title: Long-term in vitro culture of bovine preantral follicles: Effect of base medium and medium replacement methods.
abstract: Two culture media and replacement methods were compared during long-term in vitro culture of secondary follicles of cattle using α-MEM(+) or TCM-199(+) as base media. The medium replacement methods were: Conventional - removal and subsequent addition of the same amount (60μl) in a 100μl aliquot (MEM-C and TCM-C), and Small Supplementation - addition of 5μl of fresh medium to an initial small aliquot (50μl), resulting in a final volume of 125μl on the last day of culture (MEM-S and TCM-S). A total of 207 secondary follicles were cultured individually for 32 days at 38.5°C in 5% CO2 and medium replacement was performed every other day. The MEM-S treatment resulted in a larger (P<0.01) follicular diameter, greater (P<0.02) growth rate, greater (P<0.02) antrum formation, as well as greater (P<0.0001) estradiol concentrations when compared with the MEM-C treatment. The medium change methods did not affect (P>0.05) the follicular and estradiol end points for TCM-199(+). The expression of the FSHR gene was greater (P<0.03) with the TCM-C than TCM-S treatment, while the relative amounts of mRNA for IGF1 was greater (P<0.02) with MEM-S than TCM-S treatments and for VEGF was greater (P<0.02) with MEM-C than TCM-C treatment. In conclusion, the type of base medium and the effect of periodic addition of medium differentially affected follicle development, estradiol production, and gene expression. Furthermore, α-MEM(+) can be used to replace TCM-199(+) for culture of preantral follicles of cattle if progressive addition of medium is used for medium change.
authors: Department of Animal Science, Food and Nutrition, Southern Illinois University, 1205 Lincoln Drive, MC 4417, Carbondale, IL, 62901, USA; Laboratory of Manipulation of Oocytes and Preantral Follicles (LAMOFOPA), Veterinary Faculty, State University of Ceará, Av. Paranjana 1700, Campus do Itaperi, Fortaleza, 60740-903 CE, Brazil.
authors: V R
authors: VR
authors: Araújo
authors: Department of Animal Science, Food and Nutrition, Southern Illinois University, 1205 Lincoln Drive, MC 4417, Carbondale, IL, 62901, USA.
authors: M O
authors: MO
authors: Gastal
authors: Department of Animal Science, Food and Nutrition, Southern Illinois University, 1205 Lincoln Drive, MC 4417, Carbondale, IL, 62901, USA.
authors: A
authors: A
authors: Wischral
authors: Laboratory of Manipulation of Oocytes and Preantral Follicles (LAMOFOPA), Veterinary Faculty, State University of Ceará, Av. Paranjana 1700, Campus do Itaperi, Fortaleza, 60740-903 CE, Brazil.
authors: J R
authors: JR
authors: Figueiredo
authors: Department of Animal Science, Food and Nutrition, Southern Illinois University, 1205 Lincoln Drive, MC 4417, Carbondale, IL, 62901, USA. Electronic address: egastal@siu.edu.
authors: E L
authors: EL
authors: Gastal
mesh_terms: D000818:Animals; D002417:Cattle; D002470:Cell Survival; D002478:Cells, Cultured; D003470:Culture Media; D004958:Estradiol / Q000378:metabolism; D005260:Female; D066298:In Vitro Techniques; D006080:Ovarian Follicle / Q000166:cytology* / Q000502:physiology; D016133:Polymerase Chain Reaction / Q000662:veterinary; D012313:RNA / Q000378:metabolism
chemical_list: D003470:Culture Media; D004958:Estradiol; D012313:RNA
keywords: Cattle; Estradiol; Folliculogenesis; In vitro culture; Secondary follicle