Efeito do fator de crscimento semelhante à insulina-I (IGF-I) e do fator de crescimento e diferenciação-9 (GDF-9) sobre o desenvolvimento in vitro de folículos pré-antrais ovinos

Submitted by Mario BC (mario@bc.ufrpe.br) on 2018-10-17T16:41:17Z No. of bitstreams: 1 Alane Pains Oliveira Monte.pdf: 2693567 bytes, checksum: 8422930b95e7585b038f585b353e920b (MD5) Made available in DSpace on 2018-10-17T16:41:17Z (GMT). No. of bitstreams: 1 Alane Pains Oliveira Monte.pdf: 2693567 bytes, checksum: 8422930b95e7585b038f585b353e920b (MD5) Previous issue date: 2018-07-30 Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq The objective of this work was to study the effect of the insulin like growth factor-1(IGF-1) and growth differentiation factor-9 (GDF-9) on the in vitro development of sheep preantral follicles. For this, three experimental phases were developed. The first phase evaluated the effect of IGF-I on the in vitro culture of sheep isolated secondary follicles. Sixteen ovaries were allocated for the immunolocalization of IGF-I protein. Isolated secondary follicles of other ovaries were cultured for 18 days in control medium (MEM+) or MEM+ supplemented with different concentrations of IGF-I (10, 50 and 100 ng/mL) or the same treatments with the addition of 750 ng/ml of FSH. At the end of the culture, fluorescence, in vitro maturation (IVM) or LH receptor (LHR) expression analyzes were performed. Phase 2 evaluated the influence of GDF-9 on the development of preantral follicles cultured in situ. For this, ovarian fragments were cultured for 7 days in MEM or in MEM supplemented with of GDF-9 (1, 50, 100, 200 or 400 ng/mL). In addition, a second culture in MEM or MEM + 50 ng/mL of GDF-9 was performed in the presence or absence of the PI3K pathway inhibitor. At the end of culture, follicular survival, activation and diameter, immunolocalization of PCNA, activated caspase-3 and pAKT proteins were evaluated. In the phase three, isolated secondary follicles were cultured for 12 days in MEM or MEM + GDF-9 (1, 10, 50 or 100 ng/mL). At the end of culture, oocytes were submitted to IVM, follow by mitochondrial activity and ROS analysis. The results of phase 1 showed that 50 ng/mL of IGF-I + FSH showed more fully-grown oocytes (P