Schiffmann D, De Boni U
Institute of Toxicology, University of Würzburg, F.R.G.
Mutat Res. 1991 Jan;246(1):113-22. doi: 10.1016/0027-5107(91)90113-3.
The in vitro micronucleus test with Syrian hamster embryo (SHE) cells assays the induction of micronuclei by chemical agents. Both chromosome fragments and lagging chromosomes can give rise to micronuclei. Nevertheless, only limited information is available on the ultrastructure of micronuclei and the mechanisms of their formation. Diethylstilbestrol (DES), a non-mutagenic carcinogen, as well as its analogue 3.3'-DES induce micronuclei in SHE cells. A comparison of the dose response of DES-induced micronucleus formation with the previously published ones for aneuploidy and transformation shows that all 3 run in parallel. Thus, a functional relationship between these endpoints, in the SHE system, may be implied. The present study is designed to address the formation of micronuclei using supravital UV microscopy, to test for the presence of defined chromosome domains within micronuclei using immunocytochemistry, and to define aspects of their ultrastructure by electron microscopy. Supravital UV microscopy showed that 3.3'-DES induces displacement of chromosomes/chromatids during prophase/anaphase and formation of micronuclei during cytokinesis. Immunocytochemistry revealed that micronuclei contain, at high frequencies, CREST antibody-reactive kinetochores, indicating the presence of whole chromosomes or centric fragments in these structures. Moreover, transmission electron microscopy showed that micronuclei exhibit ultrastructural details typical of interphase nuclei. Specifically, micronuclei exhibited morphological evidence of a nuclear lamina and segregation of karyoplasm into euchromatic and heterochromatic regions. All micronuclei examined were enclosed by a nuclear envelope of normal morphology and showed nuclear pore complexes. Together the findings provide evidence that DES interferes with the mitotic apparatus as early as prophase, resulting in the formation of micronuclei and, as a consequence, in the loss of chromatids or chromosomes.
用叙利亚仓鼠胚胎（SHE）细胞进行的体外微核试验可检测化学试剂诱导微核形成的情况。染色体片段和落后染色体都可产生微核。然而，关于微核超微结构及其形成机制的信息有限。己烯雌酚（DES）是一种非诱变致癌物，其类似物3,3'-DES可在SHE细胞中诱导微核形成。将DES诱导微核形成的剂量反应与先前发表的非整倍体和转化的剂量反应进行比较，结果表明这三者呈平行关系。因此，在SHE系统中，这些终点之间可能存在功能关系。本研究旨在利用活体紫外线显微镜观察微核的形成，利用免疫细胞化学检测微核内特定染色体区域的存在，并通过电子显微镜确定其超微结构的各个方面。活体紫外线显微镜观察显示，3,3'-DES在前期/后期诱导染色体/染色单体移位，并在胞质分裂期间诱导微核形成。免疫细胞化学显示，微核中高频含有抗着丝点蛋白抗体反应性着丝粒，表明这些结构中存在整条染色体或着丝粒片段。此外，透射电子显微镜显示，微核呈现出典型的间期核超微结构细节。具体而言，微核呈现出核纤层的形态学证据，以及核质分为常染色质和异染色质区域的现象。所有检查的微核都被形态正常的核膜包围，并显示有核孔复合体。这些研究结果共同证明，DES早在前期就干扰有丝分裂装置，导致微核形成，进而导致染色单体或染色体丢失。
