The measurement of DNA strand breaks in rat colonic mucosa by fluorometric analysis of DNA unwinding.

We describe the detection of DNA strand breaks in the rat colonic mucosa (in vivo and in vitro) by fluorometric analysis of DNA unwinding. DNA strand breaks were detected, in a dose-dependent manner, 2 h following an intracaecal dose of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG, 5-100 mg/kg). Levels of double-stranded DNA in the colonic mucosa of control animals, after 60 min unwinding time, were 62.8 +/- 3.4% (n = 6) and were significantly reduced at 5 mg/kg to 52.1 +/- 1.9% (n = 3). DNA strand breaks were also detected in isolated rat colonic mucosal cells following 15 min treatment with MNNG (0.1-1 mM) in vitro. The percentage of double-stranded DNA, after 60 min unwinding time, was significantly reduced (13.6 +/- 5.4%, n = 3) from control levels following treatment with 0.1 mM. A post-treatment incubation (30 min) of cells treated at a MNNG concentration of 0.25 mM resulted in 78.2 +/- 24.0% (n = 3) of these lesions being repaired.