Timing of Inhibitory Actions of Gossypol on Cultured Bovine Embryos

Culture of bovine preimplantation embryos with gossypol, a polyphenolic pigment in cottonseed, inhibits development. Neither stage at which embryos are most sensitive to gossypol, nor the mechanism by which development is blocked is known. Our objectives were to characterize stages at which gossypol inhibits embryonic development and evaluate involvement of apoptosis in actions of gossypol. When presumptive 1-cell embryos were cultured continuously in medium containing gossypol at concentrations of 0, 2.5, 5, and 10 μg/mL, cleavage rate was not reduced by any concentration of gossypol, but percentages of 1-cell embryos that became blastocysts 8 d after insemination was reduced by the 10 μg/mL dose of gossypol. Culture of presumptive 1-cell embryos with gossypol at 10 μg/mL for 24 h was not sufficient to block development. Furthermore, gossypol did not affect development to the blastocyst stage when 2-cell embryos were cultured with gossypol at 10 μg/mL for 24 h or 7 d. Culture of embryos ≥16 cells with gossypol at 10 μg/mL for 24 h failed to reduce cell number 24 h later or increase blastomere apoptosis. Results indicate that embryonic development can be disrupted by long-term exposure to gossypol at 10 μg/mL and that exposure at the 1-cell stage is required. Thus, it is likely that the deleterious effects of gossypol involve disruption of events at the 1-cell stage and such effects are reversible if gossypol is removed. After the 1-cell stage, gossypol does not affect development because the critical event that gossypol disrupts occurs at the 1-cell stage only or the embryo develops cytoprotective mechanisms after the 1-cell stage that limit actions of gossypol.