Table 2. Demographic Statistics of the Leiden Longevity Study and the Leiden 85-Plus Study

Leiden Longevity Study Leiden 85-Plus Study Partners of Offspring Offspring of Octogenarian Participants (Controls) Nonagenarian Sib Pairs Nonagenarian Sib Pairs Young Leiden Controls From Leiden City 251 525 (238 sibships) 379 (164 sibships) 244 655 (SD) 59.5 (7.4) 60.6 (6.8) 93.8 (3.1) 31.0 (5.6) 87.5 (3.1) Male 45.0% 43.8% 32.5% 55.3% 32.7%

Note: SD ¼ standard deviation.

increases with the age of the individuals (30) and that the siblings in our study are approximately 5 years younger than those in the study by Puca and colleagues (7). A test for linkage in 13% of our participants who match the criteria of Puca and colleagues (7) did not reveal any linkage but is also simply underpowered to allow any conclusion. Possibly the 4q25 locus is confined to exceptional longevity and the familial effects on nonagenarian longevity in our study (19) are explained by other loci. Alternatively, the genetic determinants of longevity in the Dutch population may differ from those of the Boston study, despite the fact that both populations are of Caucasian descent.

Meta-Analysis

Recently, several studies (8,28,29) reported on the putative effect of the MTP T-Q haplotype on longevity. In a meta-analysis of all published studies, we analyzed 4915 long-lived cases older than 85 years and 3002 younger controls. When a heterogeneity test was performed that included the first and second tier of the U.S. study and the French, German, Danish, Leiden Longevity, and Leiden 85- Plus Studies, significant heterogeneity was observed ( p ¼ .039). The pooled odds ratio with random effects was 0.93 (95% CI, 0.77–1.14). When the two tiers were replaced by the U.S. proactively matched cases and controls, no heterogeneity could be observed among the studies ( p ¼ .256) and the pooled odds ratio with fixed effects was the same as with random effects: 1.00 (95% CI, 0.89–1.13) (Figure 3).

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Because association analysis is more sensitive than linkage analysis (31), we might have missed the linkage signal even in the presence of association. Therefore, we tested two polymorphisms in the MTP gene for association with longevity in the Leiden Longevity Study and the two cohorts of the prospective Leiden 85-Plus Study. This gene is located at chromosome 4q25, and a haplotype of these two SNPs has previously been identified as a marker for human life span (8) showing association in U.S. centenari- ans compared to U.S. controls. Our analyses, however, provided no evidence for association of (cid:1)493G/T, Q95H, or their haplotypes with longevity. This may have been explained if this 4q25 locus is simply not involved in the nonagenarian longevity trait of the Leiden Longevity population. Also, recent studies (28,29) describe the absence of association between the two-SNP haplotype of the MTP gene and human longevity in a cross-sectional design.

D ISCUSSION

To investigate whether the previously identified locus on chromosome 4q25, involved in exceptional human longev- ity, also plays a role in becoming a nonagenarian, we performed linkage analysis in 379 individuals belonging to 164 nonagenarian sib pairs from the Leiden Longevity Study. We found no evidence for linkage of nonagenarian longevity in this chromosomal region, despite previous observations that 4q25 was linked to healthy aging in a small study of 70-year-old twins (18).

A major reason for the absence of linkage in our study may be that the power to detect linkage with longevity

Table 3. Frequencies (%) of the (cid:1)493 T and 95 Q Microsomal Transfer Protein (MTP) Alleles and Their Haplotype in the Leiden Longevity Study and the Leiden 85-Plus Study

(cid:1)493 T 95 Q T-Q Haplotype Study Study Groups Frequency p Value Frequency p Value Frequency p Value Leiden Longevity Partners of offspring (Controls) 24.7 94.6 19.3 Offspring of nonagenarian sib pairs 27.1 .41* 93.0 .32* 17.5 .61y Nonagenarian sib pairs 26.2 .60z 93.7 .57z 19.5 .93§ Leiden 85-Plus Young Leiden Controls 24.0 93.4 17.9 Octogenarian participants born in Leiden 24.9 .64jj 95.3 .43jj 19.8 .46{

Notes: *p value of the z-test between the offspring of the nonagenarian sib pairs (n ¼ 525) and the partners of the offspring (n ¼ 251) as age- and environment- matched controls. yp value of the odds ratio of the T-Q haplotype in the offspring of the nonagenarian sib pairs (n ¼ 197) compared to the partners of the offspring (n ¼ 251) as age- and environment-matched controls. zp value of the z-test between the nonagenarian sib pairs (n ¼ 379) and the partners of the offspring as young controls (n ¼ 251). §p value of the odds ratio of the T-Q haplotype in the nonagenarian sib pairs (n ¼ 164) compared to the partners of the offspring as young controls (n ¼ 251). jjp value of the z-test between the octogenarians of the Leiden 85-Plus Study (n ¼ 655) and the young Leiden controls (n ¼ 244). {p value of the odds ratio of the T-Q haplotype in octogenarians of the Leiden 85-Plus Study (n ¼ 655) compared to the young Leiden controls (n ¼ 244).

Notes: *p value of the z-test between the offspring of the nonagenarian sib pairs (n ¼

) and the partners of the offspring (n ¼

) as age- and environment- matched controls. yp value of the odds ratio of the T-Q haplotype in the offspring of the nonagenarian sib pairs (n ¼

) compared to the partners of the offspring (n ¼

) as age- and environment-matched controls. zp value of the z-test between the nonagenarian sib pairs (n ¼

) and the partners of the offspring as young controls (n ¼

). §p value of the odds ratio of the T-Q haplotype in the nonagenarian sib pairs (n ¼

) compared to the partners of the offspring as young controls (n ¼

). jjp value of the z-test between the octogenarians of the Leiden

-Plus Study (n ¼

) and the young Leiden controls (n ¼