Analysis of chromosomal damage after in vivo exposure to ⁵⁶ Fe ions by means of mFISH and micronucleus methods

In this study, we examined the induction of chromosomal damage at day 7 following whole-body exposure of male CBA/CaJ mice to different doses of 1 GeV/amu ⁵⁶ Fe ions (0, 0.1, 0.5, and 1.0 Gy) or ¹³⁷ Cs γ rays as the reference radiation (0, 0.5, 1.0, and 3.0 Gy, using a GammaCell40). Two cytogenetic assays were used to evaluate dose-response relationships for the induction of chromosomal damage. These include: (1) the whole-genome multi-color fluorescence in situ hybridization (mFISH) technique to examine chromosomal damage in metaphases prepared from bone marrow (BM) cells and (2) the mouse in vivo micronucleus (MN) assay to evaluate chromosomal damage (induced in the bone marrow) in blood erythrocytes. By means of the mFISH method, we detected all types of aberrations from mice exposed to either ⁵⁶ Fe ions or ¹³⁷ Cs γ rays. These were translocations (Robertsonian, reciprocal and incomplete one-way types), dicentrics and breaks (both chromatid- and chromosome-types). Each type of radiation-induced significant dose-dependent increases (ANOVA, p < 0.01) in the frequencies of chromosomal damage (including the numbers of abnormal cells). Our data indicated that the ⁵⁶ Fe ions were more effective (per unit dose) than ¹³⁷ Cs γ rays in inducing damage: about four times for abnormal cells or breaks (both chromatid- and chromosome-types), and 1.6 times for exchanges (all types). Complex chromosome rearrangements were also found in BM cells of mice exposed to 1.0 Gy of ⁵⁶ Fe ions or 3.0 Gy of ¹³⁷ Cs γ rays, but their frequencies were low. Moreover, the frequencies of complex exchanges found at day 7 after exposure of mice to 1.0 Gy of ⁵⁶ Fe ions or 3.0 Gy of ¹³⁷ Cs γ rays were similar. By means of the blood MN assay, we detected dose-dependent increases in the frequencies of MN in normochromatic erythrocytes (NCE or mature red blood cells) at day 7 following in vivo exposure to ⁵⁶ Fe ions or ¹³⁷ Cs γ rays. In contrast, only a slight increase in the frequency of MN in polychromatic erythrocytes (PCE or immature red blood cells) was detected at day 7 in blood samples of mice exposed to ⁵⁶ Fe ions, but not ¹³⁷ Cs γ rays. The MN-NCE data showed that ⁵⁶ Fe ions were approximately four times more efficient in inducing chromosomal damage than ¹³⁷ Cs γ rays. The results demonstrated the sensitivity of the mouse in vivo MN assay in detecting chromosomal damage that was induced in the bone marrow of mice exposed in vivo to varying doses of ⁵⁶ Fe ions, when appropriate cell types were used for the analysis at a specific time post-irradiation. However, the MN assay is incapable of detecting stable-type chromosomal damage (e.g., translocation) making the MN assay less informative than the mFISH method.