Semester

Summer

Date of Graduation

2000

Document Type

Thesis

Degree Type

MS

College

Davis College of Agriculture, Natural Resources and Design

Department

Physiology, Pharmacology & Neuroscience

Committee Chair

Michael R. Miller.

Abstract

This study examines the effect of acetaminophen (APAP) on the estrogen-responsive gene alkaline phosphatase in Ishikawa endometrial cancer cells. Basal levels of alkaline phosphatase activity were significantly inhibited in cells treated with 0.03, 0.1, 0.3 and 1mM APAP compared to control. Alkaline phosphatase activity induced with 1nM estradiol was significantly inhibited in a concentration dependent manner by 0.0.1, 0.3 and 1mM APAP. APAP did not directly inhibit the activity of alkaline phosphatase at any of the concentrations tested. Increasing estradiol concentrations (0.01, 0.1 and 1nM) did not alter the level of inhibition of alkaline phosphatase activity by 0.1, 0.3 or 1mM APAP. This is in agreement with other studies, which have concluded that APAP does not compete with estradiol for binding at the estrogen receptor (Harnagea-Theophilus a et al., 1999; Isenhower et al., 1986; Miller et al., 1999). Protein concentrations were significantly decreased in Ishikawa cells with 0.3 and 1mM APAP alone or in the presence of 0.01, 0.1 to 1nM estradiol. Inhibition of alkaline phosphatase activity by high levels (0.3 and 1mM) of APAP may be in part due to toxic effects of APAP as determined by LDH release. Fluorescent microscopy using calcein staining techniques show that Ishikawa cells treated with 1nM E2+0.3mM APAP are visibility misshapen; cells treated with 1nM E2+1mM APAP are no longer attached to the plate indicating toxicity. The final part of this study looked at the interaction between antiestrogen-4-hydroxy tamoxifen (OHT) and APAP on alkaline phosphatase activity in Ishikawa cells. While OHT inhibited alkaline phosphatase activity ∼50 percent in estradiol-induced cells, addition of 0.03-1mM APAP increased inhibition significantly in a concentration dependent manner. Results of this study indicate that APAP decreases estrogen-responsive alkaline phosphatase activity in Ishikawa cells in vitro, and that this inhibition is not attributed to direct inhibition of enzyme activity.

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