Paranko J, Salonen I
Department of Anatomy, University of Turku, Finland.
Reprod Fertil Dev. 1995;7(1):97-105. doi: 10.1071/rd9950097.
Calicin, a basic cytoskeletal protein has been proposed to be involved in the formation and maintenance of the highly regular organization of the postacrosomal perinuclear theca, the calyx of mammalian spermatozoa. The fate of human sperm calicin in the zona-free hamster egg fertilized in vitro has been analysed at the light and electron microscope level using polyclonal mouse anti-calicin antibodies. Calicin was localized in the postacrosomal dense lamina of the capacitated acrosome-reacted spermatozoa attached on the surface of the egg as well as in the spermatozoa at an early stage of ooplasmic incorporation. As determined by the aid of the DNA-specific fluorescent dye 4,6-diamidino-2-phenylindole (DAPI), the pattern of staining with anti-calicin changed from a funnel-shape into a ring soon after the onset of the nuclear decondensation. Later, no anti-calicin labelling could be detected around the more decondensed sperm nuclei. Because the residual, ring-like accumulation of calicin was associated with the least decondensed chromatin, it appears that the degradation of calicin-containing perinuclear theca is intimately involved with the posteriorly advancing nuclear disintegration. The ring formation also suggests that the calyx of human spermatozoa is not structurally homogeneous at least in terms of sensitivity to ooplasmic degradation. Calicin appears to be unaffected by lytic enzymes of the acrosome. The present study further shows that DAPI can be effectively used to analyse sperm nuclear decondensation in vitro.
钙结合蛋白是一种碱性细胞骨架蛋白，有人提出它参与了顶体后核周鞘（哺乳动物精子的顶体杯）高度规则组织的形成和维持。利用多克隆小鼠抗钙结合蛋白抗体，在光学显微镜和电子显微镜水平上分析了人精子钙结合蛋白在体外无透明带仓鼠卵受精后的命运。钙结合蛋白定位于附着在卵表面的获能顶体反应精子的顶体后致密层以及卵质掺入早期的精子中。借助DNA特异性荧光染料4,6-二脒基-2-苯基吲哚（DAPI）确定，在核去浓缩开始后不久，抗钙结合蛋白的染色模式从漏斗形变为环形。后来，在去浓缩程度更高的精子核周围未检测到抗钙结合蛋白标记。由于钙结合蛋白的残留环状积累与去浓缩程度最低的染色质相关，似乎含钙结合蛋白的核周鞘的降解与向后推进的核解体密切相关。环状结构的形成还表明，至少就对卵质降解的敏感性而言，人精子的顶体杯在结构上并非均匀一致。钙结合蛋白似乎不受顶体裂解酶的影响。本研究进一步表明，DAPI可有效地用于体外分析精子核去浓缩。
