Faculty of Veterinary Medicine, LAMOFOPA, PPGCV, State University of Ceara, Fortaleza, CE, Brazil.
Mol Reprod Dev. 2010 Mar;77(3):231-40. doi: 10.1002/mrd.21138.
The aims of this study were to investigate steady-state level of Kit Ligand (KL) mRNA and its effects on in vitro survival and growth of caprine preantral follicles. RT-PCR was used to analyze caprine steady-state level of KL mRNA in primordial, primary, and secondary follicles, and in small (1-3 mm) and large (3-6 mm) antral follicles. Furthermore, ovarian fragments were cultured for 1 or 7 days in Minimal Essential Medium (MEM(+)) supplemented with KL (0, 1, 10, 50, 100, or 200 ng/ml). Noncultured (control) and cultured fragments were processed for histology and transmission electron microscopy (TEM). RT-PCR demonstrated an increase in steady-state level of KL mRNA during the transition from primary to secondary follicles. Small antral follicles had higher steady-state levels of KL mRNA in granulosa and theca cells than large follicles. After 7 days, only 50 ng/ml of KL had maintained the percentage of normal follicles similar to control. After 1 day, all KL concentrations reduced the percentage of primordial follicles and increased the percentage of growing follicles. KL at 10, 50, 100, or 200 ng/ml increased primary follicles, compared to MEM(+) after 7 days. An increase in oocyte and follicular diameter was observed at 50 ng/ml of KL. TEM confirmed ultrastructural integrity of follicles after 7 days at 50 ng/ml of KL. In conclusion, the KL mRNAs were detected in all follicular categories. Furthermore, 50 ng/ml of KL maintained the integrity of caprine preantral follicle cultured for 7 days and stimulated primordial follicle activation and follicle growth.
本研究旨在探讨 Kit 配体（KL）mRNA 的稳态水平及其对山羊腔前卵泡体外存活和生长的影响。采用 RT-PCR 分析原始卵泡、初级卵泡和次级卵泡，以及小（1-3mm）和大（3-6mm）腔前卵泡中 KL mRNA 的山羊稳态水平。此外，将卵巢片段在添加 KL（0、1、10、50、100 或 200ng/ml）的最小必需培养基（MEM(+))中培养 1 或 7 天。未培养（对照）和培养的片段进行组织学和透射电镜（TEM）处理。RT-PCR 表明，KL mRNA 的稳态水平在从初级卵泡向次级卵泡过渡期间增加。小腔前卵泡的颗粒细胞和膜细胞中的 KL mRNA 稳态水平高于大卵泡。7 天后，只有 50ng/ml 的 KL 维持正常卵泡的百分比与对照相似。培养 1 天后，所有 KL 浓度均降低了原始卵泡的百分比并增加了生长卵泡的百分比。与 MEM(+)相比，10、50、100 或 200ng/ml 的 KL 在第 7 天增加了初级卵泡。在 KL 50ng/ml 下观察到卵母细胞和卵泡直径增加。TEM 在 KL 50ng/ml 下培养 7 天后证实了卵泡的超微结构完整性。综上所述，在所有卵泡类别中均检测到 KL mRNAs。此外，50ng/ml 的 KL 维持了培养 7 天的山羊腔前卵泡的完整性，并刺激了原始卵泡的激活和卵泡生长。
