1141160720 Does the anti‐proliferative action of ovine uterine serpin involve apoptosis?

Abstract

Ovine uterine serpin (OvUS) is the major progesterone‐induced protein in the uterus of the pregnant sheep. This protein is a member of the serine proteinase inhibitor superfamily and it has been proposed to down‐regulate uterine immune function during pregnancy to protect the fetus. In vitro experiments have shown that both the native and the recombinant (r) form of the protein can inhibit mitogen‐induced lymphocyte proliferation and growth of canine primary osteogenic sarcoma cells, mouse lymphoma cells, human prostatic adenocarcinoma cells (PC‐3 cell line) and bovine preimplantation embryos. The mechanism by which OvUS inhibits cell proliferation is still unknown. Accordingly, experiments were conducted to test whether rOvUS exerts its anti‐proliferative action through apoptosis. In the first experiment, the anti‐proliferative effect of rOvUS on PC‐3 cells was tested to determine the minimal concentration effective at inhibiting proliferation. Proliferation was inhibited by concentrations of OvUS at 8 μg/mL and higher. The incorporation of [3H]thymidine was 4043, 3998, 3464, 2785, 2827, 2310, and 2332 dpm for 0, 0.5, 2, 8, 32, 125, and 250 μg/mL, respectively. In the second experiment, PC‐3 cells were cultured for 48 hr with 50, 100 and 200 μg/mL of rOvUS or ovalbumin. Cells were then fixed and apoptotic cells identified using the TUNEL assay to detect DNA fragmentation. There was no effect of OvUS at any concentration tested on the percent of cells that were apoptotic. Percent apoptosis was 1.3% for control cells, 3.8%, 4.8% and 2% for cells cultured with 50, 100 and 200 μg/mL rOvUS, and 2% for cells cultured with 200 μg/mL ovalbumin. Results confirm that OvUS inhibits proliferation of PC‐3 cells and indicate that inhibition does not involve induction of apoptosis. Further experiments are warranted to elucidate the anti‐proliferative mechanism of action of OvUS.