Crypt fission is a physiologic mechanism of crypt reproduction. It increases in pathophysiologic situations where intestinal regeneration is required (eg, radiation injury). Polyamine metabolism is important in the regulation of intestinal growth and recovery from injury in response to a variety of stimuli. Our aim was to determine whether inhibition of polyamine synthesis by difluoromethylornithine (DFMO) influenced crypt fission. Forty-eight rabbits underwent patch enteroplasty in the terminal ileum. One group served as a control group and the other took 2% DFMO orally. Animals (n= 6) from each group were killed at 7, 14, 21, and 28 days. Normal ileum adjacent to the enteroplasty was studied. Crypt dissection was performed 2 hours after vincristine was administered intravenously to determine crypt cell production rate, crypt depth, and proportion of bifurcating crypts (fission). DFMO administration decreased crypt fission (4±2% vs. 11±2% and 13±1% vs. 34±4% at 7 and 14 days) compared to control animals. There was a corresponding increase in crypt depth at 14 and 21 days. Crypt cell production rate was similar in both groups and did not change with time. Mucosal ornithine decarboxylase activity (11.9±2.2 vs. 1.2±0.3 specific activity at 21 days; P< 0.05) and polyamine content (323±32 vs. 17±8 and 382±89 vs. 160±47 pmol/mg at 14 and 21 days, control vs. DFMO; P< 0.05) were significantly lower in the DFMO group. The following conclusions were drawn:(1) DFMO administration inhibits crypt fission in stimulated intestinal epithelium;(2) this effect correlates temporally with reduced polyamine production; and (3) reduced crypt fission is another potential mechanism of inhibition of intestinal growth by altered polyamine metabolism.