Interaction of Sirtuin 1 (SIRT1) candidate longevity gene and particulate matter (PM2.5) on all-cause mortality: a longitudinal cohort study in China

doi:10.21203/rs.3.rs-77022/v2

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Interaction of Sirtuin 1 (SIRT1) candidate longevity gene and particulate matter (PM2.5) on all-cause mortality: a longitudinal cohort study in China

https://doi.org/10.21203/rs.3.rs-77022/v2

This work is licensed under a CC BY 4.0 License

Journal Publication

published 14 Mar, 2021

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Background The SIRT1 gene was associated with the lifespan in several organisms through inflammatory and oxidative stress pathways. Long-term air particulate matter (PM) is detrimental to health through the same pathways.

Methods We used the Chinese Longitudinal Healthy Longevity Survey (CLHLS) to investigate whether there is a gene-environment (G×E) interaction of SIRT1 and air pollution on mortality in an older cohort in China. Among 7,083 participants with a mean age of 81.1 years, we genotyped nine SIRT1 alleles for each participant and assessed PM_2.5 concentration using 3-year average concentrations around each participant's residence. We used Cox-proportional hazards models to estimate the independent and joint effects of SIRT1 polymorphisms and PM_2.5 exposure on all-cause mortality, adjusting for a set of confounders.

Results There were 2,843 deaths over 42,852 person-years. The mortality hazard ratio (HR) and 95% confidence interval (CI) for each 10 μg/m³ increase in PM_2·5 was 1.08 (1.05-1.11); for SIRT1_391 was 0.77 (0.61, 0.98) in the recessive model after adjustment. In stratified analyses, participants carrying two SIRT1_391 minor alleles had a significantly higher HR for each 10 μg/m³ increase in PM_2.5 than those carrying zero minor alleles (1.323 (95% CI: 1.088, 1.610) vs. 1.062 (1.028, 1.096) p for interaction = 0.03). Moreover, the interaction of SIRT1 and air pollution on mortality is significant among women but not among men. We did not see significant relationships for SIRT1_366, SIRT1_773, and SIRT1_720.

Conclusion We found a gene-environment interaction of SIRT1 and air pollution on mortality, future experimental studies are warranted to depict the mechanism observed in this study.

Environmental Engineering

Longevity gene

Air pollution

Sex difference

CLHLS

SIRT1 is a gene well-documented to be associated with aging and longevity[1]. It is encoded by the SIRT1 gene is the most conserved mammalian nicotinamide adenine dinucleotide (NAD+) dependent histone deacetylase, it also has been demonstrated as regulating lifespan in many model organisms such as yeast, Caenorhabditis elegans, and rodents through several biopathways, including anti-inflammation, regulation of metabolism, hypoxic responses, and circadian rhythms [2, 3]. In humans, SIRT1 improves healthy aging and affects human life expectancy through its protective role in a variety of biological processes related to age-related diseases ranging from metabolic disorders, cellular senescence, cardiac aging, oxidative stress, neurodegeneration, inflammatory signaling, and placental cell survival [1, 4]. Human aging is characterized by a chronic, low-grade inflammation level, and ambient air pollution may be involved in developing degenerative diseases such as aging arteries and brains through shared inflammatory pathways [5]. Biologically, SIRT1 can modulate inflammatory genes such as NF-κB and NLRP3 and further leads to delayed onset of age-related symptoms and pathologies [6, 7]. To date, a few observational studies have reported that SIRT1 (i.e., SNP rs7896005, rs12778366, rs4746720) was linked to long-term survival and longevity in the human population, their results were generally restricted by either small population size or cross-sectional design [8-11].

Air pollution is one of the biggest environmental risks to human health, attributing to one in every nine deaths annually, and has been identified as a global priority in the sustainable development agenda [12]. Despite efforts on improving air quality in many cities globally, increasing epidemiological studies reported the association of air pollutants with adverse health outcomes, for example, all-cause of mortality [13, 14]. According to an analysis of data from the 2015 Global Burden of Disease Study, exposure to ambient fine particulate matter (PM_2.5: particulate matter (PM) with aerodynamic diameters<2.5 μm) is the fifth leading risk factor for death, accounting for 7.6% of total global deaths and 4.2% of global DALYs, with China accounting for a large share of these burdens [15]. China’s coal-based energy-intensive development path has led to a steep increase in emissions of PM_2.5 and other pollutants [16], estimated to have led to 1.6 million deaths from heart and lung diseases or stroke, approximately accounting for one in six premature deaths in China [17]. Thus, the comprehensively health impact of air pollutions in Low- and Middle-income countries such as China remains as one of the major health issues, and further attention as well as joint approaches are needed.

Increasingly, we have seen that air pollution does not affect everyone equally, with some populations more susceptible to its detrimental effects. Genetic susceptibility is likely to play a vital role in response to air pollution [18]. Inflammation and oxidative stress are documented to play a role in the mechanistic pathways, including nuclear factor kappa B cells (NF-kB) signaling, Krüppel-like Factor 2 (Klf2) mediated immune response, nuclear factor E2-realest factor 2 (Nrf2)-mediated oxidative stress response, NLR family pyrin domain containing 3 (NLRP3) inflammasome activation, glutathione metabolism, coagulation system, endogenous reactive oxygen species (ROS) production, and other cytokines signaling, between air pollution exposures and adverse health outcome including mortality [19]. Notably, biological studies indicated that SIRT1 can be modulated through most of those pathways, including NF-kB [20]. Klf2 [21], Nrf2 [22], NLRP3 [7], and ROS [23]. Additionally, previous experimental studies observed that air pollution and SIRT1 have an interactive effect on pulmonary diseases [24, 25], cardiovascular diseases [26]. Although the associations between exposure to air pollutants and SIRT1 gene were observed in in vitro studies, less effort has been put forth in the investigation on a population level, especially for the vulnerable older adults. Additionally, epidemiologic evidence and rodent model showed the effect of SIRT1 on disease and longevity vary by inflammatory levels [11, 27] and can be double-edged sword: lower levels of SIRT1 at an early time (short term exposure of toxicants) accentuate acute inflammation-related autotoxicity by increasing NFκB RelA/p65 activity, but prolonged upgrading in SIRT1 during late inflammation are associated with immunosuppression and increased mortality [28]. Given the SIRT1 and PM_2.5 shared several common biological pathways containing inflammation and oxidative stress on mortality and they are tended to interplay with each other [26, 29], we hypothesized a potential synergistic effect between PM_2.5 exposure and SIRT1 polymorphisms on mortality.

To test our hypotheses, we used a nationally representative cohort of individuals aged 65 and older from the Chinese Longitudinal Healthy Longevity Study (CLHLS). First, we aim to estimate the independent and joint effects of SIRT1 polymorphisms and PM_2.5 exposure on all-cause mortality. Second, we take advantage of the sample size to study the interaction effect of SIRT1 and air pollution on mortality. Considering the “male-female health-survival paradox” in longevity research, and there are marked sex differences in genetic associations with longevity; thus we also assessed the three-way interaction by sex.

Study population

We used data from the Chinese Longitudinal Healthy Longevity Study (CLHLS), which are publicly available from Peking University Open Research Data (https://opendata.pku.edu.cn/dataverse/CHADS). The baseline and follow-up surveys were conducted in 1998, 2000, 2002, 2005, 2008–2009, 2011–2012, and 2014 in a randomly selected half of the counties and cities in 23 out of 31 provinces in China. The study was the first national longitudinal survey on the determinants of healthy aging among the oldest old individuals in China. Details of descriptions of the CLHLS including the rationale and design have been described previously [30]. With 631 cities and counties randomly selected as the sample sites, the population in 23 selected provinces represents 85% of the total population in China (Figure 1). CLHLS was approved by the Institutional Review Board, Duke University (Pro00062871), and the Biomedical Ethics Committee, Peking University (IRB00001052–13074). All participants or their legal representatives signed written consent forms to participate in the baseline and follow-up surveys. This study followed the Strengthening the Reporting of Observational Studies in Epidemiology (STROBE) reporting guidelines.

There was a total of 34394 participants in 5 waves of CLHLS, recruited from 2000 to 2011. We excluded 25220 participants without available SIRT1 genotypic data or did not meet the selection criteria of sample filtering, 545 participants aged 64 or younger, 212 participants with missing PM_2.5 values, 567 participants were not of Han ethnicity (according to the ID card or household registry) or having a missing value in covariates; 767 participants lost to follow-up at the first follow-up survey. The samples were required to meet 3 selection criteria: (1) a genotype calling rate >90%; (2) no existing population stratification according to multidimensional scaling (MDS) procedure implemented in PLINK v1.07, based on which individuals deviating from the main population cluster were removed; and (3) no inclusion of duplicates or first-degree relatives when evaluating pairwise through identity by descent (IBD). Accordingly, the final sample that met inclusion criteria for this study was 7083 participants (Figure S1). The sample consisted of 3677 women and 3406 men; 3272 participants were 65 to 79 years of age, 1840 were 80-89 years of age, 1305 were 90-99 years of age, and 667 were 100 years of age or older. To test the possibility of potential selection bias, gender, age, and residence were compared between participants who lost to follow-up (767 participants) or not (7083 participants) at the first follow-up survey; the significant difference for ages (86.0 vs 81.1) and residence (rural: 53.8% vs 66.7%) between two groups, while no significant difference for sex (female: 52.7% vs 51.9%).

Assessment of PM_2.5exposure

We estimated ground-level concentrations of PM_2.5from the Atmospheric Composition Analysis Group based on participants’ residential addresses [31]. It combines remote sensing from National Aeronautics and Space Administration’s Moderate Resolution Imaging Spectroradiometer, Multiangle Imaging Spectroradiometers, and Sea-viewing Wide field-of-view Sensor satellite instruments; vertical profiles derived from the GEOS-Chem chemical transport model; and calibration to ground-based observations of PM_2.5 using geographically weighted regression. Annual PM_2.5 estimates were calculated from 2000 to 2014, at 1 km x 1 km spatial resolution, which was the longest and the highest resolution exposure dataset available [32, 33]. Additionally, our estimations were highly consistent with out-of-sample cross-validated concentrations from monitors (R²=0·81) and another exposure dataset in China (R²=0·79) [31]. A previous study found that the three-year average PM_2.5 before death or the end of the study had the strongest association with mortality among old adults in China [32]. Therefore, we used a three-year average PM_2.5 to reflect ambient air pollution in this study.

The SIRT1 genotypes

We customize an SNP chip containing 27,656 selected longevity and disease-related SNPs for targeted genotyping [34]. We selected candidate SNPs from the public database of the National Center for Biotechnology Information (NCBI; http://www-ncbi-nlm-nih-gov/gene/23411) to cover the SIRT1 gene region in equally spaced intervals [35]. The minor allele frequencies (MAF) of the polymorphisms required to be > 10% [36]. The selected and genotyped SIRT1 SNPs were rs12778366 (promoter), rs3758391 (promoter), rs2273773 (exon), rs2236319 (intron), rs1885472 (intron), rs7069102 (intron), rs10823112 (intron), rs3818291 (intron), and rs4746720 (intron), which have covered major functional SNPs in SIRT1 gene. Among the 9 SNPs, SIRT1_366 (rs12778366), SIRT1_391 (rs3758391), SIRT1_773 (rs2273773), and SIRT1_720 (rs4746720) were annotated and regarded as tagging SNPs due to the high linkage disequilibrium, which were used as proxies for the rest of 5 SNPs in the analysis (Table S1). The Hardy-Weinberg equilibrium of the 9 SIRT1 SNPs was tested with the GENEPOP package (version 1.2). To determine the SIRT1 carrier, we coded the genotype based on the minor allele number [37]. For each SNP, three different inheritance models (additive, dominant, and recessive model) were tested. In the additive model, the genotype that contains zero, one, or two copies of minor allele was coded as 0, 1 or 2; In the dominant model, the genotype that contains at least one copy of minor allele was coded as 1, otherwise it was coded as 0; In the recessive model, the genotype that contains two copies of minor allele was coded as 1 and otherwise it was coded as 0.

All-cause mortality

The primary outcome was all-cause mortality. Mortality information was obtained from the follow-up survey done in 2011 and 2014. The date of death would be validated by death certificates when available - otherwise, the close family member’s report was used.

Covariates

Covariates were chosen as potential confounders between exposures and outcomes or predictors of outcomes. All self-reported information was collected through a face-to-face home interviews by trained research staff members. Interviewees were encouraged to answer as many questions as possible. If they were unable to answer questions, a close family member or another proxy, such as a primary caregiver, provided answers. We included baseline age, gender, marital status, residence, education, occupation, smoking status, drinking status, physical activity, and the wave of first interview as covariates. We classified marital status into two categories: currently married and living with spouse as married, and widowed/separated/divorced/never married/married but not living with spouse as not married. We divided residence into urban and rural areas based on governmental administrative categories. We used the schooling year to evaluate education level. We categorized the occupation into two groups: professional and technical personnel, governmental, institutional, or managerial personnel as non-manual, and agriculture, forest, animal husbandry, fishery worker, industrial worker, and others as manual. We divided the regular exercise, smoking, and alcohol drinking status into three categories: “Current”, “Former”, and “Never”. For example, participants were asked “do you do exercise regularly at present (planned exercise like walking, playing balls, running and so on)?” and/or “did you do exercise regularly in the past?”. We defined the regular exercise status as “Current” for participants who answered “Yes” to the first question, “Former” for those who answered “No” to the first question and “Yes” to the second question, and “Never” for those who answered “No” to both two questions. We categorized the participants into two geographical regions: South China (Guangdong, Guangxi, Hainan, Chongqing, Sichuan, Anhui, Hubei, Fujian, Jiangxi, Jiangsu, Shanghai, and Zhejiang) and North China (Beijing, Shandong, Heilongjiang, Jilin, and Liaoning, Hebei, Henan, Shanxi, Tianjin, and Shaanxi).

Statistical analysis

We used the cox proportional hazard model for every SIRT1 SNP and three-year average PM_2.5 (as continuous variables or category variables by quartiles) separately to evaluate their single effect on mortality. We added the interaction term of SNP and three-year average PM_2.5 in the cox model to investigate the interaction of SIRT1 and three-year average PM_2.5. The genotype can be defined as low-risk genotype (G0) and high-risk genotype (G1) based on the genotypes’ hazard ratio for mortality risk. The three-year average PM_2.5 was discretized into two categories as low PM_2.5 exposure (E0) versus high PM_2.5 exposure (E1) using the median of three-year average PM_2.5 as the cut-off point. The gene effect in different environment exposure and environment effect among participants with different genotypes can be evaluated by comparing the difference among the four categories (G0×E0 stands for low-risk genotype under low PM2.5 exposure, G0×E1 stands for low-risk genotype under high PM2.5 exposure, G1×E0 stands for high-risk genotype under low PM2.5 exposure, and G1×E1 stands for high-risk genotype under high PM2.5 exposure) of the combined term. To investigate the gender-specific G×E effect, we adopt an integrated statistical model of three-way interaction to assess varied effect magnitude in the hazard ratio of mortality between those who have different combinations of sex, SIRT1 genotypes, and exposure to PM_2.5 [38].

We measured the survival time in months from the first interview date to the recorded death date or last interview date. All models adjusted for age, gender, marital status, residence area, education, occupation, smoking status, drinking status, and physical activity. We calculated hazard ratios (HRs) and 95% CIs to indicate the effect magnitude of SIRT1 and PM_2.5 on mortality. We analyzed for effect modification by potential modifier variables, then did stratified analyses by sex, urban or rural residence, financial status, smoking status, and North or South geographical regions.

We did additional analyses to verify the results. We compared baseline characteristics between the included participants and all participants in the five waves of CLHLS to assess the sample's representativeness. We adjusted the regression models by using more informative covariates and by excluding covariates with missing values. We also plotted the geographical distribution of the included participants.

We used R 3.6.1 (R Foundation for Statistical Computing) and SAS university edition to perform all the analyses. All p values were from 2-sided tests and results were deemed statistically significant at p < .05 for all analyses.

Among the 7083 participants, 48.1% were men and 51.9% were women, with a mean (SD) age of 81.1 (11.5) years. More than half of the participants were illiterate (57.1%), residents in rural (66.7%), responded to no engage in regular exercise (64.5%), never smoked (64·0%), and never drank alcohol (68.5%) (Table S2). The mean 3-year PM_2.5 was 51.0 μg/m³ (13.5), with populations in rural regions and southern regions experiencing lower ambient air pollution. Four tag SNPs (SIRT1_366, SIRT1_391, SIRT1_773, and SIRT1_720) were used to represent the 9 SIRT1 SNPs as they had high Linkage Disequilibrium (r² ≥ 0.975; Figure S2). The distributions of the tag SNPs of SIRT1 are similar across varied baseline characteristics including sex, age, and education (Table S2), indicating SIRT1 SNPs were randomly distributed across population characteristics. While participants who were older, illiterate, responded of no regular exercise, reside in northern China tended to live in areas with higher PM_2.5 exposure (Table S2). The final sample has a lower average of age, more likely to be men and rural residents, had a higher proportion of currently married status, than those who were excluded; no significant difference has been detected regarding the education level, exercise habit, smoking and drinking status between the two groups (Table S3).

The mean follow-up time (SD) was 6.1 (3.5) years (range: 0 to 14 years). In the 42852 person-years of follow-up, we saw 2843 mortality events (40.1%) between 2002-2014. The mortality rate was 6.6 per 100 person-years for our entire study population. In the additive model, SIRT1_391, SIRT1_472, and SIRT1_102 minor allele carriers had lower mortality than their counterparts. The recessive model showed similar results to the addictive model, yet no statistical significance was observed in the dominant model (Table 1). We also did not see a significant association of SIRT1_366, SIRT1_773, and SIRT1_720 with mortality.

Table 1. The association between SIRT1 SNPs with mortality

Minor allele copy number	n	HR (95% CI)	p value
Minor allele copy number	n	HR (95% CI)	p value
SIRT1_366 (n=7055)
Additive model
0	5281	Ref
1	1635	1.050 (0.963,1.146)	0.269
2	139	1.209 (0.905,1.615)	0.199
Dominant model
0	5281	Ref
1 or 2	1774	1.059 (0.973,1.153)	0.182
Recessive model
0 or 1	6916	Ref
2	139	1.195 (0.895,1.595)	0.227
SIRT1_391 (n=7077)
Additive model
0	5006	Ref
1	1887	1.023 (0.942,1.111)	0.589
2	184	0.769 (0.603,0.981)	0.035
Dominant model
0	5006	Ref
1 or 2	2071	0.997 (0.92,1.081)	0.947
Recessive model
0 or 1	6893	Ref
2	184	0.764 (0.599,0.974)	0.030
SIRT1_773 (n=7071)
Additive model
0	3795	Ref
1	2743	0.993 (0.919,1.072)	0.854
2	533	0.960 (0.825,1.117)	0.597
Dominant model
0	3795	Ref
1 or 2	3276	0.988 (0.918,1.064)	0.748
Recessive model
0 or 1	6538	Ref
2	533	0.963 (0.831,1.116)	0.616
SIRT1_720 (n=7046)
Additive model
0	2292	Ref
1	3383	1.045 (0.961,1.137)	0.302
2	1371	1.013 (0.910,1.127)	0.814
Dominant model
0	2292	Ref
1 or 2	4754	1.036 (0.957,1.121)	0.383
Recessive model
0 or 1	5675	Ref
2	1371	0.986 (0.898,1.083)	0.774

Note: All models were adjusted for age at baseline, sex, education,

marriage, occupation, residence, exercise, smoking, and alcohol consumption.

In the fully adjusted model, the all-cause mortality hazard ratio (HR) and 95% CI for each 10 μg/m³ increase in PM_2.5 was 1.082 (1.053, 1.111). We also conducted our analysis by quartiles, to account for a possible nonlinear dose-response relationship between PM_2.5 exposure and risk of longevity. Compared with the participants who resided in areas with the lowest quartile of PM_2.5, the HR of mortality for 2nd quartile, 3rd quartile, and 4th quartile were 1.203 (1.078, 1.343), 1.431 (1.287, 1.593), 1.297 (1.167, 1.442), respectively (Table 2). In the stratified analyses by tags of SIRT1 genotypes, participants who carry zero or one SIRT1_391 minor allele had an HR of 1.078 (95% CI: 1.049,1.107) on mortality for each 10 μg/m³ increase in PM_2.5, while the HR was 1.336 (95% CI: 1.079, 1.653) in those who carry two SIRT1_391 minor alleles (data not shown).

Table 2. The association between three-year average PM_2.5 with mortality

PM_2.5	n	HR (95% CI)	p value
Quartiles of PM_2.5
Quartile 1	1,786	1.00 (reference)	/
Quartile 2	1,761	1.203 (1.078, 1.343)	0.000939
Quartile 3	1,765	1.431 (1.287, 1.593)	4.42E-11
Quartile 4	1,771	1.297 (1.167, 1.442)	1.37E-06
10-μg/m³ change in PM_2.5	/	1.082 (1.053, 1.111)	1.20E-08

Note: All models were adjusted for age at baseline, sex, education, marriage, occupation,

residence, exercise, smoking, and alcohol consumption.

According to an addictive model with interaction terms of SIRT1 SNPs and PM_2.5, the detrimental effect of PM_2.5 exposure on mortality for the participants carrying two SIRT1_391 minor alleles was stronger than those carrying zero alleles. Similar results were found in the SIRT1_472 and SIRT1_102, of which were in LD with SIRT1_391. For instance, the test for the interaction between SIRT1_391 and PM_2.5 in additive models revealed a significant gene-environment interaction (p for interaction=0.03). The HR of 10-μg/m³ 3-year average PM_2.5 on mortality for the participants carrying two SIRT1_391 minor alleles was e^(0.06+0.220)=1.323 (95% CI: 1.088, 1.610) and for those carrying zero minor allele would be e^0.06= 1.062 (1.028, 1.096) (Table 3). We also tested the interaction effect of SIRT1 SNPs and PM_2.5 in the recessive model, the results are not meaningfully different from findings based on the additive model (Table S4). As for the dominant model, we found there was on difference of PM_2.5 on mortality between participants carrying one or two alleles and those carrying zero allele (Table S5).

Table 3. The interaction between PM_2.5 and SIRT1 SNPs (additive model) on mortality

Additive model	Model without interaction term		Model with interaction term
	HR (95% CI)	p value	β	SE	p value
Carrying SIRT1_366 minor allele status
0 copy	Ref	/	/	/	/
1 copy	1.059 (0.970,1.155)	0.200	0.248	0.171	0.147
2 copies	1.245 (0.931,1.663)	0.139	0.011	0.545	0.984
10-μg/m³ unit of PM_2.5	1.083 (1.054,1.113)	7.04E-09	0.088	0.016	3.50E-08
Interaction term
zero SIRT1_366 minor allele×PM_2.5	/	/	/	/	/
one SIRT1_366 minor allele×PM_2.5	/	/	-0.037	0.032	0.249
two SIRT1_366 minor alleles×PM_2.5	/	/	0.043	0.105	0.685

Carrying SIRT1_391 minor allele status
0 copy	Ref	/	/	/	/
1 copy	1.026 (0.944,1.114)	0.548	-0.255	0.167	0.127
2 copies	0.768 (0.602,0.980)	0.034	-1.452	0.577	0.012
10-μg/m³ unit of PM_2.5	1.082 (1.053,1.112)	1.07E-08	0.06	0.016	2.64E-04
Interaction term
zero SIRT1_391 minor allele×PM_2.5	/	/	/	/	/
one SIRT1_391 minor allele×PM_2.5	/	/	0.054	0.031	0.081
two SIRT1_391 minor alleles×PM_2.5	/	/	0.220	0.101	0.030

Carrying SIRT1_773 minor allele status
0 copy	Ref	/	/	/	/
1 copy	1.001 (0.926,1.081)	0.986	0.109	0.153	0.478
2 copies	0.951 (0.817,1.106)	0.513	-0.379	0.318	0.234
10-μg/m³ unit of PM_2.5	1.082 (1.053,1.112)	1.19E-08	0.083	0.019	1.33E-05
Interaction term
zero SIRT1_773 minor allele×PM_2.5	/	/	/	/	/
one SIRT1_773 minor allele×PM_2.5	/	/	-0.021	0.029	0.464
two SIRT1_773 minor alleles×PM_2.5	/	/	0.06	0.056	0.286

Carrying SIRT1_720 minor allele status
0 copy	Ref	/	/	/	/
1 copy	1.044 (0.960,1.135)	0.319	0.381	0.168	0.023
2 copies	1.001 (0.899,1.113)	0.992	0.201	0.223	0.366
10-μg/m3 unit of PM_2.5	1.081 (1.052,1.111)	1.93E-08	0.117	0.024	1.45E-06
Interaction term
zero SIRT1_720 minor allele×PM_2.5	/	/	/	/	/
one SIRT1_720 minor allele×PM_2.5	/	/	-0.065	0.031	0.036
two SIRT1_720 minor alleles× PM_2.5	/	/	-0.038	0.041	0.345

Note: All models were adjusted for age at baseline, sex, education, marriage, occupation, residence, exercise, smoking, and alcohol consumption.

Further, we conducted a three-way interaction analysis to examine whether there is a sex difference in the effect of G×E interaction. Figure 2 demonstrates exposure to higher PM_2.5 does not affect mortality significantly in male and female who carry zero or one SIRT1_391 allele, but PM_2.5 exposure substantially increases the mortality risk among male and female carriers of two SIRT1_391 alleles. It is noted that among the participants with exposure to the high concentration of PM_2.5, females carry two SIRT1_391 alleles have a significantly excess risk of mortality than those with zero or one SIRT1_391 allele; but the interaction is not significant among male participants. Specifically, in females who carrying two SIRT1_391, exposure to higher level of PM_2.5 is associated with a 47.7% higher risk of mortality, while the effect size is 16.8% in men.

Our study indicated/replicated the detrimental effect of air pollution on all-cause mortality. We found there was a protective effect of some alleles of SIRT1 on all-cause mortality, in concurrence with several prior animal studies and population studies. An added evidence of our study is finding an interaction between PM_2.5 induced premature mortality and carriers of SIRT1_391, SIRT1_472, and SIRT1_102 genotypes, which carriers of 2 alleles of SIRT1_391 can counteract the detrimental effect of PM_2.5 and reduce 26.1% risk of premature mortality. Although none of the association between SNPs and mortality would be statistically significant after the most stringent Bonferroni correction (α=0.05/4≈0.0125), we used candidate SNPs tested in previous studies instead of random SNPs so that the need for stringent correction may not be necessitated. Our study used a large old cohort to better elucidate evidence of several cellular and molecular pathways studies on the protective role of SIRT1. Prior studies indicate SIRT1 is involved in the pathway of PM-induced airway inflammation with the activation of SIRT1 to prevent airway disorders [24, 25]. On the cellular and molecular side, using in vivo models of airway inflammation and in vitro culture of human bronchial epithelial (HBE) cells exposed to PM_2.5 and resveratrol (SIRT1 activator), SIRT1 expression was decreased in HBE cells and lung tissues after PM_2.5 exposure, suggesting that SIRT1 is involved in the pathogenesis of PM-induced airway inflammation [29]. Second, a plethora of animal studies investigating PM_2.5 haas documented pathways of induce oxidative stress to trigger inflammation and thrombosis [39]. It was hypothesized that SIRT1 as a member of class III histone deacetylase, controls lung inflammation and coagulation after PM_2.5 exposure. SIRT1 knock-out mice exhibited aggravated lung vascular leakage and inflammation after PM_2.5 exposure, correlated with increased NF-κB acetylation and activation. This indicates that SIRT1 functions as a suppressor of coagulation after particulate matter exposure [29]. Further molecular evidence also implicates the role of SIRT1 in protein/histone deacetylase and as a protective factor of the development of pulmonary emphysema [40]. PM_2.5 exposure in animal models was also demonstrated to introduce the repression activity of SIRT1 in the mice liver [41].

Our study builds on these animal models and is the largest population-based study on the role of SIRT1 in air pollution-induced mortality. Previously, a series of genes involved in oxidative stress and inflammatory pathways were studied for interaction with air pollutants, including GSTM1, GSTP1, NQO1, and TNF. There were both positive and null findings [42]. Studies indicate genetic susceptibility is likely to play a role in response to air pollution, especially on cardiovascular and respiratory outcomes [29]. A recent review indicated that SIRT1 plays in toxicological damage caused by environmental toxicants such as PM_2.5, the PM-induced injury affect SIRT1 expression, which then affects the expression and activity of downstream proteins, resulting in toxic damage [43]. In addition, one molecular biology finding suggested that PM_2.5 can upregulate MicroRNA-146a-3p and induces the inflammatory macrophage polarization by targeting SIRT1 [44]. A recent population study also found that exposure to long-term air pollution, even at a low level, can alter gene expression and in turn to impact individual’s health [18]. Various studies have indicated that aging and lifespan are modulated by genetic and environmental factors as well as their interactions. Extant basic and translational studies have reported the interaction effects of air pollution and SIRT1 on the incidence or progression of pulmonary diseases [25, 29, 45] and cardiovascular diseases [26]. As we do not have a complete understanding of the synergistic health effects of air pollution and SIRT1 in humans, and prior studies are mainly focused on animal models, our finding of an interactive effect needs validation.

We also observe significant gender differentials in the interaction effect of PM_2.5 and SIRT1 genotypes on mortality. The association of PM_2.5 exposure with a reversal of the negative effects of carrying two SIRT1_391 alleles on mortality is much stronger in females than in males. In females who carrying two SIRT1_391, exposure to higher level of PM_2.5 is associated with a 47.7% higher risk of mortality, while the effect size is 16.8% in men. Underlying mechanisms gender difference may be due to sex hormones and distinct innate immune systems [46]. First, estrogen can downregulation of SIRT1, which in turn reduces the anti-oxidative anti-inflammatory effect on PM-induced toxicants [47]. Although production of estrogen declined dramatically in females after menopause status, the estrogen remained at moderately low level. SIRT1, possibly through the AKT and ERK signaling pathways, plays a crucial role in estrogen in protecting arteries from senescence and atherosclerosis [48]. Resveratrol also functions as a suppressor of PM-induced inflammatory signaling pathways by inhibiting COX-2/PGE2 expression [49]. Second, sex-specific analyses of longevity gene indicate “innate immune system in men and of the tryptophan and PGC-1 pathways in the regulation of immune-related pathways in women suggests that women and men have different approaches for longevity, in which the SIRT1 deacetylates PGC-1α and enhances PGC-1α activity, insufficient NAD⁺ availability and SIRT1 enzymatic activity may be contributing factors [50]. Moreover, another sex-specific study on human heart reveals a female sex-specific downregulation of SIRT1 and SIRT3 in aged hearts, as well as a decline in mitochondrial anti-oxidative defense and a pro-inflammatory shift in old female hearts but not in male hearts. The ex-specific downgrade of SIRT1 in females than in males might predispose females in a more susceptible condition when exposed to PM-derived toxicants [51].

Our study has several limitations. The biggest limitation is that our cohort did not ascertain reliable cause-specific mortality which means we could not determine the possible pathways of air-way induced mortality. Second, we used satellite-derived residential area-level PM_2.5 measurement, which has been shown to be reliable but may not be as precise as ground-level monitors and personal monitors. However, we do not expect differential misclassification that biases our effect estimates. In addition, we used 3-year average PM_2.5, so we can only study long-term air pollution difference. Third, our study did not have biomarkers or dietary determinants of nicotinamide mononucleotide (NMN), an NAD+ precursor, to assess SIRT1 activation activity; however, we do not expect these factors to varing with air pollution levels. Lastly, we also did not have biomarker data or clinical diagnosis of pulmonary function of the participants to ascertain differences based on SIRT1 genotype status. Thus, our study cannot distinguish the possible mechanism of the interaction between SIRT1 and air pollution on neuroprotection, metabolism, and cell survival. Lastly, we currently do not possess epigenetic markers in this cohort to measure gene expression.

Our study has many strengths. First, it is one of the largest population studies on the role of SIRT1 in human populations. Second, we had over one decade of follow-up on the survival status of participants. We had a highly diverse group of individuals from within China, including all climatic and geographical regions covering 23 provinces. Third, as our cohort was designed to study socioeconomic determinants of health, we had a wide range of possible confounders to adjust for. Fourth, conducting this study in a developing country, we had high heterogeneity of a range of low and high ambient air pollution levels so that we can see a dose-response relationship. Lastly, a relatively large sample size gave us statistical power for interaction variables and stratified analyses.

In conclusion, our study suggested a gene-environmental interaction of SIRT1 genotype and air pollution in a large population-based cohort, provided epidemiological human evidence to validate the many animal studies. If SIRT1 does protect against PM_2.5 exposure, those living in areas with high levels of air pollution may benefit from induced SIRT1 activity through supplementation or interventions. Our findings indicate future clinical trials are needed to validate this hypothesis. If successful, this may be public health and clinical intervention through targeting mechanisms to protect against air pollution insult on the human body.

Availability of supporting data

The data that support the findings of this study are available from the corresponding author, upon reasonable request.

Ethics declarations

Ethics approval and consent to participate

CLHLS was approved by the Institutional Review Board, Duke University (Pro00062871), and the Biomedical Ethics Committee, Peking University (IRB00001052–13074). All participants or their legal representatives signed written consent forms to participate in the baseline and follow-up surveys. This study followed the Strengthening the Reporting of Observational Studies in Epidemiology (STROBE) reporting guidelines.

Consent for publication

Written informed consent for publication was obtained from all participants.

Competing interests

None reported

Funding

This work was supported by Bill & Melinda Gates Foundation, National Institute of Health of the United States (2P01AG031719), National Key R&D Program of China (2018YFC2000400) and National Natural Sciences Foundation of China (71490732, 81903392, 81941021), China Postdoctoral Science Foundation funded project (2019M650359).

Acknowledgements

We are grateful to the CLHLS study, which provided the data in this research. The CLHLS was supported by funds from the US National Institute on Aging (NIA), the China Natural Science Foundation, the China Social Science Foundation, and the United Nations Fund for Population Activities (UNFPA) and was managed by the Center for Healthy Aging and Development Studies, Peking University.

Author information

Yao Yao, MD,¹ Linxin Liu, MMed,² Guang Guo, PhD,³ Yi Zeng, PhD,^1,4 John S. Ji, ScD^2,5

Center for Healthy Aging and Development Studies, National School of Development, Peking University, Beijing, China

E-mail: [email protected]

Environmental Research Center, Duke Kunshan University, Kunshan, China

Email: [email protected]

Department of Sociology, Carolina Population Center, and Carolina Center for Genome Sciences, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA

Email: [email protected]

Center for the Study of Aging and Human Development, Duke Medical School, Durham, NC, USA

E-mail: [email protected]

Nicholas School of the Environment, Duke University, Durham, NC, USA

E-mail: [email protected]

Contributions

Yao Yao drafted the original manuscript, made review and edit. Linxin Liu performed the analysis, made review and edit. Guang Guo and Yi Zeng made review and edit. John S. Ji conceptualized the idea, designed the methodology, made review and edit.

Corresponding author

Correspondence to John S. Ji

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Interaction of Sirtuin 1 (SIRT1) candidate longevity gene and particulate matter (PM2.5) on all-cause mortality: a longitudinal cohort study in China

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