Growth and antrum formation of bovine primary follicles in long-term culture in vitro |
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| Authors: | Jing Sun Xiangdong Li |
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| Institution: | 1. Faculty of Veterinary Medicine, Laboratory of Manipulation of Oocytes and Preantral Follicles (LAMOFOPA), Veterinary Science Post Graduation Program (VSPGP), State University of Ceara, Av. Paranjana, 1700, Campus do Itaperi, Fortaleza, CE 60740-000, Brazil;2. Laboratory of Animal Physiology, Department of Animal Science, Federal University of Ceara, Av. Mister Hull S/N, Campus Pici, Fortaleza, CE CEP: 60021-970, Brazil;3. Potiguar University, Laureate International Universities, Av. Senador Salgado Filho, 1610, Lagoa Nova, Natal, RN CEP: 59056-000, Brazil;4. Faculty of Veterinary Medicine, Laboratory of Carnivore Reproduction (LRC), State University of Ceara, Av. Paranjana, 1700, Campus do Itaperi, Fortaleza, CE CEP: 60740-000, Brazil;1. Laboratory of Animal Reproduction, Faculty of Veterinary Medicine, Federal University of Uberlândia, 38400-902, Uberlândia, Minas Gerais, Brazil;2. Universidad de Murcia, Murcia, Spain;3. Nanobiotechnology Laboratory, Institute of Biotechnology, Federal University of Uberlândia, Uberlândia Minas Gerais, Brazil;4. Laboratory of Biology of Reproduction, Institute of Biomedical Sciences, Federal University of Uberlândia, Uberlândia Minas Gerais, Brazil;5. Department of Medical Microbiology and Immunology, University of California Davis, Davis CA, USA |
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| Abstract: | Successful antral formation in vitro from bovine preantral follicles (145–170 μm) has been described previously, but antrum formation from the primary follicle (50–70 μm) has not yet been achieved in vitro. The aim of the study was to establish an optimal culture system supporting the growth and maturation of bovine primary follicles (50–70 μm) in vitro. Bovine primary follicles were cultured in a three-dimensional culture system for 13 or 21 days in alpha-minimum essential medium. Various treatments including follicle stimulating hormone (FSH), luteinizing hormone (LH), 17β-estradiol (E2), basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF) were tested. The follicular diameter and antrum formation rate were recorded, and follicular maturation markers (P450 aromatase, CYP19A1; anti-Mullerian hormone, AMH; growth differentiation factor-9, GDF9; bone morphogenetic protein-15, BMP15; and type III transforming growth factor β receptor, TGFβR3) were analyzed by real-time RT-PCR. After 21 days of culture under each treatment condition, the follicular diameter was significantly enlarged in the presence of FSH + LH + E2 + bFGF or FSH + LH + E2 + bFGF + EGF (p < 0.05). An addition of 50 ng/ml bFGF or bFGF + 25 ng/ml EGF initiated antrum formation by day 19 and day 17 of culture, and the antral cavity formation rate was 16.7% and 33.3% by 21 days of culture, respectively. The expression of follicular maturation markers (CYP19A1, AMH, GDF9, BMP15 and TGFβR3) was significantly altered. We conclude that addition of 50 ng/ml bFGF + 25 ng/ml EGF to media containing FSH + LH + E2 turned out to be the most effective optimized culture conditions to support the growth and maturation of bovine primary follicles in vitro. |
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| Keywords: | Bovine Primary follicle Hormones Growth factors Antrum formation |
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