摘要
三丁基锡(TBT)作为一种生物灭杀剂,广泛应用于船舶防污涂料中。我们前期的研究表明,将热带爪蟾胚胎在环境浓度下的TBT中暴露24 h、36 h和48 h后,胚胎产生特异的畸形现象,如眼睛畸形、泄殖腔膨大、窄鳍和皮肤色素消失等,而导致这些畸形表型产生的原因尚未明确。
为了进一步研究TBT诱导热带爪蟾胚胎产生特异性畸形表型的原因,我们将热带爪蟾胚胎暴露于50、100和200 ng·L-1TBTCl中,36 h和72 h后观察畸变部位组织结构的变化。以揭示TBT诱导的热带爪蟾胚胎内部组织结构的变化,及其与畸形表型之间的联系。
结果表明,TBT导致热带爪蟾胚胎眼睛畸形、躯干膨大、泄殖腔膨大、尾鳍变窄甚至消失。进一步的组织学观察显示,晶状体和视网膜层很少分化甚至不分化,色素层既不连续也不平滑;外鳃未被吸收,内鳃未分化完全;软腭板形态畸形,甚至咽部没有形成软腭板;膨大的腹腔内部充满了包含大量卵黄的未分化的内脏细胞;大量富含卵黄颗粒的细胞填充在呈肿块状或棒状隆起的泄殖腔内部,堵塞了出口甚至外翻至体外,有的泄殖腔外表皮膨胀;在躯干和尾部的背鳍和腹鳍均变窄甚至完全消失。
组织结构的变化揭示了畸形表型的内在原因。所有胚胎早期的细胞均含有卵黄颗粒,这些卵黄颗粒随着胚胎的发育而在细胞内逐渐被吸收。浓度组腹腔内细胞富含卵黄颗粒,说明细胞未完全分化,导致肠道没有进一步拉长和盘绕,进而使得腹腔膨大。在TBT暴露组中,泄殖腔壁细胞富含卵黄颗粒,细胞类型与大肠壁细胞类似。由此可见,增厚的泄殖腔壁仍然来自于原有未分化的肠道组织。泄殖腔上表皮细胞未分化、外表皮扩张和背、腹鳍的消失导致了泄殖腔的膨大。
热带爪蟾尾鳍的变窄甚至消失可能与TBT诱导的细胞凋亡有关。在TBT导致的胚胎畸形中,尾鳍的畸形和泄殖腔的膨大总是同时发生,而泄殖腔和尾鳍的发育均与骨形成蛋白信号(BMP)有关。总之,组织学观察揭示了TBT诱导的爪蟾胚胎特异性畸形产生的内在原因,并为机制的解析提供了依据。
Tributyltin (TBT) has been widely used as a biocide in antifouling paints. In our previous study, we found that unique malformations are induced in Xenopus tropicalis embryos by TBT at environmentally relevant concentrations after 24,36 and 48 h of exposure. The most obvious alterations were abnormal eyes, enlarged proctodaeums, narrow fins and skin hypopigmentaion. The reasons leading to the external changes, however, are not well known.
Based on the findings of morphological malformations above, we exposed embryos of X. tropicalis to 50,100 and 200 ng·L-1 tributyltin chloride (TBTC1) for 36 and 72 h, and the histological changes were observed on the main phenotypes of malformations. Our aim was to determine the histological abnormalities induced by TBT and to reveal the internal relationship between histological changes and the external malformations.
The embryos treated with TBT showed multiple malformations such as abnormal eyes, enlarged trunks, enlarged proctodaeums and narrow fins. The histological changes were further observed on these main malformations induced by TBT. The lens and the retinal layers of abnormal eyes were slightly or barely differentiated, and that the pigment epithelium was neither continuous nor smooth. The external gills were not absorbed, and the inner gills were not well developed. The velar plates developed in a deformed way, what is worse, some were absent in the pharynx. The abdomens were full of undifferentiated gut tissue with yolk-rich inclusions in the tadpoles with enlarged trunks. The proctodaeums formed a bump-like or columnar structure. The mass of yolk-rich cells occupied the lumen, blocked the opening and even turned inside out of the proctodaeum. Both the ventral and dorsal fins in trunks and tails became narrow or even disappeared totally.
Our results suggest that great changes of histology took place corresponding to the unique phenotypes. All the embryonic cells inherit yolk platelets from the egg cytoplasm and consumed them intracellularly during embryogenesis. The yolk-rich mass in TBT treatment tadpoles suggest that the gut tissue was poorly differentiated, which led to the failed elongation of the guts and subsequently the enlarged trunks. In TBT treatment groups, both the large intestine and proctodaeum were rich in yolk granules, sharing the similar cell types. Therefore, the thick proctodaeums still came from the original undifferentiated gut tissues. The enlarged proctodaeums were due to the undifferentiation of inner layer, the expansion of outer epidermal part and the absence of fins around them.
The narrow fins or absence of fins in X. tropicalis larvae might be due to the apoptosis induced by TBT. The fin defects were always companied by enlarged protodaeums in this study. The development of proctodaeums and ventral fins are closely related to bone morphologenetic protein (Bmp) signaling. In brief, the histological observations provided insights into the reason of the unique external malformations in some degree.
引文
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