n-3 Fatty acids decrease arterial low-density lipoprotein cholesterol delivery and lipoprotein lipase levels in insulin-resistant mice

Abstract

Objective: To determine whether n-3 fatty acids (n-3) influence arterial cholesterol delivery and lipoprotein lipase (LpL) levels in insulin-resistant mice.

Methods and results: Insulin resistance contributes to risk of cardiovascular disease. It was previously reported that saturated fat (SAT) diets increased, but n-3 diets decreased, arterial low-density lipoprotein (LDL) cholesterol deposition from LDL total and selective uptake; this was associated with increased or decreased arterial LpL, respectively. Insulin receptor transgenic knockout mice (L1) were fed a chow, SAT, or n-3 diet for 12 weeks. Double-fluorescent boron dipyrromethene (BODIPY)-cholesteryl ester (CE) and Alexa dye-labeled human LDL were injected to separately trace LDL-CE and LDL-apolipoprotein B whole particle uptake. In contrast to SAT, n-3 diets markedly reduced all plasma lipids, ameliorating progression of insulin resistance. As opposed to SAT, n-3 reduced arterial LDL uptake, CE deposition, and selective uptake. Disparate patterns of CE deposition between diets were comparable with arterial LpL distribution; SAT induced high LpL levels throughout aortic media; LpL was limited only to intima in n-3-fed mice.

Conclusions: n-3 diets diminish arterial LDL-cholesterol deposition in mice with insulin resistance, and this is associated with changes in arterial LpL levels and distribution.

Figure 1

Effects of diet on mouse body weight in L1 mice fed a chow (■), SAT (▲), or n-3 (●) diet for 12 weeks. Each point represents the mean±SD of 10 mice. * and # denote significant differences between chow and SAT and between chow and n-3, respectively. P<0.001.

Figure 2

Plasma lipid profiles of L1 mice fed a chow (■), SAT (▲), or n-3 (●) diet. A through C, Blood samples were determined weekly for FFA (A), TG (B), and T-Chol (C) levels, as described in the Methods section. The results are expressed as the mean±SD (n>8). *P<0.05, chow vs SAT or n-3; #P<0.05, n-3 vs SAT.

Figure 3

Plasma lipoprotein profiles in L1 mice fed a chow, SAT, or n-3 diet. A and B, TG (A) and Chol (B) levels were determined for each fraction after separating by fast-performance liquid chromatography. Elution fractions 2 to 10, 12 to 22, and 25 to 35 represent elution zones for very LDL, LDL, and high-density lipoprotein, respectively.

Figure 4

A, Comparison of arterial LDL-CE (BODIPY) and LDL-apoB (Alexa) uptakes in L1 mice and wild-type (WT) littermates. Merged 2-channel images (right) were used to assess colocalization of dye (Alexa) and BODIPY, with yellow fluorescence indicating colocalization (magnification ×20). B, Ratios of arterial uptake of total LDL-CE to whole LDL particle uptake determined for L1 mice and WT littermates, as described in the Methods section. Data are given as the mean±SE. *P<0.05.

Figure 5

A, Effects of diet on fluorescent LDL-CE (BODIPY) and apoB (Alexa) uptakes in aortas. B, Ratios of arterial uptake of total LDL-CE to whole LDL particle uptake determined for each diet group. Data are given as the mean±SE. Ratios were normalized to the same unit area for each diet. A ratio >1 indicates SU (dotted line). *P<0.05, chow vs SAT or n-3; #P<0.05, SAT vs n-3.

Figure 6

Effects of insulin resistance and diet on arterial LpL levels. A, Immunohistochemical staining of arterial LpL. Reddish brown staining shows the presence of LpL. B, Representative images of a Western blot for LpL from pooled mouse artery homogenates (n=3). C, Immunofluorescence of arterial LpL from mice fed a chow, SAT, or n-3 diet. Magnifications are ×20 (A) and ×40 (C). WT indicates wild type.

Figure 7

Arterial macrophages and LpL in the aorta. A, Representative images of chow-, SAT-, or n-3–fed L1 mouse aortas stained for ECs (blue), LpL (green), and macrophages (red) (magnification ×40). B, Immunofluorescence of macrophages measured for each group (n>3). C, Percentage of macrophages colocalizing with LpL determined for each group (n>3), as described in the Methods section. *P<0.05, SAT vs chow; #P<0.05, SAT vs n-3.