There are large differences in ApoE allele frequencies among geographical areas and populations (Hallman et al., 1991; Singh et al., 2006), e.g., with high relative frequencies of ApoE ε4 in Northern Europe (15–21%), intermediate frequencies in Central Europe (11–13%), and low frequencies in Southern Europe (4–6%) (Lewis and Brunner, 2004). The ε4-allele frequency in our Spanish (10%) and Italian (8%) young controls was relatively higher than that previously reported by Lewis and Brunner (2004) in Southern Europe. Whereas, the ApoE ε4-allele frequency in Spanish and Italian centenarians was 4.9 and 3.2% respectively, which is within the range of previously published frequencies (1–10%) for European centenarians (Blanche et al., 2001; Capurso et al., 2004; Castro et al., 1999; Gerdes et al., 2000; Louhija et al., 1994; Maruszak et al., 2012; Panza et al., 1999; Panza et al., 2003; Schachter et al., 1994; Seripa et al., 2006). On the other hand, al- lele frequencies in the Japanese centenarians studied here (ε2 = 7.3%;

There are several biological mechanisms to postulate ApoE, and particularly the ε4-allele, as a candidate to influence longevity. The

Table 5

Genotype/allele distributions of the apolipoproteinE (ApoE) gene and results of binary logistic regression in the Japanese cohort.

Cases (centenarians) Controls Cases vs. controls Fisher's test P-value Cases vs. controls Binary logistic regression n % n % OR(95%CI) P-value Genotype ε2/ε2 1 0.1 0 0.0 1.000 – – ε2/ε3 102 14.0 39 7.8 0.001 1.86 (1.26, 2.75) 0.002 ε2/ε4 2 0.3 2 0.4 1.000 0.68 (0.09, 5.00) 0.709 ε3/ε3 562 77.1 350 70.3 0.008 1.34 (1.04, 1.74) 0.024 ε3/ε4 59 8.1 98 19.7 b0.001 0.34 (0.24, 0.89) b0.001 ε4/ε4 3 0.4 9 1.8 0.018 0.22 (0.60, 0.83) 0.025 Allele ε2 106 7.3 41 4.1 0.002 1.81 (1.25, 2.63) 0.002 ε3 1285 88.1 837 84.0 0.004 1.43 (1.13, 1.81) 0.003 ε4 67 4.6 118 11.9 b0.001 0.35 (0.26, 0.57) b0.001

Significant P-values and associations are in bold. 95% CI, 95% confidence interval; OR, odds ratio (adjusted by

Table 6

Subgroup comparisons of genotype/allele distributions of the apolipoprotein E (ApoE) gene and results of binary logistic regression in Japanese centenarians.

Oldest centenarian Rest of centenarians Oldest vs. the rest Oldest vs. the rest (oldest quartile) Fisher's test P-value n % n % OR (95%CI) P-value Genotype ε2/ε2 1 0.7 0 0.0 0.210 – – ε2/ε3 23 15.0 79 13.7 0.694 1.17 (0.70, 1.94) 0.546 ε2/ε4 0 0.0 2 0.3 1.000 – – ε3/ε3 121 79.1 441 76.6 0.588 1.28 (0.83, 1.98) 2.58 ε3/ε4 7 4.6 52 9.0 0.094 0.45 (0.20, 1.02) 0.056 ε4/ε4 1 0.7 2 0.3 0.507 1.73 (0.16, 19.22) 0.655 Allele ε2 25 8.2 81 7.0 0.535 1.21 (0.75, 1.93) 0.433 ε3 272 88.9 1013 87.9 0.692 1.12 (0.75, 1.66) 0.593 ε4 9 2.9 58 5.0 0.127 0.53 (0.26, 1.09) 0.083

Abbreviations: 95% CI, 95% confidence interval; OR, odds ratio (adjusted by sex).

ε4-allele is a well-established risk factor for age-related conditions such as mainly CVD and AD. A meta-analysis including 121 studies showed that this allele is associated with higher CVD risk, coupled with higher plasma cholesterol levels (Bennet et al., 2007). The association with CVD may be the consequence of higher LDL cholesterol and triglyc- eride levels in the plasma of ε4-carriers (Dallongeville et al., 1992). A recent meta-analysis of 21 studies on ApoE polymorphisms and AD concluded that the ε4-allele is strongly associated with a significantly higher risk of sporadic late onset AD (Sadigh-Eteghad et al., 2012). ApoE-lipoproteins bind to several cell-surface receptors to deliver lipids, andalsotohydrophobicamyloid-β(Aβ)peptide,whichisthoughttoini- tiate toxic events that lead to synaptic dysfunction and neurodegenera- tion in AD (Liu et al., 2013). ApoE isoforms differentially regulate Aβ aggregation and clearance in the brain, and have important functions in regulating brain lipid transport, glucose metabolism, neuronal signal- ing, neuroinflammation, and mitochondrial function (Liu et al., 2013), with the ε4-allele being associated with higher Aβ cellular uptake, lead- ing to more Aβaggregationcompared toε3 (Liu et al., 2013; Smith2002; Verghese et al., 2013).

of dendritic spines and accelerating memory impairments in AD (Hashimoto et al., 2012; Wu et al., 2010). With regards to this, a recent study showed that the levels of Aβ oligomers in AD patients were lowest in those carrying the ε2-allele (Hashimoto et al., 2012). Theε2-allele also seems to play a protective role against CVD (Bennet et al., 2007; Gerdes etal.,2000;Kolovouetal.,2002),thoughtheevidenceisprobablyweaker than for negative effect of the ε4-allele (Drenos and Kirkwood, 2010). In addition, previous large population-based longitudinal studies, i.e., the Rotterdam study (Slooter et al., 2001) and the Danish 1905 birth cohort (Lindahl-Jacobsen et al., 2013) reported no favorable effect of ε2-allele on mortality in the elderly, despite protecting against cognitive decline in the oldest old (≥93 years) (Lindahl-Jacobsen et al., 2013).