Variability in repair efficiencies of radiation-induced DNA strand breaks in two DBA/2 mouse lymphoma cell lines

M. Gomolka, G. Lüke, E. Konhauser, K. Hetzl, C. Schindewolf, K. Lobenwein, S. Hornhardt, R. Balling, M. Hrabé De Angelis, T. Jung

Research output: Contribution to journalArticlepeer-review

2 Scopus citations

Abstract

In the framework of a large genetic mouse screen, we are searching for mice bearing deficiencies in their DNA repair system for radiation-induced strand breaks. The alkaline version of the comet assay was chosen as a phenotypic detection system. In order to establish positive and negative controls in the genetic screen, we analysed two DBA/2 mouse cell lines with known different radiation sensitivities. According to cell survival analysis, the L5178YS (LYS) cell line is twice as sensitive to X-ray irradiation as the resistant cell line (LYR). In addition, the two cell lines display differences in their DNA repair efficiency. Cells were g-irradiated with doses ranging form 1 to 4 Gy to establish a dose-effect relationship. Repair efficiency was tested after irradiation with 2 Gy on ice and repair times ranging from 30 min to 120 min at 37°C. Slides were evaluated by a new software developed in collaboration with Till Photonics (Munich, Germany) and Narendra Singh (Seattle, USA). The comet assay demonstrated significant differences between the two cell lines in initial radiation damage and repair in various parameters. The most informative parameters were Olive tail moment, tail extended moment (Kent) and tail integrated intensities. Damage after different doses of irradiation, expressed in relative tail moments, was always higher in LYR cells than in LYS cells. Repair efficiencies for the two cell lines showed a dependence on the growth time of the cell culture; this was probably due to the cell cycle phase of the cell culture. After 72 h of growth, followed by irradiation, repair in LYS cells was more efficient during the first 30 minutes than repair in LYR cells. In contrast, 24 h growth of the cell cultures resulted in a more efficient repair in LYR cells. In addition, relative tail moments decreased also for control cells after incubation on ice or at 37°C. After 60 minutes incubation, necrotic and apoptotic cells appeared in controls as well as in irradiated cell samples. In general, we conclude that the experimental system and the software we utilised are capable of detecting DNA damage in correlation to the irradiation doses and differences in repair can be demonstrated between the cultured cell lines. Since the cell lines differ in background DNA damage, owing to culture conditions and cell isolation, and in cell cycle distribution compared to freshly isolated mouse lymphocytes they are unsuitable positive and negative controls for our screening assay.

Original languageEnglish
Pages (from-to)50-52
Number of pages3
JournalNeoplasma
Volume46
Issue numberSUPPL. 1
StatePublished - 1999
Externally publishedYes

Keywords

  • Alkaline comet assay
  • DBA/2 mouse cell lines
  • DNA repair system

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