Statistical analysis of association between longevity and ACE polymorphism.ACE genotypes (DD genotype more frequent in centenarians): p = 0.01 andPp<10- when all genotyped contro! populations are pooled. ACE alleles (D allelemore frequent in centenarians than in controls): p< 0.01 when all genotypedcontrol populations are pooled. Populations are in Hardy-Weinberg equilibriumfor ACE alleles.

APOB polymorphismsme. CANN? .{ 

p<0.01 and p<10° using the pooled controls, Table 2).The odds ratio was 1.9 for individuals with the DD genotypecompared to those with an I allele. Taking this value as ameasure of relative risk, 18.6% of the centenarians couldbe attributable to the DD genotype.

The association of APOE alleles with longevity promptedus to investigate polymorphisms in the apolipoprotein B(ApoB) gene, since ApoE is involved in the metabolism ofApoB-containing lipoproteins’. We typed twopolymorphisms in APOB by DNA amplification" in ourpopulations. A threonine 2488 neutral substitutiondetectable by Xbal and the ins/del polymorphism in thesignal-peptide region, which have been associatedrespectively with significant effects on total cholesterol,LDL-cholesterol and triglyceride levels (for Xbal) andwith the severity of global coronary atherosclerosis (forins/del)'*. For both of these polymorphisms, the allelefrequencies were similar in the centenarian and the controlpopulations (Table 6). 

Associations in a selected subgroup 

A subset of the centenarians included in this study hadbeen independently selected through a different protocol,on the basis of their belonging to long-lived sibships. Thelong-lived sibships were defined as comprising at leasttwo siblings older than 95 years, if female or older than 90,if male. Thirty-five members of long-lived sibships whoall happened to be older than 99 years were pooled withthe directly selected population of centenarians. In thissubgroup, the APOE and ACE genotypes displayed thesame associations with an even more marked trend (Table3).

Other polymorphismsWAT. oe ed ne De eb

We typed random subsamples of our populations at twoother loci. A bi-allelic marker at D4595 on 4p16.3chromosome (the Huntington’s disease region), probe674 with Acdl (ref. 20), gave a frequency of 0.33 for the rareallele in controls (n=158) and centenarians (n=164) anda dinucleotide repeat within the B-cell leukaemia/lymphoma 2 (BCL-2) gene on chromosome 18q21.3 (ref.21) (GenBank accession number: M15701) with onefrequent allele and eight rare alleles gave a frequency of0.825 and 0.838 for the frequent allele in controls (n=103)and centenarians (#=108), respectively. Thus, for both ofthese polymorphisms there was no difference betweencontrols and centenarians.

Gender effects and interaction between lociNo Dk en a 

Our - population of centenarians was mainly composed offemales, its male to female ratio of 1:8 (see Table 1)reflecting the sex differential in longevity in France.  

Table 3 APOE and ACE alleles in centenarians with long-lived siblings

Discussion 

The potential overlap between cases and controls (thefrequency of controls that might become centenarians), issmall enough — less than 2.4% and 0.5% of women andmen respectively at age 70, based on 1980 estimates takenfrom a Caucasian population” — as to remain negligible.The frequencies found in our control group for APOEandACE alleles agree well with those reported in Frenchpopulations from Paris”, Lille and Strasbourg” and (for 

The differences reported in Tables 1 and 2 are all enhanced when controls arecompared with the selected subgroup of centenarians. However, statisticalsignificance is diminished due to the small sample size. P1, population of all thecentenarians. P2, population of controls.

 Table 5 interaction between APOEand ACE in centenariansDD ID ul22 29 (0.115) 35 (0.132) 19 (0.182)63 210 (0.833) 219 (0.823) 78 (0.750)24 13 (0.052) 12 (0.045) 7 (0.067)

functions besides its role on the renin-angiotensin andkallikrein-kinin systems. Its ability to cleave neuropeptidessuch as enkephalin, substance P and LHRH” and itsregional distribution in the brain” point toneuroendocrine functions. Furthermore, an increase, withage, in brain cortical ACE activity” suggests that an adaptiveresponse to increasing needs may occur during ageing.ACE may also function as an immunomodulator byhelping to process endogenous peptides within the MHCclass I complex in cytotoxic T lymphocytes™, and its levelis also associated with the I/D polymorphism in thesecells*®. These data indicate potential roles of ACE, outsidethe cardiovascular system, that may wellinfluence survival.Insofar as the quantity of ACE enzyme is a significantparameter in these other functions, they providea rationalefor understanding the differential influence on survival ofthe ACE-I/D polymorphism. However, the associationwith longevity may also stem from linkage disequilibriumwith another influential polymorphism in a nearby gene.In this regard, itis notable that the gene for human growthhormone, which may have an important role insenescence’, shows strong linkage to ACE, onchromosome 17q23 (ref. 26).Tha came nuectinne arica far Anan which lice in tha

The frequencies of APOE alleles are shown in parenthesisfor each genotype at the ACE locus. The visible trend ofnegative correlation between the e2 and D alleles does notreach significance (p=0.09 for a comparison of the €2 allelefrequency between DD and II genotypes),