EFFECTS OF 5-FLUOROURACIL ON EMBRYONIC RAT PALATE INVITRO - FUSION INTHE ABSENCE OF PROLIFERATION

5-Fluorouracil (5-FU) inhibits the enzyme thymidylate synthetase (TS) which results in inhibition of DNA synthesis. 5-FU is teratogenic in m any species, inducing cleft palate, limb, and tail defects. In the pre sent study, gestation day (GD) 14 embryonic rat craniofacial explants were exposed to 5-FU in organ culture with increasing concentrations a nd durations of exposure. Palates exposed to 5-FU were morphologically abnormal and craniofacial shape, size, and palatal fusion pattern wer e affected with the severity of effects dependent on concentration and duration of exposure. Cleft palate was induced in vitro as opposing p alates overlapped in a narrowed oral cavity. Palates exposed to higher levels of 5-FU were growth inhibited, but fused even though prolifera tion ceased and few cells were available to participate in elevation a nd fusion. This was demonstrated as a biphasic concentration-response profile for palatal fusion in which 0.05 to 0.15 mug 5-FU/ml produced decreasing rates of palatal fusion, while exposure to 0.15 to 3.0 mug/ ml resulted in progressively increasing rates of fusion. The effects o f 5-FU were detected biochemically as a reduction in TS activity which was concentration and time dependent during the first 12 hours, with a return to control levels by 24 hours. During the first day, 5-FU did not alter protein levels, but DNA levels significantly decreased at t he high concentration, 2.0 mug/ml. After 5 days in culture, both DNA a nd protein decreased with increasing 5-FU concentration and duration o f exposure. Also by the end of the culture period, H-3-TdR incorporati on had decreased in a concentration dependent manner. It is concluded that progressive inhibition of proliferation and growth in organ cultu re results in two different morphological outcomes: cleft palate resul ting from a narrowed oral cavity and increased incidence of anterior p alatal fusion under conditions of strong growth reduction. This study demonstrates that elevation and fusion can occur in the absence of gro wth and proliferation. Based on these observations, severe inhibition of growth or proliferation would not necessarily be sufficient to indu ce cleft palate.