Abstract: Effects of Glucose or Fructose and Metabolic Regulators (DNP, PES, Cerulenin) on Bovine Embryo Development and Lipid Accumulation

M. Barcelo Fimbres and G.E. Seidel Jr.
Animal Reproduction and Biotechnology Laboratory
Colorado State University, Fort Collins, CO 80523 USA

Slaughterhouse oocytes (n=1800) were matured and fertilized in vitro by standard procedures. Zygotes were cultured at 38.5ºC in 5%CO2/5%O2/90%N2 in CDM-1 (similar to SOF) with nonessential amino acids, 10µM EDTA, 0.5% fatty acid-free BSA, and 0.5mM glucose. After 48h, 8 cell embryos were cultured 120h in CDM-2 (CDM-1+essential amino acids and 2 mM hexose). A factorial design with glucose or fructose; and 5 treatments (0.3µM phenazine ethosulfate (PES); 0.3µM PES+30µM dinitrophenol (DNP); cerulenin (fatty acid synthase inhibitor) 1µg/mL (C1) and 3µg/mL (C3); and control) was replicated 6 times. Day 7.5 blastocysts were fixed and stained with Sudan Black B to quantify lipid droplets in the cytoplasm. A digital photo at 600x of the equatorial part of the embryo was evaluated by classifying lipophilic droplet diameters as small (<2 µm), medium (2 to 6 µm), or large (>6 µm), reported as number of lipid granules per 1,000 µm2. Blastocyst rates per oocyte matured, and per 8 cell embryo (B/E) were higher (P<0.05) for fructose than glucose (30.4% vs 26.5% and 52.8% vs 47.6%). There were differences in B/E for PES, C1, PES+ DNP, and control vs C3 (P<.05): 35.8%, 32.8%, 32%, 32.8%, and 13.6%. Numbers of small droplets were different for PES, C1, and PES+DNP, vs C3 (32, 34, 38, and 55, respectively) (P<.05), but the control did not differ from other groups (P>.14). Medium size lipid droplets were lower (P<.05) for PES, and PES+DNP than C1, C3 and the control (21, 18, 28, 34, and 34, respectively). Large droplets were similar for PES, and PES+DNP; PES was lower (P<.05) than control, C1 and C3, but no difference was found between the control and PES+DNP. C3 was not different from the control, but higher than the other treatments (P<.05). There were no effects of glucose or fructose on small or large lipid droplet sizes (P>.16), but Fructose was lower for the medium size (23.6 vs 30.4) (P<.01). In conclusion, fructose produced more blastocyst and less medium size droplets than glucose. PES reduced lipid accumulation, and cerulenin did not.


Revised: August 3, 2005

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