Lysosomal enzymes from polymorphonuclear leukocytes and proteinase inhibitors in patients with cystic fibrosis

In serum and sputum samples from 15 patients with cystic fibrosis (CF) suffering from chronic Pseudomonas aeruginosa lung infections, concentrations and/or activities of elastase derived from polymorphonuclear leukocytes (PMN), cathepsin G, myeloperoxidase (MPO), and superoxide dismutase (SOD), as well as concentrations of the proteinase inhibitors alpha 1-proteinase inhibitor (alpha 1-PI) and alpha 2-macroglobulin (alpha 2-M), were determined. High enzyme concentrations compared with those in normal control subjects were found for PMN elastase (mean, 96.1 +/- 91.7 micrograms/ml), cathepsin G (mean, 5.9 +/- 6.0 micrograms/ml), and MPO (mean, 138.0 +/- 177 micrograms/ml) in patients' sputum samples. Superoxide dismutase was not detectable in any of the sputum specimens (below 1 ng/ml). Proteinase inhibitor concentrations were elevated in serum samples (alpha 1-PI: mean, 3,457 +/- 1,084 micrograms/ml; alpha 2-M: mean, 4,835 +/- 1,334 micrograms/ml). Means of 61 +/- 38 micrograms/ml alpha 1-PI and 29 +/- 31 micrograms/ml alpha 2-M were present in the sputum specimens. Both proteinase inhibitors were functional in the serum samples. However, sputum alpha 1-PI was proteolytically degraded, as shown by western blot technique, and was not able to bind 125I-labeled PMN elastase, as shown by autoradiography. Only 10.9 +/- 8.5% of the total alpha 1-PI in the sputum samples was complexed to PMN elastase and 3.6 +/- 3.2% to cathepsin G. On the other hand, 96.2 +/- 96.8% of the total PMN elastase and 78.0 +/- 100% of cathepsin G were unbound in the sputum samples. The study suggests that the imbalance between PMN proteinases and their inhibitors is due to inactivation of alpha 1-PI in the sputum caused by proteolytic or oxidative attack from PMN enzymes.     

Sputum processing for evaluation of inflammatory mediators

Neutrophil-dominated inflammation is prominent in the cystic fibrosis (CF) and chronic bronchitis (CB) airways. We assessed the degree of airway inflammation by measuring the sputum concentrations of interleukin (IL)-8, myeloperoxidase (MPO), and deoxyribonucleic acid (DNA). We determined the relationship among the concentrations of these mediators and investigated methodological problems that may be responsible for reported variability in measurements. Sputa obtained from 31 patients were solubilized with phosphate-buffered saline, dithiothreitol (DTT) (0.1% or 1%), or dornase alfa (0.2 mg/mL). The sputum concentration of IL-8 and MPO was measured by enzyme-linked immunosorbent assay (ELISA), and DNA was measured using microfluorimetry. There was a significant relationship among sputum IL-8, MPO, and DNA. For MPO (means +/- SD), CF was 1,392 +/- 771 vs. CB at 75 +/- 65 mcg/mL; P < 0.0001. For IL-8: CF was 239 +/- 154 vs. CB at 121 +/- 108 ng/mL; P = 0.0002. For DNA, CF was 1.707 +/- 1.25 vs. CB at 0.184 +/- 0.272 mg/mL; P < 0.0001. The MPO concentration in CF sputum was approximately double after in vitro treatment with dornase alfa (P < 0.0001). There is a greater concentration of IL-8, MPO, and DNA in CF than in CB sputa. There is a significant relationship among these inflammatory markers in sputum. DNA polymers bind myeloperoxidase in the sputum, and we speculate that treatment with dornase alfa may remove a source of MPO inhibition.     

Effect of clarithromycin on airway obstruction and inflammatory markers in induced sputum in cystic fibrosis: a pilot study.

To determine whether macrolide antibiotics improve pulmonary function and decrease airway inflammation in cystic fibrosis (CF), we treated 10 patients (females; aged 19-26 years, all colonized with P. aeruginosa, none with atypical Mycobacteria) with 3 weeks of placebo, followed by 6 weeks of clarithromycin (500 mg BID) in a single-blind prospective study. We also determined the safety of sputum induction and the reproducibility of assessing inflammatory markers in induced sputum. Subjects performed spirometry and underwent sputum induction (12-min inhalation of 3% saline) at 3-week intervals. We found that sputum induction was well-tolerated. We also found that the reproducibility was high for neutrophil (PMN) number (R = 0.87, P = 0.009), interleukin (IL)-8 (R = 0.73, P < 0.05, free neutrophil elastase (NE) (R = 0.82, P < 0.05), and myeloperoxidase (MPO) levels (R = 0.86, P < 0.05), but was less so for tumor necrosis factor (TNF)-alpha (R = -0.15, P = 0.7). We found no significant difference in pulmonary function after 6 weeks of treatment with clarithromycin (FEV(1) (% predicted) (mean +/- SEM), 2.2 +/- 0.9 (60 +/- 24%) vs. 2.3 +/- 1 (61 +/- 29%)), and no significant differences in any of the inflammatory indices measured. The median (and range) values before and after treatment for indices of airway inflammation in the induced sputum samples were: for PMNs, 8 (1-326) and 21 (0.2 -175) x 10(6) cells/mL sputum; for IL-8, 156 (24-656) and 202 (16-680) ng/mL; for free NE, 260 (31-1,264) and 237 (49-1,048) microg/mL; for TNF-alpha, 20 (7-128) and 35 (17-87) pg/mL; and for MPO, 169 (13-960) and 195 (14-816) microg/mL. We conclude that clarithromycin is not uniformly effective in improving airway obstruction or in decreasing airway inflammation in patients with CF.     

Evidence for excessive bronchial inflammation during an acute exacerbation of chronic obstructive pulmonary disease in patients with alpha(1)-antitrypsin deficiency (PiZ)

Patients with homozygous (PiZ) alpha(1)-antitrypsin (AAT) deficiency have not only low baseline serum AAT levels (approximately 10 to 15% normal) but also an attenuated acute phase response. They are susceptible to the development of premature emphysema but may also be particularly susceptible to lung damage during bacterial exacerbations when there will be a significant neutrophil influx. The purposes of the present study were to assess the inflammatory nature of acute bacterial exacerbations of chronic obstructive pulmonary disease (COPD) in subjects with AAT deficiency, to compare this with COPD patients without deficiency, and to monitor the inflammatory process and its resolution following appropriate antibacterial therapy. At the start of the exacerbation, patients with AAT deficiency had lower sputum AAT (p < 0.001) and secretory leukoprotease inhibitor (SLPI; p = 0.02) with higher elastase activity (p = 0.02) compared with COPD patients without deficiency. Both groups had a comparable acute phase response as assessed by C-reactive protein (CRP) but the AAT-deficient patients had a minimal rise in serum AAT (to < 6 microM). After treatment with antibiotics, in patients with AAT deficiency, there were significant changes in many sputum proteins including a rise in SLPI levels, and a reduction in myeloperoxidase (MPO) and elastase activity (p < 0. 005 for all measures); the sputum chemoattractants interleukin-8 (IL-8) and leukotriene B(4) (LTB(4)) fell (p < 0.01), and protein leak (sputum/serum albumin ratio) became lower (p < 0.01). The changes were rapid and within 3 d of the commencement of antibiotic therapy the biochemical markers had decreased significantly, but took a variable time thereafter to return to baseline values. In conclusion, patients with AAT deficiency had evidence of increased elastase activity at the start of the exacerbation when compared with nondeficient COPD patients which probably reflects a deficient antiproteinase screen (lower sputum AAT and SLPI). The increased bronchial inflammation at presentation resolved rapidly with 14 d of antibiotic therapy.     

Elevation of prostate-specific markers after cardiopulmonary resuscitation

BACKGROUND-Prostate-specific antigen (PSA), acid phosphatase (AP), and prostatic acid phosphatase (PAP) are serum markers for adenocarcinoma of the prostate gland. Previous studies indicated that prostatic ischemia may also produce elevations of PSA. Cardiopulmonary resuscitation (CPR) is frequently associated with profound tissue hypoperfusion. The present study investigated whether PSA, AP, and PAP are influenced by prolonged CPR. METHODS AND RESULTS-PSA, AP, and PAP were assessed immediately, 12 hours, 24 hours, 2 days, 3 days, 5 days, and 7 days after prolonged CPR (>5 minutes) in 14 male and 5 female patients. No changes were noted in women. In men, serum levels increased significantly after CPR and gradually decreased to near baseline values after 7 days. PSA, AP, and PAP values above the normal range were observed in 63%, 71%, and 64% of all patients, respectively. Compared with survivors, nonsurvivors exhibited higher peak serum levels of PSA (98.6+/-14.3 versus 1.1+/-2.2 mcg/L; P<0.03), AP (57.0+/-71 versus 8.6+/-8.8 U/L; P<0.05), and PAP (47.0+/-62 versus 5.7+/-8.0 U/L; P=NS). Patients with poor neurological outcome exhibited higher peak serum levels of PSA (86.4+/-135.5 versus 12.0+/-23.8 mcg/L; P<0.05), AP (50.9+/-68.1 versus 8.7+/-9.6 U/L; P=NS), and PAP (41.6+/-59.5 versus 5.8+/-8.8 U/L; P=NS) than patients with good neurological outcome. CONCLUSIONS-Prolonged CPR is frequently associated with increases of PSA, AP, and PAP serum levels. Therefore, PSA cannot be used for diagnosis of adenocarcinoma of the prostate during the first weeks after CPR. Further evaluation of these parameters as additional prognostic markers after CPR is warranted.     

Biochemical and pathologic evidence for proteolytic destruction of lung connective tissue in cystic fibrosis

The risk for proteolysis of lung connective tissue was evaluated in patients with cystic fibrosis (CF) with chronic, severe lung infections by measuring uninhibited elastase activity in sputum samples and urinary excretion of desmosines (cross-linking amino acids in elastin). Of the 16 patients included in the study, 11 were infected with Pseudomonas aeruginosa, 2 with Pseudomonas cepacia, and 2 with both P. aeruginosa and P. cepacia. Uninhibited elastase activity (0.34 to 20.2 micrograms elastin degraded/mg protein/30 min) was detected in the sputum samples from each of 13 patients tested. Serine elastase activity was detected in the sputum of each of 12 patients, and metalloelastase (P. aeruginosa elastase and possibly macrophage elastase) activity was detected in the sputum of 11 of 12 patients tested. Male patients with CF excreted significantly more elastin cross-links (desmosines) in their urine than did control male subjects (3.6 +/- 1.7 micrograms/kg/24 h versus 1.5 +/- 0.6 micrograms/kg/24 h; p less than 0.01), and there was a significant correlation (p less than 0.05) between urine desmosine excretion and the severity of lung disease in the patients with CF as indicated by chest roentgenogram score. In 3 autopsied patients, abnormal elastin fibers were seen by light microscopy in all lung compartments. Fragmented and exfoliated elastin, evidence of active elastolysis, was noted in bronchial ulcers and abscesses. The results of this study suggest that proteolytic destruction of lung connective tissue is an ongoing process in the chronically infected CF lung and that this proteolysis contributes to the pathologic changes observed in airways and alveolar parenchyma.     

哮喘诱导痰中弹性蛋白酶和α_1-抗胰蛋白酶水平与气道炎症气流受限的关系

目的:探讨中重度支气管哮喘(哮喘)患者诱导痰中弹性蛋白酶和α1-抗胰蛋白酶水平及其与气道炎症和气流受限的关系。方法:测定12例慢性持续期中重度哮喘患者(哮喘组),12例稳定期中度慢性阻塞性肺疾病(COPD)患者和10例健康对照者的(健康对照组)肺功能。并对其诱导痰进行炎性细胞分类计数,测定诱导痰上清液中IL-8、弹性蛋白酶浓度和α1-抗胰蛋白酶的活性。结果:哮喘和COPD患者诱导痰中弹性蛋白酶浓度和α1-抗胰蛋白酶活性与健康对照组比较差异有统计学意义,且与中性粒细胞数、IL-8浓度呈正相关,但与FEV1/FVC百分比值呈负相关。哮喘和COPD患者诱导痰中弹性蛋白酶浓度/α1-抗胰蛋白酶活性比值与健康对照组比较差异有统计学意义,且与FEV1/FVC百分比值呈正相关。结论:中重度哮喘患者诱导痰中存在弹性蛋白酶与α1-抗胰蛋白酶间失平衡,增高的弹性蛋白酶与其气道炎症和气流受限有关(与COPD患者相类似),这在其气道重塑的发病机制中可能发挥重要作用。IL-8参与了中重度哮喘患者气道炎症和气道重塑的过程。     

慢性阻塞性肺疾病和哮喘患者痰中降钙素基因相关肽的变化

目的 探讨慢性阻塞性肺疾病（ＣＯＰＤ）和支气管哮喘患气道分泌物中,感觉神经肽降钙素基因相关肽（ＣＧＲＰ）释放在气道慢性炎症性疾病发病中的作用。方法 选择稳定期ＣＯＰＤ患１９例,支气管哮喘患１４例,正常对照组１０名,通过面罩吸入超声雾化的４％向张盐水诱导痰液,将获得的诱导痰涂片进行瑞氏－姬姆萨染色计数细胞成分的变化,用放射免疫方法测定诱导痰中ＣＧＲＰ免疫反应性物质（ＣＧＲＰ－ＬＩ）的含量改变,     

Impaired activity of serum alpha-1-antitrypsin in diabetes mellitus

Alpha-1-antitrypsin (AAT) is the prototypic member of the serine protease inhibitor (serpin) superfamily of proteins, which has a major role in inactivating neutrophil elastase and other proteases to maintain protease-antiprotease balance. In this study, the serum AAT was measured using enzymatic assay in diabetic patients. The serum trypsin inhibitory capacity (s-TIC) was determined in 144 outpatients with uncontrolled insulin-dependent diabetes mellitus (n=47) and non-insulin dependent diabetes mellitus (n=97). The s-TIC values were 1.960+/-0.399, 2.002+/-0.4304 and 2.867+/-0.395 micromol/min/ml in IDDM, NIDDM and healthy subjects, respectively. The diabetics individual had a significantly lower s-TIC than controls (P<0.0001). It was found that there was a negative correlation between the s-TIC and the duration of diabetes (r=-0.5420; P<0.0001). There was no correlation between s-TIC and age of patients (P=0.865). Thus, diabetes is associated with reduced trypsin inhibitory capacity of plasma.     

Inflammatory and microbiologic markers in induced sputum after intravenous antibiotics in cystic fibrosis

Induced sputum has been used to study airway inflammation. We sought to determine whether markers of infection and inflammation in induced sputum were a useful and safe outcome measure in cystic fibrosis. We hypothesized that bacterial density and inflammatory content of induced sputum would decrease after antibiotic therapy. Induced sputum was assayed for bacterial density, cell count, and differential and inflammatory markers before and after treatment with intravenous antibiotics. Fifty-five of the 72 subjects enrolled (mean age +/- SD 18.2 +/- 7.9 years) completed the study. FEV1 increased by an average 0.3 +/- 0.3 L (10.4 +/- 8.7% predicted FEV1), p<0.0001; density of Pseudomonas aeruginosa and Staphylococcus aureus decreased by 2.4 +/- 3.1 log10 cfu/g (p<0.0005) and 4.0 +/- 2.3 log10 cfu/ml (p<0.0001), respectively; neutrophil count decreased by 0.4 +/- 0.6 log10 cells/ml (p<0.0001), interleukin-8 concentration by 0.5 +/- 1.3 log10 pg/ml (p<0.05), and neutrophil elastase by 0.4 +/- 0.7 log10 microg/ml (p<0.005). Seven of 127 (6%) sputum induction procedures showed a decrease in FEV1 of 20% or more. We conclude that markers in induced sputum may be useful, noninvasive outcome measures to assess response to therapies in cystic fibrosis studies.     

The effect of augmentation therapy on bronchial inflammation in alpha1-antitrypsin deficiency

alpha1-Antitrypsin (AAT) deficiency predisposes to bronchitis and emphysema associated with neutrophilic airway inflammation. The efficacy of augmentation therapy has not been proven clinically or by demonstrating an effect on airway inflammation. We treated 12 patients with four infusions of Prolastin (60 mg/kg) at weekly intervals and monitored both the serum and secretion concentrations of AAT as well as markers of neutrophilic inflammation, including myeloperoxidase, elastase, and the neutrophil chemoattractants interleukin-8 and leukotriene B(4). Serum AAT rose and was maintained above the protective threshold. In addition, AAT concentrations in the sputum rose from a mean of 0.17 microM (SEM +/- 0.04) before therapy to concentrations similar to nondeficient subjects (0.43 +/- 0.12) 1 week after the first infusion (p < 0.01). This was associated with a reduction in elastase activity (p < 0.002) and the chemoattractant leukotriene B(4) (p < 0.02), which fell from a median baseline value of 13.46 nM (range, 4.17-55.00) to 8.62 nM (4.23-21.59) the day following the last infusion. Although median values for myeloperoxidase and interleukin-8 also fell, the changes failed to achieve statistical significance. In summary, short-term therapy with AAT increased lung secretion concentrations and was associated with a fall in leukotriene B(4), which is thought to be central to the airway inflammation of AAT deficiency.     

慢性阻塞性肺疾病患者诱导痰和血浆中白三烯B4的变化以及茶碱对其的影响

目的探讨白三烯B4(LTB4)在慢性阻塞性肺疾病(COPD)患者诱导痰和血浆的变化以及茶碱对其的影响.方法这个试验是前瞻性随机对照非盲法研究.40例COPD稳定期患者(C组),随机分为CA组(口服茶碱0.2 g/次,每天2次,1个月)与CB组(不用任何茶碱类药物).15名健康非吸烟者为正常对照组(H组).评定各组基线水平以及C组随访1个月后的临床症状与生活质量评分,测定肺功能指标、诱导痰细胞计数与分类,并用酶联免疫吸附测定(ELISA)与酶免疫试验(EIA)方法分别测定诱导痰和血浆白细胞介素8(IL-8)与LTB4浓度.结果 (1)C组诱导痰及血浆的LTB4水平分别为(794±305)pg/mg·pro、(5 219±1 185)ng/L,均显著高于H组[(155±64)pg/mg·pro、(2 283±489)ng/L,P均＜0.05].(2)诱导痰的LTB4水平与中性粒细胞、IL-8水平呈显著正相关(r分别为0.453、0.364,P均＜0.05).(3)CA组在茶碱治疗前、后诱导痰LTB4水平分别为(812±592)、(657±459)pg/mg·pro,血浆水平为(5 422±935)、(4 589±1 057)ng/L;CB组试验前、后诱导痰LTB4水平分别为(776±227)、(860±194)pg/mg·pro,血浆水平为(5 074±1 850)、(6 063±2 450)ng/L;这两组治疗前、后比较差异均无统计学意义(P均＞0.05).结论 COPD患者LTB4水平增高,参与气道炎症过程.茶碱的治疗不能显著降低诱导痰与血浆的LTB4水平.     

Safety and efficacy of recombinant alpha(1)-antitrypsin therapy in cystic fibrosis

Neutrophil elastase (NE) is thought to be the most important protease which damages the cystic fibrosis (CF) lung. Attempts have been made to suppress this activity using the plasma-derived inhibitor, alpha(1)-antitrypsin (AAT). In this pilot study, the safety and efficacy of inhaled recombinant human AAT (rAAT) as a treatment for CF were investigated. Thirty-nine patients participated in a prospective, double-blinded, randomized, placebo-controlled phase II trial to examine the effect of rAAT (500, 250, and 125 mg) on sputum NE activity. Sputum myeloperoxidase (MPO), interleukin-8, tumor necrosis factor receptors, sputum and plasma NE/AAT complexes, and safety parameters were also measured. Subjects were randomized to receive nebulized treatment once a day for 4 weeks, followed by 2-4 weeks with no study treatment, and then a 2-week rechallenge phase. Trends toward a reduction in NE activity were observed in patients treated with 500 mg and 250 mg of rAAT compared to placebo. Sputum NE/AAT complex and MPO levels were lower on rAAT compared to placebo. No major adverse events and, in particular, no allergic reactions to rAAT were observed. Although significant differences between rAAT and placebo for sputum NE activity were not observed, some improvements were found for secondary efficacy variables. This study demonstrated that nebulized rAAT is safe and well-tolerated, but has a limited effect on NE activity and other markers of inflammation.     

Neutrophil inflammation and activation in bronchiectasis: comparison with pneumonia and idiopathic pulmonary fibrosis

BACKGROUND: Pulmonary inflammation in bronchiectasis, pneumonia and idiopathic pulmonary fibrosis (IPF) is dominated by neutrophils. Pathophysiologic differences are seen in the degree of airway and tissue destruction. Neutrophil activation and neutrophil proteolytic activity might differ between bronchiectasis, pneumonia and IPF. OBJECTIVE: The aim of this study was to determine whether levels of inflammatory and protective markers in bronchoalveolar lavage (BAL) differed among cases of bronchiectasis, pneumonia and IPF. METHODS: We studied 11 bronchiectasis patients (group 1), 30 pneumonia patients (group 2), 15 IPF patients (group 3) and 12 healthy volunteers (group 4). In the bronchoalveolar lavage fluid, concentrations of alpha(1)-proteinase inhibitor, myeloperoxidase (MPO) and elastase-alpha(1)PI complex were determined using immunoluminometric assays. Elastase inhibition capacity (EIC) and elastase activity were determined using a colorimetric assay. RESULTS: No EIC, but free elastase activity, was found in 82% of group 1, 20% of group 2, 20% of group 3 and 0% of group 4. Median MPO concentration was highest in group 1: 7,951 ng/ml (16th-84th percentile [16-84%]: 256-36,342) vs. 692 ng/ml (106-2,279; group 2), 332 ng/ml (98-1,657; group 3), and 0.12 ng/ml (0.08-0.26; group 4). Bronchiectasis patients with bronchial Pseudomonas infection showed higher amounts of neutrophils (p < 0.01) and higher elastase activity (p < 0.05) than patients with sterile lavage. CONCLUSION: Bronchiectasis patients show a severe imbalance between neutrophil activity and protective molecules leading to possible lung destruction. Chronic Pseudomonas infection might trigger neutrophil activation. Future research and treatment strategies should focus on increased bacterial clearance and inhibition of neutrophil toxicity.     

Prone position improves expiratory airway mechanics in severe chronic bronchitis.

Based on lung parenchyma-airways' interdependence, the present authors hypothesised that prone positioning may reduce airway resistance in severe chronic bronchitis. A total of 10 anaesthetised/mechanically ventilated patients were enrolled. Partitioned respiratory system (RS) mechanics during iso-flow experiments (flow = 0.91 L x s(-1), tidal volume (VT) varied within 0.2-1.2 L), haemodynamics, gas-exchange, expiratory airway resistance (Raw,exp), functional residual capacity (FRC), change in FRC (DeltaFRC), end-expiratory lung volume (EELV), expiratory airway resistance at EELV (Raw,exp,EELV), intrinsic positive end-expiratory pressure (PEEPi), and mean end-expiratory flow were determined in baseline semirecumbent (SRBAS), prone, and post-prone semirecumbent (SRPP) postures. Pronation versus SRBAS resulted in significantly reduced Raw,exp (at VT > or =0.8 L), Raw,exp,EELV (18.3+/-1.4 versus 31.6+/-2.6 cm H2O x L(-1) x s(-1)), inspiratory airway resistance (at VT > or =1.0 L), static lung elastance (at VT < or =0.6 L), "additional" RS/lung resistance (at a range of VTs), DeltaFRC (0.35+/-0.03 versus 0.47+/-0.03 L), EELV (4.92+/-0.49 versus 5.65+/-0.65 L), RS/lung PEEPi (6.7+/-1.1/5.4+/-0.6 versus 8.9+/-1.7/7.8+/-1.1 cm H2O), mean end-expiratory flow (63.9+/-4.2 versus 47.9+/-4.0 mL x s(-1)), and shunt fraction (0.16+/-0.03 versus 0.21+/-0.03); benefits were reversed in SRPP. In severe chronic bronchitis, prone positioning reduces airway resistance and dynamic hyperinflation.     

Plasma factor VII and thrombin–antithrombin III levels indicate increased tissue factor activity in sickle cell patients

Although the mechanisms involved in the persistent clinical complications of sickle cell disease have not yet been fully delineated, previous studies suggest that sickle cell (HbSS) patients have a disposition to generate more thrombin and plasma in vivo than normal subjects. The reasons for the impaired regulation of haemostasis in HbSS patients is poorly understood. We report studies evaluating the extent to which in vivo coagulation and fibrinolysis are altered in HbSS patients in steady state. The concentrations of total factor VII (F(VII)t), factor VII zymogen (F(VII)z), thrombin-antithrombin III (TAT), fibrinopeptide A(FPA), and fibrin D-dimer in plasmas of 50 normal controls (HbAA) and 45 HbSS steady state patients, were measured using sensitive and specific enzyme-linked immunoassays. The average plasma concentration of F(VII)t, in sickle cell plasma was significantly lower than that of the control subjects (0.70 +/- 0.19 U/ml versus 1.16 +/- 0.41 U/ml), whereas F(VII)z in the patients and controls were 0.47 +/- 0.15 U/ml and 1.15 +/- 0.33 U/ml respectively, P < 0.001. Both measures of factor VII suggest a higher factor VII turnover in sickle cell disease. The mean concentration of TAT in the plasma of HbSS patients were significantly higher than those of HbAA controls (371 +/- 44 pM versus 42 +/- 2 pM) (P < 0.001), a difference that is strongly indicative of higher rates of in vivo thrombin generation by HbSS patients. Plasmas of HbSS patients had significantly higher concentrations of FPA compared to those of the control subjects (12.85 +/- 1.96 ng/ml versus 4.22 +/- 0.37 ng/ml) (P < 0.001). The D-dimer levels were also higher in the HbSS than control plasmas (1029.6 +/- 58.6 ng/ml versus 224.3 +/- 27.6 g/ml) (P < 0.001), with the patients' values being indicative of enhanced fibrinolysis. These results strongly suggest accelerated in vivo coagulation and fibrinolysis in HbSS patients even during steady state. They are consistent with the hypothesis that haemostasis is less tightly regulated in the HbSS patients than in HbAA controls. The altered regulation of haemostasis may contribute to the initiation of vaso-occlusive processes associated with sickle cell painful episodes.     

Increased CD40 ligand and platelet-monocyte aggregates in patients with type 1 diabetes mellitus

BACKGROUND: Diabetes mellitus is a major risk factor for cardiovascular disease and is associated with a proinflammatory and prothrombotic state. We investigated whether CD40 ligand (L) expression and platelet-monocyte aggregation are increased in patients with type 1 diabetes. METHODS: Serum C-reactive protein (CRP) and soluble (s) CD40L concentrations, platelet surface CD40L expression and platelet-monocyte aggregates were measured in 22 patients with uncomplicated type 1 diabetes and 22 age- and sex-matched non-diabetic control subjects. RESULTS: In comparison to controls, patients with type 1 diabetes had higher serum CRP concentrations (3.29 +/- 0.9 mg/L versus 0.99 +/- 0.2mg/L, P = 0.01), serum sCD40L concentrations (10.0 +/- 1.4 ng/mL versus 4.6 +/- 0.6 ng/mL, P = 0.006), and platelet surface expression of CD40L (13.8 +/- 0.9% versus 8.5 +/- 1.1%, P < 0.001). Platelet-monocyte aggregates were also significantly elevated in type 1 diabetes (35.9 +/- 3.3% versus 26.4 +/- 2.9%, P = 0.005; n = 10). We also observed a significant correlation between plasma glucose and serum CRP (r = 0.53, P = 0.01) as well as platelet-monocyte aggregates (r = 0.69, P = 0.03). CONCLUSIONS: Type 1 diabetes is associated with increased CD40L expression and platelet-monocyte aggregation, which may contribute to the proinflammatory and prothrombotic state as well as the accelerated atherogenesis associated with this disorder.     

Increased response of blood eosinophils to various chemotactic agents in quiescent Crohn disease

BACKGROUND: The number of eosinophils is increased in the mucosae of the digestive and the respiratory tracts in Crohn disease, even clinically quiescent. The mechanisms underlying this panmucosal eosinophilia are unknown. METHODS: The response of blood eosinophils to various chemotactic agents was assessed in 15 patients with clinically quiescent Crohn disease. The results were compared with 15 healthy controls. After purification, eosinophils were placed in Boyden microchambers and the chemotactic effect of PAF (10(-7) M), RANTES (50 ng/ml), IL-5 (0-20 ng/ml), IL-8 (0-50 ng/ml), Eotaxin (0-50 ng/ml) was evaluated. The number of eosinophils in induced sputum of these Crohn disease patients and controls was also assessed and the correlation between chemotaxis and eosinophil count in induced sputum was studied. RESULTS: PAF and RANTES induced a chemotactic effect both in Crohn disease patients and controls. The chemotactic index was significantly higher in Crohn than controls for PAF (2.09+/-0.24 versus 1.37+/-0.14; P < 0.05) but not RANTES. With IL-5, IL-8 and Eotaxin, there was no detectable chemotactic effect in controls while in Crohn, we observed a significant dose-dependent chemotactic effect. Furthermore, with Eotaxin 50 ng/ml, the chemotactic index was significantly higher in Crohn disease patients than controls (2.42+/-0.18 versus 1.56+/-0.28; P < 0.05). A significant increase in sputum eosinophil count and a significant decrease in sputum macrophage count in Crohn disease were observed. However, there was no correlation between eosinophil chemotaxis and sputum eosinophil count in individual patients. CONCLUSION: There is an increased response of blood eosinophils to various chemotactic agents, mainly PAF and Eotaxin, in clinically quiescent Crohn disease. This may participate in the mucosal infiltration by eosinophils in this disease.     

Airways inflammation in chronic bronchitis: the effects of smoking and alpha1-antitrypsin deficiency.

Airways inflammation in chronic bronchitis is thought predominantly to be a direct consequence of neutrophil recruitment and release of elastase in response to factors such as cigarette smoke. The aims of this study were to assess the role of smoking and determine whether the serum elastase inhibitor alpha1-antitrypsin (alpha1AT) influenced the process. Airways inflammation was compared between patients with chronic obstructive bronchitis with (n=39) and without (n=42) severe alpha1AT deficiency. The authors assessed the sputum concentration of the neutrophil chemoattractants interleukin-8 (IL-8) and leukotriene (LT)B4, myeloperoxidase (MPO) as a marker of neutrophil influx, neutrophil elastase activity and its natural inhibitors, alpha1AT and secretory leukoprotease inhibitor (SLPI). Finally serum alpha1AT was measured to determine the degree of protein leakage (sputum sol serum alpha1AT ratio). Compared to current smokers, the exsmokers had a lower concentration of the chemoattractant IL-8 (p<0.05) and a lower MPO concentration, although this failed to reach conventional statistical significance (p=0.06). Patients with alpha1AT deficiency had greater inflammation in the larger airways with increased LTB4 (p<0.005), MPO (p<0.001), neutrophil elastase activity (p<0.01), protein leak (p<0.001), and were found to have a lower anti-proteinase screen with both reduced sputum alpha1AT (p<0.001) and SLPI concentrations (p<0.05). The reduction in sputum interleukin-8 levels in exsmokers may decrease neutrophil influx and thus explain the slower rate of neutrophil mediated progression of lung disease compared to subjects who continue to smoke. Patients with alpha1-antitrypsin deficiency had greater inflammation suggesting that alpha1-antitrypsin plays an important role in protecting the larger airways from the inflammatory effects of elastase activity and may explain their more rapid progression of disease.     

Rise of plasma myeloperoxidase during interferon therapy

Abstract The plasma myeloperoxidase (MPO) level was evaluated using a specific radio-immunoassay (RIA) for MPO in α2b-interferon (IFN)-treated patients with chronic viral hepatitis. The plasma MPO was checked before and after the initial 2 weeks use of IFN at a dose of 6 × 10⁶ U/day. The mean concentration of plasma MPO was found to be markedly higher after IFN therapy than that before the therapy (421.7 ± 34.3 vs 242.9 ± 23.0 ng/mL, P < 0.001). The plasma MPO negatively correlated with the granulocyte count ( r = -0.37, P < 0.02) and the platelet count ( r = 0.49, P < 0.01), while it positively correlated with serum alkaline phosphatase (ALP; r = 0.41, P < 0.03). The plasma MPO also showed a strong correlation with plasma polymorphonuclear granulocyte elastase (PMN elastase; r = 0.73, P < 0.001). Our study thus suggests that the increased release of MPO from destroyed granulocytes is responsible for the high concentrations of the plasma MPO in patients during IFN therapy, because the plasma MPO, PMN elastase and ALP abundant in granulocytes all increased in spite of a decrease in the granulocyte count. Granulocytopenia during IFN therapy may therefore be due to the increased destruction of granulocytes in addition to a direct suppression of the bone marrow by IFN.