Moreno R D, Ramalho-Santos J, Chan E K, Wessel G M, Schatten G
Oregon Regional Primate Research Center, 505 NW 185th Avenue, Beaverton, Oregon, 97006, USA.
Dev Biol. 2000 Mar 15;219(2):334-49. doi: 10.1006/dbio.2000.9606.
The acrosome is an acidic secretory vesicle containing hydrolytic enzymes that are involved in the sperm's passage across the zona pellucida. Imaging of the acrosomal vesicle and the Golgi apparatus in live rhesus monkey spermatids was accomplished by using the vital fluorescent probe LysoTracker DND-26. Concurrently, the dynamics of living spermatid mitochondria was visualized using the specific probe MitoTracker CMTRos and LysoTracker DND-26 detected the acrosomal vesicle from its formation through spermatid differentiation. LysoTracker DND-26 also labeled the Golgi apparatus in spermatogenic cells. In spermatocytes the Golgi is spherical and, in round spermatids, it is localized over the acrosomal vesicle, as confirmed by using polyclonal antibodies against Golgin-95/GM130, Golgin-97, and Golgin-160. Using both live LysoTracker DND-26 imaging and Golgi antibodies, we found that the Golgi apparatus is cast off from the acrosomal vesicle and migrates toward the sperm tail in elongated spermatids. The Golgi is discarded in the cytoplasmic droplet and is undetectable in mature ejaculated spermatozoa. The combined utilization of three vital fluorescent probes (Hoechst 33342, LysoTracker DND-26, and MitoTracker CMTRos) permits the dynamic imaging of four organelles during primate spermiogenesis: the nucleus, the mitochondria, the acrosomal vesicle, and the Golgi apparatus.
顶体是一种酸性分泌囊泡,含有水解酶,这些酶参与精子穿过透明带的过程。通过使用活性荧光探针溶酶体示踪剂DND - 26,实现了对恒河猴活精子细胞中顶体囊泡和高尔基体的成像。同时,使用特异性探针线粒体示踪剂CMTRos观察活精子细胞线粒体的动态变化,溶酶体示踪剂DND - 26则检测顶体囊泡从其形成到精子细胞分化的过程。溶酶体示踪剂DND - 26还标记了生精细胞中的高尔基体。在精母细胞中,高尔基体呈球形,在圆形精子细胞中,高尔基体位于顶体囊泡上方,这一点通过使用针对高尔基体蛋白95/GM130、高尔基体蛋白97和高尔基体蛋白160的多克隆抗体得以证实。通过活细胞溶酶体示踪剂DND - 26成像和高尔基体抗体的联合使用,我们发现高尔基体从顶体囊泡上脱离,并在伸长的精子细胞中向精子尾部迁移。高尔基体被丢弃在细胞质滴中,在成熟的射出精子中无法检测到。三种活性荧光探针(Hoechst 33342、溶酶体示踪剂DND - 26和线粒体示踪剂CMTRos)的联合使用,使得在灵长类动物精子发生过程中能够对四种细胞器进行动态成像:细胞核、线粒体、顶体囊泡和高尔基体。