A Western-fed diet increases plasma HDL and LDL-cholesterol levels in apoD-/- mice

Abstract

Objective: Plasma apolipoprotein (apo)D, a ubiquitously expressed protein that binds small hydrophobic ligands, is found mainly on HDL particles. According to studies of human genetics and lipid disorders, plasma apoD levels positively correlate with HDL-cholesterol and apoAI levels. Thus, we tested the hypothesis that apoD was a regulator of HDL metabolism.

Methods & results: We compared the plasma lipid and lipoprotein profiles of wild-type (WT) C57BL/6 mice with apoD-/- mice on a C57BL/6 background after receiving a high fat-high cholesterol diet for 12 weeks. ApoD-/- mice had higher HDL-cholesterol levels (61±13-apoD-/- vs. 52±10-WT-males; 37±11-apoD-/- vs. 22±2 WT-female) than WT mice with sex-specific changes in total plasma levels of cholesterol and other lipids. Compared to WT, the HDL of apoD-/- mice showed an increase in large, lipid-rich HDL particles and according to size various quantities and sizes of LDL particles. Plasma levels of lecithin:cholesterol acyltransferase in the control and apoD-/- mice were not different, however, plasma phospholipid transfer protein activity was modestly elevated (+10%) only in male apoD-/- mice. An in vivo HDL metabolism experiment with isolated Western-fed apoD-/- HDL particles showed that female apoD-/- mice had a 36% decrease in the fractional catabolic rate of HDL cholesteryl ester. Hepatic SR-BI and LDLR protein levels were significantly decreased; accordingly, LDL-cholesterol and apoB levels were increased in female mice.

Conclusion: In the context of a high fat-high cholesterol diet, apoD deficiency in female mice is associated with increases in both plasma HDL and LDL-cholesterol levels, reflecting changes in expression of SR-BI and LDL receptors, which may impact diet-induced atherosclerosis.

Figure 1. Up-regulation of plasma apoD levels in Western-fed WT mice.

Western blot and semi-quantitative analyses of apoD levels in WT mice fed a 12-week chow and Western diets. Data is representative of both sexes, n = 4–5 per group, mean±SD, *p<0.05.

Figure 2. ApoE-HDL enrichment and lipids content changes in HDL particles.

Size exclusion chromatography lipid profiles (total cholesterol, free cholesterol, phospholipids) of pooled plasma from A) female and B) male WT and apoD−/− mice after 12 weeks on a Western diet.

Figure 3. No change in plasma apoAI, but changes in apolipoproteins content in HDL-size exclusion chromatography fractions.

A) Western blot analysis of apoAI levels from the SEC-HDL fractions. B) Semi-quantitative analysis of plasma apoAI levels. C) SimpleStain and Western blotting for apolipoprotein levels of pooled SEC-HDL fractions (10–20 µg).

Figure 4. Increased PLTP activity.

A) Plasma LCAT and B) PLTP activities from female and male WT and apoD−/− mice on a Western diet for 12 weeks. n = 7–14, values are mean±SD, *p<0.05.

Figure 5. Decrease in the relative protein expression of SRBI and LDLR in female apoD−/− mice but opposite pattern for mRNA expression.

Liver levels of A) Cholesterol and B) Triglyceride C) Relative expression of genes and D) Protein expression levels of LDLR and SRBI after 12 weeks on a Western diet. n = 5–7 per group, mean±SD, *p<0.05, **p<0.01.

Figure 6. A decrease in ³H-CE HDL fractional catabolic rate in female apoD−/− mice only.

Mice were fed a Western diet for 12 weeks and ³H-cholesteryl oleate-labeled apoD−/−HDL was injected. A) Changes in plasma ³H-CE-HDL and calculated fractional catabolic rate (FCR), B) ³H-cholesterol (% of injected dose) in the liver, C) ³H-sterol and bile acids (% of injected dose) in feces. The data represent n = 4–6 per group, mean±SD, * indicates statistical significant, p<0.05.