Cloning and Sequence Analysis of Killer Whale (Orcinus orca) Interleukin-10 (IL-10)
IAAAM Archive
Jean Marie Herrnian; Don P. King; David A. Ferrick
U.C. Davis, School of Veterinary Medicine, Department of Pathology, Microbiology and Immunology, Davis, CA

Abstract

Cytokines are regulatory proteins secreted by activated cells of the immune system and a variety of other cells in the body. The pleiotropic effects of cytokines include numerous effects on cells of the immune system and modulation of inflammatory responses. Cytokines act in regulatory networks and a single cytokine may act as both a negative signal and a positive signal depending on the target cell type.

IL 10 in the mouse and human is produced by T cell subsets, B lymphomas, activated mast cells, epithelial cells, activated macrophages and keratinocytes. IL10's ability to block activation of cytokine synthesis and several accessory functions of macrophages enables it to act as a potent suppressor of macrophages T cell subset and NK cell activity, consequently driving the immune response toward humoral rather than cell mediated immunity. IL 10 inhibits the production of the inflammatory cytokines IL 1, TNF alpha, and IL 6 by activated macrophages. These cytokines, when excessively elevated in serum of humans and mice have been shown to correlate with septicemia and septic shock accompanied by high mortality rates. Mice rendered deficient in the expression of the IL 10 gene are profoundly more sensitive to endotoxin induced shock, develop chronic intestinal inflammation and are unable to limit immunopathologic damage upon the application of cutaneous irritants. Since both the gastrointestinal tract and skin are sites of continuous exposure to potential antigens, IL 10 has proven to play a prominent role regulating cellular immune hyperresponsiveness.

The study of health issues in captive and free ranging cetaceans has suggested that reduced immune competence as well as opportunistic and chronic infectious diseases to be problematic. Understanding the basic function of the cetacean immune system is central to the study of cetacean health and illness, and the major obstacles for evaluating immune functions in cetaceans are due to the lack of availability of cetacean specific reagents. Our previous work demonstrated the ability to utilize the recombinant human cytokines IL 2 and IL 10 to assess the roles of these cytokines in the cetacean immune system. Both were shown to function in a similar fashion and at comparable levels in killer whale peripheral blood mononuclear cells (PBMC) and human control PBMC.

While the cytokines themselves tend to remain functional through evolution by retaining the basic structure and physical properties of their receptor binding regions, the antigenic regions to which antibodies bind are not usually well conserved. Therefore, most antibodies to the cytokines are very species specific, and a species specific antibody would then be necessary to evaluate of the levels of a cytokine in biological samples. The cetacean cytokine proteins and reagents made to them will be useful to understand the involvement of cytokines in the regulation of cetacean immune responsiveness.

In order to produce reagents to the endogenous killer whale IL 10 protein and to have a cetacean specific reagent to better characterize the function of killer whale IL 10 in vivo it was necessary to first clone the killer whale IL 10 molecule. This was done by reverse transcription of mRNA isolated from mitogen stimulated killer whale PBMC into cDNA. The cDNA was then cloned by PCR using degenerate primers designed from conserved areas of known IL 10 sequences. The kw IL 10 sequence will be presented and analyzed for its evolutionary relatedness to other known IL 10 sequences. Pertinent biology of IL 10 and its potential use as a diagnostic and therapeutic agent in cetacean medicine will be discussed.

Speaker Information
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Jean Marie Herrman


MAIN : Immunology : Killer Whale IL-10
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