Zinc deficiency causes apoptosis but not cell cycle alterations in organogenesis-stage rat embryos: effect of varying duration of deficiency.

Zinc deficiency is teratogenic in all species in which it has been examined. Zinc is an essential component of enzymes involved in DNA synthesis and cell proliferation, and may play an as yet undetermined role in apoptosis. To further our understanding of the role of zinc in normal development, we examined cell death and cell cycle parameters in embryos of pregnant rats fed a zinc-deficient diet for 2 to 10 days (0.5 microgram zinc/g diet; zinc-adequate diet was 25 micrograms zinc/g). To elucidate sensitive periods of development and susceptible cell populations, dams were fed the zinc-deficient diet from gestation day 1, 3, 7, or 9 and killed on day 11. Embryos were examined for morphology and developmental stage. From each litter, 2-3 embryos were stained with Nile blue sulfate (NBS) to visualize cell death, 3 embryos were frozen for flow cytometric cell cycle analysis and cell counts, and selected embryos were preserved for histological examination. Dams fed the zinc-deficient diet for more than 3 days reduced their food intake through gestation day 8 but increased food intake on day 9. Maternal plasma zinc dropped to 10-25% of control levels in the zinc-deficient groups. Zinc deficiency from gestation day 1 or 3 resulted in two categories of affected litters on day 11. One category had embryos which were morphologically normal but displayed extensive NBS staining in the visceral arches, neural tube, and somites. The second category had developmentally retarded or maldeveloped embryos which showed little NBS staining. Zinc deficiency from gestation day 7 produced cell death in the posterior dorsal midline in the area of premigratory neural crest cells, which was confirmed by histological examination. Zinc deficiency from gestation day 9 did not affect morphology or NBS staining. Percentages of cells in the G0/G1, S, and G2M phases of the cell cycle on gestation day 11, determined by flow cytometry, were similar to controls in all groups. This study shows that as few as 4 days of maternal zinc deficiency can produce excess embryonal cell death, and that neural crest cells may be particularly sensitive.