Candidate: Susan J. Lentz

Degree of: Doctor of Philosophy

Department: Biological Sciences

Title: Cellular Proliferation, Death and Histology of Gambusia Affinis Livers After Exposure to 2-Aminofluorene and Benzidine

Committee:
Dr. Jay Means, Chair
Dr. Charles Ide
Dr. Robert Eversole
Dr. Robert Buck

Date: Tuesday, May 27, 2003, 1:00 p.m. - 3:00 p.m.
2902 Wood Hall

Abstract: A fundamental need exists for aquatic animals in the biomonitoring of aquatic ecosystems for environmental carcinogens. G. affinis, a small fish species, is selected for this study due to its hardiness, non-migratory nature, geographic diversity and ability to develop tumors. Molecular tools for cellular proliferation and death are examined for use with G. affinis. Additionally, these tools are used to investigate the early carcinogenic effects of 2-aminofluorene and benzidine, both individually and combined, on G. affinis livers.
Antibodies to proliferating cell nuclear antigen (PCNA), p53, cleaved caspase-3 and cleaved poly(ADP-ribose) polymerase (PARP) were examined for use in G. affinis. While p53 and cleaved caspase-3 and PARP results were inconclusive, PCNA was successful in detecting cellular proliferation in G. affinis. Additionally, the terminal deoxynucleotidyl transferase-mediated dUTP-biotin neck end labeling (TUNEL) method was examined and found to be a successful indicator of cell death.

Therefore, cellular proliferation, death, and their difference in G. affinis livers were assayed after dietary exposure to 0.069 mM and 6.9 mM doses of 2AF, BZ and 2AF/BZ for 4, 8 and 12 weeks. Hematoxylin and eosin ( H&E) staining was used for histological examination. All treatment groups had similar responses at differing degrees. These results from G. affinis exposure to 2AF, BZ and 2AF/BA indicate:

· A transient increase in cellular death at 4 weeks that decreased at 8 and 12 weeks, some below control levels.

· A substantial increase in cellular proliferation throughout the 4, 8 and 12 exposures above control levels.

· A net increase in cellular growth above control levels.

· Increases in oval cell proliferation, altered foci and tumor development.

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