The Journals of Gerontology Series A: Biological Sciences and Medical Sciences 62:507-511 (2007)
© 2007 The Gerontological Society of America
Association of Interleukin-19 Gene Polymorphisms With Age
Naoko Okayama,
Yutaka Suehiro,
Yuichiro Hamanaka,
Junji Nakamura and
Yuji Hinoda
1 Department of Laboratory Medicine, Yamaguchi University Graduate School of Medicine, Japan.
2 Division of Clinical Laboratory, Yamaguchi University Hospital, Japan.
Address correspondence to Yuji Hinoda, MD, PhD, Department of Laboratory Medicine, Yamaguchi University Graduate School of Medicine, 1-1-1, Minami-Kogushi, Ube, Yamaguchi 755-8505, Japan. E-mail: hinoda{at}yamaguchi-u.ac.jp
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Abstract
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We previously demonstrated a significant association of the interleukin-10 (IL-10) –819 T/C polymorphism with age. IL-19 stimulates the production of IL-10, and the IL-19 gene is located adjacent to the telomere side of the IL-10 gene. To explore the relationship between IL-19 single nucleotide polymorphisms (SNPs) and age, we genotyped 500 Japanese individuals (mean age: 56.7 years, range 19–100) for IL-19 Ser175Phe (T/C), –513 T/C, 1098 G/T (intron 1) and 5420 G/C (5'-untranslated region). Three of four SNPs (Ser175Phe, –513 T/C and 1098 G/T) exhibited a weak but significant association with age by chi-square test and logistic regression analysis (p <.05). IL-19 Ser175Phe was in linkage disequilibrium with –513 T/C and 1098 G/T, but not with IL-10 –819 T/C. These data suggest that IL-19 polymorphisms may be associated with age in a Japanese population.
A previous investigation by our group demonstrated that the interleukin-10 (IL-10) promoter polymorphisms –819 T/C and –592 A/C were associated with age in a Japanese population (1). However, it remains unknown whether these polymorphisms are susceptibility loci for aging or are merely markers for a nearby susceptibility gene. The IL-19 gene is located adjacent to the telomere side of the IL-10 gene on 1q32 and belongs to the IL-10 family (2). IL-19 is produced by monocytes, especially under activated conditions, and to a lesser extent by B cells (3,4). Although little is known about the biological function of IL-19, it has been demonstrated that IL-19 stimulates the production of IL-6 and tumor necrosis factor-alpha (TNF-
) from monocytes in vitro (5), suggesting that IL-19 might be a proinflammatory cytokine. On contrast, inflammation is now considered to be an important risk factor for aging (6,7). Of note, the increased plasma levels of IL-6 and TNF- may be strong predictors of all-cause mortality risk in elderly cohorts (2). If IL-19 plays a role in the regulation of IL-6 and TNF- in vivo, individual differences in IL-19 production may be connected with the aging process. In this study, we evaluated the relationship between four IL-19 polymorphisms, including a nonsynonymous single nucleotide polymorphism (SNP), with age and the linkage disequilibrium (LD) of these SNPs with IL-10 –819 T/C.
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PARTICIPANTS
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Peripheral blood samples were obtained from 500 unrelated Japanese individuals (252 men, 248 women; mean age: 56.7 years, range 19–100 years) after obtaining written informed consent. Number and sex ratio (male/female) of the 500 participants in each decade were as follows: 5 (0/5) in 10s, 133 (70/63) in 20s, 39 (23/16) in 30s, 43 (22/21) in 40s, 30 (11/19) in 50s, 28 (22/6) in 60s, 79 (57/22) in 70s, 100 (35/65) in 80s, 42 (12/30) in 90s, and 1 (0/1) in 100s. Among these, 337 were healthy volunteers including students, hospital staff members, and elderly people who visited the Yamaguchi University Hospital for regular physical examinations (187 men, 150 women; mean age, 44.3 years; range 19–98 years), and 163 were patients treated in the Hospital for a variety of chronic disorders including ischemic heart disease, essential hypertension, diabetes mellitus, peptic ulcer, and liver disease (65 men, 98 women; mean age, 82.3 years; range, 64–100 years) from January 2001 through October 2004. Patients with malignancy or with severe organ failure were not enrolled in this study. Participants were drawn from Yamaguchi Prefecture and the surrounding Prefectures assuming similar environmental and social factors, and all were native Japanese. The experimental protocol was approved by the institutional ethics committee of the Yamaguchi University Graduate School of Medicine.
Genotyping
Genomic DNA was extracted from peripheral blood, treated with EDTA-2Na as an anti-coagulant, using a conventional NaI method (8). IL-19 –513 T/C (rs1028181), 1098 G/T (intron 1) (rs4845143), and 5420 G/C (5'-untranslated region) (rs2243158) were genotyped using a TaqMan assay, and Ser175Phe (T/C) (rs2243191) was genotyped by polymerase chain reaction–restriction fragment length polymorphism (PCR–RFLP), as described previously (1). For genotyping of IL-10 –819 T/C, tetra-primer amplification refractory mutation system (ARMS) PCR was performed with validation by PCR–RFLP, as described previously (9). The primers, probes, and restriction enzymes used in this study are shown in Table 1.
Statistical Analysis
The occurrence of the four SNPs tested in this study was consistent with Hardy–Weinberg equilibrium when the samples (n = 500) were examined prior to further analysis (p >.05). Association of the genotypes with age was evaluated by the chi-square test. Multivariate logistic regression analysis was performed to obtain odds ratios and 95% confidence intervals. The presence or absence of a given SNP genotype was used as a dependent variable, and age and gender were considered independent variables according to Tan and colleagues (10,11). Values of p <.05 were considered statistically significant. Hardy–Weinberg equilibrium, LD, and haplotype frequency were evaluated using SNPAlyze version 2.2 (DYNACOM Co., Ltd., Tokyo, Japan). For all other analyses, the StatView statistical software system (version 5; SAS Institute, Cary, NC) was used.
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RESULTS
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We genotyped IL-19 Ser175Phe (T/C), which is the only SNP that results in a nonsynonymous amino acid substitution, and three other SNPs, –513 T/C, 1098 G/T, and 5420 G/C, with allele frequencies similar to that of Ser175Phe. Five hundred participants were divided into a higher age group (
57 years) and a lower age group (<57 years), and the relationship between age or gender and the genotype distribution of each SNP was evaluated by a conventional chi-square test. As shown in Table 2, three of four SNPs (Ser175Phe, –513 T/C, and 1098 G/T) exhibited a weak, but significant association with age (p <.05). This association remained significant after multivariate logistic regression analysis was performed according to Tan and colleagues (10,11) (odds ratio for an age interval for 1 year, 1.008–1.009) (Table 3). To assess the effect of disorders found in 163 patients (see Participants), three major diseases including essential hypertension (n = 58), ischemic heart disease (n = 40), and diabetes mellitus (n = 20) were included in logistic regression analysis. Although the association of IL-19 SNPs with age remained statistically significant for Ser175Phe and –513 T/C (or borderline significant for 1098 T/G), there were no associations of IL-19 SNPs with any of these diseases (Table 4).
The LD for these IL-19 SNPs and IL-10 –819 T/C was calculated (Table 5). IL-19 Ser175Phe was in LD with –513 T/C and 1098 G/T, but not with IL-10 –819 T/C. The LD between IL-19 –513 T/C and 1098 G/T (D = 0.2378) was much stronger than that between Ser175Phe and –513 T/C (D = 0.1512) or between Ser175Phe and 1098 G/T (D = 0.1503). The frequency of each haplotype, consisting of IL-19 Ser175Phe, –513 T/C, and 1098 G/T, was calculated and compared between higher and lower age groups. As shown in Table 6, no significant difference was observed between higher and lower age groups when the haplotypes were divided into two groups, i.e., the most common haplotype T-T-G and the other six haplotypes. These results are likely due to a modest LD between Ser175Phe and –513 T/C or 1098 G/T.
To further evaluate the association of these IL-19 SNPs with age and gender, the frequencies of IL-19 Ser175Phe T/T, –513 T/T, and 1098 G/G genotypes in each two decades of 500 participants are shown in Table 7. In the Ser175Phe T/T genotype, both men and women showed a similar tendency that the genotype frequency increased with age. The frequency of –513 T/C or 1098 G/T genotype also increased with age between 19 and 80 years, but it tended to decrease in the highest age group (81–100 years). No particular change was seen in a female group of the 6th and 7th decades (61–80 in Table 7), in which the percentage of women was lower than that of the entire Japanese population (see Participants).
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Table 7. Frequencies of IL-19 Ser175Phe T/T, –513 T/T, and 1098 G/G Genotypes in Each Two Decades of 500 Participants Aged 19–100 Years.
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DISCUSSION
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To our knowledge, this is the first investigation to demonstrate an association of IL-19 polymorphisms with age. With regard to the population tested, there was no significant difference of genotype distribution and allele frequency on IL-10-819 T/C between our population (n = 500) and other Japanese populations previously reported (1). Furthermore, the frequencies of genotype distribution of seven SNPs including TNF-, IL-6, IL-1ß, IL-18, transforming growth factor-beta (TGF-ß), matrix metallopeptidase-1 (MMP-1), and MMP-3 were also similar to those in other Japanese populations (12–16). Although our population included 163 elderly patients from the hospital setting, we previously compared the genotype distribution of nine SNPs between 163 patients and healthy controls 64 years old (n = 85; mean age 77.9 years; range 64–98 years). No significant difference in the genotype distribution of those SNPs between these two groups was observed (1). In addition, no significant difference was seen when the genotype distribution of IL-19 SNPs including Ser175Phe, –513 T/C, and 1098 G/T was compared between these two populations (Table 8).
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Table 8. Comparison of the Genotype Distribution of IL-19 SNPs between Patients With Chronic Diseases and Healthy Adults (Aged 64 Years).
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We demonstrated a significant association of IL-19 Ser175Phe, –513 T/C, and 1098 G/T with age by using a conventional chi-square test (Table 2) and logistic regression analysis (10,11) (Tables 3 and 4). To further evaluate the association of these IL-19 SNPs with age and gender, the frequencies of IL-19 Ser175Phe T/T, –513 T/T, and 1098 G/G genotypes in each two decades of 500 participants were compared between men and women (Table 7). These genotype frequencies tended to increase with age between 19 and 80 years both in men and women. However, in the highest age group (81–100 years) Ser175Phe T/T showed further increase whereas –513 T/T and 1098 G/G tended to decrease, especially in men. As shown in Table 8, it is not plausible that chronic disorders may affect the genotype distribution of these SNPs in a higher age group. A strong LD (Table 5) and a very similar genotype distribution (Table 2) between –513 T/C and 1098 G/T suggest that these two SNPs may be associated with age in a manner different from Ser175Phe. Further studies with a larger sample size are required to confirm this hypothesis.
IL-10 –819 T/C was not in LD with these three IL-19 SNPs (Table 5), indicating that the IL-10 and IL-19 SNPs tested could be on different haplotype blocks and might be associated independently with age. This assumption may be supported by the finding that there are no haplotype blocks spanning both the IL-10 and IL-19 genes in the present HapMap database (National Center for Biotechnology Information [NCBI] 35) (SNPbrowser, version 3.1.29; Applied Biosystems, Tokyo, Japan).
The effect of the SNPs evaluated in this study on the function of IL-19 remains to be elucidated. According to PolyPhen, a computer software package for calculating the predicted impact of nonsynonymous SNPs on protein structure and function (http://genetics.bwh.harvard.edu/pph/data/), IL-19 Ser175Phe was predicted to be benign. In contrast, Chang and colleagues (17) resolved the receptor-binding site of IL-19 by using the crystal structure of the complex of IL-10 with its soluble nonglycosylated mutant of IL-10R1 and suggested that Ser175, which is in the C-terminal ß-strand of IL-19, is one of the residues within 5 Å of the receptor site. The IL-19 –513 T/C SNP is located in the promoter region. Liao and colleagues (5) described the promoter sequence of the IL-19 gene (
2 kb from the transcription start site) and potential transcription factor-binding sites, but did not show any consensus sequences for transcription factor-binding sites around nucleotide position –513. Also, we were not able to find those consensus sequences using computer software (data not shown).
Although there has been no evidence to date on the relationship of IL-19 with aging, it is becoming clear that IL-19 cooperates with IL-10 for immune function. Jordan and colleagues (18) recently demonstrated that IL-19 induces the production of IL-10 and IL-19 from peripheral blood mononuclear cells, which is negatively regulated by IL-10. Furthermore, IL-10 has also been shown to be produced by monocytes and monocyte-derived dendritic cells (DCs) (18), suggesting that IL-19 might be involved in the induction of IL-10-producing regulatory DCs that can induce T-cell anergy (19). IL-10 also modulates DC maturation, enabling DCs to induce regulatory T cells (20). In this context, there is increasing evidence for age-dependent development of regulatory T cells (21). Further studies will be required to reveal the effects of IL-10 and IL-19 polymorphisms on immune function.
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Footnotes
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Decision Editor: Huber R. Warner, PhD
Received January 15, 2007
Accepted January 17, 2007
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Abstract
Participants
