Developing methods for in vitro toxicity testing with Atlantic cod tissues

• Main Author: Skartsæterhagen, Maria Kristine
• Format: Master Thesis
• Language: English
• Published: 2019
• Subjects: atlantic cod; Gadus morhua
• Online Access: http://hdl.handle.net/10852/71993; http://urn.nb.no/URN:NBN:no-75120

Two in vitro methods have been applied and tentatively optimized for the use with Atlantic cod (Gadus morhua), in order to gain more knowledge about responses for this ecologically and economically important species. The Oslofjord was the chosen study area, where cod popoulations have been at an all-time low since 2002. In the inner parts of the fjord elevated concentrations in sediment and biota have been found for several contaminants, among them bisphenols. The outer Oslofjord is in comparison relatively unpolluted. Hepatocyte isolation was succesfully performed on wild caught cod from the outer Oslofjord with the two-step perfusion method. Wild cod hepatocytes were exposed to copper, single compound bisphenol type A, B, E, G, TMC or Z, or mixtures of bisphenols. Bisphenol exposure was not very cytotoxic at the here applied concentrations up to 100 uM, and EC50 values could not be obtained. Based on the applicable EC20 values, mixture 1 (containing bisphenol G, TMC, Z, B and E in ratios previously found in cod liver) was the most cytotoxic, followed by bisphenol G, TMC and Z. The other bisphenols did not appear to cause cytotoxicity at the concentrations tested. The alkaline comet assay was validated for use on four tissues from cod from the inner and outer Oslofjord. The comet scores displayed a typical non-normal, right-skewed distribution. Baseline responses showed higher DNA damage in whole blood and cells from the intestinal mucosa, compared to lymphocytes and kidney cells. Samples from the inner Oslofjord cod displayed less DNA damage increase when treated with hydrogen peroxide than those from the outer Oslofjord, indicating a possible adaption to repeated oxidative stress in the inner Oslofjord cod. Baseline responses from the two areas were similar, with the exception of kidney cells, where cod from the inner Oslofjord had a higher response. Biometric parameters such as length, weight, hepatosomatic index, Fulton's condition index or sex did not correlate with DNA damage in any of the tissues. Because cod is frequently kept in aquaculture, different methods for hepatocyte isolation was attempted with aquaculture cod. This was not successful. The likely explanation for this, is the fatty livers of these specimens, compared to the wild cod.