Development of an ex vivo adipose tissue model to study the impact of multiple stressors in arctic marine mammals

Details

Supervisors

Debier, Cathy ; Spinel, Alexandra

Faculty

Faculté des bioingénieurs

Degree label

Master [120] : bioingénieur en sciences agronomiques, à finalité spécialisée

Abstract

Arctic marine mammals are exposed to a wide range of multiple stressors that could result in the observed decline of their populations. In particular, their subcutaneous adipose tissue, blubber, was shown to be an appropriate target for toxicological studies because of its physiological interaction with stressors, its involvement in multiple metabolic pathways and its ease of sampling on free-ranging individuals. Most toxicological studies on marine mammals are based on correlative approaches that do not allow to establish cause-and-effect relationships. To address this problem, an ex vivo model of precision-cut adipose tissue slices (PCATS) has been developed in our research group to assess the impact of multiple stressors on adipose tissue functions. The goal of the present research was to develop this ex vivo model in arctic marine mammals. To do so, cetacean species (killer whale: Orcinus orca; humpback whale: Megaptera novaeangliae; pilot whale: Globicephala melas; sperm whale: Physeter macrocephalus) and polar bear (Ursus maritimus) were sampled in Northern Norway. Cetaceans were sampled remotely at sea using an air gun while polar bear was sampled directly on land. In parallel, pig was used as an easy-access species in Belgium, for which PCATS model was previously settled, to investigate the possible impact of sampling conditions specific to field work. The results of our experiments showed only partial success of the PCATS model and highlighted additional challenges on the newly tested arctic species. Indeed, functionality of the tissue, quantified by glycerol release upon lipolytic treatment, could only be confirmed in one humpback whale, three pilot whales and one polar bear. Cetacean PCATS showed low response to lipolytic stimulus, even when significant, after 12 and 24 h lipolysis induction by either 1 or 10 µM isoproterenol (ISO) or epinephrine (EPI). On the other hand, polar bear showed higher functionality with both ISO 10 µM and EPI 10 µM after 12 h lipolysis induction but not after 24 h. Experiments on pig showed that PCATS functionality remained good even after remote sampling and suggested that adipose tissue can be stored in the form of biopsy for up to seven days while remaining highly functional. Histological analysis showed that blubber of cetaceans is mainly composed of fibers and muscles with only a few adipocytes distributed across layers and is therefore not compatible with lipolysis experiment to assess functionality of the tissue. On the other hand, the blubber of polar bear is mainly composed of adipocytes and can therefore show better response to lipolytic stimulus. Finally, viability of the tissue was difficult to assess using lactate dehydrogenase assay due to the difficulty in lysing the tissue to obtain a positive control of cell death. Overall, functionality and viability assays should therefore be adapted for future investigations.