Durante, M.

Korf, H.-W.

Müller, M.

Natale, F.

Stöcker, H.

GSI Repository

GSITemplate2007 Response of organotypic slice cultures to ionizing radiation Mareike Müller1,2,3,*, Marco Durante2,3,4, Francesco Natale2, Horst Stöcker2,3, Horst-Werner Korf1 1Dr. Senckenbergisches Chronomedizinisches Institut, Goethe-Universität Frankfurt/Main 2GSI Darmstadt, Germany 3Frankfurt Institute for Advanced Studies, Frankfurt/Main 4Technische Universität Darmstadt.The aim of this work was to evaluate the response of liver and pancreas tissue to sparsely and densely ionizing ra-diation. For this, organotypic slice cultures (OSC) of liver (OLSC) and organotypic explant cultures (OEC) of pan-creas (OPEC) were prepared from C3H wildtype (wt) mice and transgenic c-myc/TGF-α mice with an inducible liver tumor. OLSC from the transgenic c-myc/TGF-α mice were prepared from healthy (OHLSC) and neo-plastic (ONLSC) parts of the liver. The pancreas from these animals was cultured as well (OTPEC). In order to evaluate a possible timedependence of the tissue response to ionizing radiation, OLSC and OPEC, OHLSC, ONLSC, and OTPEC were prepared at two different times of day: at the middle of the subjective day and at the middle of the subjective night. Samples were cultured in a membrane-based culturing system with a liquid-air interface for several days. OLSC and OPEC from C3H wt-mice were irradiated with X-rays at doses of 2 Gy, 5 Gy, or 10 Gy. OHLSC, ONLSC, OTPEC were irradiated with 12C-ion extended Bragg peaks at the same doses. Mock-irradiated samples served as controls. All samples were fixed 1 h and 24 hrs post-irradiation, respectively, and immunohistochemically analyzed for markers of proliferation (Ki67), apoptosis (Caspase3), and DNA double-strand breaks (γH2Ax). While the pancreas samples, unfortunately, did not pro-duce any meaningful results with regard to the evaluated parameters, healthy liver tissue showed distinct day-night differences with regard to all three analyzed parameters: the proliferation rate was significantly increased at the middle of the subjective day compared to the middle of the subjective night. Contrariwise, the apoptosis rate and rate of DNA double-strand breaks was significantly in-creased at the middle of the subjective night. These day-night differences were not detected in ONLSC. Regard-less of the radiation type and dose, irradiation of healthy liver tissue did not influence the evaluated parameters. In ONLSC, however, the rate of DNA double-strand breaks increased dose-dependently.  The effects of ionizing radiation on the circadian clockwork were further examined in tissue samples of transgenic Per2luc-mice [1]. Per2luc-mice express the en-zyme luciferase under the control of the Per2-promoter, an important element of the circadian clockwork. There-fore, the analysis of these animals allowed to record the circadian rhythm of the molecular clockwork in liver and other tissues via realtime recordings of the luciferase-activity. As could be shown in OLSC and OEC from ad-renal glands, ionizing radiation leads to a dose-dependent phase advance of the circadian clockwork.  The results of this project lead to the conclusion that ionizing radiation alter the circadian clockwork, but bare-ly influence proliferation and apoptosis in healthy liver tissue.  References [1] [Yoo2004]Yoo SH, Yamazaki S, Lowrey PL,  Shimomura K, Ko CH, Buhr ED, Siepka SM, Hong HK, Oh WJ, Yoo OJ, Menaker M, Takahashi JS „PERIOD2::LUCIFERASE real-time reporting of circadian dynamics reveals persistent circadian oscil-lations in mouse peripheral tissues“ Proc Natl Acad Sci U S A. 2004; 101(15): 5339–5346   * M. Müller is supported by a Puschmann scholarship.  GSI SCIENTIFIC REPORT 2012 HEALTH-45469

Response of organotypic slice cultures to ionizing radiation

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Response of organotypic slice cultures to ionizing radiation

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