A Comparison of the In Vivo and In Vitro Response of Rat Embryos to 5-Fluorouracil.

Abstract

ABSTRACT. This study serves to further define the capabilities of the whole embryo culture system using the well-known teratogen, 5-fluorouracil (5-FU), an antineoplastic agent. An initial in vivo study was performed whereby pregnant rats were injected intraperitoneally with 10-30 mg/kg 5-FU on day 9 of gestation. On day 20 of gestation, the effects of this drug on the growth and development of embryos were evaluated. The number of externally malformed fetuses increased in a dose-related manner, and the most common defect was micro-/anophthalmos in fetuses of dams treated with 5-FU. Growth retardation was also noted in the 5-FU treated groups. An in vitro study was performed in which drug concentrations were varied (0.15-0.30 µg/ml). Externally abnormal embryos were observed in whole embryo culture system from embryonic day 9 to 11. The most common defect was hypoplastic optic vesicles. In the whole embryo culture system, crown-rump length, somite number, protein contents, and morphological score were decreased in a dose-dependent fashion. Finally, histological evaluation and observation of the pattern of cell death of the optic vesicle of 11-day-old embryos in in vivo and in vitro were performed. These parameters revealed no differences in response between in vivo and in vitro embryos treated with 5-FU, suggesting that the whole embryo culture system was an appropriate model for developmental toxicity studies of 5-FU. -KEY WORDS: cell death, 5-fluorouracil, histological evaluation, optic vesicle, whole embryo culture.J. Vet. Med. Sci. 60(1): 93-99, 1998 order to compare the development of the optic vesicle in vivo and in vitro. MATERIALS AND METHODSChemicals: 5-Fluorouracil (5-FU, Sigma Chemical Co., MO, U.S.A.) was dissolved in saline for treatment of pregnant females or dissolved in Tyrode's buffer (Sigma) for exposure of embryos. Methionine (Sigma) and D (+)-glucose anhydrous (E. Merck. AG., Darmstadt, Germany) were added to the culture medium.Animals: Sprague-Dawley spf rats (Charles River, Atsugi, Japan) were used under conditions of constant temperature (24 ± 1°C) and relative humidity (55 ± 5%), and a defined light-dark cycle (dark period from 7:00 P.M. to 7:00 A.M.). Food (CA-1, Clea, Japan) and water were provided ad libitum. Nine-week-old virgin females after being acclimatized for 1 week, were mated overnight with a male; the following morning was designated as day 0 of gestation if sperm were detected in the vaginal smear.Teratological evaluation: In utero experiment, at a defined time (12:00 A.M.) on day 9 of gestation, pregnant females were treated intraperitoneally with 5-FU at dose levels of 10, 15, 17, 20 and 30 mg/kg. Animals serving as the control received saline (5 ml/kg). All females were killed by deep ether anethesia and hysterectomized on day 20 of gestation. The number of early and late resorptions and viable fetuses was recorded. Viable fetuses were weighed, sexed, and examined for gross external malformations including abnormality of oral cavity under a dissecting microscope.In the...

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