POSTIRRADIATION EXPRESSION OF LETHAL MUTATIONS IN AN IMMORTALIZED HUMAN KERATINOCYTE CELL-LINE

The quantification of the extent of delayed cell death and the rate an d pattern of its occurrence in relation to the cell division cycle is important in radiotherapy and also in radiation transformation studies related to protection and dose limits. Here the numbers of lethal mut ations occurring over 45 population doublings (clonal expansion to abo ut 10(13) cells per cell originally surviving irradiation) was measure d in an HPV 16 immortalized human keratinocyte cell lines used for tra nsformation studies. The results showed that when postirradiation (dos e range 1-6 Gy) growth curves were constructed, the difference in slop es could be accounted for entirely by correcting for the non-clonogeni c fraction in the cell count, excluding a longer cell generation time as an explanation. When the cell loss was examined over the entire gro wth period of 6 weeks (about 45 doublings of the cell population), it was found to be dose dependent for the first two passages, but then to become more independent of dose. The mean loss for the population ove r the entire 45 cell doublings was about 15% at each cell division. Th ere is some indication from the data that the numbers of lethal mutati ons actually begin to increase between passage 6 and 7 at every dose t ested but this requires further investigation. The results allow a tim e/cell generation dependent factor to be derived for the cell line and used in survival curve equations where effects of radiation are being measured at times distant from the original exposure.