2</sup> (1.26 Gy) for diffuse irradiation and after seven nuclear particle traversals (1.2 Gy) for targeted irradiation. Gene expression of NF-κB-regulated genes was analysed using quantitative real time qRT-PCR. For diffuse irradiation, a time-dependent expression pattern was observed for gadd45β, nfkbia, and bcl-2. For targeted irradiation the gene expression patterns were less pronounced. As NF-κB activation is discussed to result in a survival advantage of irradiated cells, the influence of NF-κB essential modulator (NEMO) on cellular survival (colony forming ability assay) after X-irradiation was analysed using wild-type and NEMOdeficient mouse embryonic fibroblasts (MEF). Results clearly show a better survival for the NF-κB proficient cell line, supporting this hypothesis. Analysing the upstream events in the NF-κB pathway it was shown that the proteasome inhibitor MG-132 abolished EGFP expression after treatment with TNF-α or with X-rays, pointing out that proteasomal degradation of IκB is involved after both treatments. Inhibition of the ataxia telangiectasia mutated (ATM) protein kinase by KU-55933 clearly demonstrated that NF-κB activation upon DNA double strand break processing depends on functional ATM after X-irradiation but not after TNF-α treatment of HEK cells. In conclusion, NF-κB activation via ATM kinase seems to be a relevant regulator of the cellular response to ionising radiation.

Document Type:Thesis (Diploma)
Title:NF-κB in the Mammalian Cellular Response to Exposure with Ionising Radiation
Authors:
AuthorsInstitution or Email of Authors
Ruscher, RolandUNSPECIFIED
Date:2008
Number of Pages:111
Status:Unpublished
Keywords:NF-κB, Space Radiation Exposure, Space Missions, Ionising Radiation, Mammalian Cells
Institution:Universität Köln
Department:Mathematisch-Naturwissenschaftliche Fakultät, Fachbereich Biologie
HGF - Research field:Aeronautics, Space and Transport (old)
HGF - Program:Space (old)
HGF - Program Themes:W FR - Forschung unter Weltraumbedingungen (old)
DLR - Research area:Space
DLR - Program:W FR - Forschung unter Weltraumbedingungen
DLR - Research theme (Project):W - Vorhaben Strahlenbiologie (old)
Location: Köln-Porz
Institutes and Institutions:Institute of Aerospace Medicine > Radiation Biology
Deposited By: Kerstin Kopp
Deposited On:19 Jun 2008
Last Modified:27 Apr 2009 14:59

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