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Patterns of induced protein synthesis and secretion in guinea-pig endometrial epithelial cell cultures in response to oestrone sulphate alone and oestrone sulphate plus progesterone were investigated. Epithelial cells were cultured for 3 days in growth medium, then washed three times in a steroid-free medium. For each experiment, anticytokeratin immunostaining was used to discriminate the epithelial cells from the stromal cells. Only experiments in which the control dishes displayed more than 80% of anticytokeratinimmunostained cells were further processed. After this period oestradiol-17β (20 nmol/l; control), oestradiol-17β (20 nmol/l) plus progesterone (0·5 µmol/l), oestrone sulphate (1 µmol/l) or oestrone sulphate (1 µmol/l) plus progesterone (0·5 µmol/l) were added to the medium for 48 h. An immunocytochemical progesterone receptor assay showed that oestradiol-17β increased the progesterone receptor content of cells, and progesterone added to cultured cells in the presence of oestradiol-17β induced a significant increase in oestrogen sulphotransferase activity assessing the hormone responsiveness of the cultured cells.

In these culture conditions and after 16 h of incubation, oestradiol-17β induced a 1·7-fold increase in [³H]thymidine incorporation into DNA, and [³⁵S]methionine incorporation into cellular proteins was linearly increased up to 8 h. Biochemical changes induced by the different hormone treatments were studied by labelling the proteins with a 6-h pulse of [³⁵S]methionine. The proteins present in the medium and in cells were analysed by two-dimensional polyacrylamide gel electrophoresis, followed by fluorography. Addition of oestrone sulphate alone or with progesterone produced a change in the patterns of cellular and secreted proteins compared with those in cells cultured with either oestradiol-17β or oestradiol-17β plus progesterone. Three cellular proteins (M_r < 14 000, isoelectric point (pI) 5·2 and 5·3; M_r 75 000, pI 4·9) and one secreted protein (M_r 155 000, pI 5·6–5·9) were specifically induced and could serve as markers of oestrone sulphate action.

Journal of Endocrinology (1989) 123, 233–241