Estradiol-promoted accumulation of receptor in nuclei of porcine endometrium cells. Immunogold electron microscopy of resting and estradiol-stimulated cells.
W D Sierralta, F Jakob, H Thole, P Engel, P W Jungblut
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引用次数: 0
Abstract
Endometrium was collected by curettage from castrated pigs, either untreated or exposed to estradiol in vivo by intrauterine injection, and processed for electron microscopy. The resin LR Gold was used for embedding, and sections were floated on droplets of 10 nm diameter gold particles, coated with the immunoglobulin-G1 (IgG1) fraction or its Fab2 fragment of a monospecific polyclonal antiserum raised in goats against the C-terminal half of the estradiol receptor. On average, only one gold particle per microns 2 became attached in the cytoplasmic area of untreated cells, whereas four were found over the nuclear area. These figures rose to 2-3/microns 2 and 15-26/microns 2, respectively, within 10 min after exposure to estradiol. The labeling intensities of nuclei in cell clusters and of coprocessed nuclei released from cells ruptured during curettage were identical in all situations. Nuclear pores were frequently tagged after estradiol treatment. The proportions of tagging densities in nuclei of untreated and estradiol-exposed cells corresponded to those of receptor contents measured in extracts of isolated nuclei by ligand binding. This correlation was not seen for the cytoplasmic compartment of untreated cells, the scarce tagging of which is interpreted by hidden antigenic determinants. Our morphological analyses support the conclusions drawn from biochemical data (Sierralta et al., 1992) of an estradiol-promoted translocation of receptor from the cytoplasm into the nucleus.
采用刮除法收集去势猪子宫内膜,分别未经处理或宫内注射雌二醇,处理后进行电镜观察。用LR Gold树脂包埋,将切片漂浮在直径为10 nm的金颗粒液滴上,涂有山羊培养的针对雌二醇受体c端一半的单特异性多克隆抗血清的免疫球蛋白g1 (IgG1)片段或其Fab2片段。在未经处理的细胞中,平均每微米只有一个金颗粒附着在细胞质区域,而在细胞核区域则有四个。在接触雌二醇后的10分钟内,这些数字分别上升到2-3/微米2和15-26/微米2。在所有情况下,细胞团核的标记强度和刮除时破裂细胞释放的共加工核的标记强度是相同的。雌二醇处理后,核孔经常被标记。未经处理和雌二醇暴露的细胞的细胞核中标记密度的比例与通过配体结合在离体细胞核提取物中测量的受体含量的比例相对应。这种相关性在未处理细胞的细胞质室中未见,其稀少的标记被隐藏的抗原决定因素解释。我们的形态学分析支持生化数据得出的结论(sierra alta et al., 1992),雌二醇促进了受体从细胞质向细胞核的易位。