Effect of estradiol and progesterone on growth of porcine myometrial smooth muscle cells and phospholipase C and adenylate cyclase signalling systems in vitro.

The objective of presented studies was to investigate whether estradiol and progesterone administered in vivo and/or added in vitro can influence the primary myometrial cell culture and how these steroid hormones can affect hCG stimulated cAMP and inositol phosphate production in the porcine uterine myocytes. Myometrial smooth muscle cells were obtained from six ovariectomized gilts pretreated (n = 4) or not (n = 2) with oestradiol benzoate and progesterone for 5 consecutive days. Immunocytochemical staining proved that the pattern of filamentous actin in the cytoplasm of the myometrial fibroblasts (basketlike network) was different from that of myometrial smooth muscle cells (long parallel fibres). The myocytes derived from steroid treated pigs and supplemented with estradiol and progesterone in vitro formed a hillock pattern on days 4-5 day of culture whereas cells obtained from not steroid pretreated gilts were smaller and did not create confluent form. Myocytes were treated in vitro with two doses of estradiol/progesterone (low - 0.2 nM/50 nM and high - 2 nM/500 nM, respectively) and two doses of hCG - 0.1 mU and 1000 mU/ml to study hCG action on the second messenger system in myocytes. The myometrial smooth muscle cells treated with low dose of estradiol and progesterone in vivo responded with much higher accumulation of inositol phosphates to strong (1000 mU/ml) hCG stimulation when compared with those receiving high dose of both steroids. The different doses of estradiol and progesterone caused a similar increase in basal cAMP accumulation as compared to control cells cultured without steroid hormones. hCG (0.1 mU/ml) had usually the additive effect on cAMP production in porcine myometrial cells. The presented paper shows that estradiol and progesterone administration in vivo followed by steroid hormone treatment in vitro affects the primary myometrial cells culture and that both steroid hormones modify the basal accumulation of the second messengers: cAMP and IP3 and their answer to hCG stimuli in pigs.