) 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

). {p value of the odds ratio of the T-Q haplotype in octogenarians of the Leiden

-Plus Study (n ¼

) compared to the young Leiden controls (n ¼

).

-493G/T‘b$20 oso sz0Ainiqeqosd Jeans 00'0100 105 1109585Age (years)Q95H HlsZz0 0s'0 scoAyiigeqoid |EAIAINS    00090 95 100 105 11085Age (years)

of the studies. This finding may indicate that the U.S. first and second tier are confounded by population stratification. It should be noted that Geesaman and colleagues found an 18% frequency of the associated haplotype in the U.S. centenarian cases of the Boston study (8) and a 29% frequency in their young U.S. control group. In all published data (28,29) as well as in the French centenarians and controls (8), this haplotype frequency is approximately 19%. Also, the MTP allele frequencies found in the Framingham Offspring study, a Caucasian U.S. population, were similar to those in European Caucasians (32). Among all pub- lished MTP allele and haplotype frequencies (11– 13,17,28,29,32,33), only the U.S. control groups show an overrepresentation of the associated alleles and haplotype.

It is, therefore, likely that the positive association found in the two U.S. tiers is due to population stratification in the control groups. The long-lived U.S. cases reflect the ethnic distribution of the United States near the beginning of the 20th century, whereas the U.S. control population reflects more recent generations (34). To minimize the problem of admixture, Geesaman and colleagues selected only the Caucasian individuals, and they proactively matched the controls to the long-lived cases using the genotypes of 60 random SNPs. However, moderate levels of stratification may remain undetected when a small number of SNPs (50– 100) are investigated (35). This implies that, even after proactively matching, residual population stratification may still have been present in the U.S. control data set (36). Because our meta-analysis including the U.S. proactively matched sample showed that this putative MTP risk haplotype has no effect on human longevity in Caucasian populations, we conclude that it is more likely that the association found by Geesaman and colleagues has its source in the stratification of (especially) the control group of the U.S. population rather than in the genetic effect of the locus on exceptional longevity.

Conclusion

Considering the absence of linkage in our study, we find it unlikely that chromosome 4q25 harbors a gene explaining the familial effects on Dutch nonagenarian longevity. Furthermore, the current evidence does not support the MTP gene at chromosome 4q25 to be the major longevity gene explaining the previously detected linkage to excep- tional longevity.

Figure 2. Kaplan–Meier survival curves per genotype in the two combined cohorts of the Leiden 85-Plus Study for the (cid:1)493G/T and Q95H microsomal transfer protein (MTP) gene single-nucleotide proteins (SNPs).

Because the first cohort of the Leiden 85-Plus study was followed for 14 years and the second cohort for 7 years, we were also able to analyze the effect of the MTP SNPs prospectively. None of the genotypes of the (cid:1)493G/T or Q95H MTP SNPs showed an effect on either overall mortality or on specific causes of death (such as cardiovas- cular disease, which would be the most likely cause of death when MTP would have an effect on mortality). This result is in accordance with the Danish longitudinal results (29).

ACKNOWLEDGMENTS

This study is supported by a grant from Innovation Oriented research Program IOP on Genomics (SenterNovem; IGE01014) and by the Centre for Medical Systems Biology (CMSB), a center of excellence approved by The Netherlands Genomics Initiative/Netherlands Organisation for Scientific Research (NWO).

We appreciate the great effort of Michiel Leijnen and J. Kate van Duijn in genotyping the microsatellite markers and the MTP SNPs in samples of the Leiden Longevity Study and the two cohorts of the Leiden 85-Plus Study. We thank Bas Heijmans for his inspiring comments.

To deal with the discrepancies reported in the literature, we performed a meta-analysis on all published data on the association of the MTP T-Q haplotype and longevity. The meta-analysis including the first and second tier of the U.S. samples showed significant heterogeneity. When, however, the U.S. proactively matched cases and controls were

Address correspondence to Marian Beekman, PhD, Leiden University Medical Center, Section of Molecular Epidemiology, P.O. Box 9503, 2300 RA Leiden, The Netherlands. E-mail: M.Beekman@LUMC.nl

US proactively matchedFranceGermanyDenmarkLeiden Longevity Leiden 85-plusPooled Odds Ratio   0.6T081.0Odds Ratio

from http://biomedgerontology.oxfordjournals.org/ by guest on June 6, 2016

Figure 3. Meta-analysis of the effects of the microsomal transfer protein (MTP) gene T-Q haplotype on longevity. The size of the black square indicates the size of the study.

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