Date of Award

1996

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

First Advisor

Patricia A. Melrose

Abstract

This project was performed in order to determine whether exercise-induced increases in plasma cortisol concentrations may be responsible for enhanced lymphokine activated killer (LAK) cell activity. Secondly, this project included preliminary experments which were designed to test the hypothesis that stimulatory effects of cortisol on LAK cell activity may be due to cross-talk between signalling systems important to glucocorticoid effects and stimuli which act through nuclear activator protein-1 (AP-1). Blood samples were collected from resting horses in order to determine whether the animals had normal plasma glucocorticoid concentrations. All horses included in these experiments had $10\sp{-7}$ to $10\sp{-8}$M cortisol concentrations and samples collected over a 24 hour period exhibited a normal diurnal rhythm. Plasma cortisol concentrations were measured before and after injection of cortisol or vehicle and the beginning of exercise stress. Radioimmunoassayable cortisol concentrations were increased (P $<$ 0.05) in cortisol treated animals. Vehicle injection did not affect cortisol concentrations. Exercise-stressed horses had higher (P $<$ 0.05) basal cortisol concentrations compared to vehicle or cortisol-treated horses. Exercise significantly increased plasma cortisol at all time points. Cortisol injections did not elevate plasma cortisol to concentrations measured in exercise-stressed horses. Extracts from PBMCs were analyzed to determine if they contained immunoreactive (ir) AP-1 protein and glucocorticoid receptors (GR). Cell extracts were resolved by SDS-PAGE and blotted onto nitrocellulose membranes. Primary GR antibody bound to a single band ($\sim$100KD) which was similar to that reported for GRs from other species. Dot blots were used to detect ir-Fos and Jun in extracts from equine PBMCs. Extracts containing at least 90 ng of protein contained specifically labeled Fos and Jun. The last experiment cultured PBMCs with media, IL-2, IL-2 plus phorbol ester (TPA) or IL-2 plus cortisol. As compared to the media control, LAK cell activity was stimulated by IL-2 and inhibited by TPA and high dose cortisol. Immunoreactive-Jun was increased by TPA and this effect was abolished by 10$\sp{-4}$M cortisol. These experiments indicate that stimulatory effects of exercise on LAK cell activity are unlikely to be due to cortisol actions or to stimuli which act through protein kinase C/AP-1 messenger systems.

ISBN

9780591035049

Pages

181

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