however, KL-VS allele status was associated significantly with incidence of stroke (P(cid:1) 0.026). Similar to the CVD risk factor data, heterozygous individuals have the fewest stroke events and homozygous KL-VS the most. Multiple logistic regression was performed incorporating traditional CVD risk factors, demonstrating that KL-VS allele status is an inde- pendent predictor of stroke (P(cid:1) 0.009; Table 4). Wild-type individuals had a 5.88-fold (95% CI, 1.18 to 29.41) increased OR for stroke and homozygous KL-VS individuals a 30.65- fold (95% CI, 2.55 to 368.00) increased OR for stroke, relative to heterozygous individuals.

Medication data were available for   50% of our samples, with   33% taking cholesterol-lowering medication and ~S 50% taking antihypertensives. To test for potential con- founding attributable to medication, we substituted pulse pressure (defined as SBP-DBP) for SBP as the outcome in our regression analysis because this measure is robust to the effects of blood pressure medication18 and does not result in a loss of sample size. The overall effect of KL-VS genotype was significant (P(cid:1) 0.039), again exhibiting a heterozygous advantage in conjunction with a homozygous disadvantage for the KL-VS allele. To examine the effects of cholesterol- lowering medication, we added a term into the regression model for medication status, and although the sample size was significantly reduced, the effect sizes for individuals homozygous for the wild-type or KL-VS alleles were in- creased, indicating that any potential confounding attributable to medication is more likely to lead to a false-negative result rather than a false-positive result.

Prospective Analysis of Survival in Probands

Probands were recruited from 1998 to 2002 and were fol- lowed prospectively for mortality status; as of January 2004, 96 of 216 (44.4%) had died. Univariate logistic regression indicates that KL-VS allele status is a significant predictor of mortality (P(cid:1) 0.016), and similar to the other phenotypes

P<0.026 StrokeFFmFVgvvVP=0.79 

Because recent work has demonstrated the importance of lipid particle size in CAD and longevity, we examined whether HDL-C and LDL-C particle size are associated with KL-VS allele status. Univariate regression analysis indicates that KL-VS allele status is a significant predictor of LDL-C particle size (P(cid:1) 0.030) and HDL-C particle size (P(cid:1) 0.023); however, in multivariate analyses, KL-VS allele status was not a significant predictor of LDL-C particle size (P(cid:1) 0.17) or HDL-C particle size (P(cid:1) 0.75; data not shown).

Analysis of CVD Events

A subset of samples had information on history of events. A total of 11.1% reported having had a stroke (30 271) and 13.4% a myocardial infarct (MI) (39 of 292). 3 depicts event data stratified by KL-VS genotype. No of KL-VS allele status on MI was observed (P(cid:1)

Figure 3. Coronary vascular disease event data stratified by KL-VS allele status. P value indicates overall significance of the KL-VS allele as determined by the (cid:3)2 test.

4. Multivariate Regression Analysis for Cardiovascular Events*

Event P † Genotype Beta‡ SE P OR/RR§ 95% CI robust   Stroke 0.009 FF 1.77 0.82 0.031 5.88 1.18 to 29.41 VV 3.42 1.27 0.007 30.65 2.55 to 368.00 Mortality 0.010 FF 0.77 0.31 0.012 2.15 1.18 to 3.91 VV 1.50 0.62 0.015 4.49 1.35 to 14.97

*Model is adjusted for age, sex, body mass index, LDL-C, HDL-C, triglycerides, SBP, DBP, and proband status.

†P refers to the overall significance of the KL-VS allele (see Materials and Methods). robust ‡Homozygous wild-type (FF) and KL-VS (VV) individuals are compared with heterozygous individuals (FV); §mortality data are prospective and so RRs are calculated.

frequency of KL-VS heterozygous individuals as a function of age in the current cohort of 525 Ashkenazi Jews demon- strated remarkable concordance with the initial study in  Czechs, exhibiting a heterozygous advantage for ages 79 years. A combined analysis was performed for overlapping age ranges, demonstrating a 1.57-fold (95% CI, 1.23 to 1.98) increased OR for survival in two independent populations. No significant trend was observed for homozygous KL-VS indi- viduals (data not shown), largely attributable to small sample size. However, the prospective analysis of mortality (dis- cussed below) confirms the homozygous KL-VS disadvan- tage for survival.

examined, heterozygous individuals have the lowest mortality and homozygous KL-VS individuals the highest (Figure 4). We performed Cox regression survival analysis, adjusting for age, sex, body mass index, SBP, DBP, HDL-C, LDL-C, and triglycerides, indicating that KL-VS allele status is associated significantly with mortality after adjusting for potential con- founders (P(cid:1) 0.01). Wild-type individuals had a 2.15-fold (95% CI, 1.18 to 3.91) increase in relative risk (RR) for mortality relative to heterozygous individuals. Homozygous KL-VS individuals displayed a 4.49-fold (95% CI, 1.35 to 14.97) increased RR compared with heterozygous individu- als. These data strongly support the cross-sectional analyses indicating a heterozygous advantage for survival late in life.