Nitric oxide as a regulator in preimplantation embryo development and apoptosis

Objective: To investigate the mechanisms of nitric oxide (NO) in the development and apoptosis of preimplantation mouse embryos. Design: Prospective, controlled study. Setting: Medical college laboratory. Subject(s): Two-cell embryos from outbred ICR mice. Intervention(s): Hyperstimulation protocol, two-cell embryos were collected, then treated with or without an NO synthase inhibitor (L-NAME) or an NO donor (SNP) and combined with a cGMP analogue (8-Br-cGMP) or a selective inhibitor of NO-sensitive soluble guanylyl cyclase (ODQ). Main Outcome Measure(s): The development of ICR mouse embryo from two cells to blastocyst stages in vitro. Result(s): The development of blastocyst was inhibited by L-NAME in a concentration-dependent manner (0.1-10 muM) and 0.1 muM SNP reversed this effect (80.5% of control). Annexin-V/propidium iodide and terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling techniques demonstrated that excessive NO (greater than or equal to 10 muM) might induce apoptosis in the mouse embryos. 8-Br-cGMP reversed the inhibitory effect of L-NAME and rescued the embryo growth. ODQ inhibited the embryo development in a dose-responsive fashion (0.1-100 muM) but had no effect in the NO-induced embryo apoptosis. P53 and Bax were found to be up-regulated during the embryo fragmentation. Conclusion(s): These results indicate that the cGMP pathway might be involved in the NO-regulated embryonic development, but not in NO-induced apoptosis, for which P53/Bax pathway might be involved. (Fertil Steril(R) 2001,75:1163-71. (C) 2001 by American Society for Reproductive Medicine.).