A randomized trial and novel SPR technique identifies altered lipoprotein-LDL receptor binding as a mechanism underlying elevated LDL-cholesterol in APOE4s

Abstract

At a population level APOE4 carriers (~25% Caucasians) are at higher risk of cardiovascular diseases. The penetrance of genotype is however variable and influenced by dietary fat composition, with the APOE4 allele associated with greater LDL-cholesterol elevation in response to saturated fatty acids (SFA). The etiology of this greater responsiveness is unknown. Here a novel surface plasmon resonance technique (SPR) is developed and used, along with hepatocyte (with the liver being the main organ modulating lipoprotein metabolism and plasma lipid levels) uptake studies to establish the impact of dietary fatty acid composition on, lipoprotein-LDL receptor (LDLR) binding, and hepatocyte uptake, according to APOE genotype status. In men prospectively recruited according to APOE genotype (APOE3/3 common genotype, or APOE3/E4), triglyceride-rich lipoproteins (TRLs) were isolated at fasting and 4-6 h following test meals rich in SFA, unsaturated fat and SFA with fish oil. In APOE4s a greater LDLR binding affinity of postprandial TRL after SFA, and lower LDL binding and hepatocyte internalization, provide mechanisms for the greater LDL-cholesterol raising effect. The SPR technique developed may be used for the future study of the impact of genotype, and physiological and behavioral variables on lipoprotein metabolism. Trial registration number NCT01522482.

Figure 1. Sensorgram of the interaction between VLDL-1 rich particles (ranging from 0 to 40 nM apoB) and the LDL-receptor immobilized on the surface of a CM5 sensor chip.

Experimental data (dotted lines) was fitted with the two-state binding model (solid line).

Figure 2. Binding of lipoproteins to the LDL-receptor.

Lipoproteins (A) bind to the LDLR (B) to form complex AB, followed by a conformational change to form a more stable complex AB*. The association rate constants are ka1 and ka2 and the dissociation rate constants are kd1 and kd2. The equilibrium constant for each binding step are K1 = ka1/kd1 and K2 = ka2/kd2. The overall equilibrium binding constants are therefore K_A = K1 (1 + K2) and K_D = 1/K_A.

Figure 3. Overview of the sensorgrams for the two state conformational change reaction (A + B ↔ AB ↔ AB*).

The output response (solid line) is the sum of the individual responses for the fast process to form AB and the conformational change to form the more stable complex AB*. Reprinted from Morton et al. (1985) with permission from the author.

Figure 4. Effects of test meal composition on the binding of postprandial VLDL-1 rich TRL compared with fasting LDL particles to immobilized LDLR on a CM5 chip.

The figure represents single pooled samples (from two study participants) post SFA, UNSAT and SFA-DHA meal consumption, run in duplicate. The lipoprotein fractions were normalized for lipoprotein particle number using apoB concentration. Abbreviations: SFA, saturated fatty acid meal; SFA-FO, SFA meal with fish oil; UNSAT, unsaturated fatty acid meal.