Determination of ApoE Phenotypes

Results

ApoE phenotyping was chosen instead of apoE genotyping because the former technique was used in studies of the reference populations (see below). We have previously found that these two techniques yield discrepant results in only 2.5% of cases.27 ApoE phenotyping was carried out with a modifi- cation28 of the method of Havekes et al28 based on isoelectric focusing of delipidated plasma followed by immunoblotting with anti-apoE antiserum. Before the isoelectric focusing the plasma was treated with cysteamine as described by Weisgra- ber et al.M

The Finnish centenarians were characterized by rel- atively low plasma total (4.6 mmol/L) and LDL choles- terol (2.4 mmol/L) levels and high HDL cholesterol (1.5 mmol/L) levels (Table 1). No significant sex differences were found in the mean plasma lipid or apolipoprotein levels. The sex difference in the phospholipid levels, even though statistically significant, was in relative terms similar to those in the HDL and LDL cholesterol levels. For subsequent analysis of plasma lipoproteins in the different genotypes, the data for the two sexes were combined.

The apoE phenotype distribution and allele frequencies in centenarians were compared with those in 615 adult (age range, 20 to 55 years) and 1577 young (age range, 3 to 18 years) Finns described previously.8" In the former study the partic- ipants were mainly from the vicinity of Helsinki, the capital of the country, but the latter study included volunteers from both

The frequency of the apoE e2 allele was significantly higher and that of the e4 allele lower among the centenarians than in the younger Finnish populations

TABLE 1. Plasma Uplds and Apollpoproteins In Finnish Centenarians

TABLE 3. Apollpoproteln E Phenotypes and Allele Frequencies in Centenarians by Sex

Genetic Grouping Women Men ApoE phenotypes, No. of subjects E2E2 1 0 E2E3 18 5 E3E3 104 20 E3E4 27 2 E4E4 1 0 E1E3 0 1 ApoE allele frequencies, % e2 6.6 8.9 e3 83.8 85.7 E4 9.6 36 ef 0 1.8

Women Men Variable (n=151) (n-28) P Triglycerides, mmol/L 1.65±0.78 1.52±0.58 NS* Cholesterol, mmol/L 4.64+0.94 4.23±0.84 NS LDL cholesterol, mmol/L 2.39±0.85 2,13±0.71 NS HDL cholesterol, mmol/L 1.50±0.41 1.42±0.38 NS HDL cholesterol, mmol/L 0.57±0.29 0.52±0.32 NS 2 HDLa cholesterol, mmol/L 0.93±0.23 0.90±0.22 NS Phospholipids, mg/dL 203.3±33.4 183.2±27.8 <.01 Apolipoproteln A-l, mg/dL 123.9±25.1 114.3±23.0 NS Apolipoprotein A-ll, mg/dL 29.3±6.53 29.4±6.5 NS Apollpoproteln B, mg/dL 81.3±16.3 76.9±18.4 NS

LDL indicates low-density lipoproteln; HDL, high-density protein. Values are mean+SD.

ApoE indicates apolipoprotein E.

studied earlier (Table 2). The most remarkable finding of the present study was the 63% reduction in the prevalence of the apoE e4 allele from the oldest Finnish age group (Table 2). There were no significant differ- ences between women and men in the prevalences of the different apoE phenotypes or alleles, loss of the e4 allele from this age group having taken place in both sexes (Table 3).

phenotypes, showed a tendency for the concentrations to be highest in the centenarians with at least one e4 allele and lowest in those with at least one e2 allele (Table 4). However, the differences reached statistical significance for HDL cholesterol only (Table 4). In 2 fact, all individuals with apoE3/E4 and apoE4/E4 phe- notypes had plasma HDL cholesterol levels higher than 1.0 mmol/L, while their total cholesterol levels ranged

The apoC-III 52 allele occurred among the centenar- ians with a relatively high frequency (12.9%) that showed a difference bordering on significance (P—.05) when compared with either of the younger reference cohorts (Table 2). In contrast, there were no significant differences in apoB Xba I allele frequencies between the centenarians and the younger populations (Table 2). Plasma total and lipoprotein cholesterol concentra-

Plasma cholesterol levels tended to increase in the order of the apoB Xba I genotypes: X1X1<X1X2 <X2X2 (Table 4). Since both the apoE E4 phenotype and the apoB X2 allele seemed to have a cholesterol- elevating effect, the combined influence of the alleles at the two loci was examined. Centenarians with the apoB X2X2 genotype and apoE E4 phenotype had a signifi- cantly higher mean plasma cholesterol level (5.24

Plasma total and lipoprotein cholesterol concentra- tions, divided into three groups according to apoE

2. Apollpoproteln E, B, and C-lll Allele Frequencies In Finnish Centenarians Study) and Younger Finnish Populations

Gene/Allele Centenarians Adult Finns* Young Flnnst ApoE e2 7.0(4.6-10.1) 4.1* (3.0-5.3) 3.9§ (3.3-46.3) e3 84.4(80.6-88.1) 73.3|| (70.9-75.8) 76.7§ (75.2-78.2) z4 8.4(5.7-11.7) 22.7||(20.3-25.1) 19.4||(18.0-20.8) ApoB X1 57.9 (52.7-62.9) 58.2 (53.1-63.4) 57.8 (53.9-61.7) X2 42.1 (37.1-47.3) 41.8 (36.6-46.9) 42.2 (38.3-46.1) ApoC-III S1 87.1 (83.7-90.6) 91.6 (88.3-94.3) 91.2 (88.7-93.3) S2 12.9(9.4-16.3) 8.4 (5.7-11.7) 8.8+(6.7-11.3)

ApoE indicates apolipoprotein E; ApoB, apolipoprotein B; and ApoC-III, apolipoprotein C-lll. are percentages (95% confidence intervals). *For apoE alleles, data from Reference 8 (n=615; age group, 20-55 years). For apoB alleles, from Reference 18 (n=176; age group, 20-66 years). ApoC-III alleles were determined from the cohort (n=179) during the present study. tFor apoE alleles, data from Reference 11 (n=i577; age group, 3-18 years). For apoB and alleles, data from Reference 17 (n=307; age group, 9-21 years). tP<.05, §P<.01, ||P<.001 compared with the centenarians.

*For apoE alleles, data from Reference 8 (n=615; age group, 20-55 years). For apoB alleles, data from Reference 18 (n=176; age group, 20-66 years). ApoC-III alleles were determined from the same

tFor apoE alleles, data from Reference 11 (n=i577; age group, 3-18 years). For apoB and apoC-III alleles, data from Reference 17 (n=307; age group, 9-21 years). tP<.05, §P<.01, ||P<.001 compared with the centenarians.