Immunocytochemistry versus binding assays of the estrogen receptor in the reproductive tract of spayed and hormone-treated macaques

Abstract

We used immunocytochemistry (ICC) with monoclonal antibodies to the estrogen receptor (ER) to localize ER in the oviducts, uteri, and cervix of untreated, estrogen-treated, and estrogen-progestin-treated spayed macaques. We also used binding assays with labeled estrogens to quantify nuclear and cytosolic ER levels in parallel samples of the same tissues. In untreated spayed animals, cytosolic ER levels were much higher than nuclear ER levels, but all specific staining was nuclear. After treatment for 14 days with estradiol (E2), the degree of staining for ER in cell nuclei in the oviduct, cervix, and endometrium had increased, and there were significant increases in both nuclear and cytosolic ER levels. In the myometrium, ER levels and ICC staining of nuclei increased minimally with E2 treatment. In animals treated for 2 weeks with E2 followed by 2 weeks with E2 and progesterone (P; sequential P treatment) the degree of nuclear ER staining in the oviduct, endometrium, and cervix greatly decreased, and cytosolic and nuclear levels of ER declined significantly. In the myometrium of such animals there was a minimal decrease in the degree of staining and a nonsignificant decline in cytosolic and nuclear ER levels. Sequential P treatment reduced the degree of nuclear staining in the oviduct and endometrium below that found in spayed animals; however, such treatment only lowered the amount of cytosolic, not nuclear, ER significantly below spayed levels in those same tissues. Some animals were treated sequentially with P and sampled 1, 3, 12, and 24 h after the onset of P treatment. By 1 h, nuclear ER levels in the endometrium were significantly suppressed, but cytosolic levels were not lowered until 3 h of treatment; ICC staining was also not substantially reduced until 3 h of P treatment. In the oviduct, nuclear ER levels were significantly reduced by 1 h of P treatment, but cytosolic levels were not lowered until after 12-24 h of P treatment; the degree of nuclear staining in the oviduct was also not substantially reduced until 12-24 h of P treatment. In myometrium, there was no significant decline in ER in nuclear or cytosolic fractions or any substantial decrease in the degree of nuclear staining at any time during this treatment. These observations suggest that the ER detected by ICC in the nuclei of target cells in frozen sections represents the total ER detectable by binding assays in cytosolic and nuclear fractions.(ABSTRACT TRUNCATED AT 400 WORDS)