Cell-surface oxytocinase inactivates oxytocin and regulates oxytocin stimulation. We reported that oxytocinase in human endometrial epithelial cells was secreted from the cell membrane in the mid-secretory phase and disappeared from the cell surface. On the other hand, the production in human endometrium of prostaglandins, which play important roles in the reproductive process, has been reported to be upregulated by oxytocin. We investigated whether progesterone affects cell-surface oxytocinase and oxytocin-induced prostaglandin E2 (PGE2) production in vitro. Progesterone induced secretion of oxytocinase into the culture medium, which resulted in a decrease in cell-surface oxytocinase. Production of PGE2 was increased slightly by oxytocin without progesterone, and significantly with progesterone. The inhibition of oxytocinase activity by amastatin had a similar effect to the loss of cell-surface oxytocinase caused by progesterone. It is therefore likely that the cell-surface oxytocinase of endometrial epithelial cells modified by progesterone plays an important role in the function of the human endometrium through PGE2.