Cell preparation

All tissue samples (hysterectomy, endometrial, and decidua samples) were collected in complete RPMI medium [supplemented with 10% fetal calf serum (FCS), 2 mM l-glutamine, and penicillin/streptomycin] and manually dissected into small pieces. Next, tissue pieces were transferred to a tube and incubated in a water bath with a magnetic stir at 37°C in RPMI containing collagenase II (0.25 mg/ml) and deoxyribonuclease (DNase; 0.2 mg/ml) until the tissue was dissolved or a maximum time of 40 min reached. The reaction was stopped with addition of fivefold the volume of complete RPMI. Then, the suspension was passed through a 40-μm mesh and remaining pieces pushed through the mesh with a syringe plunger. Last, mononuclear cells were isolated with density gradient centrifugation, washed with phosphate-buffered saline (PBS), and either frozen down or directly used for experiments. Peripheral blood cells were isolated with density gradient centrifugation and washed with PBS before being frozen down or used for experiments. Menstrual blood was processed as previously described (14).