Induction and persistence of large gammaH2AX foci by high linear energy transfer radiation in DNA-dependent protein kinase-deficient cells

BRACALENTE C; IBAÑEZ IL; MOLINARI B; PALMIERI M; KREINER A; VALDA A; J. DAVIDSON; DURAN H

INTERNATIONAL JOURNAL OF RADIATION ONCOLOGY BIOLOGY PHYSICS

ELSEVIER SCIENCE INC

Lugar: Amsterdam; Año: 2013 vol. 87 p. 785 - 794

0360-3016

Purpose: The aim of this study was to evaluate the cell response to DNA double-strand breaks (DSBs) induced by low and high linear energy transfer (LET) radiations when the catalytic subunit of DNA-PK (DNA-PKcs), an essential protein of the non-homologous end-joining repair pathway, lacks kinase activity. Methods and Materials: CHO10B2, a Chinese hamster ovary cell line, and its derived radiosensitive mutant cell line, irs-20, lacking DNA-PKcs activity, were evaluated after 0-3 Gy of gamma-rays, plateau and Bragg peak protons and lithium beams by clonogenic assay and, as a measurement of DSB, phosphorylated H2AX (gamma-H2AX) foci number and size were quantified by immunocytofluorescence. Results: Irs-20 exhibited greater radiosensitivity and higher amount of gamma-H2AX foci than CHO10B2 at 6 h after irradiation for all type of radiations. Remarkably, CHO10B2 and irs-20 maintained their difference in radiosensitivity after high-LET radiation. Six hours after low-LET radiations, irs-20 did not reach basal levels of gamma-H2AX at high doses, whereas CHO10B2 recovered basal levels for all doses. After high-LET radiation, only CHO10B2 exhibited a reduction in gH2AX foci, but it never reached basal levels. Persistent foci in irs-20 confirmed a repair deficiency. Interestingly, after 30 minutes of high-LET radiation both cell lines exhibited large foci (size >0.9 mm2) related to the damage nature, whereas at 6 hours irs-20 showed a higher amount of large foci than CHO10B2, with a 7-fold increase at 3 Gy, that could also be associated to radiosensitivity.