Research Scientist from the Department of Obstetrics and Gynecology, New York University School of Medicine, 550 First Avenue, NBV 9N1, New York, NY, USA.
Human Reproduction Division, Department of Obstetrics and Gynecology, Faculty of Medicine of Ribeirao Preto, University of Sao Paulo, Ribeirao Preto, SP, Brazil.
J Assist Reprod Genet. 2019 Sep;36(9):1877-1890. doi: 10.1007/s10815-019-01530-w. Epub 2019 Jul 22.
To identify and characterize amyloid-like substance (ALS) in human and mouse oocytes and preimplantation embryos.
An experimental prospective pilot study. A total of 252 mouse oocytes and preimplantation embryos and 50 immature and in vitro matured human oocytes and parthenogenetic human embryos, from 11 consenting fertility patients, ages 18-45. Fluorescence intensity from immunofluorescent staining and data from confocal microscopy were quantified. Data were compared by one-way analysis of variance, with the least square-MEANS post-test, Pearson correlation coefficients (r), and bivariate analyses (t tests). ALS morphology was verified using transmission electron microscopy.
Immunostaining for ALS appears throughout the zona pellucida, as well as in the cytoplasm and nucleus of mouse and human oocytes, polar bodies, and parthenogenetic embryos, and mouse preimplantation embryos. In mouse, 2-cell embryos exhibited the highest level of ALS (69000187.4 ± 6733098.07). Electron microscopy confirmed the presence of ALS. In humans, fresh germinal vesicle stage oocytes exhibited the highest level of ALS (4164.74088 ± 1573.46) followed by metaphase I and II stages (p = 0.008). There was a significant negative association between levels of ALS and patient body mass index, number of days of ovarian stimulation, dose of gonadotropin used, time between retrieval and fixation, and time after the hCG trigger. Significantly higher levels of ALS were found in patients with AMH between 1 and 3 ng/ml compared to < 1 ng/ml.
We demonstrate for the first time the presence, distribution, and change in ALS throughout some stages of mouse and human oocyte maturation and embryonic development. We also determine associations between ALS in human oocytes with clinical characteristics.
鉴定和描述人卵母细胞和植入前胚胎中的类淀粉样物质（ALS）。
这是一项实验性前瞻性试点研究。共检测了 11 名年龄在 18-45 岁之间的有生育意愿的患者的 252 个小鼠卵母细胞和植入前胚胎以及 50 个不成熟和体外成熟的人类卵母细胞和孤雌胚胎的类淀粉样物质。通过免疫荧光染色进行荧光强度定量，利用共聚焦显微镜获取数据。采用单因素方差分析，最小二乘法均值检验、皮尔逊相关系数（r）和双变量分析（t 检验）进行数据比较。利用透射电子显微镜验证 ALS 的形态。
免疫组化染色显示，在小鼠和人卵母细胞、极体和孤雌胚胎以及小鼠 2 细胞胚胎的透明带以及细胞质和细胞核中均有 ALS 存在。在小鼠中，2 细胞胚胎的 ALS 水平最高（69000187.4±6733098.07）。电镜证实了 ALS 的存在。在人类中，新鲜的生发泡期卵母细胞表现出最高水平的 ALS（4164.74088±1573.46），其次是中期 I 期和中期 II 期（p=0.008）。ALS 水平与患者的体重指数、促排卵天数、促性腺激素用量、取卵和固定时间间隔以及 hCG 触发后的时间呈显著负相关。与 <1ng/ml 的 AMH 相比，1-3ng/ml 的 AMH 患者的 ALS 水平显著更高。
我们首次证明了 ALS 存在于小鼠和人卵母细胞成熟和胚胎发育的某些阶段，并在其中分布和变化。我们还确定了人类卵母细胞中 ALS 与临床特征之间的关联。
