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In this study, we systematically compared the morphological, functional and molecular characteristics of granulosa cells and oocytes obtained by a three-dimensionalin vitromodel of ovine ovarian follicular growth with those of follicles recoveredin vivo. Preantral follicles of 200 µm diameter were recovered and cultured up to 950 µm over a 20-day period. Compared within vivofollicles, thein vitroculture conditions maintained follicle survival, with no difference in the rate of atresia. However, thein vitroconditions induced a slight decrease in oocyte growth rate, delayed antrum formation and increased granulosa cell proliferation rate, accompanied by an increase and decrease inCCND2andCDKN1AmRNA expression respectively. These changes were associated with advanced granulosa cell differentiation in early antral follicles larger than 400 µm diameter, regardless of the presence or absence of FSH, as indicated by an increase in estradiol secretion, together with decreased AMH secretion and expression, as well as increased expression ofGJA1,CYP19A1,ESR1,ESR2,FSHR,INHA,INHBA,INHBBandFST. There was a decrease in the expression of oocyte-specific molecular markersGJA4,KIT,ZP3,WEE2andBMP15 in vitrocompared to thatin vivo. Moreover, a higher percentage of the oocytes recovered from cultured follicles 550 to 950 µm in diameter was able to reach the metaphase II meiosis stage. Overall, thisin vitromodel of ovarian follicle development is characterized by accelerated follicular maturation, associated with improved developmental competence of the oocyte, compared to follicles recoveredin vivo.