Background Fatty degeneration of thymus (or thymus involution) has long been considered a normal ageing process. However, there is emerging evidence that thymic involution is linked to T cell aging, chronic inflammation and increased morbidity. Other factors, aside from chronological age, have been proposed to affect the involution rate. In the present study, we investigated the imaging characteristics of thymus on computed tomography (CT) in a Swedish middle-aged population. The major aims were to establish the prevalence of fatty degeneration of thymus and to determine its associations with demographic, lifestyle and clinical factors, as well as inflammation, T cell differentiation and thymic output. Results In total, 1 048 randomly invited individuals (aged 50-64 years, 49% females) were included and thoroughly characterized. CT evaluation of thymus included measurements of attenuation, size and a 4-point scoring system, with scale 0-3 based on the ratio of fat and soft tissue. A majority, 615 (59%) showed complete fatty degeneration, 259 (25%) predominantly fatty attenuation, 105 (10%) half fatty and half soft-tissue attenuation, while 69 (6.6%) presented with a solid thymic gland with predominantly soft-tissue attenuation. Age, male sex, high BMI, abdominal obesity and low dietary intake of fiber were independently associated with complete fatty degeneration of thymus. Also, fatty degeneration of thymus as well as low CT attenuation values were independently related to lower proportion of naive CD8(+) T cells, which in turn was related to lower thymic output, assessed by T- cell receptor excision circle (TREC) levels. Conclusion Among Swedish middle-aged subjects, nearly two-thirds showed complete fatty degeneration of thymus on CT. This was linked to depletion of naive CD8(+) T cells indicating that CT scans of thymus might be used to estimate immunological aging. Furthermore, our findings support the intriguing concept that obesity as well as low fiber intake contribute to immunological aging, thereby raising the possibility of preventive strategies.

OBJECTIVES: To investigate the association between type I collagen α1 chain (COL1α1) levels and coronary artery disease (CAD) by using absolute quantification in plasma. Also, to investigate the correlates of COL1α1 to clinical characteristics and circulating markers of collagen metabolism.

DESIGN: Life conditions, Stress and Health (LSH) study: prospective cohort study, here with a nested case-control design.Assessing Platelet Activity in Coronary Heart Disease (APACHE) study: prospective cohort study.

SETTING: LSH: primary care setting, southeast Sweden.APACHE: cardiology department, university hospital, southeast Sweden.

PARTICIPANTS: LSH: 1007 randomly recruited individuals aged 45-69 (50% women). Exclusion criteria was serious disease. After 13 years of follow-up, 86 cases with primary endpoint were identified and sex-matched/age-matched to 184 controls.

APACHE: 125 patients with myocardial infarction (MI), 73 with ST-elevation MI and 52 with non-ST-elevation MI.

EXCLUSION CRITERIA: Intervention study participation, warfarin treatment and short life expectancy.

PRIMARY AND SECONDARY OUTCOME MEASURES: Primary outcome was the association between baseline COL1α1 and first-time major event of CAD, defined as fatal/non-fatal MI or coronary revascularisation after 13 years. Secondary outcomes were the association between the collagen biomarkers PRO-C1 (N-terminal pro-peptide of type I collagen)/C1M (matrix metalloproteinase-mediated degradation of type I collagen) and CAD; temporal change of COL1α1 after acute MI up to 6 months and lastly, correlates between COL1α1 and patient characteristics along with circulating markers of collagen metabolism.

RESULTS: COL1α1 levels were associated with CAD, both unadjusted (HR=0.69, 95% CI=0.56 to 0.87) and adjusted (HR=0.55, 95% CI=0.41 to 0.75). PRO-C1 was associated with CAD, unadjusted (HR=0.62, 95% CI=0.47 to 0.82) and adjusted (HR=0.61, 95% CI=0.43 to 0.86), while C1M was not. In patients with MI, COL1α1 remained unchanged up to 6 months. COL1α1 was correlated to PRO-C1, but not to C1M.

CONCLUSIONS: Plasma COL1α1 was independently and inversely associated with CAD. Furthermore, COL1α1 appeared to reflect collagen synthesis but not degradation. Future studies are needed to confirm whether COL1α1 is a clinically useful biomarker of CAD.

Purpose: To compare acute effects of moist snuff with or without nicotine and red wine with or without alcohol on prandial hormones and metabolism.Basic procedures and methods: Two deciliters of wine, with or without alcohol, were taken together with a standardized supervised meal in 14 healthy women and men. All participants also combined the meal with usage of with moist snuff, with or without nicotine. The snuff was replaced hourly at each of the four settings, i.e. snuff with or without nicotine combined with red wine with or without alcohol, that started at 0800 oclock and were finished at noon.Main findings: We found ghrelin levels to be more efficiently suppressed when drinking red wine with alcohol compared to non-alcoholic wine by analyzing area under the curve (AUC). AUC for regular wine was 370 +/- 98 pg/ml x hours and 559 +/- 154 pg/ml x hours for de-alcoholized red wine, p < 0.0001 by general linear model. The postprandial metabolic rate was further elevated following alcohol containing red wine compared with nonalcoholic red wine (p = 0.022). Although glucose levels were not uniformly lower after alcoholic red wine, we found lowered glucose levels 3 h after the meal (mean glucose wine: 4.38 +/- 0.96 mmol/l, non-alcoholic wine: 4.81 +/- 0.77 mmol/l, p = 0.005). Nicotine-containing moist snuff (AUC: 1406 +/- 149 nmol/ml x hours) elevated the levels of serum cortisol compared with nicotine-free snuff (AUC: 1268 +/- 119 nmol/ml x hours, p = 0.005). We found no effects of nicotine or alcohol on feelings of satiety.Conclusions: Alcohol in red wine augmented the postprandial suppression of ghrelin and it also lowered postprandial glucose 3 h post-meal. These effects are in line with observational trials linking regular intake of moderate amounts of red wine with lower risk for diabetes.

Background: Inflammation and oxidative stress form a vicious circle in atherosclerosis. Oxidative stress can have detrimental effects on T cells. A unique subset of CD4(+) T cells, known as regulatory T (T-reg) cells, has been associated with atheroprotective effects. Reduced numbers of T-reg cells is a consistent finding in patients with chronic coronary syndrome (CCS). However, it is unclear to what extent these cells are sensitive to oxidative stress. In this pilot study, we tested the hypothesis that oxidative stress might be a potential contributor to the T-reg cell deficit in CCS patients. Methods: Thirty patients with CCS and 24 healthy controls were included. T-reg (CD4+CD25+CD127(-)) and conventional T (CD4+CD25(-), T-conv) cells were isolated and treated with increasing doses of H2O2. Intracellular ROS levels and cell death were measured after 2 and 18 h, respectively. The expression of antioxidant genes was measured in freshly isolated T-reg and T-conv cells. Also, total antioxidant capacity (TAC) was measured in fresh peripheral blood mononuclear cells, and oxidized (ox) LDL/LDL ratios were determined in plasma. Results: At all doses of H2O2, T-reg cells accumulated more ROS and exhibited higher rates of death than their T-conv counterparts, p &lt; 0.0001. T-reg cells also expressed higher levels of antioxidant genes, including thioredoxin and thioredoxin reductase-1 (p &lt; 0.0001), though without any differences between CCS patients and controls. T-conv cells from CCS patients were, on the other hand, more sensitive to oxidative stress ex vivo and expressed more thioredoxin reductase-1 than T-conv cells from controls, p &lt; 0.05. Also, TAC levels were lower in patients, 0.97 vs 1.53 UAE/100 mu g, p = 0.001, while oxLDL/LDL ratios were higher, 29 vs 22, p = 0.006. Conclusion: T-reg cells isolated from either CCS patients or healthy controls were all highly sensitive to oxidative stress ex vivo. There were signs of oxidant-antioxidant imbalance in CCS patients and we thus assume that oxidative stress may play a role in the reduction of T-reg cells in vivo.

Vulnerability to stress-induced inflammation has been linked to a dysfunctional hypothalamus–pituitary–adrenal (HPA) axis. In the present study, patients with known or suspected coronary artery disease (CAD) were assessed with respect to inflammatory and HPA axis response to acute physical exercise. An exercise stress test was combined with SPECT myocardial perfusion imaging. Plasma and saliva samples were collected before and 30 min after exercise. Interleukin (IL)-6 and adrenocorticotropic hormone (ACTH) were measured in plasma, while cortisol was measured in both plasma and saliva. In total, 124 patients were included of whom 29% had a prior history of CAD and/or a myocardial perfusion deficit. The levels of exercise intensity and duration were comparable in CAD and non-CAD patients. However, in CAD patients, IL-6 increased after exercise (p = 0.019) while no differences were seen in HPA axis variables. Conversely, patients without CAD exhibited increased levels of ACTH (p = 0.003) and cortisol (p = 0.004 in plasma, p = 0.006 in saliva), but no change in IL-6. We conclude that the IL-6 response to acute physical exercise is exaggerated in CAD patients and may be out of balance due to HPA axis hypoactivity. It remains to be further investigated whether this imbalance is a potential diagnostic and therapeutic target in CAD.

Identification of biomarkers can help monitor and prevent cardiovascular disease (CVD) risk. We performed an exploratory analysis to identify potential biomarkers for coronary heart disease (CHD) in participants from the Life Conditions, Stress, and Health study. A total of 1,007 participants (50% women), randomly selected from the general population, were followed for incident CHD at 8 and 13 years of follow-up. Plasma levels of 184 CVD-related biomarkers were measured in samples collected at baseline in 86 cases with CHD and 184 age- and sex-matched controls by proximity extension assay. Biomarker levels were presented as normalized protein expression values (log 2 scale). After adjusting for confounding factors, 6 biomarkers showed significant association with incident CHD at 13 years. In a sensitivity analysis, this association remained significant at 8 years for 3 biomarkers; collagen alpha-1(I) chain (COL1A1), bone morphogenetic protein-6 (BMP-6), and interleukin-6 receptor alpha chain (IL-6R alpha). When entering these biomarkers in the full adjustment model simultaneously, their association with incident CHD at 13 years remained significant, hazards ratio being 0.671, 0.335, and 2.854, respectively per unit increase in normalized protein expression values. Subjects with low COL1A1, low BMP-6, and high IL-6R alpha levels had a hazards ratio of 5.097 for incident CHD risk (p = 0.019), compared with those without. In conclusion, we identified COL1A1, BMP-6 and IL-6Ra as biomarkers for incident CHD over a long-term follow-up in this exploratory analysis. For COL1A1 and BMP-6 this has not been previously reported. Further studies are needed to confirm our findings and establish their clinical relevance. (C) 2019 Elsevier Inc. All rights reserved.

Background Low-grade systemic inflammation is a predictor of recurrent cardiac events in patients with coronary artery disease (CAD). Plasma proteins such as matrix metalloproteinase (MMP)-9 and myeloperoxidase (MPO) have been shown to reflect basal as well as stress-induced inflammation in CAD. Measurements of MMP-9 and MPO in saliva might pose several advantages. Therefore, we investigated whether salivary levels of MMP-9 and MPO corresponded to plasma levels in patients with coronary artery disease (CAD), both at rest and after acute physical exercise. Methods A bicycle ergometer test was used as a model for stress-induced inflammation. Twenty-three CAD patients performed the test on two occasions 3-6 months apart. Whole unstimulated saliva was collected before, directly after and 30 min after exercise while plasma was collected before and after 30 min. MMP-9 and MPO in saliva and plasma were determined by Luminex. Results MMP-9 and MPO levels were 2- to 4-fold higher in saliva than in plasma. Amongst the saliva samples, and also to a great extent amongst the plasma samples, the levels of both types of protein showed strong intercorrelations between the levels at rest and after exercise during the two visits. However, there were no (or weak) correlations between salivary and plasma MMP-9 and none between salivary and plasma MPO. Conclusion We conclude that salivary diagnostics cannot be used to assess systemic levels of MMP-9 and MPO in CAD patients, neither at rest nor after acute physical exercise.

Objective: Mechanisms behind sustained inflammation in patients with coronary artery disease (CAD) are not clarified but hypothalamus-pituitary-adrenal (HPA) axis dysfunction may have a role. Here, we investigated whether inflammatory status of peripheral blood mononuclear cells (PBMCs) was associated with altered glucocorticoid sensitivity in CAD patients. Methods: In 55 CAD patients and 30 controls, mRNA levels of GR-alpha, GR-beta, NF-kappa B, I kappa B alpha, MMP-9 and TIMP-1 were measured in PBMCs. Suppressive effects of dexamethasone on GR-alpha, GR-beta, NF-kappa B, I kappa B alpha, MMP-9 and TIMP-1 mRNA levels were assessed in PBMCs ex vivo. Salivary cortisol was repeatedly measured over 3 days. Results: GR-alpha mRNA levels were higher in CAD patients than in controls, 0.50 (0.38-0.59) versus 0.26 (0.18-0.37), pamp;lt;.001, while GR-beta mRNA levels were equally low in both groups. GR-alpha mRNA expression was associated with inflammatory gene expression and, also, with flatter diurnal cortisol rhythm. In both patients and controls, dexamethasone suppressed gene expression of NF-B, IB, MMP-9 and TIMP-1 (p amp;lt; .001). Dexamethasone also reduced GR-alpha mRNA levels (p amp;lt; .001), while LPS increased it (p amp;lt; .001). Conclusions: PBMCs from CAD patients displayed an inflammatory gene expression profile. This was not explained by reduced glucocorticoid sensitivity. Instead, inflammation was associated with increased expression of GR-alpha mRNA, suggesting a hypocortisolemic state.

Background and aims: The expression of FOXP3 isoforms affects regulatory T (Treg) cell function. Reduced Treg cell function has been associated with coronary artery disease (CAD). However, alternative splicing of FOXP3 in CAD has not been investigated. Methods: FOXP3 splice variants and IL17A transcripts in peripheral blood mononuclear cells from stable CAD patients and healthy controls were quantified, and FOXP3 isoform expression in response to T cell receptor (TCR) stimulation or LDL was analyzed by flow cytometry. Results: Compared to healthy controls, CAD patients expressed significantly more FOXP3 transcripts that included exon 2, whereas alternative splicing of exon 7 in correlation with IL17A expression was reduced. Moreover, TCR stimulation, as well as exposure to LDL, decreased alternative splicing of FOXP3 in CD4+ T cells in vitro. Conclusions: Our results demonstrate that blood mononuclear cells in stable CAD patients express a ratio of FOXP3 isoforms that is characteristic for activated CD4+ T cells. (C) 2017 Elsevier B.V. All rights reserved.

Annexin A1 (AnxA1) is a key player in resolution of inflammation and a mediator of glucocorticoid actions. In atherosclerotic tissue, increased expression of AnxA1 has been associated with protective plaque-stabilizing effects. Here, we investigated the expression of AnxA1 in peripheral blood mononuclear cells (PBMCs) from patients with coronary artery disease (CAD). Blood was collected from 57 patients with stable CAD (SCAD) and 41 healthy controls. We also included a minor group (n = 10) with acute coronary syndrome (ACS). AnxA1 mRNA was measured in PBMCs. Expression of AnxA1 protein (total and surface-bound) and glucocorticoid receptors (GR) were detected in PBMC subsets by flow cytometry. Also, salivary cortisol, interleukin(IL)-6 and IL-10 in plasma, and LPS-induced cytokine secretion from PBMCs, with or without dexamethasone, were assessed. AnxA1 mRNA was found to be slightly increased in PBMCs from SCAD patients compared with controls. However, protein expression of AnxA1 or GRs in PBMC subsets did not differ between SCAD patients and controls, despite SCAD patients showing a more proinflammatory cytokine profile ex vivo. Only surface expression of AnxA1 on monocytes correlated with dexamethasone-mediated suppression of cytokines. In ACS patients, a marked activation of AnxA1 was seen involving both gene expression and translocation of protein to cell surface probably reflecting a rapid glucocorticoid action modulating the acute inflammatory response in ACS. To conclude, surface expression of AnxA1 on monocytes may reflect the degree of glucocorticoid sensitivity. Speculatively, "normal" surface expression of AnxA1 indicates that anti-inflammatory capacity is impaired in SCAD patients.

BACKGROUND AND AIMS: Many coronary artery disease (CAD) patients exhibit chronic low-grade inflammation. Carotenoids are anti-oxidants with potential anti-inflammatory properties. Here, we first assessed relationships between interleukin (IL)-6 and individual carotenoids in plasma from CAD patients. Based on the results, we proceeded to assess anti-inflammatory effects of one carotenoid, lutein, in peripheral blood mononuclear cells (PBMCs) from CAD patients.

METHODS: Lutein + zeaxanthin (isomers with lutein being dominant), β-cryptoxanthin, lycopene, α- and β-carotene and IL-6 were measured in plasma from 134 patients with stable angina (SA) and 59 patients with acute coronary syndrome. In 42 patients, plasma measurements were also performed 3 months after coronary intervention. PBMCs from SA patients were pre-treated with lutein (1, 5 and 25 μM) for 24 h followed by 24 h incubation ± lipopolysaccharide (LPS). Cell pellets were collected for IL-6, IL-1β and TNF mRNA and intracellular lutein. Cytokine secretion was measured in cell media.

RESULTS: Only lutein + zeaxanthin were inversely correlated with IL-6 in SA patients at baseline (r = -0.366, p < 0.001) and follow-up (r = -0.546, p < 0.001). Ex vivo, lutein was taken up by PBMCs from SA patients in a dose- and time-dependent manner. Pre-treatment with lutein dose-dependently lowered LPS-induced secretion of IL-6, IL-1β (p < 0.01) and TNF (p < 0.05), and also reduced IL-6, IL-1β and TNF mRNA expression (p < 0.05).

CONCLUSIONS: Clinical findings highlighted the inverse association between lutein and IL-6 in CAD patients. Anti-inflammatory effects of lutein in PBMCs from CAD patients were consolidated in ex vivo experiments. Taken together, these results show that lutein has the potential to play a role in resolution of chronic inflammation in CAD patients.

Psychological stress is thought to be an important trigger of cardiovascular events, yet the involved pathways and mediators are largely unknown. Elevated systemic levels of the pro-inflammatory alarmin S100A8/A9 correlate with poor prognosis in coronary artery disease (CAD) patients. Here, we investigated the links between S100A8/A9 release and parameters of anti-inflammatory glucocorticoid secretion in two different cohorts subjected to a psychological stress test. In the first cohort of 60 CAD patients, psychological stress induced a rapid increase of circulating S100A8/A9. This rapid S100A8/A9 response strongly correlated with elevated evening saliva cortisol levels, suggesting an association with a dysregulatedhy pothalamic-pituitary-adrenal (HPA) axis. In the second cohort of 27 CAD patients and 28 controls, elevated S100A8/A9 levels were still detectable 24 h after stress in 40% of patients and 36% of controls, with a tendency for higher levels in patients. The sustained S100A8/A9 response was associated with a poor rapid cortisol release after stress in patients, but not in the control group. Our findings reveal for the first time that acute psychological stress induces elevated levels of S100A8/A9. We also provide hypothesis-generating evidence that dysregulated cortisol secretion in CAD patients might be associated with an exaggerated pro-inflammatory S100A8/A9 response.

ObjectiveRegulatory T cells (Tregs) are considered atheroprotective, and low levels have been associated with the acute coronary syndrome (ACS), particularly non-ST elevation (NSTE)-ACS. However, the functional properties as well as homeostasis of Tregs are mainly unknown in coronary artery disease (CAD). Here, we investigated the composition and functional properties of naive (n) and memory (m)Tregs in patients with NSTE-ACS and in patients 6-12months post-ACS. MethodsBased on the expression of CD25, FOXP3, CD127, CD45RA, CD39 and CTLA-4, Tregsubsets were defined by flow cytometry in whole blood or isolated CD4(+) T cells. The functional properties of nTregs and mTregs were examined in terms of proliferative capacity and modulation of cytokine secretion. To understand the potential consequences of Treg defects, we also investigated correlations with lipopolysaccharide (LPS)-induced cytokine secretion and ultrasound-defined carotid atherosclerosis. ResultsBoth NSTE-ACS and post-ACS patients exhibited reduced levels of nTregs (P&lt;0.001) compared with healthy control subjects, but without compensatory increases in mTregs. Both nTregs and mTregs from patients showed significantly lower replicative rates and impaired capacity to modulate T-cell proliferation and secretion of interferon-gamma and IL-10. The Treg defect was also associated with LPS-induced cytokine secretion and increased burden of carotid atherosclerosis. ConclusionOur results demonstrate a functional and homeostatic Treg defect in patients with NSTE-ACS and also in stabilized patients 6-12months after ACS. Moreover, this defect was associated with a subclinical proinflammatory and atherogenic state. We believe that the failure to preserve Treg function and homeostasis reflects a need for immune-restoring strategies in CAD.

Stress and inflammation are both important risk factors for coronary artery disease (CAD). However, the susceptibility to stress-induced inflammation and its determinants have been little explored in patients with CAD. Here, our aim was to study the stress-induced inflammatory response, more precisely the early release of matrix metalloproteinase (MMP)-9, and its association with cortisol response in patients with CAD. Sixty-four patients underwent a standardized laboratory stress test. The stress-induced release of MMP-9 was closely associated with the release of other neutrophil-associated proteins, MMP-8 and myeloperoxidase (MPO). It also showed a large variation among patients, as did cortisol. Twenty minutes after stress, a negative association between changes in MMP-9 and cortisol was seen (p amp;lt; 0.01). In vitro, dexamethasone reduced the IL-8-mediated release of MMP-9 from neutrophils, indicating that glucocorticoids may exert rapid effects on neutrophil activation. Further characterization of patients revealed that stress-induced release of MMP-9 was related to leukocyte telomere shortening and increased ultrasound assessed plaque occurrence in the carotid arteries, but not to other characteristics such as age, gender or psychological background factors. The susceptibility to stress-induced release of MMP-9 may thus have impact on disease phenotype. Stress tests can be useful to identify CAD patients in need of novel prevention and treatment strategies. (C) 2016 Published by Elsevier Ltd.

BACKGROUND: Inflammation may play an important role in type 2 diabetes. It has been proposed that dietary strategies can modulate inflammatory activity.

METHODS: We investigated the effects of diet on inflammation in type 2 diabetes by comparing a traditional low-fat diet (LFD) with a low-carbohydrate diet (LCD). Patients with type 2 diabetes were randomized to follow either LFD aiming for 55-60 energy per cent (E%) from carbohydrates (n = 30) or LCD aiming for 20 E% from carbohydrates (n = 29). Plasma was collected at baseline and after 6 months. C-reactive protein (CRP), interleukin-1 receptor antagonist (IL-1Ra), IL-6, tumour necrosis factor receptor (TNFR) 1 and TNFR2 were determined.

RESULTS: Both LFD and LCD led to similar reductions in body weight, while beneficial effects on glycaemic control were observed in the LCD group only. After 6 months, the levels of IL-1Ra and IL-6 were significantly lower in the LCD group than in the LFD group, 978 (664-1385) versus 1216 (974-1822) pg/mL and 2.15 (1.65-4.27) versus 3.39 (2.25-4.79) pg/mL, both P < 0.05.

CONCLUSIONS: To conclude, advice to follow LCD or LFD had similar effects on weight reduction while effects on inflammation differed. Only LCD was found significantly to improve the subclinical inflammatory state in type 2 diabetes.

An imbalance between pro- and anti-inflammatory actions is believed to drive progression of atherosclerosis. Annexin A1 (AnxA1) is a key player in resolution of inflammation and a mediator of anti-inflammatory effects of glucocorticoids. Here, we investigated whether expression of AnxA1 in peripheral blood mononuclear cells (PBMCs) was altered in patients with coronary artery disease (CAD) and also related findings to glucocorticoid sensitivity ex vivo.

We included 57 patients 6-12 months after acute coronary syndrome (ACS), 10 patients with ACS, and healthy controls. AnxA1 mRNA was measured in PBMCs and AnxA1 protein was assessed in monocytes and lymphocyte subsets by flow cytometry. In post-ACS patients and controls, glucocorticoid sensitivity was determined by measuring inhibitory effects of dexamethasone on LPS46 induced cytokine secretion.

AnxA1 mRNA levels in PBMCs were higher in patients compared with controls, although most pronounced in ACS patients. AnxA1 protein was most abundant in the monocyte fraction. ACS patients exhibited the highest levels of cell surface-associated AnxA1 protein while levels in post-ACS patients and controls were similar. Ex vivo assays showed that PBMCs from post-ACS patients were more prone to release IL-6. Glucocorticoid sensitivity correlated with cell surface-associated AnxA1 protein in peripheral monocytes. Dexamethasone also induced upregulation of AnxA1 mRNA.

AnxA1 expression in PBMCs is closely associated with glucocorticoid actions and cell surface associated AnxA1 appears to be a marker of glucocorticoid sensitivity. Although still speculative, a “normal” expression of cell surface-associated AnxA1 in post-ACS patients may suggest that glucocorticoid actions in vivo are insufficient to provide adequate anti-inflammatory effects in these patients.

Background: Increased proportions of circulating proinflammatory CD56+ T cells have been reported in patients with coronary artery disease (CAD). Yet, little is known about regulation of these cells. In the present study, we investigated the expression and potential role of the glucocorticoid-mediated protein annexin A1 (AnxA1) in CD56+ and CD56-T cell subsets, with focus on CAD.

Methods and Results: We included totally 52 healthy individuals, 28 patients with acute coronary syndrome (ACS) and 57 patients with a history of ACS. AnxA1 mRNA expression was assessed in peripheral blood mononuclear cells. AnxA1 protein expression (total and cell surface-associated) was measured by whole blood flow cytometry in circulating CD56⁺ and CD56- T cell subsets. Furthermore, inhibitory effects of dexamethasone and/or recombinant AnxA1 on cytokine secretion by CD56⁺ and CD56^- T cells were explored in vitro. We found that CD56⁺ T cells (the majority CD8⁺), expressed higher levels of AnxA1 mRNA and protein than did CD56^- T cells. When comparing CAD patients with healthy controls, significantly higher levels of cell surface-associated AnxA1 expression were seen in patients, most pronounced in ACS patients. In vitro, dexamethasone reduced cytokine secretion by CD56⁺ T cells, whereas AnxA1 alone had no effect, and AnxA1 combined with dexamethasone abolished the dexamethasone-induced suppressive effects.

Conclusion: AnxA1 was expressed more abundantly in proinflammatory CD56⁺ T cells. Patients with ACS exhibited increased levels of cell surface-associated AnxA1, thus indicating increased activation of the AnxA1 pathway. Our data further suggested that AnxA1 might counteract glucocorticoid mediated anti-inflammatory effects in T cells.

Background: Matrix metalloproteinase (MMP)-9 may play a central role in the development and progression of atherosclerosis. Emerging evidence also indicates an association between MMP-9 and depressive symptomatology. Here, we investigated whether expression of MMP-9 and its inhibitors in blood mononuclear cells and plasma were related to depressive symptoms in patients with a recent myocardial infarction (MI). Methods and Results: Blood sampling was performed between 6 and 18 months after MI in 57 patients. Forty-one clinically healthy subjects were included as controls. Gene expression of MMP-9 and its main tissue inhibitors TIMP-1 and -2 were analyzed in freshly isolated or cultured blood mononuclear cells. Corresponding protein levels were assessed in cell supernatants and plasma. In post-MI patients, mRNA levels of MMP-9 and TIMP-1 and -2 were significantly higher than in controls while protein levels in cell supernatants and plasma did not differ between groups. The Center for Epidemiological Studies - Depression (CES-D) scale was used to assess depressive symptomatology. Repeated assessments during the first 18 months after MI showed significantly higher CES-D scores in patients compared with controls. However, there were no relationships between depressive mood and any of the measurements of MMP-9 or TIMPs. Conclusion: Our findings indicate that overexpression of MMP-9 and TIMPs in blood mononuclear cells and elevated depressive symptoms represent two unrelated phenomena after MI.

We studied the effects of probiotic treatment on the innate immune system during infancy. The study included a subgroup of infants recruited to the pilot study testing the feasibility of probiotics intervention in infants with genetic risk of type 1 diabetes (T1D). A mixture of Lactobacillus rhamnosus GG (5 x 109 cfu), Lactobacillus rhamnosus LC705 (5 x 109 cfu), Bifidobacterium breve Bbi99 (2 x 108 cfu) and Propionibacterium freudenreichii ssp. Shermani JS (2 x 109 cfu) was given to the infants beginning one to three weeks after birth until the age of 6 months. Blood samples were drawn at the age of 6, 12 and 24 months for the analyses of beta-cell autoantibodies and the phenotype and stimulation response of monocytes with flow-cytometry, including surface markers on circulating CD14+ monocytes and expression of co-stimulatory markers on CD14+ monocytes as response to stimulation with lipoteichoic acid (LTA) and lipopolysaccharide (LPS). Also gene expression of toll-like receptor (TLR) signalling molecules was studied in the peripheral blood mononuclear cell (PBMC) population.

In the children who received probiotics the number of circulating CD14+ monocytes expressing CD58 was reduced at the age of 6 months, and a tendency for a decreased induction of CCR5, CD80 and CD58 expressing monocytes as response to LTA was seen when compared to the children who received placebo. At the age of 12 months, the number of monocytes expressing CCR5 was decreased in the probiotic group, and a decreased spontaneous expression of TNFRSF1A and an increased spontaneous expression of TLR9 was observed in the PBMC from the children treated with probiotics. In the whole study group, the numbers of circulating monocytes expressing CD80 increased with age as well as the induction of CCR5, CD80 and CD58 on monocytes as response to stimulation. By the age of 24 months one child in both groups developed multiple autoantibodies.

We demonstrated that probiotics modulated the activation stage and stimulation response of monocytes, and that prolonged effects of the treatment were seen at the age of 12 months. The findings suggest that early microbial exposure may program the function of the innate immune system for later life.

Objective: Apoptosis of natural killer (NK) cells is increased in patients with coronary artery disease (CAD) and may explain why NK cell levels are altered in these patients. Soluble forms of Fas and Fas ligand (L) are considered as markers of apoptosis. Here, we investigated whether plasma levels of Fas and FasL were associated with NK cell apoptosis and NK cell levels in CAD patients. Methods: Fas and FasL in plasma were determined by ELISA in 2 cohorts of CAD patients; one longitudinal study measuring circulating NK cells and apoptotic NK cells by flow cytometry 1 day, 3 months and 12 months after a coronary event and one cross-sectional study measuring NK cell apoptosis ex vivo. Both studies included matched healthy controls. Fas and FasL were also determined in supernatants from NK cells undergoing cytokine-induced apoptosis in cell culture. Results: In the 12-month longitudinal study, plasma FasL increased by 15% (p less than 0.001) and NK cell levels by 31% (p less than 0.05) while plasma Fas did not change. Plasma FasL and NK cell levels were significantly related at 3 months and 12 months, r = 0.40, p less than 0.01. Furthermore, plasma FasL, but not plasma Fas, correlated with NK cell apoptosis ex vivo in CAD patients, r = 0.54, p less than 0.05. In vitro, cytokine-induced apoptosis of NK cells resulted in abundant release of FasL. Conclusion: In CAD patients, FasL in plasma is associated with both apoptotic susceptibility of NK cells and dynamic changes in circulating NK cells. NK cells are also themselves a potential source of soluble FasL. Our findings link NK cell status to a soluble marker with possible atheroprotective effects thereby supporting a beneficial role of NK cells in CAD.

Bakgrund: Under de senaste decennierna har förekomsten av allergiska sjukdomar ökat i västvärlden. En av möjliga förklaringar kan vara en minskad eller förändrad mikrobiell exponering under uppväxttiden. Mikrobiella stimuli under de första levnadsåren tros vara viktiga för utmognaden av immunsystemet och utveckling av tolerans. Exakt hur mikroorganismers påverkan på immunförsvaret är kopplat till utvecklingen av allergiska sjukdomar är dock ännu okänt. Genetiska polymorfier kan påverka immunsvar mot mikrobiella komponenter. En sådan polymorfi, TLR4 Asp299Gly, har observerats i genen som kodar för receptorn TLR4 som känner igen lipopolysackarid (LPS) från Gramnegativa bakteriers cellvägg, och har föreslagits vara associerad med en förändrad förmåga att svara immunologiskt mot LPS.

Syfte: Syftet med studierna var att studera immunsvar mot LPS i relation till specifika genetiska polymorfier, allergisk sjukdom samt mikrobiellt tryck i form av endotoxinnivåer.

Studiepopulationer: Denna avhandling baseras på resultat från tre studiegrupper: estniska och svenska barn som är följda från födseln upp till fem års ålder, en grupp svenska skolbarn 8 och 14 år gamla samt en grupp unga vuxna.

Metoder: Två genetiska polymorfier, TLR4 Asp299Gly och CD14/-159, analyserades. Mononukleära celler isolerades från perifert blod och odlades tillsammans med LPS från två olika Gramnegativa bakteriestammar, _{Salmonella enterica} serotype Typhimurium (Serotype Typhimurium) och Escherichia coli ( E. coli). Cytokin- och kemokinsekretion analyserades i cellsupernatanter med Luminex eller ELISA. Ytmarkörer på monocyter i helblod studerades med flödescytometri. Intracellulära signaleringsproteiner, som är inblandade i TLR4s signalvägar analyserades med Luminexteknik. mRNA uttryck av proteiner som är relaterade till LPS signalering och markörer för regulatoriska T celler analyserades med realtids-PCR.

Resultat: TLR4 Asp299Gly polymorfin var associerad med lägre fosforylering av det intracellulära signaleringsproteinet IκBα och lägre utsöndring av cytokinerna IL-12 och IL-10 efter cellstimulering med LPS hos skolbarn och unga vuxna. Skillnader i cellsvar mellan individer med och utan polymorfin kunde påvisas när cellerna odlats med LPS från Serotype Typhimurium men inte med LPS från E. coli. Polymorfin var också associerad med astma och särskilt atopisk astma.

Flera skillnader i immunsvar mot LPS observerades mellan allergiska och ickeallergiska individer. Skolbarn med astma hade lägre LPS inducerad IL-10 och IL-12 cytokinproduktion. Vuxna allergiker hade lägre LPS inducerad IκBα fosforylering. Svenska barn som vid fem års ålder hade utvecklat allergisk sjukdom hade lägre mRNA uttryck av TLR2 vid födseln.

Estniska barn hade generellt lägre LPS inducerade cytokinsvar än svenska barn vid födseln och vid 3 och 6 månaders ålder. mRNA uttrycket av de T-regulatoriskt associerade markörerna Foxp3 och Ebi3 var vid födseln högre hos de estniska jämfört med de svenska barnen.

Slutsats: Genetiska förutsättningar kan påverka immunsvar mot LPS och kan möjligen ha en betydelse för utveckling av astma. De generellt lägre LPS inducerade cytokinsvaren och högre uttryck av markörer för Treg celler hos estniska jämfört med svenska barn skulle kunna bero på att deras uppväxtmiljö med ett högre mikrobiellt tryck påverkar den tidiga utvecklingen av immunförsvaret och kan möjligen vara en bidragande förklaring till den lägre allergifrekvens som ses i Estland jämfört med Sverige.

Introduction Among sensitized infants, those with high, as compared with low levels, of salivary secretory IgA (SIgA) are less likely to develop allergic symptoms. Also, early colonization with certain gut microbiota, e.g. Lactobacilli and Bifidobacterium species, might be associated with less allergy development. Although animal and in vitro studies emphasize the role of the commensal gut microbiota in the development of the immune system, the influence of the gut microbiota on immune development in infants is unclear.

Objective To assess whether early colonization with certain gut microbiota species associates with mucosal and systemic immune responses i.e. salivary SIgA and the spontaneous Toll-like receptor (TLR) 2 and TLR4 mRNA expression and lipopolysaccharide (LPS)-induced cytokine/chemokine responses in peripheral blood mononuclear cells (PBMCs).

Methods Fecal samples were collected at 1 week, 1 month and 2 months after birth from 64 Swedish infants, followed prospectively up to 5 years of age. Bacterial DNA was analysed with real-time PCR using primers binding to Clostridium difficile, four species of bifidobacteria, two lactobacilli groups and Bacteroides fragilis. Saliva was collected at age 6 and 12 months and at 2 and 5 years and SIgA was measured with ELISA. The PBMCs, collected 12 months after birth, were analysed for TLR2 and TLR4 mRNA expression with real-time PCR. Further, the PBMCs were stimulated with LPS, and cytokine/chemokine responses were measured with Luminex.

Results The number of Bifidobacterium species in the early fecal samples correlated significantly with the total levels of salivary SIgA at 6 months. Early colonization with Bifidobacterium species, lactobacilli groups or C. difficile did not influence TLR2 and TLR4 expression in PBMCs. However, PBMCs from infants colonized early with high amounts of Bacteroides fragilis expressed lower levels of TLR4 mRNA spontaneously. Furthermore, LPS-induced production of inflammatory cytokines and chemokines, e.g. IL-6 and CCL4 (MIP-1β), was inversely correlated to the relative amounts of Bacteroides fragilis in the early fecal samples.

Conclusion Bifidobacterial diversity may enhance the maturation of the mucosal SIgA system and early intense colonization with Bacteroides fragilis might down-regulate LPS responsiveness in infancy.

Altered microbial exposure is a possible explanation for the increase of allergies in the Western world. However, genetic factors influence microbially induced immune responses. We have investigated the TLR4(Asp299Gly) gene polymorphism and its possible association with receptor expression of circulating peripheral blood monocytes and the in vitro cytokine responses and phosphorylation of intracellular signaling proteins in peripheral blood mononuclear cells (PBMC) stimulated with lipopolysaccharide (LPS) from Escherichia coli and Salmonella enterica serotype Typhimurium. We studied 34 of the predominant haplotype TLR4 Asp299 (AA) and 8 heterozygote Asp299Gly (AG) individuals. TLR4 expression levels were similar in the two genotype groups. Serovar Typhimurium LPS induced interleukin-12p70 from PBMC, and the degree of phosphorylation of the intracellular signaling protein I kappa B alpha in PBMC was lower in the AG than the AA group (P = 0.03 and P = 0.04, respectively). These results were not seen, however, when PMBC were stimulated with E. coli-derived LPS. Based on these results, we propose that TLR4(Asp299Gly) gene polymorphism and the bacterial origin of LPS should be considered when environmental LPS exposure is evaluated in disease risk or protection.

Optimal activation of T lymphocytes requires a costimulatory signal provided by the interaction of molecules on the surface of T cells with their ligands expressed on dendritic cells (DC). We investigated whether DC differentiated from monocytes from healthy and birch allergic asthmatic individuals and further maturated by stimulation with cat and birch allergens and LPS differ in their phenotypic receptor expression. Similar expression of DC surface markers, including HLA-DR, CD80, CD86, CD83, CD1a and CD11c, was detected in monocyte-derived DC from allergic and healthy individuals. Cells from healthy donors stimulated either antigen showed a similar activation of the CD80 and double CD80/CD86 costimulatory molecules when compared with non-stimulated cells. In the case of cells from allergic individuals, birch allergen was unable to produce the same increased expression of CD80 alone or in combination with CD80/CD86, in comparison with cells stimulated with cat and LPS. Levels of IL-6, IL-8, IL-10, MCP-1/MCAF and MIP-1β were similar in the supernatant of non-stimulated DC from both groups of subjects. By contrast, the spontaneous secretion of IL-12p70 and TNF-α was higher in the supernatant of DC from healthy subjects when compared with that from allergic individuals. Stimulation with birch and LPS resulted in an increased secretion of IL-12p70 in samples from healthy when compared with that in allergic individuals. The results suggest an impaired specific maturation of DC from birch allergic individuals in association with birch-specific immune responses. Lower secretion of IL-12p70 from birch-stimulated DC from allergic individuals suggests that not only maturation, but also the specific Th1 function of these cells seems to be affected in those individuals.

Background

Bacterial signals play an important role in the maturation of the immune system. Polymorphisms in genes coding for receptors to bacterial components can alter the immune responsiveness of the host to microbial agents and may indicate the development of aberrant immune responses that are associated with immune-mediated diseases such as atopic diseases.

Objective

The study's objective was to investigate the relationship between TLR4 and CD14 gene polymorphisms, the LPS responsiveness of PBMCs, and the presence of asthma and allergic rhinoconjunctivitis in children.

Methods

The TLR4 (Asp299Gly) and CD14/−159 polymorphisms were determined in 115 Swedish children aged 8 and 14 years. LPS-induced IL-12(p70), IL-10, and IFN-γ responses of PBMCs from 69 of the children were analyzed by means of ELISA. The levels of soluble CD14 in serum samples were analyzed by means of ELISA, and the total IgE levels were analyzed by means of UniCAP Total IgE (Pharmacia Diagnostics, Uppsala, Sweden).

Results

Decreased LPS-induced IL-12(p70) and IL-10 responses were associated with the TLR4 (Asp299Gly) polymorphism and independently with asthma, especially atopic asthma. The TLR4 (Asp299Gly) polymorphism was associated with a 4-fold higher prevalence of asthma in school-aged children (adjusted odds ratio 4.5, 95% CI 1.1-17.4) but not to allergic rhinoconjunctivitis.

Conclusion

A TLR4 polymorphism modifies innate immune responses in children and may be an important determinant for the development of asthma. This may influence the outcome of intervention studies that use microbial stimuli as immune modulators.

Type 1 diabetes (T1D) has been associated with an aberrant maturation of dendritic cells (DC). We studied the maturation of monocyte-derived DC in children with newly diagnosed T1D and in healthy children with genetic risk for T1D. Peripheral blood monocytes from children with newly diagnosed T1D (n = 12; mean age 13.2 years), children with human leukocyte antigen (HLA)-risk genotype of T1D (n = 7; mean age 12.7 years) and healthy children (n = 14; mean age 11.2 years) were in vitro differentiated into DC. Expression of HLA-DR, CD80/86 and CD11c and secretion of interleukin (IL)-12, tumor necrosis factor (TNF)-α, IL-6 and IL-10 were measured using flow cytometry. Lower percentage of DC expressed CD11c and HLA-DR, and decreased production of TNF-α was found in children with newly diagnosed T1D and in children at genetic risk when compared to healthy children. Children with risk genotype also had decreased IL-12 production by DC. Children with T1D and children at genetic risk of T1D appear to have similar aberrancies in the maturation of DC, which may predispose to β-cell autoimmunity.

Background: Allergies have increased during the last decades with the highest prevalences in countries with a Westernised lifestyle. As microbial activation might be required for an accurate maturation of the immune system, a reduced or altered microbial exposure during infancy has been suggested as a possible cause for the increase of allergic diseases in affluent countries.

Aim: The aim of this study was to investigate the gene expression of TLR2, TLR4, Foxp3, Ebi3 and SOCS3 involved in LPS signaling pathways and immune regulation in two prospectively followed birth cohorts, one from Estonia and one from Sweden, in relation to domestic endotoxin levels and allergy development.

Material and methods: Twenty-three children from Estonia and 52 from Sweden were followed from birth to five years of age regarding allergy development using questionnaires, clinical examinations and skin prick tests. RNA was isolated from unstimulated peripheral blood mononuclear cells (PBMC) in samples collected at birth, 3, 6, 12 and 24 months of age. mRNA expression of TLR2, TLR4, SOCS3, Ebi3 and Foxp3 was analysed with real-time RTPCR. The mRNA expression of the genes were compared to previously obtained data of LPS/IFN-γ induced cytokine and chemokine PBMC secretion at the corresponding time-points and also to domestic endotoxin levels.

Results: The Estonian infants had higher mRNA expression of the two regulatory T cell (Treg) associated markers Foxp3 and Ebi3 at birth than the Swedish infants. The Foxp3 and Ebi3 expression correlated at all time-points. The mRNA expression of TLR2, TLR4 and SOCS3 was similar in Estonian and Swedish infants at all ages. None of the genes were associated with endotoxin levels.TLR4 mRNA expression correlated positively with cytokines that were upregulated by LPS/IFN- γ stimulation, but was negatively correlated to CCL2 secretion which was downregulated by the stimulation. Low expression of TLR2 mRNA at birth was found in Swedish children who were allergic at 5 years of age. Sensitisation up to 5 years of age was associated with low Ebi3 expression at birth.

Conclusion: Estonian children are born with enhanced Ebi3 and Foxp3 expression compared to Swedish children, suggesting an increased capacity for early immune regulation among infants from a country with a low prevalence of allergic disease. An increased early capacity to respond to TLR2 ligands, i.e. Gram positive bacteria, may protect against allergy development.

Background: Psychological stress and inflammation are both important risk factors for coronary artery disease (CAD). Susceptibility to mental stress-induced inflammation has been little explored in patients with CAD. Here, we investigated whether stress-induced inflammatory response, more precisely neutrophil activation, was associated with cortisol reactivity, leukocyte telomere length (TL) and carotid atherosclerotic burden in CAD.

Methods: Sixty-four patients with stable CAD underwent a laboratory stress test. Matrix metalloproteinase (MMP)-9, MMP-8, tissue inhibitors (TIMP)-1 and -2, myeloperoxidase (MPO) and salivary cortisol were measured before and 20 min after stress. Leukocyte TL was assessed as well as basal cortisol levels, background psychological factors and atherosclerosis in carotid arteries.

Results: The variation in stress-induced release of neutrophil markers was substantial. Patients were therefore divided into lower and upper tertiles depending on changes in serum MMP-9, T1: -12 %, T3: +27 %, with corresponding changes in MMP-8 and MPO. Clinical or psychological characteristics did not differ between groups, neither did basal levels of neutrophil markers or cortisol. Cardiovascular reactivity during stress was similar in T1 and T3, while cortisol declined after stress only in T3 (-30 %). Leukocyte TL was shorter in T3 than in T1, 0.78 vs 0.88, p = 0.006. Moreover, presence of plaques in right carotid artery differed between T1 and T3, 66 % vs 100 %, p = 0.004.

Conclusion: Stress-induced neutrophil activation in CAD patients was associated with altered cortisol reactivity, leukocyte telomere attrition and increased subclinical atherosclerosis. Data suggest that mental stress testing can identify high-risk patients in need of novel prevention and treatment strategies.

Background: Allergic diseases have increased in the last decades, particularly in countries with an affluent lifestyle, possibly due to a reduced or altered microbial exposure during infancy.

Objective: The aim of this study was to follow lipopolysaccharide (LPS) induced immune responses prospectively in infants from Estonia and Sweden, i.e. two countries with different frequencies of allergic disease and a different domestic endotoxin exposure. '

Methods: The study included 14 Estonian and 36 Swedish infants who were followed prospectively from birth up to two years of age regarding development of allergy using questionnaires, clinical examinations and skin prick tests. Isolated cord blood mononuclear cells (birth) and peripheral blood mononuclear cells (obtained at 3, 6, 12 and 24 months) were cultured for 24h with LPS in combination with IFN-γ. The secretion of IL-6, CXCL8 (IL-8), IL-10, IL12p70, IL-17, IL-1β, CCL2 (MCP-1), CCL4 (MIP-1β) and TNF was analysed with a Luminex assay.

Results: The Swedish, as compared to Estonian children, had higher levels of LPS/IFN-γ induced CCL4 and IL-6 at birth and of IL-1β, IL-12p70, IL-17 and TNF at 3 months as well as IL-6 at 6 months. Also, the levels of CCL2 at 3 and 6 months of age and CCL4 and TNF at 6 months of age were higher in Swedish than Estonian infants in unstimulated cultures. Sensitised Swedish infants had higher levels of LPS/IFN-γ induced IL-10 at 3 and 12 months of age compared to non-sensitised Swedish infants.

Conclusion: The enhanced LPS/IFN-γ induced cytokine and chemokine secretion in Swedish, compared to Estonian infants may support a less rapid induction of immune regulation in an affluent society. This could possibly be due to a reduced microbial pressure on Swedish children during early childhood.

Introduction Function and frequency of CD4⁺CD25^high T cells in type 1 diabetes (T1D) patients has been studied in cross-sectional studies. We wanted to investigate the changes in number and function of CD4⁺CD25^high T cells in children with T1D during follow-up.

Material and Method The study includes in total 35 children with T1D and 17 healthy children. The number of CD4⁺CD25^high regulatory T cells and their CTLA-4 expression was analyzed by flow cytometry and the function as the effect of CD4⁺CD25^high T cells on proliferative and cytokine response after mitogen stimulation.

Results We found no differences between healthy and diabetic children in the amount of CD4⁺CD25^highCTLA-4+ regulatory T cells and the numbers did not change during 18 months follow-up. We observed higher median fluorescence intensity of intracellular CTLA-4 on CD4⁺CD25^high T cells in the diabetic children at diagnos compared to healthy children. Proliferation response to PHA was higher in the CD25depleted PBMC than in the whole PBMC population in diabetic and healthy children. Children with T1D showed a higher spontaneous secretion of IL-10 and TGF-β and higher expression of IFN-γ specific mRNA among CD25depleted PBMC than was seen in healthy children.

Conclusion Our data indicate that children with T1D have similar amounts of circulating CD4⁺CD25^high regulatory cells as healthy children. Our data further suggest an increased activity among regulatory T cells in children with T1D.

Context: Coronary artery disease (CAD) is characterized by low-grade chronic inflammation including leukocyte-derived overexpression of matrix metalloproteinase (MMP)-9 and its tissue inhibitors (TIMPs). The mechanisms behind this overexpression are not clarified but may involve dysregulated glucocorticoid regulation of MMP-9.

Objective: We hypothesized that the increased expression of MMP-9 and TIMPs in peripheral blood mononuclear cells (PBMCs) from CAD patients was associated with reduced glucocorticoid sensitivity.

Setting: This was an observational study conducted at the Outpatient Cardiology Clinic at the University Hospital, Linköping, Sweden.

Participants: CAD patients with a history of non-ST-elevation myocardial infarction were consecutively included. Healthy control subjects were randomly recruited from the Swedish Population Register.

Main outcome measures: We measured mRNA and protein levels of glucocorticoid receptors (GR)-α and -β in PBMCs in vivo and further investigated the effects of dexamethasone on MMP-9, TIMP, GR-α and -β expression ex vivo.

Results: The GR-α mRNA levels were markedly increased in PBMCs from CAD patients, whereas GR-β mRNA levels did not differ between patients and controls. Ex vivo, the inhibitory effects of dexamethasone on MMP-9 and TIMP mRNA and protein expression were equal to or slightly higher in patients. Moreover, the dexamethasone treatment resulted in significantly reduced levels of GR-α mRNA in PBMCs.

Conclusion: In contrast to our original hypothesis, we found evidence for increased glucocorticoid sensitivity in PBMCs from CAD patients. It may be suggested that the overexpression of MMP-9 and TIMPs in patients is associated with a state of relative hypocortisolism, thus contributing to a systemic low-grade inflammation.