Martins F S, Celestino J J H, Saraiva M V A, Matos M H T, Bruno J B, Rocha-Junior C M C, Lima-Verde I B, Lucci C M, Báo S N, Figueiredo J R
Laboratory of Manipulation of Oocytes Enclosed in Preantral Follicles, Veterinary Faculty, State University of Ceará, Fortaleza, CE, Brazil.
Reprod Fertil Dev. 2008;20(8):916-24. doi: 10.1071/rd08108.
The aim of the present study was to investigate the effects of growth and differentiation factor-9 (GDF-9) on the survival and activation of preantral follicles, as well as their subsequent progression to secondary follicles, using goat ovarian cortical culture in vitro. Pieces of ovarian cortex were cultured for 1 and 7 days in minimum essential medium (MEM) with or without different concentrations of GDF-9 (1-200 ng mL(-1)). On Day 0 and after 1 and 7 days of culture, cortical pieces were fixed for histological and transmission electron microscopy evaluation. Preantral follicles were classified according to their development stage (primordial, intermediate, primary and secondary) and on the basis of morphological features (normal or degenerated). In addition, follicular and oocyte diameters were determined before and after culture. The results showed that, compared with non-cultured cortical tissue (Day 0), the culture of ovarian tissue significantly reduced (P < 0.05) the percentage of normal follicles in all media tested, except for tissue cultured in the presence of 200 ng mL(-1) GDF-9. Furthermore, in all media tested, the percentage of primordial follicles was significantly reduced (P < 0.05), with a concomitant increase in the percentage of developing follicles. The highest percentage of secondary follicles was observed after 7 days of culture in MEM plus 200 ng mL(-1) GDF-9. At all concentrations of GDF-9 tested, follicular diameter increased significantly after 7 days of culture compared with non-cultured cortical tissue. In conclusion, the results of the present study indicate that 200 ng mL(-1) GDF-9 maintains the survival of preantral follicles and promotes activation of primordial follicles. Furthermore, GDF-9 stimulates the transition from primary to secondary follicles, maintaining ultrastructural integrity of the follicles.
本研究的目的是利用山羊卵巢皮质体外培养，研究生长分化因子9（GDF-9）对窦前卵泡存活和激活以及随后发育为次级卵泡的影响。将卵巢皮质块在添加或不添加不同浓度GDF-9（1-200 ng mL(-1)）的最低必需培养基（MEM）中培养1天和7天。在培养第0天以及培养1天和7天后，将皮质块固定用于组织学和透射电子显微镜评估。根据窦前卵泡的发育阶段（原始、中间、初级和次级）以及形态特征（正常或退化）对其进行分类。此外，在培养前后测定卵泡和卵母细胞直径。结果表明，与未培养的皮质组织（第0天）相比，卵巢组织培养显著降低了（P < 0.05）所有测试培养基中正常卵泡的百分比，但在添加200 ng mL(-1) GDF-9的培养基中培养的组织除外。此外，在所有测试培养基中，原始卵泡的百分比显著降低（P < 0.05），同时发育中卵泡的百分比增加。在添加200 ng mL(-1) GDF-9的MEM中培养7天后，观察到次级卵泡的百分比最高。在所有测试的GDF-9浓度下，培养7天后卵泡直径与未培养的皮质组织相比显著增加。总之，本研究结果表明，200 ng mL(-1) GDF-9可维持窦前卵泡的存活并促进原始卵泡的激活。此外，GDF-9刺激初级卵泡向次级卵泡的转变，维持卵泡的超微结构完整性。
