Peptides extracted from vero cell cultures overcome the blastocyst block of mouse embryos in a serum-free medium

Purpose: The aim of this work was to evaluate the effect of a Vero cell coculture system on the development of mouse embryos. Methods: Mouse embryos were randomly divided and cultured in human tubal fluid (HTF) medium with/without Vero cell monolayers, conditioned medium (CM) obtained from Vero cell cultures, and HTF medium supplemented with peptides extracted from CM. The concentrated CM was examined by SDS/PAGE. Results: The development of mouse embryos was blocked at the blastocyst stage in pure HTF medium (1.4% hatching at day 5). This "blastocyst block≓ was overcome by coculture with Vero cell monolayers (48.1% hatching at day 5; 1.4 vs 48.1%; P<0.001). CM and the addition of 5% fetal bovine serum (24.1 and 34.9% hatching, respectively, at day 5) were also able to enhance the process of hatching. In the other experiment, the addition of peptides extracted from Vero cell cultures also overcame the blastocyst block (12.5%) compared with pure HTF medium (2.1%) (P<0.05). Electrophoretic separation revealed several classes of polypeptides consistently secreted into CM obtained from Vero cell cultures. Most peptides occurred in the M r range between 6.5 kd and 35.9 kd. Conclusion: A developmental block (blastocyst block) of mouse embryos in a serum- and protein-free medium (HTF) was discovered in this study. This block was effectively overcome by HTF plus serum and coculture with Vero cell monolayers and also by the peptides extracted from Vero cell-conditioned medium. We speculate that certain factors secreted or converted by Vero cells may be critical in hatching of mouse embryos. Further study of these factors may be helpful in delineating its mechanism.