Changes in epidermal radiosensitivity with time associated with increased colony numbers

Epidermal clonogenic cell survival and colony formation following irradiati on were investigated and related to radiosensitivity. A rapid in vivolin vi tro assay was developed Tor the quantification of colonies arising from sur viving clonogenic cells in pig epidermis after irradiation. Bromodeoxyuridi ne (BrdU)-labelled cells in full thickness epidermal sheets M were visualiz ed using standard immunohistochemistry. In unirradiated skin, approximate t o 900 BrdU-positive cells mm(-2) were counted. In a time sequence experimen t, BrdU-positive cell numbers increased from an average of 900 cells mm(-2) to approximately 1400 cells mm(-2) after BrdU-labelling for 2-24 h. In irr adiated skin, colonies containing greater than or equal to 16 BrdU-positive cells were seen for the fil st time at days 14/15 after irradiation. The n umber of these colonies per cm(2) as a function of skin surface dose yielde d a cell survival curve with a D-0-value (+/- SE) of 3.9 +/- 0.6 Gy. This r elatively high Do-value is possibly due to a rapid fall off in depth dose d istribution for the iridium-192 source and consequently a substantial contr ibution of hair follicular epithelium to colony formation. At 14/15 days af ter irradiation, the ED50 level of 33.6 Gy for the in vivo response of mois t desquamation corresponded with 2.7 colonies cm(-2) Surprisingly, the numb er of colonies increased with time after irradiation with an estimated doub ling time of approximate to4 days, while the Do-value remained virtually un changed. This increase in colony numbers could be due to migration of clono genic cells, to the recruitment of dormant clonogenic cell survivors by ele vated levels of cytokines, or to both. Although frequent biopsying caused i ncreased cytokine levels, which had a systemic effect on unirradiated skin, it had no influence on colony formation in irradiated skin. Smaller coloni es, containing 4-8 cells or 9-15 cells, were abundant, particularly after h igher doses, which resulted in higher Do-values. The majority of these smal l colonies were abortive and did not progress to larger colonies. There was no statistical evidence for significant variations in the interanimal resp onses.