DNA damage and survival in bystander human intestinal cells treated with conditioned medium from tritium-labeled cells

Abstract

Background: Tritium exposure could be one of the radiation hazards in case of accidental exposure with intestine as one of the major target organs. In the cells, low-energy beta emitted from tritium would traverse a very short distance (a few microns). Hence, the intestinal epithelial cells with nuclear localization of tritium would exert its radiobiological effect also through bystander mechanism. In the present study, the effect of conditioned medium obtained from tritiated thymidine-labeled human normal intestinal epithelial (INT407) cells was studied on respective bystander cells in terms of magnitude of survival and induction of DNA damage. Materials and Methods: The survival and proliferation of bystander INT407 cells treated with control/irradiated conditioned medium were studied using clonogenic and 5-bromo-2-deoxyuridine (BrdU)-labeling assays. The magnitude of DNA double-strand break was measured by immunofluorescence of η-H2AX by confocal microscopy. Intracellular nitric oxide (NO) in these cells was measured using 4,5-diaminofluorescein diacetate fluorescent dye. Results: Bystander cells treated with conditioned medium from tritiated thymidine-labeled cells showed increased clonogenic survival and BrdU labeling. Cells labeled with tritiated thymidine showed attenuation of η-H2AX foci at longer period (24 and 48 h) of labeling than at 15 h. Moreover, the bystander cells treated with irradiated conditioned medium showed a higher magnitude of η-H2AX foci at 24 h. However, compared to 24 h, 48-h treatment of irradiated conditioned medium resulted in a decrease in η-H2AX foci in the bystander cells. Increased level of intracellular NO was observed in the bystander cells treated with irradiated conditioned medium. Conclusions: Bystander cells treated with conditioned medium obtained from tritiated thymidine-labeled cells showed increased clonogenic survival and proliferation, which was correlated with an increase in DNA double-strand break and NO production in these cells.

Keywords

• bystander effect
• dna damage
• intestinal cells
• proliferation
• tritium