Abstract
Our previous study showed that n-3 polyunsaturated fatty acid (PUFA) and estrogen (E) had synergistic hypocholesterolemic effects by inhibiting cholesterol synthesis and enhancing bile acid synthesis. The purpose of the present study was to investigate the hypothesis that α-linolenic acid (ALA), eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA) decrease low-density lipoprotein cholesterol (LDL-C), synergistically with E, via hepatic cholesterol synthesis and clearance. Rats were fed a diet with either 0% n-3 PUFA or 1% ALA, EPA, or DHA, relative to total energy consumption, for the entire 12-week study. After ovariectomy, rats were injected with either corn oil or E every 4 days for the last 3 weeks of the study. In combination with E, dietary supplementation with EPA or DHA increased the phosphorylated adenosine monophosphate-activated protein kinase/adenosine monophosphate-activated protein kinase ratio and LDL receptor expression, and it decreased the expression of 3-hydroxy-3-methylglutaryl coenzyme A reductase, sterol regulatory element-binding protein-2, and proprotein convertase subtilisin/kexin type 9 in the liver. In addition, dietary supplementation with EPA or DHA increased hepatic expression of cholesterol 7α-hydroxylase, sterol 12α-hydroxylase, and sterol 27-hydroxylase. However, E decreased the expression of cholesterol 7α-hydroxylase and sterol 12α-hydroxylase and increased the expression of estrogen receptor α and β in the liver. ALA had no significant effects on cholesterol metabolism. In conclusion, the present study suggests that dietary supplementation with EPA and DHA decreased LDL-C synthesis and increased bile acid synthesis and LDL-C clearance by LDL receptor, synergistically with E.
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Key Points
α-Linolenic acid (18:3n3, ALA), an n-3 PUFA present in high concentrations in flaxseed oil and perilla plants, can be converted into a longer-chain n-3 PUFA. The effect of ALA on LDL-C is debated and the mechanisms whereby ALA affects cholesterol synthesis and clearance are uncertain. Estrogen (E) has LDL-C–lowering effects by downregulating cholesterol synthesis and upregulating LDLR while decreasing bile acid synthesis. Previous study showed that E, alone, has hypocholesterolemic effects in addition to synergistic effects with n-3 PUFA. The purpose of the present study was to investigate the hypothesis that ALA, EPA, and DHA decrease LDL-C synergistically with E via hepatic cholesterol synthesis and clearance.
In the present study, the research hypothesis that EPA and DHA decreased LDL-C synergistically with E was accepted, but the hypothesis of a synergic effect of ALA and E is rejected. EPA and DHA, but not ALA, in combination with E had synergistic LDL-C–lowering effects by downregulating cholesterol synthesis and upregulating bile acid synthesis and cholesterol clearance. EPA and DHA lowered serum TC and LDL-C levels by inhibiting cholesterol synthesis and enhancing bile acid synthesis. In the present study, ALA (≤4% of total energy intake) had no effect on serum levels of TC and LDL-C or on enzymes related to hepatic cholesterol synthesis, such as HMGCR, SREBP-2, and p-AMPK/AMPK, or bile acid synthesis, such as CYP7A1, CYP8B1, and CYP27A1. However, other studies reported that ALA (>4% of total energy intake) lowers TC and LDL-C through the increase of bile acid synthesis by increasing the activity of CYP7A1, not by decreasing cholesterol synthesis. This suggests that it was unclear whether the effect of ALA on LDL-C was due to its conversion to EPA and DHA or due to the direct effects of ALA itself. This present study showed that supplementation with ALA, EPA, and DHA increased serum levels of 18:3n-3, 20:5n-3, and 22:6n-3, respectively. n-3 PUFA and E synergistically elevated serum levels of 22:5n3 and 22:6n3.
There are some limitations in this study. The ethyl ester form of n-3 PUFA was used rather than the triglyceride form because the ethyl ester form of n-3 PUFA is more stable to peroxidation than the triglyceride form. In addition, rats are commonly used for lipoprotein research, but animal data cannot be directly extrapolated to humans.