Research Project: Metabolism and Health Effects of Phospholipid and Triglyceride Forms of Omega-3 Polyunsaturated Fatty Acids

Location: Immunity and Disease Prevention Research

2019 Annual Report

Objectives
Objective 1: Determine the differential metabolism and uptake of n-3 fatty acids from dietary phospholipid and triglyceride forms in different tissues. -Sub-objective 1A: Determine the differential uptake and accretion of PL and TG forms of DHA in different mouse tissues. - Sub-objective 1B: Compare the metabolism of PL and TG forms of DHA in different mouse tissues. - Sub-objective 1C: Determine whether the differential accretion of DHA in brain from the PL and TG forms will result in differences in mouse behavior. Objective 2: Compare the anti-inflammatory and immune-modulating ability of n-3 fatty acids from dietary phospholipid and triglyceride forms in different tissues. -Sub-objective 2A: Compare the anti-inflammatory and immune-modulating ability of PL and TG forms of DHA by using experimental allergic encephalopathy (EAE)as a mouse model of brain inflammation. - Sub-objective 2B: Compare the anti-inflammatory and immune-modulating ability of PL and TG forms of DHA in the prevention of diet-induced NAFLD in mice.

Approach
Proposed experiments will compare accretion, metabolism and health effects n-3 fatty acids from the phospholipid (PL) and the triglyceride (TG) forms in different mouse tissues. For objective 1, we will determine the dose dependent accretion and metabolism of the PL-DHA and TG-DHA in the mouse tissues and their effects on the behavioral responses. We hypothesize that tissue accretion of DHA will increase with its increased intake, and at a given dose it will be greater in the PL group than in the TG group for tissues which express mfsd2a transporter (brain and liver) and will be similar in tissues (heart, muscle, and adipose tissue) that do not express this transporter. We also hypothesize that concentration of DHA in different lipid classes and PL subclasses, and that of DHA metabolites will be a function of tissue DHA concentration, and that DHA will improve learning and memory, and decrease fear and anxiety induced stress; effects of PL-DHA will be greater than that of TG-DHA on the behavioral responses tested. For objective 2, we will compare anti-inflammatory and immune-modulating effects of single equivalent dose (based on the results of objective 1) of the PL- and TG-DHA on brain and liver. For studies with brain we will use the mouse model of experimental allergic encephalomyelitis (EAE), and for liver we will use the high fat and high sucrose diets fed mouse model of nonalcoholic fatty liver disease (NAFLD). We hypothesize that both forms of DHA will delay the onset and severity of EAE and NAFLD when compared with the control groups, and PL-DHA will be more effective than TG-DHA. We anticipate our results will support the proposed hypotheses, however, if DHA does not have any effect on the response variables tested, we may repeat some experiments by increasing the duration of feeding. Whether or not the two forms of DHA differ in their effects on the responses tested will be equally useful information. A variety of behavioral, biochemical, molecular, immunohistochemistry, flowcytometery, and other analytical methods will be used. The mouse models and the methods proposed here have been previously used in our or our collaborator’s laboratories.

Progress Report
This is the final report for project 2032-51530-024-00D, which expired in March 2019, and was replaced by project 2032-51530-026-00D, "Impact of Diet on Intestinal Microbiota, Gut Health and Immune Function." During the performance period of this project, research was conducted under Objective 1 to determine if there was differential metabolism and uptake of dietary docosahexaenoic acid (DHA), an n-3 polyunsaturated fatty acid (PUFA) typically of marine origin, from two different forms found in the diet. These experiments were performed in mice. This is of interest because diets rich in DHA are associated with health benefits, including decreased inflammation and decreased plasma triglyceride levels. Dietary DHA is either found as a component of triglyceride (TG), as in most seafood, or found as a component of phospholipids (PL), as in some supplements (including Krill oil). The goal of this objective was to determine if purified DHA-TG and DHA-PL were absorbed or metabolized differently and whether there are differences in cognitive functioning. Researchers found that accretion of dietary DHA in mouse tissues including liver, adipose tissue, heart, eye and brain, did not differ between its purified PL and TG forms. This finding does not support the claim that DHA-PL is more bioavailable than DHA-TG and this finding was reported in a peer-reviewed publication. Because DHA is abundant in the brain, the researchers examined effects of the two forms of DHA on cognitive function, as intake of n-3 PUFA in humans may prevent cognitive decline in the elderly. The researchers found that mice fed DHA-PL showed enhanced emotion-based learning compared to mice fed DHA-TG and results were published in a peer-reviewed journal. Researchers also performed a second set of experiments in mice to determine whether any differential accumulation of DHA-PL and DHA-TG in the brain affected progression of a model inflammatory neurodegenerative disease, experimental autoimmune encephalomyelitis (EAE). EAE is a model of the human disease multiple sclerosis (MS). MS is a neurologic autoimmune disease, which is the leading cause of non-traumatic neurologic disability in young adults in the U.S. and Europe. n-3 PUFA are reported to mitigate severity of this disease. This research was successfully completed, and the results were reported in a peer-reviewed publication in the current year of the project. Briefly, researchers found that DHA of either dietary form reduced the severity of EAE compared to a diet low in n-3 PUFA. This work demonstrates that DHA, a specific n-3 PUFA, has health benefits in this mouse model and thus supports observational data from humans suggesting that n-3 PUFA may be of benefit in those at risk of neurologic inflammatory diseases, such as MS.

Accomplishments
1. DHA ameliorates onset and severity of experimental autoimmune encephalomyelitis in mice. Multiple sclerosis (MS) is a neurologic autoimmune disease, which is the leading cause of non-traumatic neurologic disability in young adults in the U.S. and Europe. Consumption of n-3 polyunsaturated fatty acids (PUFA) are reported to mitigate severity of this disease. ARS researchers in Davis, California, conducted an experiment using a mouse model of MS called experimental autoimmune encephalomyelitis, demonstrating that intake of the specific n-3 PUFA, docosahexaenoic acid (DHA), decreased the severity of this disease. This work provides support for the apparent preventive effects of n-3 PUFA intake for similar neurodegenerative diseases in humans.

Review Publications
Laugero, K.D., Adkins, Y.C., Kelley, D.S., Mackey, B.E. 2017. Emotion-Based cognition in mice is differentially influenced by dose and chemical form of dietary docosahexaenoic acid. Nutrients. 9(9):993. https://doi.org/10.3390/nu9090993.
Adkins, Y.C., Soulika, A.M., Mackey, B.E., Kelley, D.S. 2019. Docosahexaenoic acid (22:6n-3) ameliorated the onset and severity of experimental autoimmune encephalomyelitis in mice. Lipids. 54(1):13-23. https://doi.org/10.1002/lipd.12130.
Adkins, Y.C., Laugero, K.D., Mackey, B.E., Kelley, D.S. 2019. Accretion of dietary docosahexaenoic acid in mouse tissues did not differ between its purified phospholipid and triacylglycerol forms. Lipids. 54(1):25-37. https://doi.org/10.1002/lipd.12115.