Prolonged cultivation of rat uterine cells with preserved estrogen responsiveness

A rat uterine cell culture was prepared as an experimental system for investigation of mechanisms of steroid hormone actions. Cells frequently supplemented with fresh medium were successfully cultured for 4 weeks through 2 successive passages. Studies of estrogen responsiveness in the primary culture as well as in it's first subculture were performed by a small scale uptake assay for determination of specific steroid binding. Scatchard analysis of specific ovarian hormone binding confirmed that cultured uterine cells preserve both estradiol and progesterone receptors. Characteristics of specific [3H]estradiol binding detected in cells of the first subculture were comparable to those obtained in the initial primary culture. The number of specific estradiol binding sites was diminished to one third of the initial values only in cells of the second subculture, 22 days after isolation of cells from tissue. In the primary culture and in it's first subculture the cells responded to estradiol with a 2-3-fold increase in progesterone receptor level. The subcellular distribution of steroid receptors was also studied; estradiol receptor complexes were detected predominantly in the nuclei whereas progesterone receptors were nearly equally distributed between nuclei and cytosol.