Identifier

etd-04152005-095943

Degree

Master of Science (MS)

Department

Renewable Natural Resources

Document Type

Thesis

Abstract

The goal of this thesis was to evaluate temperature effects on cell proliferation of eastern oyster somatic tissues for the development of an oyster cell line. Understanding the in vivo cell proliferation of an organism is essential for the development of cell culture. Cell proliferation can be measured by identifying nuclear cellular proteins involved in growth regulation and cellular transformation. The primary objectives of this study were to: 1) develop an assay to evaluate cell proliferation, 2) develop an assay to analyze nuclear RNA content, and 3) evaluate temperature effects on cell proliferation in somatic tissues. Proliferating cell nuclear antigen was detected in all the phases of the cell cycle of the eastern oyster. The concentrations of PCNA increased from G0/G1 phase into the S phase, where it peaked in mid-S phase, and decreased in G2/M phase. The immunohistochemically stained slides were analyzed using image analysis and compared to the flow cytometric detection of PCNA. Overall, flow cytometry was superior to immunohistochemistry in time and cost efficiency, and in sensitivity to the detection of PCNA in oyster somatic tissues. A flow cytometric assay was developed to detect nuclear RNA in oyster somatic tissues. Nuclear RNA was detected in all the phases of the cell cycle of the eastern oyster. The amount of nuclear RNA increased from G0/G1 phase into the S phase, where it peaked in mid-S phase, and decreased in G2/M phase. Temperature effects on cell cycle, proliferation, and cell metabolism of oyster somatic tissues were evaluated. The cell cycle analysis of the heart and labial palps cells showed that the G0/G1-phase cells decreased as the S-phase and G2/M-phase cells increased. The cell cycle analysis of both tissues also showed the opposite effect, as G0/G1-phase cells increased the S-phase and G2/M-phase cells decreased. Heart and labial palps cells for oysters held between 10°C and 25°C increased in cell proliferation after one week of temperature fluctuation. The nuclear RNA/DNA ratio at all phases of the cell cycle for heart and labial palps cells for oysters between 15°C and 20°C was higher than at all other temperature regimes. Temperature fluctuations affected the cell cycle, proliferation, and the metabolic condition of oyster somatic cells.These results support the use of flow cytometry for the analysis of cell cycle, cell proliferation, and cell metabolism. The results also suggested that temperature fluctuation has an effect on cell cycle, cell proliferation, and cell metabolism. Future research needs to focus on improving the current in vivo study and adapt the assays for cultured cells.

Date

2005

Document Availability at the Time of Submission

Release the entire work immediately for access worldwide.

Committee Chair

Jerome F. La Peyre

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