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Expression patterns of cyclin D1, D2, and D3 in the first three cell cycles in preimplantation embryo developmentPowers, Tiffany M. January 2004 (has links)
Cell-cycle progression in mammalian cells is coordinated by a series of control points. The D-type cyclins are a family of key cell cycle regulators that are controlled largely by mitogens and their association with and activation of cdk 4 and 6 at the G1 phase of the cell cycle. This study seeks to first analyze cyclins D1, D2, and D3 expression patterns in preimplantation mouse embryos using in vivo studies and then analyze the effects of Dilantin on the cyclin D1 expression pattern in cultured embryos. Antibody staining against cyclin D1, D2, and D3 via indirect immunofluorescence using a Zeiss Confocal Microscope and analysis of individual embryo staining intensities using Zeiss computer software were employed to evaluate expression patterns throughout the first three cell cycles. The data showed that all three D cyclins were present throughout the first three cell cycles. Cyclin D1 had peak average fluorescence intensity at the G2 phase of the second cell cycle with a decrease at the G1 in the third cell cycle. Cyclin D2 had a consistent increase of fluorescence intensity throughout all three cell cycles. Cyclin D3 had peak average fluorescence intensity at the G2 phase of the second cell cycle with an immediate decrease at the Gl phase in the third cell cycle. Cyclin D1 was localized to the nucleus in G1 phases of the cell cycle. In contrast, cyclin D2 was found in the nucleus during G2 phases of the cell cycle rather than in G1. Cyclin D3 was not localized to the nucleus in either cell cycle phase throughout the first three cell cycles. These unique nuclear staining patterns seen by D1, D2, and D3 may reflect a function in the cell cycle. Embryos cultured in the presence of l0gg/ml of Dilantin were found to be slowed in development indicated by the absence of transition from the one-cell to the two-cell stage when compared to the controls. Since the Dilantin cultured embryos never reached G1 of the second cell cycle the increase in fluorescence intensity seen was still considered to be a representation of the G2 phase of the first cell cycle. Cyclin Dl's fluorescence intensity was affected by Dilantin and accompanied with unstained nuclei during the G2 phase of the first cell cycle. The peak average fluorescence intensity occurred during the G1 phase of the second cell cycle for cyclin D1 stained CZB control, while the vehicle control, 0.001N NaOH, remained constant. Both CZB and 0.001N NaOH had similar expression patterns seen previously in the cyclin D1 in vivo data. The information gained from the in vivo and in vitro experiments will help to better understand what causes the problems associated with exposure to Dilantin, and also the effects Dilantin has on the cell cycle. / Department of Biology
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In vivo Dilantin treatment alters expression levels and nuclear localization of cyclins A and B1 during mouse preimplantation embryo developmentTolle, Michelle D. January 2009 (has links)
Access to abstract permanently restricted to Ball State community only / Access to thesis permanently restricted to Ball State community only / Department of Biology
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Dilantin alters levels of DNA polymerase [delta symbol] in preimplantation mouse embryos during G1 and S phase in vivo / Dilantin alters levels of DNA polymerase in preimplantation mouse embryos during G1 and S phase in vivoCornielle Dipre, Aide R. 08 July 2011 (has links)
Dilantin (DPH) is a common anticonvulsant drug used to prevent seizures. It is known to be a human teratogen causing fetal hydantoin syndrome (FHS). FHS is characterized by multiple developmental and growth related abnormalities and mental retardation. Previous studies demonstrated that DPH slowed growth and division in preimplantation mouse embryos in vivo and in vitro. DHP exposure in utero decreased the crown to rump length and weight of 25-35% of embryos and reduced the rate of endochondral bone conversion from cartilage. In vitro preimplantation mouse embryos treated with DPH at 5, 10 and 20 μg/ml showed a reduction of 25-35% in their development, and block at 2-cell or 3-4-cell stages. These embryos also showed a prolonged DNA synthesis (S) phase during the second cell cycle. Nuclear localization and concentration levels of
cyclin A , the S phase cyclin, were also altered in vivo in 2-cell DPH treated embryos compared with NaOH control embryos during G1, S phase and G2 of the first, second and third cell cycles. DPH altered patterns of expression of cyclin A were associated with cell cycle disregulation during preimplantation development. The purpose of the current study was to determine whether DPH also affects the concentration of DNA pol δ catalytic subunit in 2-cell preimplantation mouse embryos at G1 and S phases, thus delaying DNA synthesis and contributing to FHS. Immunofluorescence and confocal microscopy were used as tools to determine relative levels and distribution of DNA pol δ (for consistency with text) in the cytoplasm and the nuclei of DPH and NaOH treated 2-cell embryos at G1 and S phase of the second cell cycle. DPH decreased DNA pol δdelta total embryo and nuclear levels by 43% and 36%, respectively, in G1 compared with NaOH controls. Similarly, nuclear levels of DNA pol δ in DPH embryos in S phase near the G2 transition of the second cell cycle increased to 144% of NaOH control levels; there was not a statistically significant difference between total embryonic levels of late S phase DNA pol δ in DPH and NaOH treated control embryos. The results indicated that DPH affects the levels of DNA pol δduring G1 and S phase near the G2 transition of the second cell cycle in preimplantation mouse embryos. The significant alteration in the levels of DNA pol δ during S phase and its probable consequent altered polymerase activity could contribute to an explanation for the extension of S phase in preimplantation embryos observed by Blosser and Chatot. Even more,
the alteration in the levels of DNA pol δ and potentially in its exonuclease activity could lead to an increase in the rate of mismatches and mutations suggesting a likely explanation for some features of FHS. / Department of Biology
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Dilantin affects the rate of DNA synthesis via cyclin A and decreased concentrations of DNA polymerase [delta] in preimplantation mouse embryosTolliver, Autumn R. 14 December 2014 (has links)
Access to abstract restricted until 12/14/2014. / Access to thesis restricted until 12/14/2014. / Department of Biology
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