Facultad de Ciencias de la Salud, Carrera de Medicina, Universidad Maimónides, Ciudad Autónoma de Buenos Aires, Buenos Aires, Argentina.
Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Ciudad Autónoma de Buenos Aires, Buenos Aires, Argentina.
PLoS One. 2018 Sep 7;13(9):e0203268. doi: 10.1371/journal.pone.0203268. eCollection 2018.
Emerging evidence has shown that oocytes from diabetic ovaries exhibit delayed maturation, mitochondrial dysfunction and meiotic defects, which are related increased apoptosis. The main objective of the present study was to analyze the apoptosis pathways activated during follicular loss at multiple time points in a diabetic mouse model. Twenty BALB/c mice were used in this study, and diabetes mellitus was induced by streptozotocin injection. Three diabetic and two control animals were sacrificed on days 15, 20, 70 and 80 posttreatment. The ovaries were then removed; one was used for follicular counting, TUNEL, immunohistochemistry and immunofluorescence, while the other was used for Western blot analysis. The proteins studied were BAX, BCL2, t-BID, FAS, FASL, active caspase 8, active caspase 9 and active caspase 3. Follicular apoptosis decreased over time, with the highest values observed at 15 days posttreatment. Granulosa cells were positive for active caspase 3, which showed constant expression levels at all time points. FAS, FASL, t-BID and active caspase 8 showed strong cytoplasmic immunostaining in the oocytes and granulosa cells of the diabetic mice, with significant increases observed at 15, 20 and 70 days posttreatment. BAX expression was slightly higher in the diabetic mouse ovaries than in the control ovaries at 15, 20 and 70 days posttreatment, whereas the highest active caspase 9 expression was at observed 20 days posttreatment. Low BCL2 protein levels were detected in the diabetic mouse ovaries at all time points. This study describes for the first time the behavior of apoptosis-related proteins in the diabetic mouse ovary and shows not only that the FAS/FASL pathway contributes to follicular loss but also that antral follicles are the most affected.
新出现的证据表明，糖尿病卵巢中的卵母细胞表现出成熟延迟、线粒体功能障碍和减数分裂缺陷，这与凋亡增加有关。本研究的主要目的是分析在糖尿病小鼠模型中多个时间点卵泡丢失过程中激活的凋亡途径。本研究使用了 20 只 BALB/c 小鼠，并通过链脲佐菌素注射诱导糖尿病。在治疗后第 15、20、70 和 80 天，分别处死 3 只糖尿病和 2 只对照动物。然后取出卵巢；一个用于卵泡计数、TUNEL、免疫组织化学和免疫荧光，另一个用于 Western blot 分析。研究的蛋白质有 BAX、BCL2、t-BID、FAS、FASL、活性 caspase 8、活性 caspase 9 和活性 caspase 3。卵泡凋亡随时间减少，治疗后 15 天观察到最高值。颗粒细胞中活性 caspase 3 呈阳性，在所有时间点均表现出恒定的表达水平。FAS、FASL、t-BID 和活性 caspase 8 在糖尿病小鼠的卵母细胞和颗粒细胞中表现出强烈的细胞质免疫染色，在治疗后 15、20 和 70 天观察到明显增加。BAX 表达在治疗后 15、20 和 70 天的糖尿病小鼠卵巢中略高于对照组，而最高的活性 caspase 9 表达在治疗后 20 天观察到。在所有时间点，糖尿病小鼠卵巢中均检测到低水平的 BCL2 蛋白。本研究首次描述了凋亡相关蛋白在糖尿病小鼠卵巢中的行为，不仅表明 FAS/FASL 途径有助于卵泡丢失，而且表明窦前卵泡是最受影响的。
