Dietary unsaturated fat increases HDL metabolic pathways involving apoE favorable to reverse cholesterol transport

Abstract

Background: HDL that contains apolipoprotein E (apoE) is a subspecies especially active in steps in reverse cholesterol transport, a process that brings cholesterol from peripheral cells to the liver. Here, we studied the effect of dietary unsaturated fat compared with carbohydrate on the metabolism of HDL containing apoE.

Methods: We enrolled 9 adults who were overweight or obese and had below-average HDL-cholesterol in a crossover study of a high-fat diet, primarily unsaturated, and a low-fat, high-carbohydrate diet. A metabolic tracer study was performed after each diet period.

Results: Dietary fat increased the secretion, metabolism, and clearance of HDL subspecies containing apoE. Dietary fat increased the rate of clearance of large cholesterol-rich HDL containing apoE and increased their conversion to small HDL containing apoE, indicating selective cholesterol ester delivery to the liver. The high-unsaturated-fat diet did not affect the metabolism of HDL lacking apoE.

Conclusion: HDL containing apoE is a diet-responsive metabolic pathway that renders HDL more biologically active in reverse cholesterol transport. This may be a mechanism by which unsaturated fat protects against coronary heart disease. Protein-based HDL subspecies such as HDL containing apoE may be used to identify additional atheroprotective treatment targets not evident in the total HDL-cholesterol measurement.

Trial registration: ClinicalTrials.gov NCT01399632.

Funding: NIH and the National Center for Advancing Translational Science.

Keywords: Lipoproteins; Metabolism.

Conflict of interest statement

Conflict of interest: FMS was a consultant to Pfizer and MedImmune-Astra Zeneca on drug development and was an expert witness in cases involving Aegerion, Abbvie, and Pfizer. FMS and JDF are inventors on patents awarded to Harvard University pertaining to HDL.

Figures

Figure 1. Overview of dietary intervention and sampling protocol.

After screening, study participants were instructed to continue on their usual diet (self-selected diet) prior to the study period, upon which they were randomized to either receive a high-unsaturated-fat or low-fat diet for 4 weeks. After 4 weeks on either diet, subjects were admitted to the hospital on the morning of day 29 for the infusion protocol. They were instructed to eat the prescribed study breakfast (B) before coming to the hospital. At 10 am (time 0), each subject received a bolus infusion of D3-leucine. Samples were collected at 0, 0.5, 1, 1.5, 2, 3, 4, 6, 8, 10, 12, 14, 16, 18, and 22 hours after infusion while the study participants were in the hospital. Participants also ate lunch (L), dinner (D), and a snack (S) in the hospital. After the 22-hour sample was collected, participants were released from the hospital with their remaining study meals and were instructed to return to the hospital at 10 am for the next 3 days for the remaining blood draws (46, 70, and 94 hours). Before receiving the alternate diet and repeating the study protocol, they were instructed to return to their usual diet for a washout period of 3 weeks. Mono, monounsaturated fat; poly, polyunsaturated fat; sat, saturated fat; carb, carbohydrate; chol, cholesterol.

Figure 2. Effect of diet on fasting concentrations of plasma lipids and apolipoproteins.

Values (mg/dl) were measured at each study participant’s screening visit (lipids only) and following completion of each dietary intervention. HFD, high-fat diet; LFD, low-fat diet. Mean values (n = 9) shown in larger red circles and dotted red lines. P values were calculated using 2-tailed t test. (A) Triglycerides. (B) Total cholesterol. (C) LDL-cholesterol. (D) HDL-cholesterol. (E) ApoA-I. (F) ApoE. (G) ApoCIII. (H) ApoB.

Figure 3. Model fit for apoA-I tracer enrichments for each HDL subspecies across 2 diets.

(A) SAAM-II compartmental model (bare-minimum model; see ref. 12). (B) Modifications to the bare-minimum model. Blue arrows and delay compartment represent additional pathways for the HDL containing apoE (E+) subspecies only. (C) Model fit for the mean (n = 9) apoA-I tracer enrichment on 4 sizes of HDL, across each HDL subspecies and diet. The percentage D3-leucine (D3-Leu) tracer enrichments (D3-leucine/[D3-leucine + D0-leucine] × 100) were generated by averaging all participants’ enrichments (n = 9) at each time point and modeling them as a single participant. Points = data; lines = model fit; a1 = α-1; a2 = α-2; a3 = α-3; preb = pre-β. FF, forcing function.

Figure 4. Dietary unsaturated fat increases the metabolism of HDL containing apoE.

(A and B) HDL containing apoE (E+) has a faster FCR than HDL lacking apoE (E–) on both high-fat and high-carbohydrate diets. (C and D) Dietary unsaturated fat increases the clearance rates of the larger sizes (α-1 and α-2) of E+. ApoA-I fractional catabolic (turnover) rates (FCRs) in pools/day are shown for each HDL subspecies during intake of 2 diets. Each data point is an individual study participant (n = 9 for all). Black bar = mean, error bars = SEM. LFD, low-fat diet; HFD, high-fat diet. P values were calculated using paired 2-tailed t test. Overall P value for comparison considering all sizes together is shown on each graph. (A) Comparison of apoA-I FCRs on subspecies of E+ or E– in the context of a high-fat diet. (B) Comparison of apoA-I FCRs on subspecies of E+ or E– in the context of a low-fat diet. (C) Comparison of apoA-I FCRs across 2 diets only among the subspecies of E+. (D) Comparison of apoA-I FCRs across 2 diets only among the subspecies of E–.

Figure 5. Dietary unsaturated fat increases the synthesis of HDL containing apoE.

Mean (n = 9) plasma apoA-I pool sizes and synthesis rates are shown in 4 sizes of HDL subspecies containing apoE (E+) or not containing apoE (E–), across 2 diets (HFD, high-fat diet; LFD, low-fat diet). Error bars = SEM. P values were calculated using paired 2-tailed t test. (A) Mean pool size of apoA-I (mg) and percentage of apoA-I on E+ HDL (numbers below bars). (B) Mean synthesis rate of apoA-I (mg/day) and percentage of apoA-I synthesized on E+ HDL per day (numbers below bars).

Figure 6. HDL containing apoE (E+): dietary unsaturated fat increases apoA-I flux through synthesis, size interconversion, and clearance pathways.

Individual values shown, with mean value (n = 9) in larger red circles and dotted red line. HFD, high-fat diet; LFD, low-fat diet. P values were calculated using paired 2-tailed t test. (A) Pathways from the source compartment to each HDL size, representing synthesis and secretion from the liver (or small intestine) and appearance in plasma. (B) Pathways between HDL sizes, representing HDL size expansion or contraction. The source size and destination size are indicated above each graph. (C) Clearance of apoA-I on each HDL size from plasma. Note that all of α-3 is converted to pre-β and is thus not directly cleared from plasma (see Figure 3, A and B).

Figure 7. Model of reverse cholesterol transport stimulated by dietary unsaturated fat (HFD) through HDL containing apoE.

In the left panel, HDL not containing apoE is secreted in all sizes by the liver, which is weakly stimulated by HFD (gray box with dashed up arrow). This HDL subspecies visits peripheral tissues (represented by the macrophage), where it may take up cholesterol but is insufficient to increase HDL to a larger size category. This HDL subspecies is cleared slowly by the liver with an average residence time of approximately 2.5 days, regardless of diet. In the right panel, the HFD significantly increases secretion of all sizes of HDL containing apoE (red box with bold red arrows). This HDL visits peripheral tissues, takes up cholesterol, and undergoes size expansion, which is stimulated by HFD. HFD significantly increases clearance of this subspecies as well as size contraction from α-3, representing lipid uptake and pre-β generation at the liver, reducing the residence time to approximately 9 hours (HFD) and 16 hours (low-fat diet, LFD). These actions together may represent enhanced reverse cholesterol transport (RCT) driven by high-unsaturated-fat diets.