Apolipoprotein E polymorphisms

Apolipoprotein E contains 299 amino acids and an arginine-rich glycoprotein. It is associated with VLDL, chylomicrons, chylomicron remnants, intermediate density lipoproteins and with some cholesterol-rich subclasses of HDL.
The structural gene for apo E is polymorphic, with 3 common alleles (e2,e3,e4) coding for three isoforms of Apo E protein: E2, E3 and E4. These differ in their amino acid sequence at positions 112 and 158. Apo E3, the prevalent allele, has cysteine at position 112 and arginine at position 158. In Apo E2 a cysteine residue replaces arginine at position 158, whilst Apo E4 has arginine is both positions.
Individuals inherit one Apo E allele from each parent and the six genotypes e2/2,e2/3e3/3,e2/4,e3/4,e4/4, encode 6 phenotypes.
Apo E isoforms differ in charge and can be phenotyped by isoelectric focusing and immunoblotting. Genotyping is performed using PCR based methods.

Clinical indications:
Homozygosity at the e2 locus is associated with the development of type III dyslipidaemia or ‘remnant disease’. More than 90% of patients with type III hyperlipidaemia express e2/e2. Approximately 1% of the population are homozygous for e2/e2 but only 1 in 50 of homozygotes develop type III hyperlipidaemia. Additional factors are required and may include the development of hypothyroidism, obesity, insulin resistance or diabetes.
Variation in apo E geno / phenotype is also believed to account for approximately 8% variation in serum cholesterol concentration at the population level, carriage of the e2 and e4 alleles being associated with lower and higher serum cholesterol respectively. The apo E geno / phenotype is also a marker of responsiveness to cholesterol-lowering dietary and drug therapies that target either cholesterol synthesis or absorption. This is largely due to an association between apo E isoforms and cholesterol synthesis and absorption; e2 carriers are high cholesterol synthesisers; e4 carriers are high cholesterol absorbers.

Interpretation:
Apo E bands are visualised by UV transillumination compared to reference primers. Patients with remnant (Type III) hyperlipidaemia are usually homozygous for the Apo E2 allele which may be demonstrated by either phenotyping or genotyping. Different degrees of protein post-translational modification may give rise to misleading results, particularly in pathological states such as diabetes mellitus, in which case genotyping is more reliable. Rare alleles may not be detected by allele specific genotyping methods. Mutations which do not alter electrophoretic mobility will not be detected by phenotyping.

Patient preparation:
A non-fasting sample is adequate

Sample preparation and transport:
EDTA plasma is preferred for phenotyping. Genotyping requires EDTA whole blood. Transport by first class post

Information required:
Age, sex, NHS/Hospital No.
Medication

References:
Utermann G, Hees M, Steinmetz A. Polymorphism of apolipoprotein E and
occurrence of dysbetalipoproteinaemia in man. Nature. 1977;269: 604-7

Havekes LM, de Knijff P, Beisiegel U, Havinga J, Smit M, Klassen E. A rapid micro method for Apolipoprotein E phenotyping directly in serum. J Lipid Res 1987; 28: 455-63

Song Y, Stampfer MJ, Liu S. Meta-analysis: apolipoprotein E genotypes and risk for coronary heart disease. Ann Intern Med. 2004; 141: 137-47

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