| Visualizing lipid metabolism in intact, functional organs remains an underdeveloped area of in vivo cellular imaging. Due to the technical hurdles faced when trying to image small and easily metabolized lipids, we lack a subcellular understanding of lipid dynamics in the normal and diseased metabolic states. The work presented in this dissertation describes a fluorescent feeding assay I have developed to improve our ability to visualize lipid metabolism in vivo and subsequent use of this technique to investigate the influence of two different environmental factors on intestinal and whole body lipid metabolism: intestinal microbes and dietary sugars.;The fluorescent feeding assay I have developed utilizes liposomes to orally deliver fluorescent fatty acids (BODIPY-FAs) to 6-days post fertilization (dpf) larvae. Following a short feed, metabolic function as well as digestive organ structure can be assessed, as the fluorescent FAs accumulate readily throughout numerous digestive organs and tissues. Different chain length FAs (short, medium and long) accumulate in distinct patterns throughout digestive organs, with each chain length suited to visualize particular larval organs and cellular structures. In addition, I have demonstrated the screening potential of this assay by feeding fatfree (ffr) and sox9b mutant larvae fluorescent liposomes. The canalicular and ductal defects in these larvae are nicely illuminated with this assay, illustrating the screening potential of this tool.;In collaboration with the Rawls laboratory I have also used the aforementioned feeding assay to explore the impact of gut microbes on intestinal FA uptake in larval zebrafish. Our work has found that the Firmicutes phylur is associated with enhanced FA absorption, as germ free larvae exhibit fewer lipid droplets in their intestinal enterocytes and hepatocytes, as well as less total body fat. Ongoing work aims to utilize the fluorescent feeding assay in.;conjunction with other methods to examine the effects of early sugar exposure on lipid metabolism during larval development. Such work may enable us to model early aspects of childhood diabetes and obesity, providing insight into the lasting impact of early sugar-exposure on cellular development and metabolic homeostasis. |
