Date of Award
Master of Science
James D. Godkin
Judy Grizzle, Hank Kattesh
During early pregnancy in sheep, implantation occurs relatively late and fetal-maternal interactions are critical to maintenance of pregnancy, embryonic survival and development. Reproductive processes are mediated in part by conceptus secretory proteins and uterine proteins. Vitamin A (retinol) plays a role in reproductive processes and must be present in precise amounts for normal embryonic development. Most retinol is stored in the liver and transported bound to retinol-binding protein (RBP). Within the liver, the levels of RBP are regulated by retinoid status. Previous studies have suggested that placental RBP may be secreted in an apo- form by the conceptus in order to buffer the teratogenic effects of retinoids. The purpose of this study was to determine if retinoids influence RBP secretion by the conceptus and to determine if conceptus RBP is secreted in an apo- or holo- form. Day 14 ovine conceptuses were obtained from uterine flushes and cultured in Dulbecco's minimal essential medium (DMEM) for 24 and 48 hours with addition of retinoids (10 µM all-trans retinol, 1 or 10 µM all-trans retinoic acid) in order to evaluate the effects of retinoid treatment on secretion and synthesis of RBP by the conceptus. Concentration of RBP was determined by an enzyme linked immunosorbant assay (ELISA) in culture medium and solubilized conceptus tissues. Retinol concentration of control conceptus cultures (no addition of retinoids) was analysed by extracting culture medium and quantifying retinol-associated fluorescence of the extracts. There were no differences in RBP concentration due to treatments in either the culture medium or conceptus tissues. Retinol was undetectable in the medium of control conceptus cultures. The results of this study demonstrate regulation of RBP in the conceptus may be different than that of the liver since physiological doses of retinoids did not have an effect on RBP secretion or synthesis in the day 14 ovine conceptus. The failure to detect retinol in medium of control conceptus cultures indicates that either RBP is secreted in an apo- form or retinol concentrations were below the sensitivity of the assay.
Jacobs, Wendy G., "Regulation of retinol-binding protein secretion and expression in the ovine conceptus and uterus by retinoids and steroids. " Master's Thesis, University of Tennessee, 1996.