It is generally accepted that the effectiveness of ionizing radiation depends on the quality of the radiation beam. Densely ionizing, high-linear energy transfer (LET) types of radiation are biologically more effective than sparsely ionizing, low-LET types of radiation at inducing cell lethality for a given absorbed dose. This increased efficiency of inactivating cells by high-LET beams compared to low-LET beams is usually described by the relative biological effectiveness (RBE).
Among the various types of DNA damage, DNA double strand breaks (DSBs) are considered the most cytotoxic lesions induced by ionizing radiation. As many types of high-LET beams, including neutrons, in general do not appear to induce more DSBs than low-LET radiation [
1‐
7], it seems likely that the differences in biological effect are associated with the type of DSBs induced by radiations of differing LET and the mechanisms involved in the processing of those DSBs. It has been described that the degree of complexity of DNA DSBs and its possible association with other types of damage varies depending on the LET characteristics; therefore the biological repairability of DSBs may vary with radiation type [
3,
8,
9].
In mammalian cells, the homologous recombination (HR) and nonhomologous end-joining (NHEJ) pathways are identified as the two main mechanisms involved in the repair of DSBs. The NHEJ pathway however is regarded as the major pathway for the repair of radiation-induced DSBs in mammalian cells [
10,
11]. One of the key-players in this pathway is the Ku heterodimer, a highly stable protein complex consisting of a 70 kDa and a 86 kDa polypeptide, better known as Ku70 and Ku80 [
12,
13]. The importance of the Ku70 and Ku80 proteins in DNA DSB repair after
low-LET radiation is well demonstrated by the profound enhancement in radiosensitivity of both Ku80-defective mutant rodent cell lines (e.g. the xrs-5 and xrs-6 cell line) [
14] and human cell lines expressing reduced levels of the Ku proteins [
15‐
23]. To date however, the knowledge regarding the importance of the Ku heterodimer and the NHEJ repair mechanism in the cellular response to
high-LET radiation, including high energy neutrons, is limited and diverging results were described when using cell survival as an endpoint to analyze radiosensitivity [
3,
5,
24‐
28]. In these reports, cellular radiosensitivity was investigated in Ku-deficient rodent cell lines with a wide variety of high-LET radiation qualities (fast neutrons, α-particles, iron ions, carbon ions). When the high-LET beams used had mean LET values inferior to 100 keV/μm, the majority of these studies reported similar RBE values in the repair-deficient and -proficient cell lines [
3,
5,
24] pointing to an involvement of the Ku protein in the repair of the radiation-induced damage. When the radiation quality of the high-LET beam was superior to 100 keV/μm, RBE values close to or equal to 1 in repair-deficient cell lines were observed [
3,
25‐
27], indicating no major involvement of the NHEJ mechanism in the repair of high-LET radiation-induced damage. However, contradictory observations [
28] and the lack of studies conducted with
human Ku-deficient cell lines suggests the importance of further research into the biological mechanisms involved in the cellular response to high-LET radiation, especially given the growing interest and use of high-LET radiation in radiotherapy [
29,
30].
In the present study, we investigated the role of the Ku heterodimer in the repair of DNA lesions induced by p(66)+Be(40) neutrons (mean LET ~20 keV/μm) and 6 MV X-rays. After knockdown of the Ku heterodimer by lentiviral-mediated RNA interference (RNAi) of Ku70 in a human mammary epithelial cell line (MCF10A), cellular radiosensitivity was measured using a crystal violet cell proliferation assay, while chromosomal radiosensitivity was evaluated using the micronucleus (MN) assay.