Skip to main content
NIHPA Author Manuscripts logoLink to NIHPA Author Manuscripts
. Author manuscript; available in PMC: 2012 Mar 1.
Published in final edited form as: Eur J Cancer Prev. 2011 Mar;20(2):86–95. doi: 10.1097/CEJ.0b013e3283429e45

Do Interleukin Polymorphisms Play a Role in the Prevention of Colorectal Adenoma Recurrence by Dietary Flavonols?

Gerd Bobe 1, Gwen Murphy 2, Paul S Albert 3, Leah B Sansbury 4, Matthew R Young 1, Elaine Lanza 1, Arthur Schatzkin 5, Nancy H Colburn 1, Amanda J Cross 5
PMCID: PMC3029494  NIHMSID: NIHMS259957  PMID: 21160427

Abstract

Chemopreventive dietary compounds, such as flavonols, may inhibit colorectal carcinogenesis partly by altering cytokine expression and attenuating inflammation. Single nucleotide polymorphisms (SNPs) in the promoter regions of genes encoding cytokines may influence flavonol-induced changes in cytokine expression and consequently cancer risk. Using logistic regression, we estimated odds ratios (OR) and 95% confidence intervals (CI) for the association between SNPs of interleukin (IL)-1β, 6, 8, and 10, alone or combined with flavonol intake or serum IL concentration changes, and adenoma recurrence in 808 participants from the intervention arm of the Polyp Prevention Trial, a 4-year intervention study evaluating the effectiveness of a low-fat, high-fiber, high-fruit and vegetable diet on adenoma recurrence.. Overall, SNPs in genes encoding IL-1β, 6, 8, and 10 were not associated with their corresponding serum concentrations or adenoma recurrence. However, individuals homozygous for IL-10 -592 C (OR = 2.23, 95% CI: 1.07–4.66, P interaction = 0.03) or IL-10 -819 C (OR = 2.18, 95% CI: 1.05–4.51, P interaction = 0.05) had an elevated risk of high risk adenoma recurrence when their serum IL-10 concentrations increased during the trial. In addition, IL-6 -174 GG in combination with above median flavonol intake (OR = 0.14, 95% CI: 0.03–0.66) or with decreased IL 6 concentrations (OR = 0.14, 95% CI: 0.03–0.65) reduced the risk of advanced adenoma recurrence, although the interaction term was not statistically significant. In conclusion, our results suggest that IL SNPs, in combination with a flavonol-rich diet or decreased serum IL, may lower the risk of adenoma recurrence.

Keywords: adenoma recurrence, flavonols, interleukins

INTRODUCTION

Flavonols are a group of bioactive polyphenols that are abundant in some fruits, vegetables, and tea [12]. In the U.S., the median flavonol intake is estimated to be approximately 10–12 mg/d with a range between 0 and 40 mg (Peterson JJ, personal communication). We reported that daily consumption of dietary flavonols above 30 mg decreases the risk of advanced and high risk colorectal adenoma recurrence [3]. Flavonols may impede growth of colorectal neoplasms by their anti-oxidative [4], anti-mutagenic [5], anti-proliferative [6], anti-inflammatory [79], and anti-cell transformation properties [10] as well as by inducing apoptosis [11], and inhibiting matrix metalloproteinases and angiogenesis [1213]. We are especially interested in the anti-inflammatory properties of flavonols for cancer prevention, as flavonols may attenuate the secretion of the pro-inflammatory and pro-carcinogenic cytokines interleukin (IL) 1β, 6, 8 [1416; 8; 1718] and inhibit the secretion of the pro-and anti-carcinogenic IL-10 [19; 15].

We reported that increases in serum IL-1β, IL-6, and IL-10 are potential indicators of advanced adenoma recurrence; moreover, that a decrease in these serum cytokines in conjunction with flavonol consumption above 30 mg/d indicated an even lower risk of advanced adenoma recurrence [3](Bobe et al., in press). Single-nucleotide polymorphisms (SNPs) in the promoter region of genes encoding IL-1β, IL-6, IL-8, and IL-10 may alter the secretion of cytokines [2024] and, as a result, the risk of adenoma recurrence generally or in response to dietary flavonols. Thus, IL SNPs may indicate individuals more likely to benefit from a chemopreventive dietary intervention.

The aim of this study was to examine whether SNPs in the promoter regions of genes encoding IL-1β, 6, 8, and 10 could influence, alone or in combination with flavonol intake or serum IL concentrations, adenoma recurrence. To our knowledge, this is the first study to investigate the joint effects of IL genotype, serum IL levels, and flavonol intake on adenoma recurrence.

MATERIALS AND METHODS

Study Design and Population

The Polyp Prevention Trial (PPT) was a large, multi-center, randomized 4-year nutritional intervention trial to evaluate the effects of promoting a high fiber, high-fruit and vegetable, low-fat diet on the recurrence of colorectal adenomas. Details of the study have been described elsewhere [2527]. In brief, study participants had at least one histologically confirmed colorectal adenoma identified by complete colonoscopy in the six months prior to study entry. Of the 1,905 participants who completed the trial by undergoing a colonoscopy at the end of year four, 958 were in the intervention arm. Our study included the 808 participants in the intervention arm with available dietary data for any of the first 3 years of the trial, IL-SNP data, and serum from baseline (T0) and from either year 1 (T1) or 3 (T3). Dietary information and serum samples from T4 were not used because some of these were taken after the final colonoscopy. All lesions were examined for histological features and degree of atypia by two independent pathologists. Recurrence outcomes were defined as having any (n = 318), advanced (≥1 adenoma of ≥1 cm in size, having at least 25% villous component, or exhibiting high-grade dysplasia; n = 45) or high risk (≥1 advanced adenoma or ≥3 pathologically confirmed adenomas; n = 88) adenomas.

Lifestyle and Flavonol Data

Participants completed an interviewer-administered questionnaire regarding demographics, family history, and use of medication or supplements at baseline (T0) and at each of the annual follow-up visits (T1, T2, T3, T4). A modified food frequency questionnaire (FFQ) eliciting information on the frequency and portion size of 119 food and beverage items consumed over the past 12 months [2829] was also completed at each of these visits. Trained, certified nutritionists reviewed all FFQs with participants. Using the 2007 U.S. Department of Agriculture (USDA) flavonoid database that contains flavonol values for 308 food and beverage items [30], 55 of the 119 food and beverage items could be matched with flavonol values, which were provided as concentrations of the individual flavonols isorhamnetin, kaempferol, myricetin, and quercetin. Flavonol intake was calculated as the sum of isorhamnetin, kaempferol, myricetin, and quercetin, which represent over 99% of dietary flavonol intake [12]. Compared with 24-hr dietary recall and four-day food record data, the FFQ slightly overestimated fat and underestimated fiber, fruit & vegetable intake and had acceptable correlations of fat (r = 0.63), fiber (r = 0.63), fruit & vegetable (r = 0.72), dry bean (r = 0.76), and other macro- and micronutrients [29; 27].

Interleukin Genotype Data

The SNPs in IL-encoding genes were selected based on the following criteria: (i) a reported association between the SNP and colorectal adenoma or cancer [3135], (ii) an association between the SNP and transcriptional activity and serum concentrations of its encoded gene product [2024], and (iii) evidence that flavonols alter the activities of transcription factors of IL-encoding genes [36; 6; 10; 37] and serum concentrations of their gene product [3839; 18]. Details of the genotyping have been previously described [40]. In brief, high-throughput genotyping was carried out by BioServe Biotechnologies, Ltd. (Laurel, MD) using a two-step PCR process and mass spectrometry (Masscode, Qiagen Genomics, Bothel, WA) [41]. Call rates for SNPs in IL-encoding genes were as follows: IL-1β -511 C>T (rs16944) 93.5%, IL-6 -174 G>C (rs1800795) 91.8%, IL-8 -251 T>A (rs4073) 92.7%, IL10 -592 C>A (rs1800872) 98.3%, IL10 -819 C>T (rs1800871) 95.0%, and IL-10 -1082 G>A (rs1800896) 95.5%. Quality control consisted of repeated assays of approximately 10% randomly selected samples as well as the inclusion of blinded controls. The concordance of duplicate samples was above 97%.

Interleukin Serum Data

Serum concentrations of IL-1β, IL-6, IL-8, and IL-10 at baseline, T1, and T3 were measured by the Clinical Support Laboratory of SAIC Frederick, Inc. (Frederick, MD) using a commercially available multiplex 96-well enzyme-linked immunoabsorbent assay kit (MS6000 Human Pro-Inflammatory 9-Plex Ultra-Sensitive Kit K11007; Meso Scale Diagnostics, Gaithersburg, MD) on a Sector Imager 6000 according to the manufacturer’s recommendation (Meso Scale Diagnostics, Gaithersburg, MD). The interassay coefficients of variation (CVs) were below 15%.

Statistical Analyses

Statistical analyses were performed using SAS, version 9.1 (SAS, Inc., Cary, North Carolina) software. Baseline characteristics, average dietary intake for the first 3 years of the trial, and serum IL concentrations were evaluated by adenoma recurrence at T4 (no vs. any, high risk, or advanced adenoma recurrence) using Wilcoxon rank-sum test for continuous variables and Fisher’s exact test for categorical variables and are shown as medians and interquartile ranges (IQRs). All SNPs were tested for, and were in agreement with, Hardy-Weinberg equilibrium.

The associations between IL SNPs and their corresponding serum concentrations were evaluated with the Kruskal Wallis test and multiple linear regression models. Potential confounders, listed in Table 1, were included in the logistic regression models if the confounder changed the association by 10% or more, was associated with both IL SNPs and their corresponding serum concentrations, and had a χ2 P value ≤ 0.20. Regular nonsteroidal anti-inflammatory drug (NSAID) use was not included in the statistical models because it was not associated with adenoma recurrence in this population (Table 1) [42; 3].

Table 1.

Proportions and medians (interquartile ranges) of participant characteristics in the intervention arm of the Polyp Prevention Trial by adenoma recurrence at T4 (n = 808)*

Characteristics Adenoma Recurrence (T4)
None Any High Risk Advanced

Median (IQR) or % Median (IQR) or % P value Median (IQR) or % P value Median (IQR) or % P value
Sample size (%) 490 (61) 318 (39) 88 (11) 45 (6)
Baseline (T0):
Gender (% male) 63 69 0.15 73 0.11 67 0.75
Race (% Caucasian) 89 90 0.73 90 0.86 87 0.63
Education (% ≤ high school) 23 28 0.13 31 0.14 29 0.36
Family history of colorectal cancer (% yes)a 27 26 0.94 31 0.52 27 1.00
Smoker (% current) 11 13 0.32 16 0.20 13 0.62
NSAID use (% yes)3 36 37 0.82 33 0.63 31 0.63
Supplement use (% yes)b 45 44 0.77 41 0.49 38 0.43
Hormone therapy (% yes)b 14 9 0.06 7 0.08 9 0.49
Age (y) 60.0 (52.0–67.0) 64.0 (56.0–70.0) <.0001 66.0 (57.0–71.0) <.0001 66.0 (57.0–71.0) 0.002
Body mass index (kg/m2) 27.5 (25.0–30.1) 27.4 (25.1–30.3) 0.78 28.3 (25.6–31.1) 0.06 29.4 (26.7–32.4) 0.01
Physical activity (hr/wk)c 8.50 (4.00–15.8) 8.38 (3.85–15.7) 0.72 8.27 (3.33–12.6) 0.22 5.75 (1.94–12.0) 0.07
Dietary flavonols (mg/d) 13.9 (9.32–20.8) 15.7 (10.1–22.9) 0.03 13.8 (9.23–19.9) 0.76 12.2 (8.00–18.1) 0.22
Serum cytokines (in pg/mL):
Interleukin 1β 0.36 (0.22–0.60) 0.36 (0.22–0.63) 0.89 0.37 (0.21–0.71) 0.93 0.34 (0.19–0.62) 0.64
Interleukin 6 1.88 (1.35–2.80) 1.89 (1.36–2.75) 0.83 1.97 (1.43–2.68) 0.39 1.77 (1.41–2.62) 0.91
Interleukin 8 10.5 (7.87–14.0) 10.3 (8.09–14.3) 0.60 11.1 (8.47–16.8) 0.12 11.4 (8.79–15.8) 0.18
Interleukin 10 3.27 (2.16–5.68) 3.28 (2.12–6.27) 0.93 3.23 (2.22–4.62) 0.47 2.80 (2.03–4.18) 0.14
Trial (T1,2,3d):
Dietary intake:
Alcohol (g/d) 0.91 (0.00–8.49) 0.96 (0.00–8.05) 0.54 0.99 (0.00–5.80) 0.64 0.54 (0.00–4.93) 0.13
Energy (1,000 kcal/d) 1.77 (1.52–2.06) 1.80 (1.54–2.07) 0.80 1.82 (1.56–2.00) 0.91 1.82 (1.57–1.99) 0.90
Fat (% kcal/d) 22.4 (18.5–26.5) 22.6 (19.6–27.8) 0.11 23.5 (20.5–29.5) 0.007 26.9 (21.2–30.6) 0.001
Fiber (g/d) 32.2 (24.3–41.1) 31.0 (23.2–39.0) 0.17 29.5 (22.1–36.2) 0.03 29.4 (20.4–37.7) 0.05
Fruits & vegetables (servings/d) 5.72 (4.43–7.20) 5.69 (4.51–7.01) 0.71 5.21 (4.38–6.53) 0.05 4.93 (4.04–6.03) 0.01
Flavonols (mg/d) 29.6 (21.2–40.5) 29.7 (21.1–38.9) 0.74 25.9 (16.5–36.4) 0.02 20.6 (14.9–30.8) 0.0004
Dry beans(g/d) 31.2 (15.4–56.2) 30.4 (14.5–49.5) 0.26 23.5 (8.79–42.1) 0.01 14.0 (7.37–37.5) 0.0004
Serum cytokines (in pg/mL):
Interleukin 1β 0.36 (0.24–0.64) 0.36 (0.23–0.58) 0.43 0.41 (0.25–0.68) 0.40 0.43 (0.25–0.71) 0.32
Interleukin 6 1.96 (1.44–2.77) 2.01 (1.53–2.80) 0.39 2.27 (1.66–3.10) 0.01 2.07 (1.66–3.07) 0.15
Interleukin 8 10.7 (8.18–15.5) 10.9 (8.28–14.7) 0.70 11.0 (8.31–16.0) 0.88 11.5 (8.78–17.1) 0.38
Interleukin 10 3.20 (2.21–5.53) 3.21 (2.31–5.98) 0.77 3.29 (1.99–5.08) 0.59 3.21 (1.87–5.77) 0.64
*

Participant characteristics stratified by adenoma recurrence have been previously presented for 872 participants from the intervention arm of the Polyp Prevention Trial [3][Bobe et al., in press].

All comparisons against the no adenoma recurrence group. P values for differences in proportions were calculated using Fisher’s exact test. P values for differences in medians were calculated using Wilcoxon rank-sum test.

a

Family history of colorectal cancer was defined as having ≥ 1 first-degree relative with colorectal cancer at baseline.

b

Regular non-steroidal anti-inflammatory drug (NSAID) use, hormone therapy, and supplement use were tested at baseline and use throughout the first 3 trial years. Regular dietary supplement use was defined as taking supplement ≥1 weekly. Regular medication use, including NSAIDs, was defined as taking medication ≥1 monthly. Hormone replacement therapy included both unopposed estrogen and estrogen/progestin combinations.

c

Physical activity was defined as self-reported time typically spent for any type of moderate or vigorous physical activity.

d

T1,2,3: mean values of the first three years of the trial for dietary variables and geometric mean of year 1 and 3 cytokine values.

We used logistic regression to calculate odds ratios (OR) and 95% confidence intervals (CI) for adenoma recurrence at T4 by IL SNPs, using the homozygote of the more frequent allele as the referent category and race, age, and gender as covariates. For linear trend testing, the referent category was the more frequent allele which was scored on an ordinal scale as 0, the heterozygote was scored as 1, and the rare homozygote was scored as 2 (co-dominant model). For the dominant model, homozygous and heterozygous carriers of the less frequent gene were combined and scored as one. The IL-10 haplotypes were composed of 3 polymorphic sites -1082 A/G, -819 C/T, and -592 C/A: IL-10 A-C-C was comprised of -1082 AA, -819 CC, -592 CC (referent haplotype); IL-10 G-C-C being -1082 G, -819 CC, -592 CC; and IL-10 G-T-A being -1082 G, -819 T, -592 A.

The combined effect of IL genotype and flavonol intake on colorectal adenoma recurrence was evaluated using the median flavonol intake during the first 3 years of the trial as the cut-point (≤ median, > median) and carrier status of the rare allele (yes, no). The group with the largest number of advanced adenoma cases was chosen as the referent category consisting of individuals with below median flavonol intake, who carried at least one copy of the less frequent allele. To examine the combined effect of IL genotype and changes in serum IL-concentrations, defined as the geometric mean of T1 and T3 minus the baseline values, we used the median IL change as the cut-point (≤ median, > median); the referent category was individuals with an increase in IL concentrations, who carried at least one copy of the less frequent allele. Effect modification was evaluated by calculating the cross product interaction terms. All p values corresponded to two-sided tests. Differences were considered to be significant at P ≤ 0.05 and borderline significant at 0.05 < P ≤ 0.10.

RESULTS

At the end of the 4-year trial, 39% of the 808 participants in the intervention arm had 1 or more adenoma, 11% had a high risk adenoma, and 6% had an advanced adenoma (Table 1). Adenoma recurrence was more common in older individuals. Flavonol consumption doubled from 14.6 mg/d at baseline to 29.7 mg/d during the first 3 years of the trial [3]. Those with any adenoma recurrence consumed more flavonols at baseline. Individuals with high risk and advanced adenoma recurrence ate more calories from fat and consumed less fruits & vegetables, fiber, flavonols, and dry beans during the first 3 years of the study. High risk adenoma recurrence was also positively associated with BMI.

The SNPs in genes encoding IL-1β, IL-6, IL-8, and IL-10 were not associated with serum concentrations of their gene product; however, baseline serum IL-10 concentrations tended to linearly increase with IL-10 -819 T allele copy number (Pfor trend = 0.07) and IL-10 G-T-A haplotype (Pfor trend = 0.06) (Table 2). Furthermore, an exploratory analysis suggested that IL SNPs may alter serum concentrations of ILs other than their own gene product, for example IL-1β -511 SNP was associated with serum IL-8, IL-8 -251 SNP was associated with serum IL-6, and IL-10 -592 SNP was associated with serum IL-1β (results not shown).

Table 2.

Medians (interquartile ranges; in pg/mL) of serum interleukin concentrations in the intervention arm of the Polyp Prevention Trial by their corresponding interleukin polymorphism (n = 808)

Interleukin Interleukin Concentrations (Median, IQR in pg/mL) P Non P for

Polymorphism Param.* Trend
IL-1β -511 C/C C/T T/T
Sample size 319 359 78
Baseline (T0) 0.35 (0.22–0.63) 0.37 (0.22–0.59) 0.38 (0.21–0.74) 0.93 0.40
Intervention (T1,3) 0.36 (0.23–0.59) 0.38 (0.24–0.65) 0.37 (0.25–0.61) 0.74 0.82
Change (T1,3-T0) −.01 (−0.15–0.14) 0.02 (−0.11–0.16) 0.03 (−0.11–0.21) 0.19 0.75

IL-6 -174 G/G G/C C/C
Sample size 284 353 105
Baseline (T0) 1.83 (1.27–2.95) 1.89 (1.36–2.72) 2.03 (1.44–2.84) 0.62 0.40
Intervention (T1,3) 1.95 (1.44–2.88) 2.00 (1.44–2.80) 1.90 (1.49–2.54) 0.85 0.82
Change (T1,3-T0) 0.02 (−0.44–0.55) 0.11 (−0.35–0.50) −.01 (−0.70–0.54) 0.52 0.72

IL-8 -251 T/T A/T A/A
Sample size 187 378 179
Baseline (T0) 10.6 (7.95–13.5) 10.4 (7.88–14.1) 10.7 (8.05–14.9) 0.63 0.23
Intervention (T1,3) 11.2 (8.39–15.7) 10.5 (7.93–15.1) 10.9 (8.41–16.2) 0.38 0.45
Change (T1,3-T0) 0.50 (−2.17–3.08) 0.19 (−2.09–2.38) 0.49 (−1.73–2.82) 0.67 0.75

IL-10 -592 C/C C/A A/A
Sample size 432 275 62
Baseline (T0) 3.16 (2.07–5.59) 3.36 (2.20–6.48) 3.35 (1.96–4.89) 0.53 0.16
Intervention (T1,3) 3.18 (2.09–5.59) 3.31 (2.14–5.66) 3.08 (2.25–5.33) 0.80 0.26
Change (T1,3-T0) −.03 (−0.82–0.56) −.16 (−0.96–0.50) 0.04 (−0.49–0.83) 0.14 0.29

IL-10 -819 C/C C/T T/T
Sample size 444 279 66
Baseline (T0) 3.14 (2.09–5.51) 3.42 (2.21–6.71) 3.35 (1.98–4.89) 0.29 0.07
Intervention (T1,3) 3.13 (2.13–5.55) 3.34 (2.23–5.66) 3.02 (2.22–5.23) 0.72 0.24
Change (T1,3-T0) 0.00 (−0.82–0.58) −.12 (−0.96–0.49) 0.00 (−0.64–0.60) 0.26 0.29

IL-10 -1082 A/A A/G G/G
Sample size 221 409 147
Baseline (T0) 3.24 (2.02–5.94) 3.44 (2.24–6.18) 3.24 (2.02–5.27) 0.29 0.29
Intervention (T1,3) 3.04 (2.15–5.58) 3.35 (2.25–5.86) 2.97 (2.00–5.09) 0.46 0.17
Change (T1,3-T0) −.03 (−0.70–0.60) −.07 (−1.01–0.58) −.05 (−0.83–0.46) 0.56 0.09

IL-10 Haplotypea A-C-C G-C-C G-T-A
Sample size 143 269 322
Baseline (T0) 3.32 (2.01–5.27) 3.18 (2.15–5.69) 3.39 (2.18–6.48) 0.38 0.06
Intervention (T1,3) 3.26 (2.35–5.78) 3.31 (2.23–5.78) 3.16 (2.01–4.88) 0.53 0.16
Change (T1,3-T0) −.05 (−0.81–0.51) −.03 (−0.93–0.62) −.09 (−0.93–0.56) 0.81 0.20
*

P values for differences in medians among the flavonol intake quartiles were calculated based on the Kruskal-Wallis test.

Median concentrations of each interleukin genotype were used to determine P for trend of the interleukin concentrations using a multiple regression model adjusting for age tertiles (<58, 58–66, >66 yrs), sex, race (Caucasian, non Caucasian), average BMI (<25, 25.0–29.9, ≥30 kg/m2), and current smoking status during the first 3 trial years.

a

Composed of 3 polymorphic sites: -1082 AA/G, -819 CC/T, and -592 CC/A.

Alone, the examined IL SNPs did not predict risk of adenoma recurrence in either a dominant or co-dominant model; except that the IL-1β -511 T and the IL-6 -174 G alleles may have a protective effect against high risk (OR = 0.62; 95% CI: 0.38–1.01; P = 0.05 for the dominant model) and advanced adenoma recurrence (Pfor trend = 0.07 for the co-dominant model), respectively (Table 3).

Table 3.

Association between interleukin polymorphisms and colorectal adenoma recurrence in participants of the Polyp Prevention Trial at the end of Year 4 (n = 808)

Interleukin Adenoma Recurrence
None Any High Risk Advanced

Polymorphism n (%) n (%) OR (95% CI)* n (%) OR (95% CI)* n (%) OR (95% CI)*
IL-1β -511
C/C 190 (59.6) 129 (40.4) 1.00 43 (13.5) 1.00 19 (6.0) 1.00
C/T 222 (61.8) 137 (38.2) 0.90 (0.66–1.24) 32 (8.8) 0.61 (0.36–1.01) 18 (5.0) 0.76 (0.38–1.52)
T/T 50 (61.1) 28 (35.9) 0.83 (0.49–1.41) 8 (10.3) 0.68 (0.29–1.57) 4 (5.1) 0.75 (0.24–2.36)
C/T + T/T 272 (62.2) 165 (37.8) 0.89 (0.66–1.21) 40 (9.2) 0.62 (0.38–1.01) 22 (5.0) 0.76 (0.39–1.47)
P-trend 0.42 0.11 0.46
IL-6 -174
G/G 181 (63.7) 103 (36.3) 1.00 30 (10.6) 1.00 13 (4.6) 1.00
G/C 208 (58.9) 145 (41.1) 1.25 (0.89–1.75) 45 (12.7) 1.40 (0.82–2.40) 22 (6.2) 1.76 (0.81–3.86)
C/C 64 (61.0) 41 (39.0) 1.19 (0.74–1.91) 9 (8.6) 1.04 (0.45–2.40) 8 (7.6) 2.44 (0.89–6.64)
G/C + C/C 272 (59.4) 186 (40.6) 1.24 (0.90–1.71) 54 (11.8) 1.33 (0.79–2.24) 30 (6.6) 1.89 (0.89–4.01)
P-trend 0.32 0.58 0.07
IL-8 -251
T/T 118 (63.1) 69 (36.9) 1.00 16 (8.6) 1.00 8 (4.3) 1.00
A/T 228 (60.3) 150 (39.7) 1.16 (0.80–1.68) 44 (11.6) 1.45 (0.77–2.71) 23 (6.1) 1.50 (0.64–3.48)
A/A 112 (62.6) 67 (37.4) 1.03 (0.66–1.60) 16 (8.9) 1.08 (0.50–2.33) 10 (5.6) 1.28 (0.47–3.49)
A/T + A/A 340 (61.0) 217 (39.0) 1.12 (0.79–1.60) 60 (10.8) 1.34 (0.73–2.45) 33 (5.9) 1.43 (0.63–3.24)
P-trend 0.87 0.82 0.63
IL-10 -592
C/C 264 (61.6) 168 (38.9) 1.00 51 (11.8) 1.00 27 (6.3) 1.00
C/A 171 (62.2) 104 (37.8) 0.92 (0.67–1.26) 26 (9.5) 0.76 (0.45–1.28) 13 (4.7) 0.69 (0.34–1.40)
A/A 33 (53.2) 29 (46.8) 1.45 (0.83–2.53) 7 (11.3) 1.17 (0.47–2.90) 3 (4.8) 0.85 (0.23–3.08)
C/A + A/A 204 (60.5) 133 (39.5) 0.99 (0.74–1.34) 33 (9.8) 0.82 (0.50–1.33) 16 (4.7) 0.72 (0.37–1.39)
P-trend 0.55 0.68 0.42
IL-10 -819
C/C 267 (60.1) 177 (39.9) 1.00 54 (12.2) 1.00 29 (6.5) 1.00
C/T 173 (62.0) 106 (38.0) 0.90 (0.66–1.23) 26 (9.3) 0.72 (0.43–1.21) 12 (4.3) 0.60 (0.29–1.22)
T/T 36 (54.5) 30 (45.5) 1.35 (0.79–2.33) 17 (10.6) 1.08 (0.44–2.67) 3 (4.5) 0.76 (0.21–2.76)
C/T + T/T 209 (60.6) 136 (39.4) 0.97 (0.72–1.30) 33 (9.6) 0.78 (0.48–1.26) 15 (4.3) 0.62 (0.32–1.21)
P-trend 0.70 0.52 0.24
IL-10 -1082
A/A 132 (59.7) 89 (40.3) 1.00 23 (10.4) 1.00 11 (5.0) 1.00
A/G 251 (61.4) 158 (38.6) 0.93 (0.66–1.31) 43 (10.5) 0.99 (0.57–1.74) 24 (5.9) 1.20 (0.56–2.56)
G/G 88 (59.9) 59 (40.1) 1.06 (0.68–1.63) 19 (12.9) 1.36 (0.69–2.69) 3 (4.5) 1.21 (0.46–3.17)
A/G + G/G 339 (61.0) 217 (39.0) 0.96 (0.70–1.34) 62 (11.2) 1.08 (0.64–1.84) 15 (4.3) 1.20 (0.58–2.49)
P-trend 0.87 0.42 0.67
IL-10 Haplotypea
A-C-C 85 (59.4) 58 (40.6) 1.00 19 (13.3) 1.00 8 (5.6) 1.00
G-C-C 161 (59.9) 108 (40.1) 0.93 (0.61–1.42) 30 (11.2) 0.76 (0.40–1.45) 18 (6.7) 1.14 (0.47–2.75)
G-T-A 197 (61.2) 125 (38.8) 0.86 (0.57–1.30) 32 (9.9) 0.66 (0.35–1.26) 15 (4.7) 0.72 (0.29–1.79)
*

All comparisons against the no adenoma group. Multivariate OR and 95% CI models were adjusted for age tertiles (<58, 58–66, >66 yrs), sex, and race (Caucasian or not). Homozygous carriers of the more frequent gene are the reference group. The dominant model is evaluated by the OR (95% CI) for carriers of the less frequent gene combined. The co-dominant model is evaluated by P-trend

a

Composed of 3 polymorphic sites: -1082 AA/G, -819 CC/T, and -592 CC/A.

We investigated whether IL SNPs in combination with changes in serum IL concentrations during the trial may affect risk of adenoma recurrence (Table 4). The analysis is based on the widespread assumption that SNPs will only be able to affect adenoma risk if they are functionally significant, meaning that they can alter secretion of their gene products, which is in this case IL secretion. An increase in serum IL-10 resulted in a greater risk of high risk adenoma recurrence when combined with IL-10 -592 CC (OR = 2.23, 95% CI: 1.07–4.66; P = 0.03 compared with individuals carrying the A allele; PInteraction = 0.03) or with IL-10 -819 CC (OR = 2.18, 95% CI: 1.05–4.51; P = 0.04; compared with individuals carrying the T allele; PInteraction = 0.05). In addition, individuals with the combination of IL-6 -174 GG and a decrease in serum IL-6 had a statistically significantly lower risk of advanced adenoma recurrence compared to those with each of the other 3 combinations of IL-6 -174 SNP and serum IL-6 change (PInteraction = 0.06).

Table 4.

Association between the combination of change in serum cytokine concentrations from baseline and interleukin polyporphisms and colorectal adenoma recurrence among intervention group participants of the Polyp Prevention Trial (n = 808)

Cytokine Changea (in pg/mL) Interleukin Polymorphism Adenoma Recurrence (T4)*
No Any High Risk Advanced

n (%) n (%) OR (95% CI)* n (%) OR (95% CI)* n (%) OR (95% CI)*
IL-1β IL-1β -511
High > 0.01 TT + CT 151 (65.7) 79 (34.3) 1.00 23 (10.0) 1.00 13 (5.7) 1.00
CC 93 (62.4) 56 (37.6) 1.15 (0.74–1.78) 18 (12.1) 1.40 (0.70–2.81) 9 (6.0) 1.19 (0.47–3.00)
Low ≤ 0.01 TT + CT 121 (58.5) 86 (41.5) 1.34 (0.90–1.98) 17 (8.2) 0.93 (0.47–1.86) 9 (4.3) 0.81 (0.33–2.01)
CC 97 (57.1) 98 (42.9) 1.41 (0.93–2.14) 25 (14.7) 1.70 (0.83–3.25) 10 (5.9) 1.28 (0.52–3.18)
Interaction P-value 0.78 0.60 0.68
IL-6 IL-6 -174
High > 0.04 CC + CG 141 (60.0) 94 (40.0) 1.00 29 (12.3) 1.00 16 (6.8) 1.00
GG 77 (58.3) 55 (41.7) 1.01 (0.64–1.58) 19 (14.4) 0.98 (0.49–1.94) 11 (8.3) 0.90 (0.37–2.18)
Low ≤ 0.04 CC + CG 131 (58.7) 92 (41.3) 1.04 (0.71–1.52) 25 (11.2) 0.82 (0.45–1.50) 14 (6.3) 0.83 (0.38–1.82)
GG 104 (68.4) 48 (31.6) 0.70 (0.45–1.09) 11 (7.2) 0.47 (0.21–1.02) 2 (1.3) 0.14 (0.03–0.65)
Interaction P-value 0.20 0.30 0.06
IL-8 IL-8 -251
High > −0.01 AA + AT 171 (61.1) 109 (38.9) 1.00 23 (8.2) 1.00 15 (5.4) 1.00
TT 68 (70.1) 29 (29.9) 0.69 (0.41–1.14) 6 (6.2) 0.74 (0.28–1.95) 3 (3.1) 0.55 (0.15–2.01)
Low ≤ −0.01 AA + AT 169 (61.0) 108 (39.0) 1.00 (0.70–1.52) 37 (13.4) 1.65 (0.92–2.94) 18 (6.5) 1.19 (0.57–2.49)
TT 50 (55.6) 40 (44.4) 1.17 (0.72–1.93) 10 (11.1) 1.32 (0.57–3.08) 5 (5.6) 1.02 (0.34–3.05)
Interaction P-value 0.13 0.90 0.61
IL-10 IL-10 -592
High > −0.04 AA + AC 102 (61.8) 63 (38.2) 1.00 12 (7.3) 1.00 7 (4.2) 1.00
CC 134 (60.9) 86 (39.1) 1.07 (0.70–1.63) 33 (15.0) 2.23 (1.07–4.66) 20 (9.1) 2.42 (0.94–6.22)
Low ≤ −0.04 AA + AC 102 (59.3) 70 (40.7) 1.12 (0.72–1.75) 21 (12.2) 1.70 (0.77–3.73) 9 (5.2) 1.29 (0.45–3.70)
CC 130 (61.3) 82 (38.7) 1.08 (0.70–1.66) 18 (8.5) 1.24 (0.56–2.78) 7 (3.3) 0.87 (0.28–2.65)
Interaction P-value 0.72 0.03 0.07
IL-10 IL-10 -819
High > −0.04 TT + TC 101 (60.8) 65 (39.2) 1.00 12 (7.23) 1.00 7 (4.2) 1.00
CC 140 (60.9) 90 (39.1) 1.00 (0.66–1.53) 35 (15.2) 2.18 (1.05–4.51) 21 (9.1) 2.32 (0.91–5.90)
Low ≤ −0.04 TT + TC 108 (60.3) 71 (39.7) 1.01 (0.65–1.57) 21 (11.7) 1.54 (0.70–3.36) 8 (4.5) 1.01 (0.34–2.97)
CC 127 (59.3) 87 (40.7) 1.08 (0.71–1.66) 19 (8.9) 1.25 (0.56–2.76) 8 (3.7) 0.94 (0.32–2.79)
Interaction P-value 0.83 0.05 0.20
IL-10 IL-10 -1082
High > −0.04 GG + GA 168 (61.8) 104 (38.2) 1.00 34 (12.5) 1.00 22 (8.1) 1.00
AA 66 (58.9) 46 (41.1) 1.16 (0.74–1.84) 11 (9.8) 0.88 (0.41–1.87) 5 (4.5) 0.62 (0.22–1.75)
Low ≤ −0.04 GG + GA 171 (60.2) 113 (39.8) 1.09 (0.77–1.54) 28 (9.9) 0.79 (0.45–1.38) 10 (3.5) 0.44 (0.20–0.98)
AA 66 (60.6) 43 (39.4) 1.05 (0.66–1.67) 12 (11.0) 0.85 (0.40–1.78) 6 (5.5) 0.63 (0.24–1.68)
Interaction P-value 0.57 0.70 0.27
IL-10 IL-10 Haplotypeb
High > −0.04 GG/GA-TT/TC-TT/TC 182 (62.1) 111 (37.9) 1.00 30 (10.2) 1.00 6 (5.5) 1.00
AA-CC-CC 40 (56.3) 31 (43.7) 1.30 (0.76–2.22) 12 (16.9) 1.91 (0.88–4.16) 17 (5.8) 2.24 (0.88–5.71)
Low ≤ −0.04 GG/GA-TT/TC-TT/TC 176 (59.1) 122 (40.9) 1.13 (0.80–1.58) 32 (10.7) 1.02 (0.58–1.78) 8 (11.3) 0.93 (0.45–1.93)
AA-CC-CC 45 (62.5) 27 (37.5) 1.08 (0.63–1.86) 7 (9.7) 1.07 (0.43–2.67) 0 (0.0)
Interaction P-value 0.43 0.33
*

Multivariate OR and 95% CI models were adjusted for age tertiles (<58, 58–66, >66 yrs), sex, average BMI (<25, 25.0–29.9, ≥30 kg/m2), race (Caucasian, non Caucasian), and current smoking status during the first 3 trial years. An increase in IL-10 concentrations was associated with a significantly greater risk of high risk adenoma recurrence in individuals homozygous with IL-10 -592 C and IL-10 -819 C compared with individuals carrying the IL-10 -592 A and IL-10 -819 T allele, respectively. The combination of decreased IL-6 concentrations and IL-6 -174 GG had a significantly lower risk estimate of advanced adenoma recurrence compared to the other 3 combinations. In individuals with IL-10 -592 CC, IL-10 -819 CC, and IL-10 -1082 AA/AG, a decrease in IL-10 concentration had a significantly lower risk estimate of advanced adenoma recurrence than an increase in IL-10 concentration. The interaction between change in IL-10 concentration and IL-10 haplotype on the risk of advanced adenoma recurrence could not be computed because no individual with a decrease in IL-10 and IL-10 AA-CC-CC haplotype had advanced adenoma recurrence.

a

Change in cytokine values is defined as difference between the geometric mean value of years 1 and 3 and baseline.

b

Composed of 3 polymorphic sites: -1082 AA/G, -819 CC/T, and -592 CC/A.

When IL SNPs were analyzed with flavonol intake (Table 5), individuals that carried IL-6 -174 GG and had above median flavonol consumption had a statistically significantly lower risk of advanced adenoma recurrence compared to each of the other 3 combinations of IL-6 -174 SNP and flavonol intake; however, the interaction term was, in contrast to the main effects for IL-6 SNP (P = 0.05) and flavonol intake (P = 0.04), not significant (P = 0.16). Furthermore, those with above median flavonol intake combined with the IL-1β -511 T allele (OR = 0.32, 95% CI: 0.11–0.94; P = 0.04), IL-10 -592 A allele (OR = 0.17, 95% CI: 0.04–0.80; P = 0.02), or IL-10 -819 T allele (OR = 0.18, 95% CI: 0.04–0.84; P = 0.03) had a lower risk of advanced adenoma recurrence compared with individuals of the same genotype and below or median flavonol intake (Table 5).

Table 5.

Association between the combination of flavonol intake during the trial and interleukin polymorphisms and colorectal adenoma recurrence among intervention group participants of the Polyp Prevention Trial (n = 808)

Flavonola (in mg/d) Interleukin Polymorphism Adenoma Recurrence (T4)*
No Any High Risk Advanced

n (%) n (%) OR (95% CI)* n (%) OR (95% CI)* n (%) OR (95% CI)*
Mean (T1,2,3) IL-1β -511
Low ≤ 29.7 TT + CT 133 (61.0) 85 (39.0) 1.00 26 (11.9) 1.00 17 (7.8) 1.00
CC 98 (60.9) 63 (39.1) 0.99 (0.65–1.52) 25 (15.5) 1.44 (0.76–2.72) 13 (8.1) 1.26 (0.56–2.84)
High > 29.7 TT + CT 139 (63.5) 80 (36.5) 0.97 (0.65–1.46) 14 (6.4) 0.59 (0.28–1.24) 5 (2.3) 0.32 (0.11–0.94)
CC 92 (56.3) 98 (43.7) 1.29 (0.87–1.91) 18 (11.4) 1.16 (0.58–2.34) 6 (3.8) 0.59 (0.21–1.63)
Interaction P-value 0.40 0.54 0.61
Mean (T1,2,3) IL-6 -174
Low ≤ 29.7 CC + CG 136 (59.1) 94 (40.9) 1.00 32 (13.9) 1.00 19 (8.3) 1.00
GG 91 (63.2) 53 (36.8) 0.81 (0.52–1.26) 19 (13.2) 0.80 (0.41–1.57) 11 (7.6) 0.74 (0.31–1.75)
High > 29.7 CC + CG 136 (59.6) 92 (40.4) 1.07 (0.72–1.59) 22 (9.6) 0.80 (0.43–1.51) 11 (4.8) 0.66 (0.29–1.52)
GG 90 (64.3) 50 (35.7) 0.88 (0.55–1.39) 11 (7.9) 0.57 (0.26–1.25) 2 (1.4) 0.14 (0.03–0.66)
Interaction P-value 0.96 0.80 0.16
Mean (T1,2,3) IL-8 -251
Low ≤ 29.7 AA + AT 170 (60.1) 113 (39.9) 1.00 37 (13.1) 1.00 21 (7.4) 1.00
TT 58 (65.2) 31 (34.8) 0.84 (0.51–1.40) 9 (10.1) 0.76 (0.34–1.73) 7 (7.9) 1.06 (0.41–2.73)
High > 29.7 AA + AT 170 (62.0) 104 (38.0) 1.05 (0.73–1.52) 23 (8.4) 0.80 (0.41–1.42) 12 (4.4) 0.70 (0.31–1.57)
TT 60 (61.2) 38 (38.8) 0.97 (0.59–1.59) 7 (7.1) 0.56 (0.23–1.38) 1 (1.0) 0.14 (0.02–1.11)
Interaction P-value 0.80 0.96 0.15
Mean (T1,2,3) IL-10 -592
Low ≤ 29.7 AA + AC 105 (60.7) 68 (39.3) 1.00 22 (12.7) 1.00 14 (8.1) 1.00
CC 129 (61.4) 81 (38.6) 1.00 (0.65–1.52) 29 (13.8) 1.04 (0.55–1.96) 17 (8.1) 1.03 (0.47–2.25)
High > 29.7 AA + AC 99 (60.4) 65 (39.6) 1.13 (0.72–1.78) 11 (6.7) 0.59 (0.26–1.31) 2 (1.2) 0.17 (0.04–0.80)
CC 135 (60.8) 87 (39.2) 1.16 (0.75–1.80) 22 (9.9) 0.95 (0.47–1.89) 10 (4.5) 0.67 (0.27–1.69)
Interaction P-value 0.92 0.40 0.13
Mean (T1,2,3) IL-10 -819
Low ≤ 29.7 TT + TC 108 (60.7) 70 (39.3) 1.00 22 (12.4) 1.00 13 (7.3) 1.00
CC 129 (60.3) 85 (39.7) 1.03 (0.68–1.56) 30 (14.0) 1.07 (0.57–2.00) 18 (8.4) 1.17 (0.53–2.56)
High > 29.7 TT + TC 101 (60.5) 66 (39.5) 1.10 (0.71–1.73) 11 (6.6) 0.57 (0.26–1.27) 2 (1.2) 0.18 (0.04–0.84)
CC 138 (60.0) 92 (40.0) 1.15 (0.75–1.77) 24 (10.4) 0.99 (0.50–1.96) 11 (4.8) 0.77 (0.31–1.91)
Interaction P-value 0.96 0.34 0.14
Mean (T1,2,3) IL-10 -1082
Low ≤ 29.7 GG + GA 161 (60.1) 107 (40.9) 1.00 36 (13.4) 1.00 22 (8.2) 1.00
AA 72 (63.2) 42 (36.8) 0.87 (0.55–1.38) 14 (12.3) 0.86 (0.43–1.73) 8 (7.0) 0.79 (0.33–1.92)
High > 29.7 GG + GA 178 (61.8) 110 (38.2) 1.03 (0.72–1.47) 26 (9.0) 0.80 (0.42–1.33) 10 (3.5) 0.46 (0.20–1.04)
AA 60 (56.1) 47 (43.9) 1.28 (0.80–2.05) 9 (8.4) 0.57 (0.32–1.68) 3 (2.8) 0.39 (0.11–1.39)
Interaction P-value 0.28 0.80 0.94
Mean (T1,2,3) IL-10 Haplotype3
Low ≤ 29.7 GG/GA-TT/TC-TT/TC 178 (61.6) 111 (38.4) 1.00 35 (12.1) 1.00 19 (8.3) 1.00
AA-CC-CC 39 (55.7) 31 (44.3) 1.40 (0.82–2.40) 13 (18.6) 1.90 (0.90–4.02) 7 (7.6) 1.71 (0.66–4.42)
High > 29.7 GG/GA-TT/TC-TT/TC 180 (59.6) 122 (40.4) 1.24 (0.87–1.76) 27 (8.9) 0.89 (0.50–1.59) 11 (4.8) 0.57 (0.26–1.28)
AA-CC-CC 46 (64.3) 27 (37.0) 1.13 (0.65–1.96) 6 (8.2) 0.83 (0.32–2.20) 2 (1.4) 0.19 (0.02–1.51)
Interaction P-value 0.27 0.26 0.16
*

Multivariate OR and 95% CI models were adjusted for age tertiles (<58, 58–66, >66 yrs), sex, race (Caucasian, non Caucasian), average BMI (<25, 25.0–29.9, ≥30 kg/m2), and average energy intake (continuous) during the first 3 trial years. Individuals that consumed a flavonol-rich diet and carried IL-6 -174 GG had a significantly lower risk estimate of advanced adenoma recurrence compared to each of the other 3 combinations. Individuals that consumed a flavonol-rich diet and carried the IL-1β -511 T, IL-10 -592 A, or IL-10 -819 T allele had a significantly lower risk estimate of advanced adenoma recurrence than individuals with lower flavonol intake. In individuals with IL-10 AA-CC-CC, a higher flavonol intake was associated with a decreased risk of advanced adenoma recurrence compared to a lower flavonol intake. The combination of high flavonol intake and IL-1β -511 TT/TC had a significantly lower risk estimate of high risk adenoma recurrence than the combination of low flavonol intake and IL-1β -511 CC.

a

Flavonol intake below or above the median of the mean intake during the first 3 trial years (T1,2,3).

b

Composed of 3 polymorphic sites: -1082 AA/G, -819 CC/T, and -592 CC/A.

DISCUSSION

Our objective was to examine whether SNPs in the promoter region of genes encoding IL-1β, IL-6, IL-8, and IL-10 may influence, alone or in combination with flavonol intake or serum IL concentrations, adenoma recurrence. Overall, we found no statistically significant associations between the SNPs in IL-encoding genes and serum concentrations of their gene products or colorectal adenoma recurrence in the intervention arm of the PPT, suggesting that IL SNPs alone may not predict adenoma recurrence. However, individuals homozygous for IL-10 -592 C or IL-10 -891 C had an elevated risk of high risk adenoma recurrence when their serum IL-10 concentrations increased during the trial. In addition, IL-6 -174 GG in combination with above median flavonol consumption or with decreases in serum IL-6 concentrations during the trial resulted in a reduced risk of advanced adenoma recurrence. Our results suggest that IL SNPs, in combination with flavonol intake or serum IL concentrations, may influence adenoma recurrence.

Growing evidence suggests that chronic inflammation, involving upregulation of both pro- and anti-inflammatory ILs, may be an important target for colorectal cancer prevention [15]. Gene and protein expression of IL-1β, IL-6, and IL-8 is elevated in the tumor microenvironment compared to normal colon tissue [43], often indicated by an increase in serum IL-1β, IL-6, and IL-10 concentrations in individuals with advanced adenoma recurrence [3](Bobe et al., in press). Thus, functional SNPs in the promoter regions of genes encoding IL-1β, IL-6, IL-8, and IL-10 may alter secretion of ILs [2024] and, as a result, cancer risk [44]. Consistent with previous studies [40; 45], we found IL SNPs alone did not predict adenoma recurrence. In previous studies, the direction of associations between IL-1β, IL-6, IL-8, and IL-10 SNPs and colorectal adenoma [31] or cancer [32; 46; 33; 47; 3435; 48] have been inconsistent.

SNPs in the promoter regions of ILs are one of many mechanisms by which IL secretion, inflammation, and further downstream colorectal neoplastic changes can be altered. Besides SNPs, IL gene and protein expression can be changed by epigenetic events, abundance and distribution of nuclear transcription factors, micro RNAs, the half-life of mRNA transcripts, and other post-transcriptional mechanisms [4950]. Environmental factors, inflammation status, and IL induction model modify IL gene and protein expression [2024]. Serum IL concentration measurements may not necessarily indicate true physiological IL bioavailability because serum IL concentrations are generally low, have a limited dynamic range, diurnal variations, and short half-lives, and lack specificity for location, strength and type of inflammation. Furthermore, the influence of IL SNPs on inflammation and cancer may be more complex than a direct association between IL SNPs and serum concentrations of their gene product (in our exploratory analysis, IL SNPs were associated with serum concentrations of ILs other than their gene product).

An increase in serum IL concentrations in combination with a specific IL genotype may indicate increased risk of adenoma recurrence. In this study, individuals with IL-10 -592 CC or IL-10 -891 CC had the greatest risk of high risk adenoma recurrence when their serum IL-10 concentrations increased during the trial. The role of IL-10 in colorectal carcinogenesis is complex [15]: IL-10 may promote or inhibit colorectal neoplastic changes depending on environmental factors such as the intestinal microbial population in the host. Elevated serum IL-10 may indicate down-regulation or up-regulation of inflammation, as IL-10 secretion is up-regulated in response to increased inflammation. Smoking, nuclear transcription factors, BMI, gender, regular NSAID use, and other IL-10 SNPS may partly determine which IL-10 allele up-regulates IL-10 gene and protein expression more strongly [5152; 22; 53]. This may explain why IL-10 SNPs are not consistently associated with increased risk of colorectal cancer [3335] indicating a complex interaction between environmental factors, IL-10 SNPs, inflammation, and colorectal cancer.

Host genetics may partly predict whether dietary bioactive compounds have chemopreventive properties. Part of the chemoprotective effect of flavonols may involve attenuating secretion of pro-inflammatory compounds [6; 10; 18], which, in part, may be regulated by SNPs in the promoter region of ILs. In support of our hypothesis, individuals with IL-6 -174 GG, in combination with higher flavonol intake, had a decreased risk of advanced adenoma recurrence compared to other combinations of flavonol consumption and IL-6 -174 SNP, although this observation is based on very few cases. The role of IL-6 -174 SNP on prevention of adenoma recurrence is not surprising given that the IL-6 -174 SNP is close to binding sites of IL-6 transcription factors [50; 54] that can be modified by flavonols [36; 10] and the fact that high flavonol intake (>30 mg/d) decreased serum IL-6 concentrations and high risk and advanced adenoma recurrence in the PPT [3].

A major strength of this study is the information on adenomas from complete colonoscopies performed at baseline, T1, and T4, as well as histologic characteristics noted by two pathologists independently, decreasing the risk of misclassification. Another strength is that the dietary questionnaire, developed specifically for this study, focused on fruit & vegetable consumption [2829; 27], and the questionnaire was linked to the recently released validated USDA flavonoid database [30]. Furthermore, registered dieticians reviewed the completed dietary questionnaires with participants, which further improved the accuracy of the questionnaire [29; 27]. Other strengths of this study included the prospective and repeated collection of both the serum and dietary data.

There are, however, several limitations to our study. Our study findings may not apply to the general population because all participants had a history of adenomas, a relative healthy diet, and most engaged in a health-promoting lifestyle. The observed associations could be due to chance, multiple comparisons, or to IL SNP being in linkage disequilibrium with other functional or regulatory SNPs [23]. Dietary measurement error related to the dietary assessment technique cannot be ruled out and could lead to attenuated risk estimates. The dietary intervention of the trial was not specific to flavonol consumption; therefore, observed changes could be the result of other chemoprotective compounds known to decrease colorectal adenoma risk, such as dry beans, fiber, and folate. However, none of the other chemopreventive compounds had a significant effect on adenoma recurrence in the PPT, except for dry beans, which have a high flavonol content. Furthermore, we had limited statistical power for testing interactions between IL SNPs, flavonol intake, serum IL concentrations, and advanced adenoma recurrence. However, this is, to our knowledge, the first study to investigate the joint effects of IL genotype, serum IL levels, and flavonol intake on adenoma recurrence.

In conclusion, our results suggest that IL SNPs, in combination with a flavonol-rich diet or with a decrease in serum IL concentrations, may decrease the risk of adenoma recurrence. Further studies are needed to examine the importance and biological role of IL SNPs for chemopreventive strategies to identify individuals more likely to benefit from chemopreventive compounds such as flavonols.

Acknowledgments

We would like to thank the Polyp Prevention Trial Study Group for their outstanding contribution to this project. The authors thank Helen Rager and Yanyu Wang from the Clinical Support Laboratory of SAIC Frederick, Inc. (Frederick, MD) for cytokine analysis of the serum samples.

Funding: This study was funded by the Office of Dietary Supplements (OD-08-007) and the Intramural Research Program, National Cancer Institute, NIH, Bethesda, MD.

Abbreviations

BMI

body mass index

CI

confidence interval

CV

coefficient of variation

FFQ

food frequency questionnaire

IL

interleukin

IQR

interquartile range

NSAID

nonsteroidal anti-inflammatory drug

OR

odds ratio

PPT

Polyp Prevention Trial

SNP

single nucleotide polymorphism

Appendix

The members of the Polyp Prevention Study Group participated in the conduct of the Polyp Prevention Trial. However, the data presented in this manuscript and the conclusions drawn from them are solely the responsibility of the above listed coauthors.

National Cancer Institute—Schatzkin, A., Lanza, E., Cross, A.J., Corle, D., Freedman, L.S., Clifford, C., Tangrea, J.; Bowman Gray School of Medicine—Cooper, M.R., Paskett, E. (currently Ohio State University), Quandt, S., DeGraffinreid, C., Bradham, K., Kent, L., Self, M., Boyles, D., West, D., Martin, L., Taylor, N., Dickenson, E., Kuhn, P., Harmon, J., Richardson, I., Lee, H., Marceau, E.; University of New York at Buffalo—Lance, M.P. (currently University of Arizona), Marshall, J.R. (currently Roswell Park Cancer Center), Hayes, D., Phillips, J., Petrelli, N., Shelton, S., Randall, E., Blake, A., Wodarski, L., Deinzer, M., Melton, R.; Edwards Hines, Jr. Hospital, Veterans Administration Medical Center—Iber, F.L., Murphy, P., Bote, E.C., Brandt-Whittington, L., Haroon, N., Kazi, N., Moore, M.A., Orloff, S.B., Ottosen, W.J., Patel, M., Rothschild, R.L., Ryan, M., Sullivan, J.M., Verma, A.; Kaiser Foundation Research Institute—Caan, B., Selby, J.V., Friedman, G., Lawson, M., Taff, G., Snow, D., Belfay, M., Schoenberger, M., Sampel, K., Giboney, T., Randel, M.; Memorial Sloan-Kettering Cancer Center—Shike, M., Winawer, S., Bloch, A., Mayer, J., Morse, R., Latkany, L., D’Amato, D., Schaffer, A., Cohen, L.; University of Pittsburgh—Weissfeld, J., Schoen, R., Schade, R.R., Kuller, L., Gahagan, B., Caggiula, A., Lucas, C., Coyne, T., Pappert, S., Robinson, R., Landis, V., Misko, S., Search, L.; University of Utah—Burt, R.W., Slattery, M., Viscofsky, N., Benson, J., Neilson, J., McDivitt, R., Briley, M., Heinrich, K., Samowitz, W.; Walter Reed Army Medical Center—Kikendall, J.W., Mateski, D.J., Wong, R., Stoute, E., Jones-Miskovsky, V., Greaser, A., Hancock, S., Chandler, S.; Data and Nutrition Coordinating Center (Westat)—Cahill, J., Hasson, M., Daston, C., Brewer, B., Zimmerman, T., Sharbaugh, C., O’Brien, B., Cranston, L., Odaka, N., Umbel, K., Pinsky, J., Price, H., Slonim, A.; Central Pathologists—Lewin, K. (University of California, Los Angeles), Appelman, H. (University of Michigan); Laboratories—Bachorik, P.S., Lovejoy, K. (Johns Hopkins University); Sowell, A. (Centers for Disease Control); Data and Safety Monitoring Committee—Greenberg, E.R. (chair) (Dartmouth University); Feldman, E. (Augusta, Georgia); Garza, C. (Cornell University); Summers, R. (University of Iowa); Weiand, S. (through June 1995) (University of Minnesota); DeMets, D. (beginning July 1995) (University of Wisconsin).

Footnotes

Conflict of interest: None

References

  • 1.Chun OK, Chung SJ, Song WO. Estimated dietary flavonoid intake and major food sources of U.S. adults. J Nutr. 2007;137:1244–1252. doi: 10.1093/jn/137.5.1244. [DOI] [PubMed] [Google Scholar]
  • 2.Bobe G, Sansbury LB, Albert PS, Cross AJ, Kahle L, Ashby J, et al. Dietary flavonoids and colorectal adenoma recurrence in the Polyp Prevention Trial. Cancer Epidemiol Biomarkers Prev. 2008;17:1344–1353. doi: 10.1158/1055-9965.EPI-07-0747. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 3.Bobe G, Albert PS, Sansbury LB, Lanza E, Schatzkin A, Colburn NH, et al. Interleukin-6 as a potential indicator for prevention of high-risk adenoma recurrence by dietary flavonols in the polyp prevention trial. Cancer Prev Res (Phila Pa) 2010;3:764–775. doi: 10.1158/1940-6207.CAPR-09-0161. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 4.Wang L, Tu YC, Lian TW, Hung JT, Yen JH, Wu MJ. Distinctive antioxidant and antiinflammatory effects of flavonols. J Agric Food Chem. 2006;54:9798–9804. doi: 10.1021/jf0620719. [DOI] [PubMed] [Google Scholar]
  • 5.Gupta C, Vikram A, Tripathi DN, Ramarao P, Jena GB. Antioxidant and antimutagenic effect of quercetin against DEN induced hepatotoxicity in rat. Phytother Res. 2010;24:119–128. doi: 10.1002/ptr.2883. [DOI] [PubMed] [Google Scholar]
  • 6.Kim WK, Bang MH, Kim ES, Kang NE, Jung KC, Cho HJ, et al. Quercetin decreases the expression of ErbB2 and ErbB3 proteins in HT-29 human colon cancer cells. J Nutr Biochem. 2005;16:155–162. doi: 10.1016/j.jnutbio.2004.10.010. [DOI] [PubMed] [Google Scholar]
  • 7.Chun OK, Chung SJ, Claycombe KJ, Song WO. Serum C-reactive protein concentrations are inversely associated with dietary flavonoid intake in U.S. adults. J Nutr. 2008;138:753–760. doi: 10.1093/jn/138.4.753. [DOI] [PubMed] [Google Scholar]
  • 8.Boots AW, Drent M, Swennen EL, Moonen HJ, Bast A, Haenen GR. Antioxidant status associated with inflammation in sarcoidosis: a potential role for antioxidants. Respir Med. 2009;103:364–372. doi: 10.1016/j.rmed.2008.10.007. [DOI] [PubMed] [Google Scholar]
  • 9.Egert S, Boesch-Saadatmandi C, Wolffram S, Rimbach G, Muller MJ. Serum lipid and blood pressure responses to quercetin vary in overweight patients by apolipoprotein E genotype. J Nutr. 2010;140:278–284. doi: 10.3945/jn.109.117655. [DOI] [PubMed] [Google Scholar]
  • 10.Lee KW, Kang NJ, Heo YS, Rogozin EA, Pugliese A, Hwang MK, et al. Raf and MEK protein kinases are direct molecular targets for the chemopreventive effect of quercetin, a major flavonol in red wine. Cancer Res. 2008;68:946–955. doi: 10.1158/0008-5472.CAN-07-3140. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 11.Jeong JH, An JY, Kwon YT, Rhee JG, Lee YJ. Effects of low dose quercetin: cancer cell-specific inhibition of cell cycle progression. J Cell Biochem. 2009;106:73–82. doi: 10.1002/jcb.21977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 12.Luo H, Rankin GO, Liu L, Daddysman MK, Jiang BH, Chen YC. Kaempferol inhibits angiogenesis and VEGF expression through both HIF dependent and independent pathways in human ovarian cancer cells. Nutr Cancer. 2009;61:554–563. doi: 10.1080/01635580802666281. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 13.Zhang W, Zhang F. Effects of quercetin on proliferation, apoptosis, adhesion and migration, and invasion of HeLa cells. Eur J Gynaecol Oncol. 2009;30:60–64. [PubMed] [Google Scholar]
  • 14.Krelin Y, Voronov E, Dotan S, Elkabets M, Reich E, Fogel M, et al. Interleukin-1beta-driven inflammation promotes the development and invasiveness of chemical carcinogen-induced tumors. Cancer Res. 2007;67:1062–1071. doi: 10.1158/0008-5472.CAN-06-2956. [DOI] [PubMed] [Google Scholar]
  • 15.Lin WW, Karin M. A cytokine-mediated link between innate immunity, inflammation, and cancer. J Clin Invest. 2007;117:1175–1183. doi: 10.1172/JCI31537. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 16.Waugh DJ, Wilson C. The interleukin-8 pathway in cancer. Clin Cancer Res. 2008;14:6735–6741. doi: 10.1158/1078-0432.CCR-07-4843. [DOI] [PubMed] [Google Scholar]
  • 17.Grivennikov S, Karin E, Terzic J, Mucida D, Yu GY, Vallabhapurapu S, et al. IL-6 and Stat3 are required for survival of intestinal epithelial cells and development of colitis-associated cancer. Cancer Cell. 2009;15:103–113. doi: 10.1016/j.ccr.2009.01.001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 18.Okoko T, Oruambo IF. Inhibitory activity of quercetin and its metabolite on lipopolysaccharide-induced activation of macrophage U937 cells. Food Chem Toxicol. 2009;47:809–812. doi: 10.1016/j.fct.2009.01.013. [DOI] [PubMed] [Google Scholar]
  • 19.Sturlan S, Oberhuber G, Beinhauer BG, Tichy B, Kappel S, Wang J, et al. Interleukin-10-deficient mice and inflammatory bowel disease associated cancer development. Carcinogenesis. 2001;22:665–671. doi: 10.1093/carcin/22.4.665. [DOI] [PubMed] [Google Scholar]
  • 20.Fishman D, Faulds G, Jeffery R, Mohamed-Ali V, Yudkin JS, Humphries S, et al. The effect of novel polymorphisms in the interleukin-6 (IL-6) gene on IL-6 transcription and plasma IL-6 levels, and an association with systemic-onset juvenile chronic arthritis. J Clin Invest. 1998;102:1369–1376. doi: 10.1172/JCI2629. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 21.Hwang IR, Kodama T, Kikuchi S, Sakai K, Peterson LE, Graham DY, et al. Effect of interleukin 1 polymorphisms on gastric mucosal interleukin 1beta production in Helicobacter pylori infection. Gastroenterology. 2002;123:1793–1803. doi: 10.1053/gast.2002.37043. [DOI] [PubMed] [Google Scholar]
  • 22.Reuss E, Fimmers R, Kruger A, Becker C, Rittner C, Hohler T. Differential regulation of interleukin-10 production by genetic and environmental factors--a twin study. Genes Immun. 2002;3:407–413. doi: 10.1038/sj.gene.6363920. [DOI] [PubMed] [Google Scholar]
  • 23.Hacking D, Knight JC, Rockett K, Brown H, Frampton J, Kwiatkowski DP, et al. Increased in vivo transcription of an IL-8 haplotype associated with respiratory syncytial virus disease-susceptibility. Genes Immun. 2004;5:274–282. doi: 10.1038/sj.gene.6364067. [DOI] [PubMed] [Google Scholar]
  • 24.Rad R, Dossumbekova A, Neu B, Lang R, Bauer S, Saur D, et al. Cytokine gene polymorphisms influence mucosal cytokine expression, gastric inflammation, and host specific colonisation during Helicobacter pylori infection. Gut. 2004;53:1082–1089. doi: 10.1136/gut.2003.029736. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 25.Lanza E, Schatzkin A, Ballard-Barbash R, Corle D, Clifford C, Paskett E, et al. The polyp prevention trial II: dietary intervention program and participant baseline dietary characteristics. Cancer Epidemiol Biomarkers Prev. 1996;5:385–392. [PubMed] [Google Scholar]
  • 26.Schatzkin A, Lanza E, Corle D, Lance P, Iber F, Caan B, et al. Lack of effect of a low-fat, high-fiber diet on the recurrence of colorectal adenomas. Polyp Prevention Trial Study Group. N Engl J Med. 2000;342:1149–1155. doi: 10.1056/NEJM200004203421601. [DOI] [PubMed] [Google Scholar]
  • 27.Lanza E, Schatzkin A, Daston C, Corle D, Freedman L, Ballard-Barbash R, et al. Implementation of a 4-y, high-fiber, high-fruit-and-vegetable, low-fat dietary intervention: results of dietary changes in the Polyp Prevention Trial. Am J Clin Nutr. 2001;74:387–401. doi: 10.1093/ajcn/74.3.387. [DOI] [PubMed] [Google Scholar]
  • 28.Block G, Hartman AM, Naughton D. A reduced dietary questionnaire: development and validation. Epidemiology. 1990;1:58–64. doi: 10.1097/00001648-199001000-00013. [DOI] [PubMed] [Google Scholar]
  • 29.Caan BJ, Lanza E, Schatzkin A, Coates AO, Brewer BK, Slattery ML, et al. Does nutritionist review of a self-administered food frequency questionnaire improve data quality? Public Health Nutr. 1999;2:565–569. doi: 10.1017/s1368980099000750. [DOI] [PubMed] [Google Scholar]
  • 30.U.S. Department of Agriculture ARS. Nutrient Data Laboratory web site. USDA; 2007. USDA database for the flavonoid content of selected foods. [Google Scholar]
  • 31.Gunter MJ, Canzian F, Landi S, Chanock SJ, Sinha R, Rothman N. Inflammation-related gene polymorphisms and colorectal adenoma. Cancer Epidemiol Biomarkers Prev. 2006;15:1126–1131. doi: 10.1158/1055-9965.EPI-06-0042. [DOI] [PubMed] [Google Scholar]
  • 32.Ito H, Kaneko K, Makino R, Konishi K, Kurahashi T, Yamamoto T, et al. Interleukin-1beta gene in esophageal, gastric and colorectal carcinomas. Oncol Rep. 2007;18:473–481. [PubMed] [Google Scholar]
  • 33.Tsigris C, Chatzitheofylaktou A, Xiromeritis C, Nikiteas N, Yannopoulos A. Genetic association studies in digestive system malignancies. Anticancer Res. 2007;27:3577–3587. [PubMed] [Google Scholar]
  • 34.Cacev T, Radosevic S, Krizanac S, Kapitanovic S. Influence of interleukin-8 and interleukin-10 on sporadic colon cancer development and progression. Carcinogenesis. 2008;29:1572–1580. doi: 10.1093/carcin/bgn164. [DOI] [PubMed] [Google Scholar]
  • 35.Tsilidis KK, Helzlsouer KJ, Smith MW, Grinberg V, Hoffman-Bolton J, Clipp SL, et al. Association of common polymorphisms in IL10, and in other genes related to inflammatory response and obesity with colorectal cancer. Cancer Causes Control. 2009;20:1739–1751. doi: 10.1007/s10552-009-9427-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 36.Hungness ES, Robb BW, Luo GJ, Pritts TA, Hershko DD, Hasselgren PO. Proteasome inhibitors activate the transcription factors C/EBP-beta and delta in human intestinal epithelial cells. Biochem Biophys Res Commun. 2002;290:469–474. doi: 10.1006/bbrc.2001.6168. [DOI] [PubMed] [Google Scholar]
  • 37.Kumamoto T, Fujii M, Hou DX. Myricetin directly targets JAK1 to inhibit cell transformation. Cancer Lett. 2009;275:17–26. doi: 10.1016/j.canlet.2008.09.027. [DOI] [PubMed] [Google Scholar]
  • 38.Comalada M, Ballester I, Bailon E, Sierra S, Xaus J, Galvez J, et al. Inhibition of pro-inflammatory markers in primary bone marrow-derived mouse macrophages by naturally occurring flavonoids: analysis of the structure-activity relationship. Biochem Pharmacol. 2006;72:1010–1021. doi: 10.1016/j.bcp.2006.07.016. [DOI] [PubMed] [Google Scholar]
  • 39.Sharma V, Joseph C, Ghosh S, Agarwal A, Mishra MK, Sen E. Kaempferol induces apoptosis in glioblastoma cells through oxidative stress. Mol Cancer Ther. 2007;6:2544–2553. doi: 10.1158/1535-7163.MCT-06-0788. [DOI] [PubMed] [Google Scholar]
  • 40.Sansbury LB, Bergen AW, Wanke KL, Yu B, Caporaso NE, Chatterjee N, et al. Inflammatory cytokine gene polymorphisms, nonsteroidal anti-inflammatory drug use, and risk of adenoma polyp recurrence in the polyp prevention trial. Cancer Epidemiol Biomarkers Prev. 2006;15:494–501. doi: 10.1158/1055-9965.EPI-05-0763. [DOI] [PubMed] [Google Scholar]
  • 41.Kokoris M, Dix K, Moynihan K, Mathis J, Erwin B, Grass P, et al. High-throughput SNP genotyping with the Masscode system. Mol Diagn. 2000;5:329–340. doi: 10.1007/BF03262094. [DOI] [PubMed] [Google Scholar]
  • 42.Hartman TJ, Yu B, Albert PS, Slattery ML, Paskett E, Kikendall JW, et al. Does nonsteroidal anti-inflammatory drug use modify the effect of a low-fat, high-fiber diet on recurrence of colorectal adenomas? Cancer Epidemiol Biomarkers Prev. 2005;14:2359–2365. doi: 10.1158/1055-9965.EPI-05-0333. [DOI] [PubMed] [Google Scholar]
  • 43.Schetter AJ, Nguyen GH, Bowman ED, Mathe EA, Yuen ST, Hawkes JE, et al. Association of inflammation-related and microRNA gene expression with cancer-specific mortality of colon adenocarcinoma. Clin Cancer Res. 2009;15:5878–5887. doi: 10.1158/1078-0432.CCR-09-0627. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 44.Yang HP, Woodson K, Taylor PR, Pietinen P, Albanes D, Virtamo J, et al. Genetic variation in interleukin 8 and its receptor genes and its influence on the risk and prognosis of prostate cancer among Finnish men in a large cancer prevention trial. Eur J Cancer Prev. 2006;15:249–253. doi: 10.1097/01.cej.0000199504.07947.e7. [DOI] [PubMed] [Google Scholar]
  • 45.Hubner RA, Muir KR, Liu JF, Logan RF, Grainge MJ, Houlston RS. Polymorphisms in PTGS1, PTGS2 and IL-10 do not influence colorectal adenoma recurrence in the context of a randomized aspirin intervention trial. Int J Cancer. 2007;121:2001–2004. doi: 10.1002/ijc.22942. [DOI] [PubMed] [Google Scholar]
  • 46.Slattery ML, Wolff RK, Herrick JS, Caan BJ, Potter JD. IL6 genotypes and colon and rectal cancer. Cancer Causes Control. 2007;18:1095–1105. doi: 10.1007/s10552-007-9049-x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 47.Zumkeller N, Brenner H, Chang-Claude J, Hoffmeister M, Nieters A, Rothenbacher D. Helicobacter pylori infection, interleukin-1 gene polymorphisms and the risk of colorectal cancer: evidence from a case-control study in Germany. Eur J Cancer. 2007;43:1283–1289. doi: 10.1016/j.ejca.2007.03.005. [DOI] [PubMed] [Google Scholar]
  • 48.Vasku A, Vokurka J, Bienertova-Vasku J. Obesity-related genes variability in Czech patients with sporadic colorectal cancer: preliminary results. Int J Colorectal Dis. 2009;24:289–294. doi: 10.1007/s00384-008-0553-6. [DOI] [PubMed] [Google Scholar]
  • 49.Schetter AJ, Heegaard NH, Harris CC. Inflammation and cancer: interweaving microRNA, free radical, cytokine and p53 pathways. Carcinogenesis. 31:37–49. doi: 10.1093/carcin/bgp272. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 50.Vanden Berghe W, De Bosscher K, Boone E, Plaisance S, Haegeman G. The nuclear factor-kappaB engages CBP/p300 and histone acetyltransferase activity for transcriptional activation of the interleukin-6 gene promoter. J Biol Chem. 1999;274:32091–32098. doi: 10.1074/jbc.274.45.32091. [DOI] [PubMed] [Google Scholar]
  • 51.Crawley E, Kay R, Sillibourne J, Patel P, Hutchinson I, Woo P. Polymorphic haplotypes of the interleukin-10 5′ flanking region determine variable interleukin-10 transcription and are associated with particular phenotypes of juvenile rheumatoid arthritis. Arthritis Rheum. 1999;42:1101–1118. doi: 10.1002/1529-0131(199906)42:6<1101::AID-ANR6>3.0.CO;2-Y. [DOI] [PubMed] [Google Scholar]
  • 52.Rees LE, Wood NA, Gillespie KM, Lai KN, Gaston K, Mathieson PW. The interleukin-10-1082 G/A polymorphism: allele frequency in different populations and functional significance. Cell Mol Life Sci. 2002;59:560–569. doi: 10.1007/s00018-002-8448-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 53.Temple SE, Lim E, Cheong KY, Almeida CA, Price P, Ardlie KG, et al. Alleles carried at positions -819 and -592 of the IL10 promoter affect transcription following stimulation of peripheral blood cells with Streptococcus pneumoniae. Immunogenetics. 2003;55:629–632. doi: 10.1007/s00251-003-0621-6. [DOI] [PubMed] [Google Scholar]
  • 54.Terry CF, Loukaci V, Green FR. Cooperative influence of genetic polymorphisms on interleukin 6 transcriptional regulation. J Biol Chem. 2000;275:18138–18144. doi: 10.1074/jbc.M000379200. [DOI] [PubMed] [Google Scholar]

RESOURCES