Lipid body function as non-classical calcium stores in immunocytes

M.S. University of Hawaii at Manoa 2014.Includes bibliographical references.Lipid bodies, found in most eukaryotic cells, are intracellular lipid storage organelles. The role of lipid bodies has been most studied in adipocytes and hepatocytes due to their role in energy storage. Comparatively, little is known of the role they play in immunocytes. Both in adipocytes/hepatocytes and in cells of the immune system, lipid body numbers are dynamically regulated and can accumulate to pathophysiological levels (steatosis). In both locales this steatotic state can be induced by nutrient overload and metabolic stress, and in the latter also under certain conditions of infection. The over-arching goals of this project are (1) to study the composition (lipid and protein) and functional contributions made by lipid bodies in immunocytes, and (2) to assess the impact of altered lipid body numbers upon cellular function. The work presented in this thesis describes three areas of progress towards these goals. First, we developed a microaspiration method for isolating highly purified lipid bodies, allowing for their ex vitro manipulation and study of lipid/protein content using microscopy. This technique was validated using fluorescence microscopy of the neutral lipid dye Oil Red O in microaspirated lipid bodies. Second, we assessed the impact of the accumulation of lipid bodies (steatosis) on transcytoplasmic calcium signaling, a major activation pathway in the model immune cell system studied here. Third, we tested a new hypothesis arising from our work on transcytoplasmic calcium signaling. The apparent ability of lipid bodies to act as long term loci for calcium accumulation, coupled with recent studies showing that lipid bodies may contain mitochondria and endoplasmic reticulum, led us to hypothesize their potential role as bona fide calcium stores. Our data revealed that the lipid body population within mast cells is not homogenous. However, some LB exhibit the ability to sequester calcium and to release it in the manner of a bona fide calcium store. These observations represent a potentially novel role for lipid bodies and indicate the possibility of their contribution to calcium dynamics in immune cells under both physiological and pathophysiological conditions