Visualizing digestive organ morphology and function using differential fatty acid metabolism in live zebrafish

Abstract

Lipids are essential for cellular function as sources of fuel, critical signaling molecules and membrane components. Deficiencies in lipid processing and transport underlie many metabolic diseases. To better understand metabolic function as it relates to disease etiology, a whole animal approach is advantageous, one in which multiple organs and cell types can be assessed simultaneously in vivo. Towards this end, we have developed an assay to visualize fatty acid (FA) metabolism in larval zebrafish (Danio rerio). The method utilizes egg yolk liposomes to deliver different chain length FA analogs (BODIPY-FL) to six day-old larvae. Following liposome incubation, larvae accumulate the analogs throughout their digestive organs, providing a comprehensive readout of organ structure and physiology. Using this assay we have observed that different chain length FAs are differentially transported and metabolized by the larval digestive system. We show that this assay can also reveal structural and metabolic defects in digestive mutants. Because this labeling technique can be used to investigate digestive organ morphology and function, we foresee its application in diverse studies of organ development and physiology.

Cannabinoid receptor signaling regulates liver development and metabolism

“…To confirm these abnormalities and visualize hepatic lipid metabolism in vivo, BODIPY C5 fluorophore was administered, which labels all organs in which metabolites accumulate (Carten et al, 2011): control livers exhibit highly branched bile ducts at 6 days post fertilization (dpf ); by contrast, both cnr1 −/− and cnr2…”

Lxr-driven enterocyte lipid droplet formation delays transport of ingested lipids

“…After extraction, lipids were resuspended in 200 µl of chloroform, and a 10 µl aliquot was run on a TLC plate (TLC silica gel 60, Merck Millipore). The standards for triacylglycerol, diacylglycerol, and phosphatidylcholine were synthesized as previously described ( 8 ). A twosolvent system was used to separate phospholipids from nonpolar lipids: the fi rst solvent mixture was run until the middle of the plate (chloroform-methanol-acetic acid-water, 50:37.5:3.5:2, v/v), and the second solvent mixture was run to 2 cm from the end of the plate (hexane-diethyl ether-acetic acid, 70:30:1, v/v).…”

“…A 5% w/v dried egg yolk (Sigma) suspension was prepared in embryo water (5 mM NaCl, 0.17 mM KCl, 0.33 mM CaCl 2 , 0.33 mM MgSO 4 , pH 7.4) as previously described ( 8,21 ). Ten 6 dpf larvae were distributed per well in 6-well plates (Nest Biotech Co. Ltd.…”

“…Human jejunal biopsies ( 6 ), prolonged, label-free lipid droplet imaging of live loops of mouse intestine ( 7 ), and fl uorescent lipid probe-based visualization of intestinal lipid droplets in zebrafi sh larvae ( 8 ) have revealed that a substantial fraction of absorbed lipids remain in the enterocyte for many hours following a meal. These fi ndings suggest that there is an intrinsic mechanism for delaying, in part, transit of absorbed lipids.…”

Live imaging epidermal cell metabolism during inflammation: a new immunometabolic role for epidermal keratinocytes?

“…Live imaging these cells using neutrophil-and macrophage-marking transgenic zebrafish lines has provided a window to explore their function within an intact transparent animal model [34][35][36][37][38][39][40][41] . In addition, this transparency has been exploited to directly visualize the metabolic fates of fluorescent FA analogs [14,[42][43][44] . Using the zebrafish system to explore the function of immunoresponsive gene 1-like (Irg1l), a paralog of Irg1 (a mitochondrial enzyme that we previously showed connected metabolism and immune cell function within macrophages [14] ), we uncovered a new mechanism where Irg1l-dependent -oxidation-fueled mROS production within epidermal cells helps drive mmp9 expression and leukocyte migration during inflammation ( Figure 1B) [31] .…”