title: Insulin improves in vitro survival of equine preantral follicles enclosed in ovarian tissue and reduces reactive oxygen species production after culture.
abstract: This study investigated the effect of insulin concentration on the in vitro culture of equine preantral follicles enclosed in ovarian tissue. Ovarian tissue samples were immediately fixed (noncultured control) or cultured for 1 or 7 days in α-MEM(+) supplemented with 0 ng/mL, 10 ng/mL, or 10 μg/mL insulin. Ovarian tissues were processed and analyzed by classical histology. Culture medium samples were collected after 1 and 7 days of culture for steroid and reactive oxygen species (ROS) analyses. The percentage of morphologically normal follicles was greater (P < 0.001) in insulin-treated groups after 1 day of culture; likewise, more (P < 0.02) normal follicles were observed after 7 days of culture in medium supplemented with 10-ng/mL insulin. Furthermore, an increase (P < 0.01) in developing (transition, primary, and secondary) follicles between Days 1 and 7 of culture was observed only with the 10-ng/mL insulin treatment. ROS production after 1 or 7 days of culture was lower (P < 0.0001) in medium with 10-ng/mL insulin than the other treatments. Ovarian tissues containing preantral follicles were able to produce estradiol and progesterone after 1 and 7 days of culture; however, treatments did not differ in steroid production. In conclusion, the use of a physiological concentration (10 ng/mL) of insulin rather than the previously reported concentration (10 μg/mL) for in vitro culture of equine preantral follicles improved follicular survival and growth and lowered oxidative stress. Results from this study shed light on new perspectives for producing an appropriate medium to improve equine preantral follicle in vitro survival and growth.
authors.affiliation: Laboratory of Manipulation of Oocytes and Preantral Follicles (LAMOFOPA), Faculty of Veterinary Medicine, State University of Ceará, Fortaleza, Ceará, Brazil.
authors.forename: F L N
authors.initials: FL
authors.lastname: Aguiar
authors.affiliation: Laboratory of Manipulation of Oocytes and Preantral Follicles (LAMOFOPA), Faculty of Veterinary Medicine, State University of Ceará, Fortaleza, Ceará, Brazil.
authors.forename: F O
authors.initials: FO
authors.lastname: Lunardi
authors.affiliation: Laboratory of Manipulation of Oocytes and Preantral Follicles (LAMOFOPA), Faculty of Veterinary Medicine, State University of Ceará, Fortaleza, Ceará, Brazil.
authors.forename: L F
authors.initials: LF
authors.lastname: Lima
authors.affiliation: Laboratory of Manipulation of Oocytes and Preantral Follicles (LAMOFOPA), Faculty of Veterinary Medicine, State University of Ceará, Fortaleza, Ceará, Brazil.
authors.forename: R M P
authors.initials: RM
authors.lastname: Rocha
authors.affiliation: Laboratory of Manipulation of Oocytes and Preantral Follicles (LAMOFOPA), Faculty of Veterinary Medicine, State University of Ceará, Fortaleza, Ceará, Brazil.
authors.forename: J B
authors.initials: JB
authors.lastname: Bruno
authors.affiliation: Biotechnology Graduate School, Potiguar University/Laureate International Universities, Natal, Rio Grande do Norte, Brazil.
authors.forename: D M
authors.initials: DM
authors.lastname: Magalhães-Padilha
authors.affiliation: Federal University of Pampa, Uruguaiana, Rio Grande do Sul, Brazil.
authors.forename: F W S
authors.initials: FW
authors.lastname: Cibin
authors.affiliation: Laboratory of Manipulation of Oocytes and Preantral Follicles (LAMOFOPA), Faculty of Veterinary Medicine, State University of Ceará, Fortaleza, Ceará, Brazil.
authors.forename: A P R
authors.initials: AP
authors.lastname: Rodrigues
authors.affiliation: Department of Animal Science, Food and Nutrition, Southern Illinois University, Carbondale, Illinois, USA.
authors.forename: M O
authors.initials: MO
authors.lastname: Gastal
authors.affiliation: Department of Animal Science, Food and Nutrition, Southern Illinois University, Carbondale, Illinois, USA. Electronic address: egastal@siu.edu.
authors.forename: E L
authors.initials: EL
authors.lastname: Gastal
authors.affiliation: Laboratory of Manipulation of Oocytes and Preantral Follicles (LAMOFOPA), Faculty of Veterinary Medicine, State University of Ceará, Fortaleza, Ceará, Brazil.
authors.forename: J R
authors.initials: JR
authors.lastname: Figueiredo
mesh_terms: D000818:Animals; D003470:Culture Media; D005260:Female; D006736:Horses; D007328:Insulin / Q000494:pharmacology*; D006080:Ovarian Follicle / Q000187:drug effects* / Q000254:growth & development / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism*; D027724:Reproductive Techniques, Assisted / Q000662:veterinary; D046509:Tissue Culture Techniques / Q000662:veterinary*
chemical_list: D003470:Culture Media; D007328:Insulin; D017382:Reactive Oxygen Species
keywords: Equine ovary; Insulin; In vitro culture; Preantral follicle; ROS