Uptake and Metabolism of Tritiated 17ß-Oestradiol and Oestrone by Rabbit Uterine Tissue in vitro

Abstract

To determine whether the established capability of rabbit uterine tissue to interconvert 17 p-oestradiol and oestrone might have some effect on the mode of action of the oestrogens in this organ, the in vitro interconversion of [3H]-17p-oestradiol and [3H]oestrone by rabbit endometrial and myometrial tissue was investigated and the identity of radiometabolites in 'soluble, 'mitochondrial-microsomal' a'nd 'nuclear' preparations was studied. Both endometrial and myometrial tissue were found to be capable of oxidoreduction of the oestrogens, the equilibrium of the reaction favouring the reduction of oestrone. Irrespective of the tissue--steroid combination studied, the greater part of the radioactivity in all fractions was associated with 17p-oestradiol. The relative proportions of oestrone varied between fractions, the nuclear preparation consistently showing a lower proportion of oestrone than the other fractions. Sephade<c fractionation of a 0'4M KCl 'nuclear extract' revealed that proportionately less oestrone than 17p-oestradiol was bound to the nuclear 'receptor'. These findings provide further evidence for 17p-oestradiol being the ovarian oestrogen which is active in the uterus, and suggest a role for uterine oxidoreduction of oestrogens in the control exercised over this organ by these steroids.

Uptake and Metabolism of Tritiated Oestradiol and Oestrone by Human Endometrial and Myometrial Tissue in Vitro

“…The 'particulate' fraction was extracted by homo¬ genization first in 10 ml ethanol : water (10:1, v/v) then twice in 10 ml ethanol and this crude extract was subjected to the phenolic extraction procedure of Engel, Slaunwhite, Carter & Nathanson (1950). The methods used for extraction were identical to those already described (Gabb & Stone, 1974). In several cases reported below, chloroform did not extract the majority of the radioactivity from the medium and 'soluble' fractions of endometrial tissue.…”

“…The techniques used for thin-layer chromatography of free oestrogens and oestrogen acetates were as previously described (Gabb & Stone, 1974). Carrier oestrogen sulphates were prepared by the sulphamic acid method (Joseph, Dusza & Bernstein, 1966).…”

“…([3H]E2) and [2,4,6,7-3H]oestrone (PHJE^(both of specific activity 370 fiCi/fig) were obtained from The Radiochemical Centre, Amersham, and purified by thin-layer chromatography as previously described (Gabb & Stone, 1974). Radioactive oestrogen solutions were prepared and stored as previously described.…”

“…Before each experiment aliquots of the radioactive oestrogen stock solutions were dried under nitrogen and dissolved in the buffer used in the experiment to give a concentration of 1 ng/ml solution. Unlabelled carrier steroids were prepared and stored as previously described (Gabb & Stone, 1974). All solvents used were either reagent grade or redistilled before use.…”

“…medium, 'soluble' and 'particulate') were pooled separately. These pooled extracts were then subjected to the acetylation, saponification and, where appropriate, the reduction procedures previously described (Gabb & Stone, 1974).…”