Harbour porpoises (Phocoena phocoena) and harbour seals (Phoca vitulina) can display high levels of zinc (Zn), mercury (Hg), polychlorinated biphenyls (PCBs) and methyl sulfonyl metabolites (MeSO2-PCBs) in their tissues. Little is known about potential imp act of these pollutants on marine mammal immune system. The proposed study aims to better define the immunotoxicological risk linked to an exposure of the harbour porpoise and the harbour seal to methyl-Hg, Zn, PCBs and MeSO2-PCBs. Harbour seal and harbour porpoise peripheral mononuclear blood cells (PMBC) will be isolated (task 1) and contaminated with methyl-Hg, Zn and PCBs (task 2). The expression of cytokines (task 3) and the study of the proteome (task 4) will be studied in control and contaminated PMB C.- Task 1. PMBC will be isolated from the blood of captive and wild harbour porpoises and harbour seals and kept on cell culture medium. Analysis of Hg, Zn, PCBs and MeSO2-PCBs will be realized in the blood and serum of wild harbour porpoises and harbour seals from the North Sea to evaluate their contamination level.- Task 2. PMBC will be contaminated in vitro with methyl-Hg, ZnCl2 and PCB solutions. The tested concentrations will be in the range of concentrations naturally occurring in the blood of wild h arbour porpoises and harbour seals. - Task 3. The expression of pro- inflammatory (IL-1ß, IL-2, Il-6 and TNF-a) and anti-inflammatory (IL-10 and TGF- ß) cytokines will be investigated using RT-PCR and real-time RT-PCR. Cytokines are glycoproteins secreted by the immune cells and their expression is one important tool for the characterisation of the cellular immune response. - Task 4. In complement to cytokine mRNA expression, we wish also to understand pollutant effects on a ¿protein¿ level. Proteomic will allow the validation of biomarkers such as cytokines but also the development of new biomarkers to evaluate the impact of contaminants on the immune system.
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