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Apolipoprotein E in Hyperlipidemia

Cristina C. Walden, MD; and Robert A. Hegele, MD
[+] Article, Author, and Disclosure Information

From St. Michael's Hospital and the University of Toronto, Toronto, Ontario, Canada. Requests for Reprints: Robert A. Hegele, MD, DNA Research Laboratory, St. Michael's Hospital, 30 Bond Street, Toronto, Ontario, Canada M5B 1W8. Acknowledgments: The authors thank Dr. Philip W. Connelly and the staff of the Lipid and DNA Research Laboratories at St. Michael's Hospital for general academic support. Grant Support: Dr. Hegele is a MacDonald Scholar of the Heart and Stroke Foundation of Canada. Dr. Walden is a Jeane B. Kempner Scholar of the University of Texas Medical Branch at Galveston.

Copyright ©2004 by the American College of Physicians

Ann Intern Med. 1994;120(12):1026-1036. doi:10.7326/0003-4819-120-12-199406150-00009
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Purpose: To review DNA analysis of apolipoprotein E used to assess patients with hyperlipidemia.

Data Sources and Study Selection: 44 basic science studies of molecular analysis; 42 basic science studies of the biochemical, cellular biological, and molecular biological features of apolipoprotein E; and 29 clinical investigational studies, meta-analyses, and case series of patients with mutations in apolipoprotein E.

Data Extraction: Methods of DNA analysis were reviewed, using specific examples in human disease, and the role of apolipoprotein E in normal and disordered lipoprotein metabolism was reviewed. Genetic analysis of apolipoprotein E in populations and particularly in persons with type III hyperlipoproteinemia is reviewed.

Data Synthesis: In the general population, common DNA variants of apolipoprotein E are consistently associated with modest differences in plasma lipids and lipoproteins. Homozygosity for the E2 isoform of apolipoprotein E predisposes some patients to the development of type III hyperlipoproteinemia, a condition that involves an additional genetic or environmental factor for full clinical expression. Rare mutations of apolipoprotein E also cause hyperlipidemia.

Conclusions: DNA variation of apolipoprotein E is one of several genetic and environmental factors that interact in a complex manner to affect plasma lipoproteins. DNA analysis of apolipoprotein E can be used in persons with hyperlipidemia to identify those with type III hyperlipoproteinemia and in relatives of affected persons to identify those who are predisposed.


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Figure 1.
Methods of DNA analysis of the apolipoprotein E (apo E) gene.

The polymerase chain reaction (PCR)–based methods shown were used to detect and characterize in detail the apo E gene mutations and to determine apo E genotype. ASO = allele-specific oligonucleotide; RFLP = restriction fragment length polymorphism; DGGE = denaturing gradient gel electrophoresis.

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Figure 2.
DNA sequence of a portion of the apolipoprotein E gene.

The sequence within brackets comprises codons 250 to 259 in exon four from a mutant allele identified by DNA sequencing. Reading stepwise from 5′ to 3′, the sequence is ATA GGC CTG CAG GCC GAG GCC TTC CAG GCC. The missense mutation involves a G-for-C substitution at codon 251, which causes the substitution of glycine for arginine. A = adenine; C = cytosine; G = guanine; T = thymine.

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Figure 3.
Exogenous lipid transport.

Dietary fats are absorbed as chylomicrons, which are composed mainly of triglycerides (TG) and a small proportion of cholesterol esters (CE). Their main apolipoprotein is apo B48. As chylomicrons enter the circulation, they acquire apo E and apo CII. Apolipoprotein CII is a cofactor for lipoprotein lipase (LPL), an enzyme that hydrolyzes triglycerides into fatty acids and glycerol. The lipolytic action of lipoprotein lipase makes chylomicrons progressively smaller, depletes triglycerides, and enriches cholesterol ester. Chylomicron remnants are taken up by the liver through the remnant receptor, for which apo E is a high-affinity ligand. VLDL = very low-density lipoprotein.

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Figure 4.
Endogenous lipid transport.

Very low-density lipoproteins (VLDL) secreted by the liver are rich in triglycerides (TG) and contain apo B100, apo E, and the apo Cs. The lipolytic action of lipoprotein lipase (LPL) generates remnant particles that are enriched in cholesterol ester (CE) and apo E and are depleted of triglycerides. Some VLDL remnants are absorbed by endocytosis mediated by the low-density lipoprotein (LDL) receptor and the remnant receptor. Other VLDL remnants are hydrolyzed by hepatic lipase (HL) to yield LDL rich in cholesterol ester and containing apo B100. Most LDL undergoes LDL receptor-mediated endocytosis in the liver or peripheral tissues. The remaining LDL is absorbed in the tissues by non-LDL receptor-mediated processes (scavenger receptor).

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Figure 5.
Apolipoprotein E (apo E) phenotype determined by isoelectric focusing.

These are the isoelectric focusing gels of three homozygous persons. Apolipoprotein E isoforms are distinguished on the basis of charge by polyacrylamide gel electrophoresis. E2 has two cysteines (Cys) and is the most negatively charged isoform. E4 has no cysteine and is the most positively charged isoform. The lighter bands constitute minor sialated forms. The top of the gel is negatively charged and the bottom is positively charged. apo C = apolipoprotein C; Arg = arginine.

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Figure 6.
Location of apolipoprotein E (apo E) mutations associated with type III hyperlipidemia with respect to the putative receptor-binding domain.Table 2

In this schematic diagram, the numbers indicate apo E amino acid residues. The seven arrowheads 3′ of position 121 represent the 7-amino acid insertion of the apo E Leiden mutation ( ).

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Figure 7.
Apolipoprotein E (apo E) genotype determined by restriction isotyping.A.B.

Electrophoretic separation of HhaI digested fragments of a polymerase chain reaction-amplified portion of apo E exon four, encompassing codons 112 and 158. The six common apo E genotypes are shown. A standard DNA molecular weight marker with fragment sizes specified in base pairs is included for reference. HhaI restriction map for the three common apo E alleles. HhaI cleavage sites are indicated by arrows. The length of the fragments generated is specified on top of the lines. Codons 112 and 158 are marked.

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