Apolipoprotein (apo) E, an argininerich glycoprotein with a molecular mass of 34 kDa, is localized on chylomicrons, VLDL, intermediate-density lipoprotein, and HDL particles. Apo E is important for the regulation of triglyceride-rich lipoproteins in humans and binds to the VLDL receptor, LDL receptor, and LDL-receptor-related protein. This process allows apo E to compensate partially for the function of defective apo B-100 in humans (1). There are three different apo E alleles, ε2 (frequency, 10%), ε3 (frequency, 75%), and ε4 (frequency, 15%), that code different apo E isoforms. Apo E 3/3 is the most commonly found phenotype. Individuals with the apo E 2/2 phenotype are usually normolipidemic, but in combination with other factors (such as hyperinsulinemia, diabetes mellitus, thyroid dysfunction, or treatment with steroids), apo E 2/2 causes most typically type III hyperlipidemia. At present, it is unclear which other lipid-related cofactors might play a role in apo E 2/2-related hyperlipidemia. Most recently, apo A5 was described (2), and subsequently, a frequent mutation of apo A5, apo A5 S19W, was shown to cause hypertriglyceridemia in humans (3). However, the apo A5 mutation alone is also not sufficient to cause hypertriglyceridemia, not even in individuals homozygous for apo A5 S19W (our own observation). To clarify the roles of apo E 2/2 and apo A5 in hypertriglyceridemia, we studied a total of 170 patients with fasting triglyceride concentrations 2 g/L. In all of these patients, we determined the apo E phenotype as well as the presence of the apo A5 S19W mutation. Apo E phenotyping was performed by isoelectric focusing with use of in-house-prepared polyacrylamide gel slides (43 50 0.45 mm; T 5%, C 3%, pH 5–8) on the PhastSystemTM as described recently (4). Apo E bands were immunoprecipitated within the gel, and silver staining was performed after extensive washing and eluting of all nonprecipitated proteins. We used denaturing gradient gel electrophoresis (DGGE)(5) to screen for the apo A5 S19W mutation in the gene. In all patients with an abnormal DGGE profile, apo A5 was sequenced, and single-nucleotide polymorphisms were confirmed by PCR with restriction fragment length polymorphism analysis. In addition, apo C-II was analyzed by isoelectric focusing (6) and the LDL receptor by DGGE (5) in all patients with apo E 2/2 and/or apo A5 S19W, as described recently. A total of 7 of our 170 screened hypertrigyceridemic patients had an apo E 2/2 phenotype; 6 of these 7 apo E 2/2 patients were also heterozygous carriers for the apo A-5 S19W mutation. The mean (SD) triglyceride concentration in these patients was 4958 (1462) mg/L, their mean total cholesterol was 2683 (886) mg/L, and their mean HDL-cholesterol was 382 (117) mg/L. This finding is interesting for several reasons. Our study confirms earlier observations that apo E 2/2 is a rare cause for hypertriglyceridemia. It was present in our study population at a rate of 4.1%. Interestingly, almost all of these (six of seven) hypertriglyceridemic apo E 2/2 individuals had also the apo A5 S19W defect. The frequency of the apo A5 S19W polymorphism has been reported to be 10.9% in the general population (7). Furthermore, we failed to identify a normolipidemic apo E 2/2 individual who had the apo A5 S19W polymorphism. From this observation we conclude that apo A5 S19W is a crucial cofactor for developing hypertriglyceridemia in patients with apo E2/2. This hypothesis clearly needs to be confirmed in larger study populations, but it is consistent with our current knowledge of hypertriglyceridemia in apo E2/2. Apo E 2/2 is found in 0.6% of our population (8), and 10% of them will …