Establishment of epidermal cell lines derived from the skin of the Atlantic bottlenose dolphin (Tursiops truncatus)

Abstract

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.

Establishment of the first humpback whale fibroblast cell lines and their application in chemical risk assessment

“…3B, inset), no humpback whale-specific positive control was available. However, cytokeratin has shown to be positively expressed in other cetacean species (Pine et al, 2004, Yu et al, 2005. These results therefore lend strong support that spindle-shaped (Fig.…”

Establishment and characterization of fibroblast cell lines from the skin of the Yangtze finless porpoise

“…The dark color of some cells was due to the considerable amount of melanin pigment in the porpoise skin (Fig. 1B) (Yu et al 2005). In the progress of cell passages, the melanin density dropped off and the cytoplasm become gradually clearer.…”

“…Currently, only a few cetacean cell culture systems have been described, such as those derived from the beluga whale (Delphinapterus leucas) and Atlantic bottlenose dolphin (Tursiops truncatus) (Carvan et al 1994;Pine et al 2004;Yu et al 2005). In vitro somatic cell cultures have been applied in toxicological (Carvan et al 1995;Pine et al 2007;Wise et al 2008), epidemiological (Kadoi et al 1992;Buck et al 1993), and cytogenetic studies (Walen and Madin 1965;Duffield et al 1991;Heinzelmann et al 2008).…”

“…In vitro somatic cell cultures have been applied in toxicological (Carvan et al 1995;Pine et al 2007;Wise et al 2008), epidemiological (Kadoi et al 1992;Buck et al 1993), and cytogenetic studies (Walen and Madin 1965;Duffield et al 1991;Heinzelmann et al 2008). With the exception of successfully cultured lines, such as DKN1 (Pine et al 2004) and DS1 (Yu et al 2005), continuous cell strains have not yet been obtained, which may restrict the use in long-term studies. To our knowledge, no reports describing a successful cell line from freshwater cetaceans have been published.…”

The vitamin D3 transcriptomic response in skin cells derived from the Atlantic bottlenose dolphin

“…Dolphin health assessments have become an indicator of detrimental changes occurring within the animal’s coastal habitat [4,5]. As a protected species, there is a need for more in vitro and molecular approaches to study the dolphin; consequently, we established cell strains and SV40-immortalized cell lines from the skin of the Atlantic bottlenose dolphin [6]. We are using these cellular models to investigate the vitamin D3 pathway and associated molecular mechanisms as biochemical platforms to correlate environmental stress with dolphin health.…”