title: Establishment of the first humpback whale fibroblast cell lines and their application in chemical risk assessment.
abstract: This paper reports the first successful derivation and characterization of humpback whale fibroblast cell lines. Primary fibroblasts were isolated from the dermal connective tissue of skin biopsies, cultured at 37 °C and 5% CO2 in the standard mammalian medium DMEM/F12 supplemented with 10% fetal bovine serum (FBS). Of nine initial biopsies, two cell lines were established from two different animals and designated HuWa1 and HuWa2. The cells have a stable karyotype with 2n=44, which has commonly been observed in other baleen whale species. Cells were verified as being fibroblasts based on their spindle-shaped morphology, adherence to plastic and positive immunoreaction to vimentin. Population doubling time was determined to be ∼41 h and cells were successfully cryopreserved and thawed. To date, HuWa1 cells have been propagated 30 times. Cells proliferate at the tested temperatures, 30, 33.5 and 37 °C, but show the highest rate of proliferation at 37 °C. Short-term exposure to para,para'-dichlorodiphenyldichloroethylene (p,p'-DDE), a priority compound accumulating in southern hemisphere humpback whales, resulted in a concentration-dependent loss of cell viability. The effective concentration which caused a 50% reduction in HuWa1 cell viability (EC50 value) was approximately six times greater than the EC50 value for the same chemical measured with human dermal fibroblasts. HuWa1 exposed to a natural, p,p'-DDE-containing, chemical mixture extracted from whale blubber showed distinctively higher sensitivity than to p,p'-DDE alone. Thus, we provide the first cytotoxicological data for humpback whales and with establishment of the HuWa cell lines, a unique in vitro model for the study of the whales' sensitivity and cellular response to chemicals and other environmental stressors.
authors.affiliation: Griffith University, Environmental Futures Research Institute, Southern Ocean Persistent Organic Pollutants Program, Brisbane, QLD, Australia; Eawag, Swiss Federal Institute of Technology, Dübendorf, Switzerland. Electronic address: Michael.burkard@eawag.ch.
authors.forename: Michael
authors.initials: M
authors.lastname: Burkard
authors.affiliation: The University of Queensland, School of Veterinary Science, Gatton, QLD, Australia.
authors.forename: Deanne
authors.initials: D
authors.lastname: Whitworth
authors.affiliation: Eawag, Swiss Federal Institute of Technology, Dübendorf, Switzerland; ETH Zürich, Institute of Biogechemistry and Pollutant Dynamics, Zürich, Switzerland; EPF Lausanne, School of Architecture, Civil and Environmental Engineering, Lausanne, Switzerland.
authors.forename: Kristin
authors.initials: K
authors.lastname: Schirmer
authors.affiliation: Griffith University, Environmental Futures Research Institute, Southern Ocean Persistent Organic Pollutants Program, Brisbane, QLD, Australia.
authors.forename: Susan Bengtson
authors.initials: SB
authors.lastname: Nash
mesh_terms: D000273:Adipose Tissue / Q000737:chemistry; D000818:Animals; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D003633:Dichlorodiphenyl Dichloroethylene / Q000378:metabolism / Q000633:toxicity; D005347:Fibroblasts / Q000166:cytology; D050640:Humpback Whale / Q000502:physiology*; D020128:Inhibitory Concentration 50; D018570:Risk Assessment; D014874:Water Pollutants, Chemical / Q000378:metabolism / Q000633:toxicity
chemical_list: D014874:Water Pollutants, Chemical; D003633:Dichlorodiphenyl Dichloroethylene
keywords: Antarctica; Cell line characterization; Karyotype; Megaptera novaeangliae; Persistent Organic Pollutants (POPs); p,p′,-DDE