Relationships between other lipoprotein traits and APOC3 (cid:1)641 CC genotype are shown in Table 1. Due to the dependency of lipoprotein traits on age and their modifi- ability with medications, we only considered offspring (n ¼ 131) and controls (n ¼ 126) not using lipid lowering drugs in this analysis. In females, triglycerides (TG), high density lipoprotein cholesterol (HDL), and their ratio, as well as low density lipoprotein cholesterol (LDL) lipoprotein particle size were significantly more favorable among those with the

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Figure 3. Genotype Distribution of APOC3 C((cid:1)641)A and Serum APOC3 Concentrations (A) APOC3 C((cid:1)641)A. (B) Serum APOC3. *p , 0.05, ** p ¼ 0.001 versus control. CC, homozygous for ((cid:1)641) C; AA, homozygous for ((cid:1)641) A; CA, heterozygous for ((cid:1)641) C/A.

 tTT TToo fF fo ee ge~ Ob HW FN Naonejndod ul aseyues.10g APOC3 (641) genotype   tT NOC wmM OU TASSs otmO (Iw/Sn) UoWEAyUI909 EIOAVOffspring ProbandsControl

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Figure 3. Genotype Distribution of APOC3 C((cid:1)641)A and Serum APOC3

females and males and their significance. As shown in Table 1 the combined group also exhibited significant associations with lipoproteins according to APOC3 CC genotype.

Given that insulin resistance is associated with lipid abnormalities, CVD, and death in the elderly [9], we estimated insulin sensitivity using the homeostatic model assessment [10]. The favorable APOC3 (cid:1)641 CC genotype was associated with greater insulin sensitivity in the combined offspring/ control group (p , 0.05, both male and female) (Figure 4A).

Ordinal arrangement of 15 SNP associated with CVD and lipoprotein metabolism, according to their position on chromosomes with LD (number in boxes) where the highest rate is represented in red and no LD in lilac. Blocks define potential haplotypes between two clustered genes. (A) APOC3 and APOA4. (B) CETP.

The most prevalent CVD marker in an aging population is progressive increase in blood pressure. Therefore, this trait was examined in our participants as a surrogate for vascular aging. The prevalence of hypertension (as defined in the VII Report of the Joint National Commission on Prevention, Detection, Evaluation, and Treatment of High Blood Pres- sure) [11] was significantly lower among those with the APOC3

CC genotype compared to the CA/AA genotypes. The same trends were observed in most of the associations in men. Since there was no significant interaction between gender and genotypes, we present also the combined group of

Table 1. Comparison of Lipoprotein Traits in Females, Males, and All (Combined Offspring and Control Group) Who Were Not on Lipid- Lowering Drugs by APOC3 C((cid:1)641)A Genotype

Phenotype APOC3 Control and Offspring Female Male Female and Male CC CA/AA p-Value CC CA/AA p-Value p-Value versus CA/AA Number 32 124 20 83 Age (y) 67.8(8.6) 68.3(9.2) 0.75 71.8(1.3) 74.3(0.9) 0.22 0.99 BMI (Kg/m2) 24.9(0.5) 25(0.3) 0.91 25(0.6) 26.3(0.4) 0.12 0.14 HDL (mg/dl) 73.8(2.77) 64.5(1.5) 0.005 56.9(4.1) 51.5(1.7) 0.18 0.004 TGL (mg/dl) 136(10.5) 132(5.3) 0.79 114(15.5) 108(4.9) 0.63 0.93 TG/HDL 1.5(0.2) 2.0(0.1) 0.03 2.22(0.4) 2.34(0.1) 0.72 0.01 VLDL (mg/dl) 68.6(9.12) 68.3(4.82) 0.98 56.6(8.3) 73(7.0) 0.23 0.49 LDL (mg/dl) 129(6.02) 134(2.82) 0.46 111(7) 107(2.7) 0.55 HDL size (nm) 9.35(0.06) 9.36(0.04) 0.93 9.13(0.1) 9.15(0.1) 0.88 0.91 LDL size (nm) 21.6(0.05) 21.4(0.06) 0.03 21.3(0.1) 21.1(0.1) 0.37 0.02 APOC3 (mg/dl) 8.27(0.88) 11.2(1.05) 0.034 7.52(1.6) 11.5(1.5) 0.12 0.03 ApoA-1 (mg/dl) 186(5.25) 190(4.02) 0.57 174(9.6) 154(5.6) 0.12 0.34 ApoB (mg/dl) 102(3.77) 101(3.3) 0.93 99.6(5.7) 93.6(2.9) 0.37 0.47

CC, homozygous for ((cid:1)641) C; CA/AA, homozygous and heterozygous for ((cid:1)641) A. ApoA-1, apolopoprotein A-1; ApoB, apolipoprotein B; BMI, body mass index; VLDL, very low density lipoproteins. DOI: 10.1371/journal.pbio.0040113.t001

(cid:1)641 CC genotype than among those with CA/AA genotypes (p ¼ 0.04) (Figure 4B). Since existing CVD can lead to reductions in blood pressure, we repeated the test after excluding all participants with CVD from the analysis. Still, hypertension was less prevalent in those with the favorable APOC3 CC genotype (prevalence of hypertension in partic- ipants carrying the APOC3 CC genotype was 28.6% compared with 44% in the CA/AA group, p ¼ 0.026). Other known variants of the APOC3 gene: C((cid:1)482)T, T((cid:1)455)C, C1100T, C3175G, and T3206G, (rs2854117, rs2854116, rs4520, rs5128, and rs4225, respectively) were not associated with TG levels, insulin sensitivity, or blood pressure.

specific way [19], and in patients with type 2 diabetes predisposition [20]. While few studies have correlated both unfavorable APOC3 promoter genotypes and high APOC3 levels with outcome [21,22], our study is the first to our knowledge to implicate a genotype with low levels of APOC3 and a favorable lipid phenotype with better outcomes related to longer lifespan. Murine models expressing one to 100 copies of the human APOC3 gene had high levels of APOC3 and were hypertriglyceridemic [23]. APOC3 knockout mice show absent or decreased APOC3 protein with reduced TG levels [24], even in the presence of streptozotocin-induced diabetes [25]. The impact on lifespan has not, however, been determined in these models.

Of the 381 participants we have genotyped since 1998, 64 had the CC genotype. To describe the relationship between genotype and death, we plotted the Kaplan-Meier survival function estimates of study participants by APOC3 genotype (Figure 5). Survival was significantly greater among CC genotype carriers (p ¼ 0.0007).