Time was defined from the examination in 1989/1990 until death, end of follow-up in 2008 or last registration if subjects were lost to follow-up. Survival curves are depicted based on these Cox models.

Stratified analyses according to gender, smoking habits (never smokers vs. ever smokers), BMI, and age (dichotomized based on a median age at visit in 1989/1990, i.e., 52 yrs) were performed. Subjects with BMI $25 kg/m2 were categorized into the overweight/obese group according to World Health Organization (WHO) criteria.

The 3 tested SNPs (rs12778366, rs7069102, and rs2273773) were in Hardy-Weinberg equilibrium. Among subjects who died, 83% had the wild type genotype of rs12778366, which is significantly higher (p=0.015) than the 76% in the subjects who were still alive.

Cox regression showed that carriers of the minor allele of rs12778366 had a significantly reduced risk of mortality compared to wild types: HR 0.69 (95% CI: 0.50 to 0.96; p=0.025; see Table 2). Survival curves according to genotypes of rs12778366 clearly show the difference in mortality risk (Figure 1a). The same directions of the effect of rs12778366 were observed within groups that have different mortality risks, i.e., females (HR=0.82 (0.50-2.1.35)) and males (HR=0.63 (0.41-2.0.96)), never smokers (HR=0.53 (0.25-2.1.11)) and ever smokers (HR=0.75 (0.52-2.1.08)), younger subjects (age #52 yrs) (HR=0.68 (0.30-2.1.54)) and older subjects (age >52 yrs) (HR=0.69 (0.48-2.0.98)), (Table 3 and Figure 1 bc, e).

Remarkably, the protective effect of rs12778366 was observed in overweight/obese subjects (HR=0.62 (0.43-2.0.91)) but not in subjects with normal weight (HR=0.95 (0.50-2.1.80). The survival curves clearly show that overweight/obese minor allele carriers of rs12778366 had survival comparable to subjects with normal weight while overweight/obese subjects with the wildtype genotype had an increased mortality risk (Figure 1d). The 2 other SNPs did not show significant associations between genotypes and mortality risk (Table 2).

Glucose Tolerance Test

A linear regression model adjusted for age at the measurement was used to evaluate the associations between SNPs in SIRT1 and glucose tolerance. P values <0.05 were considered statistically significant (tested 2-sided). All statistical analyses were performed using SPSS version 18.0 for Windows.

We analyzed the available data from 535 male subjects from the current study (aged at the measurement 18-61 years) who underwent the glucose tolerance test (OGTT). We foundThe advantage of our current study is that we did not use a selection criteria based on age but investigated the whole population in a longitudinal manner. Actually, up till now, only one study in humans showed associations between SIRT1 variants and healthy aging, which the authors defined as normal brain function and verbal fluency test; laboratory findings for hemogram, peripheral smear, urine, electrolytes, chest X-ray, kidney, pulmonary function, echocardiography, and ECG were normal at the age of 60 or higher [32]. In the light of recent uncertainty about the role of sirtuins in longevity [1], our results importantly provide new evidence in favor of a role of the gene in longevity.

Strengths and limitations:

The major strength of the current study is the longitudinal design. We were able to follow participants for 18 years, providing a wide time window for evaluating survival in the cohort. A strength of our study is also the number of subjects (n = 1,390), sampled from the general population. Additionally, the high follow-up rate is a major strength of the study since 98.6% of the included subjects could be traced back. A limitation of our study is the limited data on metabolic profile because the oral glucose tolerance test was performed only in male subjects and was not accompanied by insulin measurements. Furthermore, the study population consisted of white individuals of Dutch descent which limits extrapolation to other ethnic groups.

In summary, this is the first study showing that SIRT1 plays a role in human lifespan in a non-selected general population cohort. The importance of SIRT1 is supported by the association of its polymorphism with long-term survival in the general population. Furthermore, linking SIRT1 polymorphisms to improved glucose tolerance stresses the impact of SIRT1 on metabolism in humans and identifies SIRT1 as a possible candidate for therapeutic purposes.

We have shown an association between the SIRT1 gene and long-term survival among minor allele carriers of SNP rs12778366 in SIRT1 in the total population. Furthermore, the directions of the effect did not change in stratified analyses according to gender, smoking habits, and age. Interestingly, stratification according to BMI showed the protective effect of rs1277836 only in overweight/obese subjects. Taking into account the increased mortality per se in obese subjects, this finding may shed new light on obesity-related burdens.

Supporting Information:

One of the physiological pathways through which SIRT1 may affect longevity might be glucose homeostasis. Indications suggesting a role of this pathway were backed up by evidence that minor allele carriers of SNP rs12778366 had better glucose tolerance as determined by the oral glucose tolerance test. Interestingly, stratified analysis according to BMI showed that the effect was more pronounced in overweight/obese subjects, whereas in subjects with normal weight, only the direction of the effect was observed.

Author Contributions:

Prepared the mortality data for analysis: JMV. Obtained the grants for the study: JMV HMB. Discussed results, proposed corrections and approved the final version of the manuscript: SMF JMV HMB. Conceived and designed the experiments: SMF JMV HMB. Performed the experiments: SMF. Analyzed the data: SMF. Wrote the paper: SMF.

References:

Accili D, de Cabo R, Sinclair DA () A certain role in longevity. Nat Med :