Characterization of Atlantic salmon head kidney leukocyte culture

Smith, Nicole C. (2021) Characterization of Atlantic salmon head kidney leukocyte culture. Doctoral (PhD) thesis, Memorial University of Newfoundland.

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Abstract

The Atlantic salmon (Salmo salar) is an economically important farmed and wild fish in several countries including Canada. Macrophages are white blood cells of the immune system of fish and other vertebrates, that are essential in fighting infection and disease. Elucidating how macrophages differentiate and function is necessary to fully understand how the fish immune system functions and to enable the development of methods to maintain healthy fish. Therefore, the objective of my Ph.D. thesis was to characterize the Atlantic salmon adherent head kidney leukocyte (HKL) culture, a macrophage-like model commonly used in fish immunological studies, using various genomic and complementary techniques. Using morphology (Giemsa stain) and functional (phagocytosis) assays, the results of this thesis showed that the Atlantic salmon adherent HKL population changes during culture time. At Day 1 of culture, the results suggest that adherent HKLs are a heterogeneous population of predominantly “monocyte-like”, cells but by Day 5 of culture, the cells become more homogenous selectively enriched with macrophages. RNA-sequencing identified a change in the microRNA (miRNA) profile of Day 1 and Day 5 adherent HKLs, as well as the extracellular vesciles (EVs) released from them. Many of the identified miRNAs are involved in macrophage function and/or differentiation in other species. Furthermore, using a 44K microarray, changes in the mRNA transciprtome were profiled. Macrophage-related transcripts, lipid-related transcripts, immune-related transcripts and transcription factors were identified as differentially expressed between the two cell populations. In addition, GO term enrichment and network analysis identified immune-related and immune-cell differentiation related terms. The results of this thesis provides evidence that the Atlantic salmon HKL culture changes to become predominantly “macrophage-like” by Day 5 of culture and this is something that should be kept in mind when using HKLs for in vitro fish immunology studies. This research provides novel insight into the genes, miRNAs and molecular pathways involved in the differentiation of Atlantic salmon adherent HKLs from monocyte-like cells to macrophage-like cells.

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