Esterification of 4β-hydroxycholesterol and other oxysterols in human plasma occurs independently of LCAT

The acyltransferase LCAT mediates FA esterification of plasma cholesterol. In vitro studies have shown that LCAT also FA-esterifies several oxysterols, but in vivo evidence is lacking. Here, we measured both free and FA-esterified forms of sterols in 206 healthy volunteers and 8 individuals with genetic LCAT deficiency, including familial LCAT deficiency (FLD) and fish-eye disease (FED). In the healthy volunteers, the mean values of the ester-to-total molar ratios of the following sterols varied: 4β-hydroxycholesterol (4βHC), 0.38; 5,6α-epoxycholesterol (5,6αEC), 0.46; 5,6β-epoxycholesterol (5,6βEC), 0.51; cholesterol, 0.70; cholestane-3β,5α,6β-triol (CT), 0.70; 7-ketocholesterol (7KC), 0.75; 24S-hydroxycholesterol (24SHC), 0.80; 25-hydroxycholesterol (25HC), 0.81; 27-hydroxycholesterol (27HC), 0.86; and 7α-hydroxycholesterol (7αHC), 0.89. In the individuals with LCAT deficiency, the plasma levels of the FA-esterified forms of cholesterol, 5,6αEC, 5,6βEC, CT, 7αHC, 7KC, 24SHC, 25HC, and 27HC, were significantly lower than those in the healthy volunteers. The individuals with FLD had significantly lower FA-esterified forms of 7αHC, 24SHC, and 27HC than those with FED. It is of note that, even in the three FLD individuals with negligible plasma cholesteryl ester, substantial amounts of the FA-esterified forms of 4βHC, 5,6αEC, 7αHC, 7KC, and 27HC were present. We conclude that LCAT has a major role in the FA esterification of many plasma oxysterols but contributes little to the FA esterification of 4βHC. Substantial FA esterification of 4βHC, 5,6αEC, 7αHC, 7KC, and 27HC is independent of LCAT.

Abstract 584: ApoE3[K146N/R147W] Acts as a Dominant Negative ApoE Form that Prevents Remnant Clearance and Inhibits the Biogenesis of HDL

Introduction: The K146N/R147W substitutions in human apolipoproteinE3 (apoE3) have been associated with a dominant form of type III hyperlipoproteinemia which is expressed at an early age. Methods: The effects of the K146N,R147W substitutions on the lipid and lipoprotein profiles and the HDL phenotypes were studied by adenovirus mediated gene transfer of the full length and a truncated apoE3[K146N/R147W]-202 mutant using different mouse models. Results: A low dose of adenovirus expressing the apoE3[K146N/R147W] mutant in apoE deficient or in apoA-I x apoE double deficient mice exacerbated the hypercholesterolemia and increased greatly plasma apoE and triglycerides levels. In apoE deficient mice, the apoE3[K146N/R147W] mutant displaced apoA-I from the VLDL/LDL/HDL region and resulted in the accumulation of discoidal apoE containing HDL particles in plasma. Similar doses of WT apoE3 cleared the cholesterol of apoE deficient mice without induction of hypertriglyceridemia and promoted formation of spherical HDL particles. A unique feature of the truncated apoE3[K146N/R147W]-202 mutant is that it prevented the induction of hypertriglyceridemia but did not correct the hypercholesterolemia. Other apoE-202 truncated mutants tested did not induce hypertriglyceridemia but corrected hypercholesterolemia. Infection of apoE deficient mice with adenoviruses expressing the apoE3[K146N/R147W] and lipoprotein lipase corrected the hypertriglyceridemia but did not prevent the formation of discoidal HDL particles. Infection of apoE deficient mice with adenoviruses expressing the apoE3[K146N/R147W] and lecithin:cholesterol (LCAT) acyltransferase corrected hypertriglyceridemia, normalized the plasma cholesteryl ester to total cholesterol ratio and generated spherical HDL particles. The combined data indicate that the K146N/R147W substitutions in the full length and the truncated apoE3[K146N/R147W] mutant prevent receptor mediated remnant clearance. Conclusion: The lipid/lipoprotein and HDL abnormalities observed in the different mouse models suggest that the apoE3[K146N/R147W] mutant acts a dominant negative ligand that exacerbates the dyslipidemia and also affects the activity of LCAT and thus inhibits the maturation of HDL.