ACRIDINE PROBE STUDY INTO SYNERGISTIC DNA-DENATURING ACTION OF HEAT AND ULTRAVIOLET LIGHT IN SQUAMOUS CELLS

Abstract

A new probe for cellular DNA configuration is described. It distinguishes between native and denatured DNA in a population of suspended cells by determining that fraction of acriflavine initially intercalated into DNA at 4°C which is released when the temperature is subsequently raised to 25°C. The relatively unstable DNA-acriflavine bonds at single-stranded sites dissociate at 25°C while those at double-stranded sites do not. Under conditions of low ionic strength and high DNA:dye molar ratios, about 70% of the dye binds to DNA at 4°C. The fluorescence of bound dye is quenched; bond dissociation restores its fluorescence, allowing simple fluorometric analysis. The probe was employed to study the DNA-denaturing potential of the heat gained by epidermal cells when they absorb solar radiation. Desquamated human buccal cells served as models for epidermal cells. They were pretreated with trypsin to remove the stabilizing influence of nucleoproteins on the condensed chromatin of these nonreplicative cells. Cell samples were irradiated by a mercury arc at 24°, 32°, and 42°C. The two latter temperature levels represented those obtained on the skin surface indoors, and in bright sunlight at 26°C north latitude, respectively. The dose response for DNA denaturation was linear, its rate increasing with increasing irradiation temperature. The results suggest that temperature levels to which the skin surface is raised in a sun belt environment can add substantially to the DNA-denaturing effect of ultraviolet on human squamous cells.