Serafim Michelle K B, Silva Gerlane M, Duarte Ana B G, Araújo V R, Silva T F P, Lima A K F, Chaves R N, Campello C C, Silva L D M, Figueiredo J R
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 60.740-000, Brazil.
Reprod Fertil Dev. 2013;25(6):927-34. doi: 10.1071/RD12074.
To determine whether the effects of different concentrations of insulin on the development of canine preantral follicles in vitro were associated or not with FSH, secondary follicles were isolated and cultured. In Experiment 1, follicles were cultured in the following media: modified minimum essential medium (CtrlMEM) alone; CtrlMEM plus 5 ng mL⁻¹ insulin (Ins5ng); CtrlMEM plus 10 ng mL⁻¹ insulin (Ins10ng); and CtrlMEM plus 10 μg mL⁻¹ insulin. In Experiment 2, follicles were cultured in the same media but in the presence of sequential FSH (i.e. CtrlFSH, Ins5ngF, Ins10ngF and 10μgF, respectively). Increasing concentrations of FSH (100, 500 and 1000 ng mL⁻¹) were added sequentially to the culture medium on Days 0, 6 and 12 of culture. Viability were assessed at the end of culture and follicular diameter and the antrum formation rate at four time points (Days 0, 6, 12 and 18). In Experiment 1, the high insulin concentration significantly increased follicular viability (P<0.05). In contrast, in Experiment 2, viability was not affected by the inclusion of insulin. In addition, viability was significantly better in follicles cultured in CtrlFSH (P<0.05). The diameter of follicles in the high-insulin group in Experiment 1 and high-insulin plus FSH group in Experiment 2 was superior to other groups tested. In experiment 2, the Ins10μg and Ins10μgF groups exhibited significantly higher antrum formation rates than the other groups. In conclusion, in the absence of FSH, high concentrations of insulin have beneficial effects on follicular viability. However, to promote the growth of canine preantral follicles in vitro, it is recommended that a combination of insulin and FSH be added to the medium.
为了确定不同浓度胰岛素对犬类腔前卵泡体外发育的影响是否与促卵泡激素（FSH）有关，分离并培养了次级卵泡。在实验1中，卵泡在以下培养基中培养：单独的改良最小必需培养基（CtrlMEM）；CtrlMEM加5 ng/mL胰岛素（Ins5ng）；CtrlMEM加10 ng/mL胰岛素（Ins10ng）；以及CtrlMEM加10 μg/mL胰岛素。在实验2中，卵泡在相同培养基中培养，但添加了序贯FSH（即分别为CtrlFSH、Ins5ngF、Ins10ngF和10μgF）。在培养的第0天、第6天和第12天，依次向培养基中添加浓度递增的FSH（100、500和1000 ng/mL）。在培养结束时评估活力，并在四个时间点（第0天、第6天、第12天和第18天）测量卵泡直径和腔形成率。在实验1中，高胰岛素浓度显著提高了卵泡活力（P<0.05）。相比之下，在实验2中，胰岛素的添加对活力没有影响。此外，在CtrlFSH中培养的卵泡活力显著更好（P<0.05）。实验1中的高胰岛素组和实验2中的高胰岛素加FSH组的卵泡直径优于其他测试组。在实验2中，Ins10μg组和Ins10μgF组的腔形成率显著高于其他组。总之，在没有FSH的情况下，高浓度胰岛素对卵泡活力有有益影响。然而，为了促进犬类腔前卵泡的体外生长，建议在培养基中添加胰岛素和FSH的组合。
