28. Chromatid Breaks Induced in Normal Human Cells by Heavy Ions at Initial, Metaphase and Interphase States

Yoshiya Furusawa, Tetsuya Kawata¹², Kerry George², Durante Marco³, Honglu Wu², Veronica Willingham², Hisao Ito¹ and Francis A. Cucinotta²
(¹Chiba Univ.; ²Johinson Space Center, NASA; ³Univ. Fedellico II)

Keywords: chromosome aberrations, high-LET radiations, initial breaks, cell-cycle, PCC-FISH

To investigate initial chromatid breaks and how cell-cycle delays in human cells affect the expression of complex chromosome damage in metaphase following high- and low-LET radiations, we irradiated whole blood or exponentially growing human fibroblast cells AG1522 in vitro with a low and a high dose of iron, neon, carbon, and silicon particles or -rays. Lymphocytes were cultured and meta-phase cells were collected at different time points after 48-84 h in culture. Interphase chromosomes were prematurely condensed using calyculin-A, either 48 or 72 h after exposure to iron particles or -rays. Cells in the first division were analyzed using a combination of FISH whole-chromosome painting and DAPI/Hoechst 33258 harlequin staining. Chromosomes of AG1522 cells were prematurely condensed using calyculin-A. Initial chromatid-type and isochromatid breaks in G2 cells were scored.

The dose-response for total chromatid breaks was linear regardless of radiation type. The relative biological effectiveness (RBE) increased with LET, and showed a maximum at 55-80 keV/µm, then decreased at higher LET. Induction of isochromatid-type breaks was linear to dose for high-LET radiations, but linear-quadratic for -rays or 13 keV/µm carbon beams. The RBE for the induction of isochromatid breaks obtained from linear components increased rapidly between 13 keV/µm and 80 keV/µm carbon beams and decreased gradually for higher LET beams. High-LET radiations were more effective in induction of isochromatid breaks, while low-LET radiations were more effective in induction of chromatid-type breaks. The densely ionizing track structures of high-LET radiation and the geometry of sister chromatids in G2 cells resulted in an increase in isochromatid breaks.

The delay of chromosome damage expression in metaphase was LET- and dose-dependant. This delay was mostly related to the late emergence of complex-type damage into metaphase. Yields of damage in PCC collected 48 h after irradiation with iron particles were similar to that obtained from cells undergoing mitosis after prolonged incubation. The yield of high-LET radiation-induced complex chromosome damage could be underestimated when analyzing metaphase cells collected at one time point after irradiation. Chemically induced PCC method might be a more accurate technique, because the problems with complicated cell-cycle delays would be avoided.

Publications:
1) George, K., Wu, H., Willingham, V., Furusawa, Y., Kawata, T. and Cucinotta F. A.: Int. J. Radiat. Biol. 77, 175-183, 2001.
2) Kawata, T., Durante, M., Furusawa, Y., George, K., Takai, N., Wu, H. and Cucinotta F.A.: Int. J. Radiat. Biol. 77, 165-174, 2001.
3) Kawata, T., Durante, M., Furusawa, Y., George, K., Ito, H., Wu, H and Cucinotta F. A.: Radiat. Res. 156, 598-602, 2001.