Apoptosis induced by uterine 24p3 protein in endometrial carcinoma cell line.

The biological functions and reaction pathways of lipocalins in mammalian system were sought. Mouse uterine 24p3 protein is a secreted lipocalin from mouse uterus. To evaluate the effect of uterine 24p3 protein on the reproductive system, endometrial carcinoma cell line (RL95-2) was an experimental target for achieving the in vitro study. The cells were treated with 0.75 microM dexamethasone (DEX) or under serum-free medium to mimic the stress environment for various time periods, then employing Western blot to measure the 24p3 protein secretion. It showed the time-dependent induction effect on 24p3 protein and suggested the level of protein secretion correlating to environmental stress. Furthermore, the supplementation of 24p3 protein to the medium accompanied the reduction of cell viability. It showed that the 24p3 protein may be a death factor under conditional media via PI and annexinV-FITC assay. Based on the autocrine hypothesis, we investigated the effect of 24p3 protein on cultured RL95-2 cells upon the 24p3 protein interaction. We have demonstrated significant increase in intracellular reactive oxygen species upon 24p3 protein interaction. While the changes of mitochondrial membrane potential and cytochrome c release from mitochondria occurred, the activities of caspase-8, -9 and -3 were found to have increased. The condensation of DNA was occurred suggesting that 24p3 protein induced irreparable DNA damage, which in turn triggered the process of apoptosis. It shows evidence for the direct effect of this protein on endometrial cells. These findings suggest that 24p3 protein creates an intracellular oxidative environment that induces apoptosis in RL95-2 cells.