Produção de embriões in vitro com adição de hormônio de crescimento bovino (bGH) ao meio de maturação

Abstract

Growth hormone (GH) is a hormone secreted by the pituitary gland and acts on the growth of various tissues, including the reproductive system. It is not a reproductive hormone, but has an important link to the development of ovarian follicles and the oocyte maturation. Its receptors are present in cumulus cells in cytoplasmic and nuclear membrane of the oocyte, and this causes the GH acts directly in its growth. In vitro production of embryos GH has been studied to increase the amount and quality of embryos, but the concentration of GH in vitro maturation is not yet defined, with a variation in the literature 10 to 1000 ng/mL. This study compared different dosages of GH added to the maturation medium in order to set an optimal dosage so that there is an increase in the quantity and quality of embryos produced, which was measured by the cleavage rates, embryo production and quantifying the number of embryonic stem cells, as well as comparing the interaction of GH with the generation of oxidative stress. Oocytes were matured in medium composed of TCM 199 with Earl's salts, supplemented with 10% fetal bovine serum, luteinizing hormone, follicle stimulating hormone, estradiol, and amikacin. Different dosages of GH were added to maturation medium these being 0 ng/mL 25 ng /mL, 50 ng/mL, 75 ng/mL and 100 ng/mL. After 24 hours of maturation, oocytes were fertilized and these were incubated for 22 hours to later pass to the culture and they were incubated for 7 days. On the third day of cultivation were evaluated cleavages and on the seventh day the production of embryos. Embryos expanded blastocyst stage were fixed on slides and stained with Panotic for embryonic cell count. The oxidative stress analysis was done by the reaction of the means of maturation, fertilization and cultivation with thiobarbituric acid. There were significant differences in quantitation of cells when used 100 ng/mL giving an improvement in the quality of embryos, and also increased the oxidative stress used when 75 and 100 ng/mL in fertilization and in vitro culture.