David J. Brull

Common Promoter Variant in Cyclooxygenase-2 Represses Gene Expression Evidence of Role in Acute-Phase Inflammatory Response

Objective—Cyclooxygenase (COX)-2 is a key regulatory enzyme in the synthesis of prostanoids associated with trauma and inflammation. We investigated the COX-2 gene for functional variants that may influence susceptibility to disease. Methods and Results—The promoter of COX-2 was screened for variants in healthy subjects by use of polymerase chain reaction-based methods. Promoter activity was investigated by using reporter expression experiments in human lung fibroblasts. Patients undergoing coronary artery bypass graft surgery, with measurements of plasma markers linked to COX-2 activity, were genotyped for association studies. A common COX-2 promoter variant, −765G>C, was found and shown to be carried by >25% of a group of healthy UK subjects. The −765C allele had significantly lower promoter activity compared with −765G, basally (28±3% lower, P <0.005) and in serum-stimulated cells (31±2% lower, P <0.005). In patients subjected to coronary artery bypass graft surgery, the magnitude of rise in levels of C-reactive protein (CRP) was strongly genotype dependent. Compared with −765G homozygotes, patients carrying the −765C allele had significantly lower plasma CRP levels at 1 to 4 days after surgery (14% lower at the peak of CRP levels on day 3, P <0.05 for all time points). Conclusions—For several acute and chronic inflammatory diseases, −765G>C may influence the variability of response observed.

Human CRP gene polymorphism influences CRP levels: implications for the prediction and pathogenesis of coronary heart disease.

Objective—C-reactive protein (CRP) concentrations are predictive of cardiovascular disease, and levels are heritable, in part. We identified novel polymorphisms in the CRP gene and assessed their influence on CRP level. Methods and Results—CRP was measured in 250 male army recruits before and after strenuous exercise and perioperatively in 193 coronary artery bypass graft (CABG) patients. Two novel polymorphisms were identified in the CRP gene, −717G>A in the promoter and +1444C>T in the 3′UTR. Among army recruits, CRP was higher in +1444TT homozygotes than +1444 C-allele carriers at baseline (1.04±0.38 versus 0.55±0.06, P =0.014) and at all time points after exercise (2.35±0.68 versus 1.07±0.12, 2.11±0.53 versus 0.88±0.09, and 1.77±0.44 versus 0.71±0.09, P =0.034, P =0.007, and P =0.013, at 2, 48, and 96 hours after exercise, respectively). In the CABG patients, mean CRP (mg/L) rose from 1.97±0.36 at baseline to 167.2±5.0 72 hours postoperatively. Genotype did not influence CRP at baseline; however, peak post-CABG CRP levels were higher in +1444TT homozygotes compared with +1444C-allele carriers (198±17 versus 164±5, P =0.03). Conclusions—The CRP gene +1444C>T variant influences basal and stimulated CRP level. These findings have implications both for the prediction and pathogenesis of coronary heart disease.

Bradykinin B2BKR receptor polymorphism and left-ventricular growth response

Angiotensin-converting-enzyme (ACE) activity regulates left-ventricular growth. The deletion (D), rather than the insertion (I), ACE gene variant is associated with increased ACE activity and kinin degradation, and the absence (-) rather than the presence (+) of a 9 bp deletion in the gene encoding the bradykinin 2 receptor (B2BKR) is associated with greater gene expression. We determined the ACE and B2BKR genotype of 109 male army recruits, and measured their physiological left-ventricular growth response to a 10-week physical training programme. Mean left-ventricular growth was 15.7 g (SE 3.5) in those with ACE genotype D/D and B2BKR genotype +9/+9, but -1.37 g (4.1) in those with ACE genotype I/I and B2BKR genotype -9/-9 (p=0.003 for trend across genotypes). These results suggest that kinins regulate left-ventricular growth, mediating some of the effects of ACE in this regard.

Impact of angiotensin converting enzyme inhibition on post-coronary artery bypass interleukin 6 release

Background: Angiotensin 1 converting enzyme (ACE) inhibitors reduce morbidity and mortality after coronary artery bypass graft surgery (CABG). This benefit may result from an anti-inflammatory action. Objective: To examine the effect of ACE inhibition on interleukin 6 (IL-6) concentrations after CABG. Patients and methods: 161 patients undergoing elective first time CABG were recruited, of whom 41 (25%) were receiving ACE inhibitor treatment; 21 patients with confounding postoperative complications were excluded. After these exclusions there were 33 patients (24%) on ACE inhibitor treatment. Plasma IL-6 was measured preoperatively and again six hours after CABG. Results: Baseline IL-6 concentrations (geometric mean (SEM)) were non-significantly lower among the patients receiving ACE inhibitors (3.7 (0.1) v 4.3 (0.1) pg/ml, p = 0.12). Overall, post-CABG IL-6 concentrations increased significantly (mean rise 177 (12) pg/ml, p < 0.0005). This response was blunted among ACE inhibitor treated patients. Median increases in IL-6 concentrations were 117 v 193 pg/ml, for treated v non-treated patients, respectively (Kruskal–Wallis, p = 0.02), with peak postoperative IL-6 concentrations lower among the subjects receiving ACE inhibitors than in untreated subjects (142 (19) v 196 (13) pg/ml, p = 0.02). The effect of ACE inhibitors remained significant after multivariate analysis (p = 0.018). Conclusions: ACE inhibitor treatment is associated with a reduction in IL-6 response to CABG. The data suggest that this class of drug may have a direct anti-inflammatory effect, which could explain some of its clinical benefit.

Statin therapy and the acute inflammatory response after coronary artery bypass grafting

Preoperative statin therapy has been shown to improve cardiovascular outcome after coronary artery bypass grafting; however, the clinical gain achieved is in excess of the reduction in lipids. We demonstrate that statin therapy exerts a direct anti-inflammatory effect mediated through reduced interleukin-6 production.

The effect of the Interleukin-6–174G > C promoter gene polymorphism on endothelial function in healthy volunteers

Aims  Atherosclerosis is a chronic inflammatory condition, manifest in its early stages by endothelial dysfunction. Interleukin‐6 (IL6) plays a key role in driving this process through stimulation of acute phase protein synthesis. We have examined the effect of the IL6 gene −174G > C promoter polymorphism on endothelial function in a group of healthy subjects.

Variation in the Interleukin-6 Gene Is Associated with Impaired Cognitive Development in Children Born Prematurely: A Preliminary Study

The pro-inflammatory cytokine IL-6 may be neurocytopathogenic, and elevated levels are associated with impaired neurological outcome among children born prematurely. However, the precise mechanisms underlying this association remain unclear. The rare C (rather than G) variant at position –572 in the IL-6 gene is associated with an increased IL-6 synthetic response. If IL-6 mediates cerebral injury, we would anticipate the –572 C allele to be associated with impaired childhood development. We have examined this hypothesis, studying 113 Caucasian children born at ≤32 wk gestation. Cognitive and motor functions were assessed using the Griffiths Developmental Scales at 2 y and British Ability Scales (2nd Ed.) and the ABC Movement Score at 51/2 y. Performance (median, interquartile range) in all three scales was worse in the 10 carriers of the C allele than for those with GG genotype: Griffiths Developmental Quotient: C allele, 92.4 (89.9–96.6) versus CG 100.9 (96.7–104.8), p = 0.002; General Cognitive Ability: C allele, 88.0 (80.3–102.8) versus GG 103.0 (92.0–112.0), p = 0.037; Movement ABC score: C allele 8.3 (6.6–20.3) versus GG 4.0 (1.0–9.5), p = 0.081. The presence of the rare (≥1) IL-6 -572 C-allele (CC+GC genotypes) is associated with impaired cognitive development among children born before 32 wk gestation. These data support a role for IL-6 in the genesis of neurologic impairment in such children.

Effect of a COL1A1 Sp1 binding site polymorphism on arterial pulse wave velocity: an index of compliance.

Reduced arterial compliance precedes changes in blood pressure, which may be mediated through alterations in vessel wall matrix composition. We investigated the effect of the collagen type I-&agr;1 gene (COL1A1) +2046G>T polymorphism on arterial compliance in healthy individuals. We recruited 489 subjects (251 men and 238 women; mean age, 22.6±1.6 years). COL1A1 genotypes were determined using polymerase chain reaction and digestion by restriction enzyme Bal 1. Arterial pulse wave velocities were measured in 3 segments, aortoiliac (PWVA), aortoradial (PWVB), and aorto-dorsalis-pedis (PWVF), as an index of compliance using a noninvasive optical method. Data were available for 455 subjects. The sample was in Hardy-Weinberg equilibrium with genotype distributions and allele frequencies that were not significantly different from those reported previously. The T allele frequency was 0.22 (95% confidence interval, 0.19 to 0.24). Two hundred eighty-three (62.2%) subjects were genotype GG, 148 (35.5%) subjects were genotype GT, and 24 (5.3%) subjects were genotype TT. A comparison of GG homozygotes with GT and TT individuals demonstrated a statistically significant association with arterial compliance: PWVF 4.92±0.03 versus 5.06±0.05 m/s (ANOVA, P =0.009), PWVB 4.20±0.03 versus 4.32±0.04 m/s (ANOVA, P =0.036), and PWVA 3.07±0.03 versus 3.15±0.03 m/s (ANOVA, P =0.045). The effects of genotype were independent of age, gender, smoking, mean arterial pressure, body mass index, family history of hypertension, and activity scores. We report an association between the COL1A1 gene polymorphism and arterial compliance. Alterations in arterial collagen type 1A deposition may play a role in the regulation of arterial compliance.

Cortical bone resorption during exercise is interleukin-6 genotype-dependent

The objective of this study was to examine the relationship between the interleukin-6 (IL-6) −174 G>C promoter polymorphism and exercise-induced femoral cortical bone resorption. Skeletal response to exercise was assessed in 130 male Caucasian army recruits. Five cross-sectional magnetic resonance images of the right femur were obtained before and after a 10-week period of basic physical training, and changes in cross-sectional cortical area were calculated. Recruits were genotyped for the −174 G>C IL-6 promoter polymorphism. Genotype frequencies (GG 36%, GC 47%, CC 22 17%) were in Hardy–Weinberg equilibrium. The mean percentage change in proximal femoral cross-sectional cortical area was strongly IL-6 genotype-dependent, with GG homozygotes losing 6.8 (3.82)% in cortical area, GC gaining +5.5 (4.88)% and CC gaining +17.3 (9.46)% ( P =0.007 for linear trend). These changes persisted throughout the right femur and were significant in the femur as a whole ( P =0.03). This study demonstrates an association between a functional polymorphism in the IL-6 gene and femoral cortical remodelling during strenuous physical exercise. Previous studies have suggested an important role for IL-6 in the regulation of bone mass in postmenopausal women, and in the invasion of bone by metastatic tumour deposits. These data extend these observations to the regulation of bone mass in healthy males, supporting a fundamental role for IL-6 in the regulation of bone mass and bone remodelling in humans.

Free Interleukin (IL)-18 Levels, and the Impact of IL18 and IL18BP Genetic Variation, in CHD Patients and Healthy Men

Objective—To investigate free interleukin-18 (fIL-18) levels, and variation within the IL-18 system genes, in heart surgery patients, and healthy men. Methods and Results—fIL-18 was calculated from IL-18 and IL-18 binding protein (BP) levels, in 421 healthy men and 196 post–coronary artery bypass graft (CABG) patients. After surgery, fIL-18 peaked at 6 hours (from 117 to 331 pg/mL) but fell to below presurgery levels at 24 hours (99 pg/mL), because of changes in total IL-18 and IL-18BP. fIL-18 24 hours postsurgery was significantly higher in those who suffered a major complication after surgery (125 versus 80 pg/mL, P<0.01). Baseline total IL-18 was also higher in healthy men who went on to suffer an MI over 17 years of prospective study (276 versus 240 pg/mL, P=0.01). Tagging SNPs for IL18 (n=5) and IL18BP (n=3) were determined, in both studies the IL18 HapIII haplotype (frequency 30%) was associated with 36% lower baseline fIL-18 levels before surgery (P<0.01), and 7% lower in healthy men (P=0.04). The frequency of HapIII was lower in CABG patients than in healthy men (20.7 versus 29.8%, P<0.01). Conclusions—IL-18 levels, which are determined in part by variation in IL18, play a role in CHD development and postsurgery outcome.