title: Morphologic characterization of isolated bovine early preantral follicles during short-term individual in vitro culture.
abstract: To provide new insights in the molecular mechanism controlling preantral follicular development and to unravel the needs to support in vitro follicular development of early-stage preantral follicles (PAFs), there is a need for alternative in vitro bovine follicle culture methods. In this study, we aimed to characterize follicular dynamics using an IVC system of isolated and individually cultured bovine early PAFs during 10 days to generate individual follicle follow-up data. Preantral follicles (<50 μm) were isolated from slaughterhouse ovaries and cultured individually for 10 days. Individual follicle morphology, growth, survival, quality, and cell proliferation were evaluated in time by combining noninvasive and invasive assessment methods. The PAFs were light microscopically evaluated during culture to assess follicular dynamics, stained with neutral red to determine follicle viability, stained with 4',6-diamidino-2-phenylindole and terminal deoxynucleotidyl transferase dUTP nick end labeling to evaluate cell proliferation and follicle quality, and processed for histologic evaluation to assess follicle morphology. On the basis of their morphology, follicles were subdivided in three categories, with category 1 follicles showing the best morphologic features. On Day 0, only category 1 follicles were selected, but follicle categories were reassigned on evaluation Days 1, 2, 4, 7, or 10. Although 67% of the follicles survived 10 days of IVC, the number of follicles exhibiting a normal morphology decreased significantly from Day 7 onward and the apoptotic index increased significantly from Day 10. Both category 1 and 2 follicles showed a significant increase in follicular diameter (Day 10: 21.80 ± 0.86 and 11.82 ± 0.80, respectively). This increase in follicular diameter showed to be correlated with an increase in the total cell number. In conclusion, this culture system showed to support follicular development until Day 10, although the proportion of follicles showing normal morphologic features and the follicular quality decreased after 10 days of IVC. Follicles maintaining their category 1 morphologic features over time seem to be of a better quality and show a higher developmental competence as compared to category 2 and 3 follicles.
authors.affiliation: Laboratory of Veterinary Physiology and Biochemistry, Department of Veterinary Sciences, Faculty of Pharmaceutical, Biomedical and Veterinary Sciences, University of Antwerp, Wilrijk, Belgium. Electronic address: Ellen.Jorssen@uantwerpen.be.
authors.forename: E P A
authors.initials: EP
authors.lastname: Jorssen
authors.affiliation: Laboratory of Veterinary Physiology and Biochemistry, Department of Veterinary Sciences, Faculty of Pharmaceutical, Biomedical and Veterinary Sciences, University of Antwerp, Wilrijk, Belgium.
authors.forename: A
authors.initials: A
authors.lastname: Langbeen
authors.affiliation: Laboratory of Veterinary Physiology and Biochemistry, Department of Veterinary Sciences, Faculty of Pharmaceutical, Biomedical and Veterinary Sciences, University of Antwerp, Wilrijk, Belgium; Department of Theriogenology, Faculty of Veterinary Medicine, Cairo University, Giza, Egypt.
authors.forename: W F A
authors.initials: WF
authors.lastname: Marei
authors.affiliation: StatUa Center for Statistics, University of Antwerp, Antwerp, Belgium.
authors.forename: E
authors.initials: E
authors.lastname: Fransen
authors.affiliation: Laboratory of Veterinary Physiology and Biochemistry, Department of Veterinary Sciences, Faculty of Pharmaceutical, Biomedical and Veterinary Sciences, University of Antwerp, Wilrijk, Belgium.
authors.forename: H F M
authors.initials: HF
authors.lastname: De porte
authors.affiliation: Laboratory of Veterinary Physiology and Biochemistry, Department of Veterinary Sciences, Faculty of Pharmaceutical, Biomedical and Veterinary Sciences, University of Antwerp, Wilrijk, Belgium.
authors.forename: J L M R
authors.initials: JL
authors.lastname: Leroy
authors.affiliation: Laboratory of Veterinary Physiology and Biochemistry, Department of Veterinary Sciences, Faculty of Pharmaceutical, Biomedical and Veterinary Sciences, University of Antwerp, Wilrijk, Belgium.
authors.forename: P E J
authors.initials: PE
authors.lastname: Bols
mesh_terms: D000818:Animals; D017209:Apoptosis; D002417:Cattle; D049109:Cell Proliferation; D005260:Female; D005456:Fluorescent Dyes; D020287:In Situ Nick-End Labeling; D007211:Indoles; D009924:Organ Culture Techniques / Q000379:methods / Q000662:veterinary; D006080:Ovarian Follicle / Q000033:anatomy & histology* / Q000254:growth & development / Q000502:physiology; D013194:Staining and Labeling; D046509:Tissue Culture Techniques / Q000662:veterinary
chemical_list: D005456:Fluorescent Dyes; D007211:Indoles; C007293:DAPI
keywords: Bovine; IVC; Individual; Preantral follicle