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Neural Tube Closure in Mouse Whole Embryo Culture
Published on: October 21, 2011
Isotretinoin teratogenicity in mouse whole embryo culture
Abstract:
Recent clinical observations strongly suggest that isotretinoin [13-cis-retinoic acid (cis RA)] is a human teratogen causing primarily heart and craniofacial malformations including ear and palatal defects. The purpose of the present study was to determine if cis RA could induce similar craniofacial malformations in mouse embryo culture. Day 8 CD-1 mouse embryos were cultured for 48 hours in rat serum in the presence or absence of various concentrations of cis RA dissolved in DMSO. DMSO by itself had no effect on embryonic development; however, cis RA at 2 X 10(-5) M (6 micrograms/ml) was clearly toxic. At 2 X 10(-6) M cis RA, growth retardation was minimal, and approximately one-third of the embryos exhibited very specific defects including a dramatic reduction in the size of the first and second visceral arches, which eventually give rise to the maxilla, mandible, and ear. Similar observations were also made with 4-oxo-13-cis RA, which is a major metabolite of cis RA in the mouse and human. These malformations would be expected to result in defects similar to those observed in the human, and preliminary observations suggest these defects are due to cis RA-induced inhibition of cranial neural crest cell migration. Using day-10 mouse embryos cultured for 48 hours in Waymouth's medium containing 50% fetal calf serum, we observed that cis RA at 2 X 10(-5) M produced a high percentage of embryos with limb defects and median cleft lip. Our results demonstrate that labeled cis RA enters the tissues of the embryo both in vivo and in vitro. Cis RA inhibited proliferation of the frontonasal mesenchyme cells in primary culture with 31% inhibition occurring at 2 X 10(-5) M cis RA.
Insights
Isotretinoin, or 13-cis-retinoic acid (cis RA), causes birth defects. Studies show cis RA induces craniofacial and limb malformations in mouse embryos, likely by inhibiting neural crest cell migration.
Area of Science:
- Developmental Biology
- Teratology
- Embryology
Background:
- Clinical observations link isotretinoin (13-cis-retinoic acid, cis RA) to human birth defects, particularly heart and craniofacial malformations.
- The teratogenic potential of cis RA necessitates further investigation into its specific effects on embryonic development.
Purpose of the Study:
- To determine if cis RA can induce craniofacial malformations in cultured mouse embryos.
- To investigate the mechanism by which cis RA may cause these developmental abnormalities.
Main Methods:
- Mouse embryos (CD-1, day 8) were cultured in vitro with varying concentrations of cis RA.
- Embryos were analyzed for malformations, growth retardation, and cell proliferation inhibition.
- Labeled cis RA was used to track its entry into embryonic tissues.
Main Results:
- cis RA at 2 X 10(-6) M induced craniofacial defects, including reduced first and second visceral arches, in approximately one-third of cultured embryos.
- Higher concentrations (2 X 10(-5) M) of cis RA were toxic and induced limb defects and median cleft lip in day-10 embryos.
- cis RA inhibited frontonasal mesenchyme cell proliferation by 31% at 2 X 10(-5) M and was shown to enter embryonic tissues.
Conclusions:
- cis RA is a potent teratogen in mouse embryos, inducing craniofacial and limb malformations similar to those observed in humans.
- cis RA-induced malformations may result from inhibited cranial neural crest cell migration and reduced cell proliferation.
- These findings support the teratogenic classification of cis RA and highlight the importance of understanding its developmental toxicity.
