Effect of fetal bovine serum in FASN expression in bovine embryos cultivated in vitro

Supplementation of the medium with fetal bovine serum (FBS) is widely used to increase the embryo rate, however presents a drawback because it is related to increased intracellular lipid accumulation in embryos (DODE, RBRA, v. 37, 145-150 , 2013). The action of FASN enzyme can be related to this process, since it is responsible for the synthesis of palmitate fatty acid from the precursors malonyl-CoA, acetyl CoA and NADPH (URSTAD-JENSEN, BBA- Mol. Cell Biol. Lipids, v. 1821, 747-753, 2011). Thus, the aim of this study is to evaluate the FASN expression in bovine embryos produced supplemented with different concentrations of FBS. Bovine ovaries were collected at a local slaughterhouse and the cumulus-oocyte complexes (COCs) were matured in vitro in TCM-199 supplemented with 10% FBS, FSH, LH and antibiotic for 22 hours at 38.5°C in 5% CO2. For IVF, COCs and sptz were co-incubated in TALP medium supplemented with FERT-penicillamine, hipotaurina, epinephrine, heparin and BSA under the same conditions mentioned for IVM. After 24 hours of co-incubation presumptive zygotes were then distributed in droplets cultivation in SOF medium supplemented with BSA and antibiotics according to the experimental groups (0, 2.5, 5 and 10% FBS). For counting the number of total cell, embryos were stained with fluorochrome Hoechst 33342. For the analysis of gene expression, the mRNA was extracted by Trizol® method (CA, USA), subjected to reverse transcription with the help of High- kit Capacity cDNA Reverse Transciption (CA, USA) and then analyzed using an assay relative quantification PCR (kit Power SYBR Green® PCR Master Mix) (CA, USA). Statistical significance was estimated by ANOVA with Tukey's post-test, adopting the significance level of 5%. Regarding the blastocyst rate, a significant difference (p < 0.05) was observed between the group without adding FBS (15% ± 7.6) compared to the groups with the addition of SFB - 2.5; 5 and 10% (39% ± 5.9; 37% ± 44% ± 3.9 and 10.2, respectively). With respect to counting the number of embryonic cells, the group supplemented with 2.5% FBS performed better than the non supplemented group with SFB (136 ± 8.5 and 108 ± 12.4, respectively) (P < 0.05), however this group did not perform significantly better than the other groups. Furthermore, in the analysis of gene expression, FASN gene regulation was not altered in in vitro cultured embryos regardless of the addition of FBS (P <0.05). Thus, it is concluded that the culture of bovine embryos with different concentrations of fetal calf serum does not affect FASN gene expression. However, supplementation contributes to embryo development in a quantitative and qualitative way, corroborating previous studies.