The effect of radiation on cellular ageing

Abstract

Humans are exposed to low and moderate doses of ionising radiation (IR) from a range of diagnostic and occupational sources which may lead to cellular effects including genomic damage, cellular senescence and cell death. Cellular senescence is interesting because it is a normal, metabolically active form of tumour-suppressive growth arrest initiated in response to intrinsic and extrinsic stressors helping to prevent the proliferation of ageing/damaged cells. However, in addition to this tumour-suppressive mechanism, senescent cells and tissues have also been linked with the development of a pro-oncogenic environment resulting from the secretion of an inflammatory phenotype. Indeed, senescence is thought to be important in a number of age-related pathologies and may influence the biological effects of radiation exposure. To assess the relationship between IR and stress-induced senescence we exposed mid and late passage primary normal human bronchial epithelial (NHBE) cells to low-LET Cobalt60 γ-rays (0-2 Gy) and fixed cells 0 min – 792 hours after exposure. DNA damage in both proliferating and senescent NHBE cells was quantified in nuclei co-stained for 53BP1 DNA damage foci (average foci/nucleus and foci size), and Ki-67 proliferation antigen, utilising manual and automated foci (AutoRIF) scoring methods. As expected, 53BP1 foci were significantly induced in mid (5.3 and 6.48 53BP1 foci/nucleus) and late (6.86 and 8.21 foci/nucleus) passage NHBE cells 30 minutes after exposure to 0.3 and 0.5 Gy respectively compared to sham (mid; 1.1 and late; 2.44 foci/nucleus); the elevated numbers of foci in the late passage cells reflect the increased background. By contrast, cells that were cultured beyond this time did not reveal any elevations in foci number and instead a decline was observed at extended times. This may result from cells with persistent DNA damage being removed from the population by apoptosis. AutoRIF analysis also determined that the proportion of medium and large foci in late passage NHBE cells was increased compared to mid passage nuclei, however no IR-induced increase in foci size was seen at late times after exposure. Additionally, approximately 30% of mid-passage NHBE cells were senescent (Ki-67 negative) compared to 64% in late-passage cells, the fractions of which were also not influenced by our single dose exposures. In conclusion, our preliminary experiments do not show any IR-modifying effects after the single acute doses examined, however the end-points studied and the base-line data generated offer an excellent platform from which to assess the long-term cellular effects of exposure to acute and fractionated exposures of IR in primary NHBE cells.