Proinflammatory cytokines Il-6 and TNF-α and the development of inflammation in obese subjects.

BACKGROUND: The development of obesity and related disorders, e.g., type II diabetes (T2D), hypertension, and metabolic disturbances is strongly related to increased levels in proinflammatory cytokines (IL-1, IL-6, and TNF-α). Both IL-6 and TNF-α are secreted by adipocytes and their concentration correlates with the percentage and distribution of fat tissue in the body. Both cytokines are the main factors responsible for the induction of acute phase proteins production (e.g., CRP) and to inflammatory state. OBJECTIVE: To compare of TNF-α and IL-6 concentrations in serum from obese subjects with those in subjects with normal BMI and to analyze the relation between TNF-α, IL-6, BMI and the inflammatory state as measured by the level of CRP. MATERIAL AND METHODS: The study included 80 obese subject (54 males and 26 females) BMI >25 kg/m⊃2. A control group consisted of 53 healthy subjects (24 males and 29 females) with BMI <25 kg/m⊃2. To determine the blood plasma concentration of IL-6 and TNF, commercial ELISA assay kits were used. RESULTS: The concentration of IL-6 was lower in the control compared with the obese patients, but a significance difference concerned only female subjects (P = 0.001). TNF-α concentration was significantly higher in all obese subjects (P<0.001). A higher level of this cytokine was also found in patients with obesity suffering from T2DM. A positive correlation was present between IL-6 and TNF-α concentrations. Only did the IL-6 level correlate with the concentration of CRP in serum. CONCLUSIONS: The study confirmed that increased inflammatory cytokines lead to the persistence of inflammation in obese subjects. However, some other factors, such as gender, may contribute to the development of obesity-related inflammatory states.

Frequency of distribution of inflammatory cytokines IL-1, IL-6 and TNF-alpha gene polymorphism in patients with obstructive sleep apnea.

Obesity is one of the most commonly identified factors for the obstructive sleep apnea syndrome (OSAS). Adipose tissue is the source of many cytokines, among them there are IL-6, IL-1, and TNF-alpha. The level of inflammatory cytokines increases in people with OSAS and obesity. The aim of this study was to evaluate the distribution of genotypes in inflammatory cytokine genes in people with obesity-related OSAS. The examined group consisted of 102 person with obesity related-OSAS and 77 normal weight person without OSAS. Genotyping of DNA sequence variation was carried out by restriction enzyme (IL-1: Taq I, IL-6: Lwe I, TNF-alpha: Nco I) analysis of PCR amplified DNA. The study revealed a significant correlation between polymorphism located in the promoter region of inflammatory cytokine genes and obesity-related OSAS.

Leptin measurement in urine is a reliable method of monitoring its secretion in patients with obstructive sleep apnea syndrome.

Leptin is believed to play a significant role in the pathogenesis of obstructive sleep apnea syndrome (OSAS) as well as progression of OSAS-related obesity. It is also known that other factors such as gender and diurnal variations in serum strongly affect the measurement results making repeated blood sampling necessary for leptin precise monitoring. Since renal metabolism and urine secretion are the main elimination mechanism for leptin, in this study we evaluated urine relevance for leptin secretion monitoring. Serum and urine (collected during the day and overnight) sampled from 169 OSAS patients and 41 controls were assayed by immunoenzymatic method specific for human leptin. Only 5 (17%) controls and 10 (5.8%) OSAS patients had undetectable urine leptin. We observed significant relationships between serum and urinary leptin in both day-time (r=0.656, P<0.001) and night-time (r=0.518, P<0.001) samples and between day and night-time urine leptin (r=0.811, P<0.001). Significance values did not alter when urinary leptin levels were expressed as the ratio to urinary creatinine. Gender-related differences in leptin concentrations were present both in serum (P<0.001) and overnight urine (P<0.01) in the OSAS group. However, mean night-time urine leptin was lower in the OSAS patients (P<0.05) and their subgroups stratified according to disease severity (P<0.01), while serum leptin levels were comparable in both groups. We conclude that assaying leptin in urine by immunoenzymatic method is a reliable and useful non-invasive alternative for its serum measurement. However, night-time urine leptin levels better reflect differences in its turnover due to gender and OSAS severity.

Frequency of distribution of leptin receptor gene polymorphism in obstructive sleep apnea patients.

Leptin is an adipocyte-derived hormone regulating energy homeostasis and body weight. Leptin concentration is increased in patients with the obstructive sleep apnea syndrome (OSAS). Leptin receptor (LEPR) is a single transmembrane protein belonging to the superfamily of cytokine receptors related by a structure to the hemopoietin receptor family. The aim of the present study was to evaluate the frequency of distribution of leptin receptor gene polymorphism GLN223ARG in OSAS patients compared with healthy controls. The examined group included 179 subjects: 102 OSAS patients (74 men and 28 women) and 77 non-apneic controls (39 men and 38 women). Genomic DNA was isolated with the use of a column method and genotyping of DNA sequence variation was carried out by restriction enzyme analysis of PCR-amplified DNA. The results revealed a significant correlation between the polymorphism of LEPR and OSAS. Carriers of Arg allele in homozygotic genotype Arg/Arg and heterozygotic genotype Gln/Arg were more often obese and developed OSAS than the group of carriers of homozygotic Gln/Gln genotype. This tendency was observed in the whole examined population and in the group of obese women. We also found the highest levels of total cholesterol, LDL, HDL, and triglycerides in the group of homozygotic Arg/Arg genotype carriers, lower in heterozygotic Gln/Arg genotype carriers, and the lowest in the group of persons carring homozygotic Gln/Gln genotype. The presence of Arg allel seems linked to a higher risk of obesity and higher lipid levels in OSAS patients. OSAS may have a strong genetic basis due to the effects from a variety of genes including those for leptin receptor.

Differences in responses upon corticosteroid therapy between smoking and non-smoking patients with COPD.

Inhaled corticosteroids have a high level of topical anti-inflammatory activity. However, in patients with COPD these drugs have been reported to exert limited effects. A reduction in histone deacetylase (HDAC) activity is suggested to prevent the anti-inflammatory action of corticosteroids. Cigarette smoke is known to reduce HDAC expression. The aim of this study is to compare the outcome of corticosteroid therapy in both smoking and non-smoking COPD patients. Twenty-three smoking patients and 18 ex-smoking patients with COPD were treated with inhaled corticosteroids for a period of 2 months. Blood and induced sputum samples were collected before and after treatment. Values of FEV(1) %-predicted did not change upon the therapy, but there was a trend to improve in the ex-smokers (63.1 -> 64.8%-pred.), compared with a decrease in the smokers (63.3 -> 61.6%-pred.). The levels of the pro-inflammatory cytokine IL-8 increased in the group of smokers from 379 +/-78 to 526 +/-118 ng/ml. Although not significant, a slight decrease from 382 +/-70 to 342 +/-62 ng/ml was observed in the group of ex-smokers. The neutrophil related elastase activity showed similar effects after steroid treatment, it went up from 36.4 +/-12.0 to 113.5 +/-9.7 nmol/l in smokers, and decreased from 346.2 +/-72.1 to 131.1 +/-6.5 nmol/l in ex-smokers with COPD. These results support the evidence that inhaled corticosteroids have no anti-inflammatory effects in COPD patients, but only when these patients are still smoking. Smoking cessation seems the best therapy for COPD patients.

Inflammatory markers in the exhaled breath condensate of patients with pulmonary sarcoidosis.

Pulmonary sarcoidosis may progress to fibrosis in some patients, so that close monitoring of its activity is essential for recommending clinical strategy. Examination of airway inflammatory markers in bronchoalveolar lavage (BAL) is one of the methods applied to assess the disease severity. Recently, the expired breath condensate (EBC) has become another source of cytokines and mediators. In sarcoidosis, except for NO and oxidative stress markers, no other mediators have yet been estimated in the exhaled air. In the present study we attempted to answer the question of whether airway inflammatory markers in pulmonary sarcoidosis patients might be assessable in EBC and to what extend these markers might reflect the disease activity in the lungs IL-6, TNF-alpha, PAI-1, and IGF-1 were measured by Elisa method in EBC and BALF samples from 9 patients with newly-diagnosed pulmonary sarcoidosis. TNF-alpha, IGF-1, and PAI-1 levels in EBC and BAL samples were comparable and closely positively correlated [TNF-alpha (r=0.79, P<0.001), IGF-1 (r=0.94, P<0.001), and PAI-1 (r=0.81, P<0.001)]. In contrast, IL-6 concentration in EBC was significantly lower compared with that in BALF, while the correlation between both materials was negative (r=-0.47, P<0.05). An important distinction in IL-6 performance, which might explain this inconsistency, is its tendency to form more complex molecular forms of a higher weight than that of other cytokines. Our study shows that EBC reflects cytokine production in the lung as effectively as BALF, providing that the characteristics of proteins evaluated allow their easy transfer into the exhaled air. Further studies are required before accepting EBC samples as an equivalent to BALF.

Smokers with airway obstruction are more likely to quit smoking.

BACKGROUND: Chronic obstructive pulmonary disease (COPD), usually caused by tobacco smoking, is one of the leading causes of morbidity and mortality. Smoking cessation at an early stage of the disease usually stops further progression. A study was undertaken to determine if diagnosis of airway obstruction was associated with subsequent success in smoking cessation, as advised by a physician. METHODS: 4494 current smokers (57.4% men) with a history of at least 10 pack-years of smoking were recruited from 100 000 subjects screened by spirometric testing for signs of airway obstruction. At the time of screening all received simple smoking cessation advice. 1177 (26.2%) subjects had airway obstruction and were told that they had COPD and that smoking cessation would halt rapid progression of their lung disease. No pharmacological treatment was proposed. After 1 year all subjects were invited for a follow up visit. Smoking status was assessed by history and validated by exhaled carbon monoxide level. RESULTS: Nearly 70% attended a follow up visit (n = 3077): 61% were men, mean (SD) age was 52 (10) years, mean (SD) tobacco exposure 30 (17) pack-years, and 33.3% had airway obstruction during the baseline examination. The validated smoking cessation rate in those with airway obstruction was 16.3% compared with 12.0% in those with normal spirometric parameters (p = 0.0003). After correction for age, sex, nicotine dependence, number of cigarettes smoked daily, and lung function, success in smoking cessation was predicted by lower lung function, lower nicotine dependence, and lower tobacco exposure. CONCLUSIONS: Simple smoking cessation advice combined with spirometric testing resulted in good 1 year cessation rates, especially in subjects with airway obstruction.

Increasing COPD awareness.

Early diagnosis and smoking cessation are the only available methods to stop the progression of chronic obstructive pulmonary disease (COPD). The aim of this study was to evaluate the effects of early detection of airflow limitation (AL) in a population with high risk for COPD, using spirometric screening. Smokers aged 40 yrs with a smoking history of 10 pack-yrs were invited to visit a local outpatient chest clinic for simple spirometry (forced expiratory volume in one second (FEV1) and forced vital capacity (FVC)). Smoking history was recorded, followed by smoking cessation advice relating the results of spirometry to the smoking behaviour. Subjects who did not fulfil the above criteria (younger and/or nonsmokers) were also screened. A total 110,355 subjects were investigated; they were aged 53.5+/-11.5 yrs and 58.2% were males. Of the total amount of subjects, 64% were current smokers, 25.1% were former smokers and 10.9% were lifelong nonsmokers. Spirometry tests were within normal values for 70.3%, and 20.3% showed signs of AL: this was mild in 7.6%, moderate in 6.7% and severe in 5.9%. The remaining 8.3% of subjects presented with a restrictive pattern of ventilatory impairment. Airflow limitation was found in 23% of smokers aged 40 yrs with a history of 10 pack-yrs. This study concluded that large-scale voluntary spirometry screening of the population with high risk for COPD detects a large number of subjects with AL.

Disturbed angiogenic activity in sera from obstructive sleep apnea patients.

It is increasingly recognized that obstructive sleep apnea (OSA) syndrome is a systematic rather than local disorder. There is also growing evidence that apart from the syndrome's major features: intermittent hypoxia and sleep fragmentation, functional activity of the immune system is altered in OSA patients, with several cytokines, such as tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) taking active part in sleep regulation. Little is known about the effects exerted by chronic intermittent hypoxia combined with persistent pro-inflammatory activity of the immune system on the vascular micro milieu in OSA. In this study we attempted to confirm the hypothesized imbalance between pro- and anti-angiogenic factors by evaluating direct and indirect angiogenic activity of OSA patients' sera in the in vivo serum-induced angiogenesis (SIA) and leukocyte-induced (LIA) assays, respectively, in mice. Both tests revealed significantly inhibited angiogenic activity of OSA patients' sera compared with healthy controls (P<0.001). Moreover, differences related to the subject's weight regarding in the mean number of newly-formed vessels were observed with a significantly greater inhibition in the normal-weighing apneic subjects than in the overweight or obese ones (P<0.01). The angiogenesis inhibition index was positively related to the serum IL-6 level (r=0.35; P<0.05) in the OSA group, but not to TNF-alpha, fasting serum leptin, or OSA syndrome severity as assessed by the AHI index. Our results demonstrate that OSA is accompanied by disturbed serum angiogenic activity, apparently resulting from an imbalance between pro- and anti-angiogenic factors, some of them being produced by the adipose tissue. The disordered angiogenic activity might be related to the pathophysiology of OSA and should be considered an important causative factor for the increased prevalence of cardiovascular diseases in OSA patients.

New developments in the treatment of COPD: comparing the effects of inhaled corticosteroids and N-acetylcysteine.

Inhaled corticosteroids (ICS) are widely used for the treatment of COPD despite of controversial statements concerning their efficacy. The use of N-acetylcysteine (NAC), a mucolytic drug with antioxidant properties, is less clear, but it may counteract the oxidant-antioxidant imbalance in COPD. The aim of this study was to evaluate whether treatment of COPD patients with ICS or NAC is able to improve inflammatory indices and to enhance lung function. ICS treatment enhanced protective markers for oxidative stress such as glutathione peroxidase (GPx) (51.2 +/-5.8 vs. 62.2 +/-8.6 U/g Hb, P<0.02) and trolox-equivalent antioxidant capacity (TEAC) (1.44 +/-0.05 vs. 1.52 +/-0.06 mM, P<0.05). NAC decreased sputum eosinophil cationic protein (318 +/-73 vs. 163 +/-30 ng/ml, P<0.01) and sputum IL-8 (429 +/-80 vs. 347 +/-70 ng/ml, P<0.05). The increased antioxidant capacity prevented an up-regulation of adhesion molecules, since the levels of intracellular adhesion molecule 1 (ICAM-1) correlated negatively with GPx (P<0.0001) and TEAC (P<0.0001). On the other hand, expression of adhesion molecules was promoted by inflammation, reflected by a positive correlation between the levels of IL-8 and ICAM-1 (P<0.0001). The effects of treatment on lung function were only reflected in the FEV(1) values. The absolute value of FEV(1), both before and after salbutamol inhalation, increased from 1690 +/-98 to 1764 +/-110 ml, and 1818 +/-106 to 1906 +/-116 ml, respectively, after ICS (P<0.05) . Ten weeks after treatment, FEV(1) values dropped to 1716 +/-120 ml post-salbutamol (P<0.05). When followed by treatment with NAC, these values decreased even further to 1666 +/-84 ml. These results suggest that ICS improved lung function in COPD patients with moderate airflow obstruction, beside a minor improvement in the oxidant-antioxidant imbalance leading to a lesser expression of ICAM-1. Treatment with NAC decreased some inflammatory parameters and had indirectly an inhibitory effect on the expression of adhesion molecules.

Effects of homogenization of induced sputum by dithiothreitol on polymorphonuclear cells.

Induced sputum represents a useful and non-invasive tool to isolate different cells from the airways. Complete homogenization of sputum is important for dispersion of cells and is usually achieved by use of dithiothreitol (DTT). However, it is not known if DTT will influence the viability and functionality of cells obtained by induced sputum. In the present study, induced sputum was processed by DTT or by PBS treatment. The obtained neutrophils were compared with neutrophils obtained from peripheral blood and from bronchoalveolar lavage fluid (BAL). These isolated neutrophils were treated in a similar way as the sputum neutrophils with DTT or PBS. All isolated cells were used for chemiluminescence tests and for the measurement of elastase and myeloperoxidase release after stimulation with fMLP. The results showed that the maximum chemiluminescence response was always significantly lower after DTT treatment: blood, 16.68 +/-1.89 vs. 2.62 +/-0.43 mV, P<0.0001; sputum, 2.96 +/-0.30 vs. 1.09 +/-0.01 mV, P<0.01; BAL, 25.47 +/-0.88 vs. 8.22+/-0.20 mV, P<0.0001. Both spontaneous and fMLP-induced release of elastase and myeloperoxidase (MPO) was in most cases enhanced after DTT-treatment (P-values range from 0.24 to <0.01). We conclude that the use of DTT to homogenize sputum for dispersion of cells is harmful to cell functions and these cells are hampered for the evaluation of their normal functional characteristics.

The interrelationship between markers of inflammation and oxidative stress in chronic obstructive pulmonary disease: modulation by inhaled steroids and antioxidant.

BACKGROUND: Chronic obstructive pulmonary disease (COPD) is accompanied by both airway and systemic inflammation and by oxidative stress. This study aimed to characterise the relationship between oxidative stress and inflammatory components in induced sputum and blood. MATERIAL & METHODS: We studied blood and sputum samples from stable COPD patients (mean FEV1 60.5+/-7.5% predicted) at baseline (no treatment) and after 10 weeks treatment with either inhaled steroid, fluticasone propionate (FP) (1000 microg/d) or 10 weeks treatment with N-acetylcysteine (600mg/d) (NAC). We assessed the inflammatory markers (IL-8, ECP, sICAM-1, NE) in sputum and serum and we compared them with blood markers of oxidative stress (SOD, GPx, TEAC, albumin, vitamin E and A). RESULTS: At baseline blood sICAM-1 correlated with IL-8 levels (P<0.01, r = 0.62) and negatively with GPx (P<0.01, r = -0.63) and with TEAC (P<0.05, r = -0.53). TEAC correlated positively with GPx (P<0.01, r = 0.70). Correlation between sICAM and IL-8 disappeared after NAC treatment. The correlation between sICAM and GPx disappeared after FP treatment. The correlation between TEAC and GPx was maintained after both NAC and FP. CONCLUSIONS: The relationship between markers of inflammation, adhesion and antioxidant capacity is significantly modulated by treatment with N-acetylcysteine or inhaled corticosteroids.

Pulmonary haemodynamics in patients with OSAS or an overlap syndrome.

BACKGROUND: Alveolar hypoxia is the most important mechanism leading to pulmonary arterial vasoconstriction, remodelling and pulmonary hypertension. Patients with Obstructive Sleep Apnoea Syndrome (OSAS) experience multiple short periods of alveolar hypoxia during apnoeic episodes. However, the question as to whether these hypoxic episodes are responsible for the development of permanent pulmonary hypertension is still debatable. We aimed to investigate the relationship between the episodes of nocturnal desaturation and pulmonary haemodynamics in two distinct group patients: with pure OSAS or an overlap syndrome. METHODS: We studied 67 patients with severe OSAS (means: age 45+/-8 years, AHI 62+/-22, FEV1 3.6+/-0.8 L = 97+/-16% of predicted PaO2 72+/-10 mmHg, PaCO2 40+/-4 mmHg) and 17 patients with an overlap syndrome (OS), means: age 51+/-5 years, AHI 64+/-19, FEV1 1.5+/-0.7 = 43+/-16% of predicted PaO2 57+/-9 mmHg). All subjects underwent pulmonary artery catheterisation with pressure and flow recordings and an overnight full sleep study. RESULTS: On average patients with OSAS had nocturnal desaturation (mean overnight SaO2 = 87+/-5%) and normal PPA (15.8+/-4.6 mmHg). Only 11 out of 67 subjects (16%) presented with pulmonary hypertension. Patients with OS had nocturnal desaturation (mean overnight SaO2 = 80.2+/-8.5%) and mild pulmonary hypertension (PPA 24.2+/-7.4 mmHg). Only three out of 17 patients had normal pulmonary arterial pressure. CONCLUSIONS: In patients with severe OSAS, pulmonary hypertension is rare (16%) and is related best to the severity of the disease and to obesity. In OS patients diurnal pulmonary hypertension is frequent but does not correlate with the severity of nocturnal desaturation.

Effects of almitrine bismesylate on arterial blood gases in patients with chronic obstructive pulmonary disease and moderate hypoxaemia: a multicentre, randomised, double-blind, placebo-controlled study.

BACKGROUND: Advanced chronic obstructive pulmonary disease (COPD) generates high costs, especially when patients require domiciliary long-term oxygen therapy (LTOT). Almitrine bismesylate has been shown to improve gas exchange in the lungs. Our hypothesis was that long-term treatment with almitrine might postpone the prescription of LTOT. OBJECTIVE: To evaluate the effects of almitrine sequential treatment on arterial blood gases in COPD patients with moderate hypoxaemia. METHODS: COPD patients with moderate hypoxaemia [partial oxygen tension in arterialised blood (PaO(2)) between 7.33 and 8.66 kPa (56-65 mm Hg)] were investigated. After a 1-month run-in period, patients were given either almitrine 100 mg per day or placebo for sequential treatment for a total of 12 months. RESULTS: 115 patients in a steady state (57 in the almitrine and 58 in the placebo group) were included. Mean age was 60 years, mean forced expiratory volume in 1 s was 34 +/- 13% of predicted and mean PaO(2) was 8.04 +/- 0.5 kPa (60.5 +/- 3.8 mm Hg). 38 patients were lost to follow-up, 23 in the almitrine and 15 in the placebo group. The majority of drop-outs were due to adverse events (AE; 16 in the almitrine and 9 in the placebo group). Almitrine treatment resulted in PaO(2) improvement of 0.43 +/- 0.88 kPa (3.2 +/- 6.6 mm Hg) (p = 0.003). The treatment effect between almitrine and placebo was 0.45 kPa (3.4 mm Hg) (p = 0.003). In the almitrine group, two distinct subgroups were observed: responders (n = 19) and non-responders (n = 38). Almitrine treatment in responders resulted in a clinically significant improvement in PaO(2) of 1.36 +/- 0.7 kPa (10.2 +/- 5.3 mm Hg) (p < 0.0001) and a reduction of partial carbon dioxide tension in arterialised blood. 31 patients experienced serious AE: 17 in the almitrine and 14 in the placebo group. Five patients died during the study (3 in the almitrine and 2 in the placebo group). Most AE occurring during the study were related to underlying disease. Clinical diagnosis of polyneuropathy resulted in the withdrawal of 5 patients in the almitrine group and 3 patients in the placebo group. Four patients in the almitrine group experienced weight loss. CONCLUSIONS: Almitrine treatment of patients with severe COPD and moderate hypoxaemia resulted in a small but significant improvement in PaO(2) over 12 months. A clinically important improvement in gas exchange was observed in 33% of treated patients. These patients may be candidates for long-term treatment.