The omnibus haplotype test showed marginal associ- ation with longevity (overall P 5 0.019). Of the 6 different haplotypes with a frequency greater than 3% for the 3 variants, haplotypes A-G-C and A-C-T were

RESULTS

Table I shows the basic characteristics of 191 long- lived people (mean age, 97 years) and 53 short-lived

III. The haplotype frequencies of the 3 variants in Hsp70 family between long-lived people and as well as the relative contribution of each haplotype to longevity

Haplotypes Long-lived people Short-lived people c2 P* OR (95% CI); P† A-G-T 0.309 0.367 1.28 0.258 Ref G-C-T 0.279 0.343 1.64 0.200 0.95 (0.56,1.61); 0.843 G-G-T 0.124 0.134 0.08 0.785 1.08 (0.55,2.23); 0.827 A-G-C 0.114 0.041 4.99 0.026 3.46 (1.29,12.05); 0.025 A-C-T 0.110 0.034 5.66 0.017 4.51 (1.53,19.34); 0.016 A-C-C 0.030 0.052 1.19 0.027 0.71 (0.24,2.37); 0.546

Abbreviations: CI, confidence interval; OR, odds ratio. The alleles in haplotype are in the order of A1267 G, G190C, and T2437C variants. *P was calculated using a c2 test from a series of 232 contingency tables by combining the other haplotypes listed. †OR (95% CI); P values were calculated by logistic regression analysis using the most common haplotype A-G-T as a statistical

Table IV. The age distributions by genotype stratification of the 3 studied variants among long-lived people

A1267 G AA (n 5 65) AG (n 5 85) GG (n 5 41) P trend Age, years 94.51 6 4.97 95.29 6 5.61 95.42 6 5.78* 0.020 G190C GG (n 5 58) GC (n 5 97) CC (n 5 36) P trend Age, years 95.11 6 4.81 96.26 6 5.89* 94.21 6 5.03 0.054 T2437C TT (n 5 130) TC (n 5 54) CC (n 5 7) P trend Age, years 98.29 6 6.40† 93.80 6 4.41 95.94 6 5.72* 0.020

†P.0.01: for A1267 G compared with AA; for G190C compared with CC; for T2437C compared with TC.

Because the G190C variant was located in the pro- moter region of HSPA1A, we assessed the promoter activity by measuring the ratio of firefly luciferase activ- ity to Renilla luciferase activity in A549 cells (Fig 2). No significant differences were observed between 190 G and 190C alleles in both control (P 5 0.562) and heat- inducible (P 5 0.463) A549 cells.

significantly higher in long-lived people than short-lived people (P 5 0.026 and P 5 0.017), whereas haplotype A-C-C was significantly lower (P 5 0.027) (Table III). When using the most common haplotype A-G-T as a sta- tistical reference, haplotypes A-G-C and A-C-T had a significant 3.46- and 4.51-fold increased tendency for longevity, respectively (P 5 0.025 and P 5 0.016, re- spectively).

Table IV compares the age distributions by Hsp70 genotypes among long-lived people. Compared with 1267AA homozygotes, carriers with 1267GG genotype had a tendency of living longer (P . 0.05). The same result was noted for 190GC heterozygotes in comparison with 190CC genotype carriers. As for T2437C variant, the different allele carriers (2437TT and CC) both had an increased tendency for longevity, especially for the

DISCUSSION

It is generally believed that all organisms at the cellu- lar level respond to stress by synthesizing heat shock proteins at the expense of other proteins and the ability of human cells to respond to heat stress decreases with aging.14,15 The possibility could not be excluded that longevity might be partly heat-stress mediated. There- fore, proteins that participate in heat-stress cascade and its regulation are potential candidates, whose encoded genes may contain sequence variation contributing to the inherited tendency for longevity. We performed a candidate gene-longevity association study as well as the promoter activity assessment to explore the possible involvement of Hsp70 genes variants in susceptibility to longevity in a Xinjiang Hetian Uygur population. The Hetian city was listed as 1 of the 4 longevity districts in the world by the Institute of International Natural Medicine.16 This special population can offer us an inter- esting opportunity to establish an ideal model to decipher genetic underpinnings of natural longevity.

ing the amount of information gained about longevity status by putting 2 variants together using the MDR function. The interaction information analysis indicated that variants G190C and T2437C, connected with a red line, had a strong synergistic effect, whereas the A1267 G variant was connected with the other by a green line, which suggests a loss of information that can be inter- preted as redundancy or correlation. The pattern of ‘‘high-risk’’ (dark gray) and ‘‘low-risk’’ (light grey) genotype combinations was nonlinear across the 9 two-locus genotype cells, which represented interaction as suggested by the interaction analysis.

Synergy A1267GAdditivityRedundency G190CT2437CWeak StrongeeInteractionG190Ccc GC GG T2437C    

results of Ross et al’s One possibility is based on the different selection criteria applied to study groups. For example, in Ross et al’s study,18 the short-lived people aged 19–45 years were chosen at random from the archival DNA bank of normal healthy Caucasian in- dividuals, whereas our short-lived people aged 65–70 years were selected on the basis of many years’ fol- low-up and stringent entrance criteria (as described in Methods). Another possibility is the relative small num- ber (n 5 53) of short-lived people in the current study, which might cover the statistical significance between genetic markers and longevity. We agree that confirma- tion in a larger, well-designed study is crucial.