Fish oil-based injectable lipid emulsions containing medium-chain triglycerides or added α-tocopherol offer anti-inflammatory benefits in a murine model of parenteral nutrition-induced liver injury

Abstract

Background: Fish oil (FO) intravenous lipid emulsions (ILEs) are used as a monotherapy to treat parenteral nutrition (PN)-associated liver disease and provide essential fatty acids (EFAs) needed to sustain growth and prevent EFA deficiency (EFAD). Studies have suggested that medium-chain triglycerides (MCTs) and α-tocopherol have anti-inflammatory properties.

Objective: The purpose of this study was to test whether FO-ILEs containing MCTs and/or additional α-tocopherol decrease the inflammatory response to an endotoxin challenge compared with FO-ILE alone and preserve the ability to prevent PN-induced liver injury in mice.

Methods: A murine model of PN-induced hepatosteatosis was used to compare the effects of ILEs formulated in the laboratory containing varying ratios of FO and MCTs, and subsequently FO- and 50:50 FO:MCT-ILE plus 500 mg/L α-tocopherol (FO + AT and 50:50 + AT, respectively). C57BL/6 mice receiving unpurified diet (UPD), PN-equivalent diet (PN) + saline, and PN + soybean oil (SO)-ILE served as controls. After 19 d, mice received an intraperitoneal saline or endotoxin challenge 4 h before being killed. Serum and livers were harvested for histologic analysis, fatty acid profiling, and measurement of systemic inflammatory markers (tumor necrosis factor-α, interleukin-6).

Results: All ILEs were well tolerated and prevented biochemical EFAD. Livers of mice that received saline and SO developed steatosis. Mice that received 30:70 FO:MCT developed mild hepatosteatosis. All other FO-containing ILEs preserved normal hepatic architecture. Mice that received FO- or SO-ILE had significantly elevated systemic inflammatory markers after endotoxin challenge compared with UPD-fed controls, whereas 50:50 FO:MCT, 30:70 FO:MCT, FO + AT, and 50:50 + AT groups had significantly lower inflammatory markers similar to those seen in UPD-fed controls.

Conclusions: Mixed FO/MCT and the addition of α-tocopherol to FO improved the inflammatory response to endotoxin challenge compared with FO-ILE alone while still preventing PN-induced liver injury and EFAD in mice. There was no synergistic relation between α-tocopherol and MCTs.

Keywords: fish oil; hepatosteatosis; injectable lipid emulsions; intestinal failure–associated liver disease; intravenous fat emulsions; medium-chain triglycerides; parenteral nutrition; parenteral nutrition–associated liver disease; α-tocopherol.

FIGURE 1

Representative H&E and Oil Red O images of livers from Experiment 1 at 40× magnification. FO, fish oil; H&E, hematoxylin and eosin; MCT, medium-chain triglyceride; PN, parenteral nutrition; S, saline; SO, soybean oil; UPD, unpurified diet.

FIGURE 2

Serum T:T ratios (A), linoleic acid (B), arachidonic acid (C), α-linolenic acid (D), EPA (E), and DHA (F) of mice from Experiment 1, which tested intravenous lipid emulsions with varying FO:MCT ratios. Fatty acids are expressed as percentage of total fatty acids. Essential fatty acid deficiency is defined as a T:T ratio >0.2. Values are mean ± SEM, n = 3/group. P values are from 1-factor ANOVA adjusted for multiple comparisons by Tukey–Kramer adjustment. FO, fish oil; MCT, medium-chain triglyceride; PN, parenteral nutrition; S, saline; SO, soybean oil; T:T, triene:tetraene; UPD, unpurified diet.

FIGURE 3

Serum IL-6 and TNF-α after saline injection or endotoxin challenge of mice from Experiment 1, which tested intravenous lipid emulsions with varying FO:MCT ratios. (A, C) Serum IL-6 after saline injection and endotoxin challenge, respectively; (B, D) TNF-α after saline injection and endotoxin challenge, respectively. Values are means ± SEMs, n = 5/group. P values are from 1-factor ANOVA adjusted for multiple comparisons by Tukey–Kramer adjustment. FO, fish oil; MCT, medium-chain triglyceride; PN, parenteral nutrition; S, saline; SO, soybean oil; UPD, unpurified diet.

FIGURE 4

Representative H&E and Oil Red O images of livers from Experiment 2 at 40× magnification. 50:50, 50% FO:50% MCT; AT, α-tocopherol; FO, fish oil; H&E, hematoxylin and eosin; MCT, medium-chain triglyceride; PN, parenteral nutrition; S, saline; SO, soybean oil; UPD, unpurified diet.

FIGURE 5

Serum T:T ratios (A), linoleic acid (B), arachidonic acid (C), α-linolenic acid (D), EPA (E), and DHA (F) of mice from Experiment 2, which tested intravenous lipid emulsions with normal or additional α-tocopherol. Fatty acids are expressed as percentages of total fatty acids. Essential fatty acid deficiency is defined as a T:T ratio >0.2. Values are means ± SEMs, n = 3/group. P values are from 1-factor ANOVA adjusted for multiple comparisons by Tukey–Kramer adjustment. 50:50, 50% FO:50% MCT; AT, α-tocopherol; FO, fish oil; MCT, medium-chain triglyceride; PN, parenteral nutrition; S, saline; SO, soybean oil; T:T, triene:tetraene; UPD, unpurified diet.

FIGURE 6

Liver T:T ratios (A), linoleic acid (B), arachidonic acid (C), α-linoleic acid (D), EPA (E), and DHA (F) of mice from Experiment 2, which tested intravenous lipid emulsions with normal or additional α-tocopherol. Fatty acids are expressed as percentages of total fatty acids. Essential fatty acid deficiency is defined as a T:T ratio > 0.2. Values are means ± SEMs, n = 3/group. P values are from 1-factor ANOVA adjusted for multiple comparisons by Tukey–Kramer adjustment. 50:50, 50% FO:50% MCT; AT, α-tocopherol; FO, fish oil; MCT, medium-chain triglyceride; PN, parenteral nutrition; S, saline; SO, soybean oil; T:T, triene:tetraene; UPD, unpurified diet.

FIGURE 7

Serum IL-6 (A) and TNF-α (B) after endotoxin challenge of mice from Experiment 2, which tested intravenous lipid emulsions with normal or additional α-tocopherol. Values are means ± SEMs, n = 10/group. P values are from 1-factor ANOVA adjusted for multiple comparisons by Tukey–Kramer adjustment. 50:50, 50% FO:50% MCT; AT, α-tocopherol; FO, fish oil; MCT, medium-chain triglyceride; PN, parenteral nutrition; S, saline; SO, soybean oil; UPD, unpurified diet.