Establishment of epidermal cell lines derived from the skin of the Atlantic bottlenose dolphin (Tursiops truncatus)
Establishment of epidermal cell lines derived from the skin of the Atlantic bottlenose dolphin (Tursiops truncatus)
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DOI：
10.1002/ar.a.20266
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发表时间：
2005-12-01
期刊：
影响因子：
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通讯作者：
Gattoni-Celli, S
中科院分区：
文献类型：
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作者：
Yu, J;Kindy, MS;Gattoni-Celli, S
大西洋海豚（Tursiops truncatus）是一种在大西洋海岸发现的海洋哺乳动物，由于过去沿着美国东南部，该物种的死亡事件越来越多，已成为人们关注的焦点。由于这些海洋哺乳动物的受保护地位，评估环境压力对白海豚（BND）的影响一直很困难。本文提出的研究集中于建立表皮细胞培养物和细胞系，作为体外评价BND皮肤上的环境应激源的工具。从大西洋BND获得的皮肤样品建立表皮细胞培养物，并进行核型分析。这些培养物进一步使用免疫组化方法表征，证明细胞角蛋白的表达。通过双向聚丙烯酰胺凝胶电泳（2D-PAGE），我们观察到BND皮肤组织样品的蛋白质组图谱与皮肤来源的培养物的蛋白质组图谱具有明显的相似性。用编码SV 40小T抗原和大T抗原以及新霉素抗性基因的质粒转染表皮细胞培养物。五个新霉素抗性克隆被分离和扩增，它们都以比未转染的BND表皮培养物更快的速度增殖，而未转染的BND表皮培养物表现出衰老的迹象。通过Western印迹分析，从两个转染的克隆制备的细胞裂解物显示表达两种SV 40肿瘤抗原。这些实验结果与表达SV 40肿瘤抗原的转染克隆代表永生化BND细胞系的概念一致。来自Tursiops truncatus的表皮细胞系将提供一个独特的工具，用于研究海豚与它们生活在最重要界面（皮肤）的环境之间相互作用的关键特征。(c)2005 Wiley-Liss，Inc.
The Atlantic bottlenose dolphin (Tursiops truncatus), a marine mammal found off the Atlantic coast, has become the focus of considerable attention because of an increasing number of mortality events witnessed in this species over the last several years along the southeastern United States. Assessment of the impact of environmental stressors on bottlenose dolphins (BND) has been difficult because of the protected status of these marine mammals. The studies presented herein focused on establishing epidermal cell cultures and cell lines as tools for the in vitro evaluation of environmental stressors on BND skin. Epidermal cell cultures were established from skin samples obtained from Atlantic BND and subjected to karyotype analysis. These cultures were further characterized using immunohistochemical methods demonstrating expression of cytokeratins. By two-dimensional polyacrylamide gel electrophoresis (2D-PAGE), we observed that the proteomic profile of BND skin tissue samples shared distinct similarities with that of skin-derived cultures. Epidermal cell cultures were transfected with a plasmid encoding the SV40 small t- and large T-antigens, as well as the neomycin-resistance gene. Five neomycin-resistant clones were isolated and expanded, and all of them proliferated at a faster rate than nontransfected BND epidermal cultures, which exhibited signs of senescence. Cell lysates prepared from two transfected clones were shown to express, by Western blot analysis, both SV40 tumor antigens. These experimental results are consistent with the concept that transfected clones expressing SV40 tumor antigens represent immortalized BND cell lines. Epidermal cell lines derived from Tursiops truncatus will provide a unique tool for studying key features of the interaction occurring between dolphins and the environment in which they live at their most crucial interface: the skin. (c) 2005 Wiley-Liss, Inc.