Materials and Reagents
Recombinant mouse-soluble VEGF receptor 1/Fc Chimera (Trap) (R&D Systems, Inc., Minneapolis, MN) was dissolved in PBS buffer with 0.1% BSA. The eCG (Novormon) was provided by Syntex S.A. (Buenos Aires, Argentina). Polyclonal primary antibodies for BAX (N-20), BCL2 (N-19), BCL2L1 (S-18), TNFRSF6 (FL-335), and FASLG (Q-20) were purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA); actin B antibody was obtained from Novus Biologicals (ab-6276; Littleton, CO). Anti-rabbit and anti-goat secondary antibodies conjugated with horseradish peroxidase were purchased from Sigma-Aldrich. All other chemicals were of reagent grade and were obtained from standard commercial sources.
In Vivo Trap Treatment and Superovulation
General care and housing of rats was carried out at the Instituto de Biologia y Medicina Experimental (IByME) in Buenos Aires. Prepubertal rats were from our colony. Immature female Sprague-Dawley rats (21-23 days) were allowed food and water ad libitum and were kept at room temperature (21-23°C) on a 12L:12D cycle. After being anesthetized with ketamine HCl (80 mg/kg; Holliday-Scott S.A., Buenos Aires, Argentina) and xylazine (4 mg/kg; Konig Laboratories, Buenos Aires, Argentina), the ovaries were exteriorized through an incision made in the dorsal lumbar region.