EFECTO DE LA ERITROPOYETINA EN LA PRODUCCION IN VITRO DE EMBRIONES BOVINOS

CONDE P.A. 1,2, HERRERA C. 1, GUTIERREZ G. 1,2, QUINTANS C. 1, GENTILE T. 2, PASQUALINI R.S. 1

Estambul, Turquia

Congreso; ALPHA 2008, 7TH CONFERENCIA BIENAL; 2008

Alpha

Erythropoietin (EPO) is known to regulate the number of circulating red blood cells. Recently, EPO and EPOr mRNA were found in organs non related to erythropoiesis such as testicle, endometrium, oviduct and ovary. The aim of our work is: Experiment 1(Exp.1): Evaluate the effect of adding EPO to the in vitro maturation medium (IVMm) of bovine oocytes on the percentage of MII oocytes, cleavage and blastocyst rates as well as blastocyst cryotolerance. Experiment 2(Exp.2): Evaluate the effect of adding EPO to the in vitro fertilization medium (IVF) on fertilization rates. Bovine oocytes were obtained from slaughterhouse ovaries and for Exp.1 they were matured in vitro for 22h in groups of 5 to 10 in 50ml droplets of IVMm (De Matos et al., Mol Reprod Dev. 62:203, 2002) with 0, 7, 14, 20 or 80mU/ml EPO. Then, they were denuded from their cumulus cells or incubated for 24h with 2x106 bull sperm/ml. Presumptive zygotes were denuded and cultured for 24h in synthetic oviduct fluid medium (SOFm) in co culture with granulosa cells and then the medium was replaced for SOFm with 1.5mM glucose. The embryos remained in this medium for 7 additional days. Blastocysts obtained were cryopreserved in 0.25ml straws, with a slow freezing curve (0.6ºC/min). After that they were plunged into liquid nitrogen. Thawing was performed by keeping straws at room temperature for 10 sec. followed by immersion in a water bath at 36ºC and culture in SOFm with 1.5mM glucose for 48hs. The maturation, cleavage and blastocyst rates were compared between the different groups. In Exp.2 the oocytes were matured as in Exp. 1, without EPO, and incubated with the same concentration of sperm with 0, 8, 40 mU/ml EPO. Cleavage and pronuclear formation rates were compared between groups. Differences among groups were analyzed by Chi2 test. The supplementation with EPO during IVM demonstrated a positive effect only on the 14mU/mL group by a significant increment on the thawed hatched blastocyst rate (71% vs. 39% of the control group p<0,05. No significant differences were obtained when EPO was added to IVF. These results demonstrate an improvement on the efficiency of bovine in vitro embryo production (IVP). To our knowledge, this is the first report on the effect of EPO on in vitro embryo development.  Further investigations will be necessary to determine the effect of adding EPO during embryo development as well as its regulation in reproductive organs.