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Nitric oxide as a regulator in preimplantation embryo development and apoptosis
H W Chen1, W S Jiang, C R Tzeng
1Center for Reproductive Medicine, Department of Obstetrics and Gynecology, Taipei Medical College, Taipei, Taiwan.
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 microM) and 0.1 microM 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 (> or =10 microM) 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 microM) 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.