Abstract:
Neural tube defects (NTDs) are the most common of all serious human birth defects. Studies have identified genetic components that may confer increased susceptibility to environmentally-induced NTDs. The SWV and C57BL/6 mice strains have shown a large disparity in sensitivity to environmentally-induced NTDs. For example, the C57BL/6 is more sensitive than the SWV following cadmium exposure. Due to its complex status of proliferation, differentiation, and cell turnover, the developing embryo is highly susceptible to metal-induced cell cycle alterations and apoptosis. We hypothesize that differential metal-induced impacts within cell cycle and apoptosis pathways at the molecular level contribute to observed differing sensitivities between these two strains. A primary in vitro model was established by isolating murine embryonic fibroblasts (MEFs) at gestational day 14. Using the neutral red assay, a cytotoxicity profile was generated after 24 hr exposure to environmentally relevant concentrations of cadmium and methyl mercury. To characterize metal-induced alterations within cell cycle pathways, cell cycle kinetics were assessed using BrdU/Hoeshst/PI and transcriptional changes of cyclin b1 and p21 were measured using RTPCR. Finally, to examine changes in apoptotic pathways, caspase 3/7 and 8 activities were measured. The SWV exhibited higher sensitivity to cadmium, but not with methyl mercury, confirmed by a significant shift in cell viability compared to the C57BL/6. We observed metal-induced cell cycle alterations, including inhibition of cell cycle progression, G2/M accumulation, and repressed cyclin B1. Additionally, we observed a dose depended increase of caspase 3/7 and caspase 8 activities in both strains with both treatment, but overall, no differences in response were evident between strains for these endpoints. In summary, although both, SWV and C57BL/6 MEFs exhibited does dependent metal-induced alterations in cell cycle and apoptotic pathways, we did not observe statistical significant differences between the SWV and C57BL/6 in cellular response. This study suggests that there are no inherent genetically determined differences in apoptotic or cell cycle-related response to metals in these two strains of mice. This may imply that differential sensitivity to metals is due to other unique pathways or biological aspects yet to be considered.