Browsing by Author "Hrubec, Theresa"

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    Dysregulated Apoptosis in Teratogen-Induced Neural Tube Defects in Mice
    Mallela, Murali Krishna (Virginia Tech, 2011-01-27)
    Dysregulation of apoptosis during development is a possible mechanism for teratogen-induced birth defects. Neural tube defects (NTDs) are the second most common fetal malformations. Non-specific stimulation of maternal immune system prevents birth defects. This study investigated the role of dysregulated apoptosis in formation of NTDs from two teratogens: valproic acid (VA) and an unknown teratogen found in tap water. Interferon- γ (IFN γ) was used to stimulate maternal immunity to evaluate the role of altered apoptosis in this protective mechanism. Apoptosis was evaluated using flow cytometry, Terminal Transferase dUTP Nick End Labeling (TUNEL) assay and gene expression changes by RT2 Profiler PCR arrays. Additionally, changes in the expression of key signal transduction pathway genes that play a role in development were determined. Increased apoptosis, suggesting involvement in VA teratogenicity, was observed along the neural tube in both normal and abnormal embryos from VA-exposed dams. Increased apoptosis in normal VA-exposed embryos suggests that VA may alter other cellular processes such as cell proliferation and differentiation in addition to apoptosis. Apoptotic percentages in embryos with NTDs from IFNγ+VA dams were similar to controls, which indicated resistance to teratogen-induced apoptosis. In IFNγ+VA-exposed embryos with NTDs, immune stimulation failed to prevent apoptosis. VA initiated both death and survival signaling in the embryos; however, upregulation of the apoptotic genes and down regulation of anti-apoptotic genes of p53 and Bcl2 family tended to shift the balance towards death signaling. This change in gene expression patterns could result in increased apoptosis and NTDs in VA-exposed embryos. Immune stimulation normalized changes in the expression of pro-apoptotic signaling molecules. These results suggest immune stimulation protects embryos from teratogenicity of VA by preventing VA-induced apoptosis. VA altered the hedgehog, Wnt, retinoic acid and fibronectin signaling pathways in embryos with NTDs. These results suggest that VA also disrupted signaling pathways required for various morphogenic events during organogenesis. Immune stimulation normalized the expression of Fn1 and Hspb1 and thus may mediate protection through these signaling pathways. In tap water exposed embryos, no change in apoptotic pattern was observed by flow cytometry, TUNEL assay and RT-PCR. Also, none of the signal transduction pathway genes tested were significantly altered in tap water-exposed embryos. This suggests that apoptosis is not a mechanism for teratogenicity resulting from exposure to the contaminant in tap water.
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    The effects of temperature, water quality and culture conditions on the immunology, hematology, and blood chemistry of hybrid striped bass
    Hrubec, Theresa (Virginia Tech, 1994-12-19)
    Sunshine and palmetto bass (different crosses of hybrid striped bass) were used to determine immunologic, hematologic and serum chemistry changes under different culture conditions. The kinetics of the humoral immune response was determined for sunshine bass acclimated to 10, 18, 24, 29°C, and to elevated ammonia (0.15 mg/L NH3) and elevated nitrate (200 mg/L). These conditions are frequently encountered in aquaculture situations. Cooler temperatures decreased both the magnitude and onset of the humoral response, being lowest at 10°C, intermediate at 18°C, the highest at 24 and 29°C. Elevated ammonia did not affect the immune response, while elevated nitrate decreased antibody production to the level of the 18°C response. Hematologic reference intervals were determined for sunshine bass in tanks and recirculating systems, and palmetto bass in tanks. Serum chemistry reference intervals were determined for sunshine bass in tanks, recirculating systems and cage systems. Greater differences were observed in reference intervals between the culture systems, than between the two types of hybrid. To determine if environmental factors influenced the differences seen in the reference, sunshine bass were acclimated to 10, 18, 24, 29°C, elevated ammonia (0.15 mg/L NH3) and elevated nitrate (200 mg/L). The hematology and serum chemistry profiles of these fish were compared with the reference intervals for sunshine bass in tanks. Leukocyte, lymphocyte and monocyte counts at 10°C, and glucose and calcium at 10 and 18°C deviated sufficiently to suggest generating separate reference intervals at these temperatures. In the nitrate treated fish, creatinine levels were elevated and chloride levels were lower than controls and outside the reference interval. These two responses were presumed to be pathologic changes associated with elevated nitrate levels due to the large deviation in the analytes and the mortalities seen in the nitrate treated fish. The remaining analytes for fish in the different environments were within or slightly outside the reference intervals. These slight changes were presumed to be due to individual variation as the reference intervals were determined for fish under relatively uniform conditions and may not be sufficiently broad to cover fish from more varied environments. With minor modification, the reference intervals should apply to sunshine bass in most situations.