Intrapleural use of urokinase and DNase in pleural infections managed with repeated thoracentesis: A comparative cohort study.

INTRODUCTION: Evacuation of infected fluid in pleural infections is essential. To date, the use of an intrapleural fibrinolytic agent such as urokinase and DNase has not yet been assessed in infections managed by repeated therapeutic thoracentesis (RTT). METHODS: We performed a retrospective comparative study of two successive cohorts of consecutive patients with pleural infections from 2001 to 2018. Between 2001 and 2010, patients had RTT with intrapleural urokinase (RTT-U). After 2011, patients received intrapleural urokinase and DNase with RTT (RTT-UD). Data were collected through a standardized questionnaire. RESULTS: One hundred and thirty-three patients were included: 93 were men and the mean age was 59 years (standard deviation 17.2). Eighty-one patients were treated with a combination of intrapleural urokinase and DNase, and 52 were treated with intrapleural urokinase only. In the RTT-UD, RTT failure occurred in 14 patients (17%) compared to 10 (19%) in the RTT-U group (P = 0.82). There was no difference between the two groups in intensive care unit admission, surgical referrals or in-hospital mortality. RTT-UD was associated with faster time to apyrexia (aOR = 0.51, 95%CI [0.37-0.72]), a reduced length of hospital stay (aOR = 0.61, 95%CI [0.52-0.73]) and a higher volume of total pleural fluid retrieved (aOR = 1.38, 95%CI [1.02-1.88]). Complications were rare with only one hemothorax in the RTT-UD group and no pneumothorax requiring drainage in either group. CONCLUSION: Compared to urokinase only, intrapleural use of urokinase and DNase in RTT was associated with quicker defervescence, shorter hospital stay and increased volumes of pleural fluid drained. Randomized controlled trials evaluating urokinase and DNase with RTT technique would be required to confirm these results.

Diagnosis of anaplastic lymphoma kinase-negative anaplastic large cell lymphoma on pleural fluid cytology.

ALK-negative anaplastic large cell lymphoma (ALCL) is an aggressive non-Hodgkin T-cell lymphoma. Compared to ALK-positive ALCL, patients with ALK-negative ALCL typically are older, present with nodal disease, and have lower survival rates. We report a unique presentation of ALK-negative ALCL in a pleural fluid. Cell block preparation enabled both confirmation of the diagnosis via immunohistochemical staining and gene rearrangement testing for prognostic information.

NOX1 Promotes Mesothelial-Mesenchymal Transition through Modulation of Reactive Oxygen Species-mediated Signaling.

Pleural organization may occur after empyema or complicated parapneumonic effusion and can result in restrictive lung disease with pleural fibrosis (PF). Pleural mesothelial cells (PMCs) may contribute to PF through acquisition of a profibrotic phenotype, mesothelial-mesenchymal transition (MesoMT), which is characterized by increased expression of α-SMA (α-smooth muscle actin) and other myofibroblast markers. Although MesoMT has been implicated in the pathogenesis of PF, the role of the reactive oxygen species and the NOX (nicotinamide adenine dinucleotide phosphate oxidase) family in pleural remodeling remains unclear. Here, we show that NOX1 expression is enhanced in nonspecific human pleuritis and is induced in PMCs by THB (thrombin). 4-Hydroxy-2-nonenal, an indicator of reactive oxygen species damage, was likewise increased in our mouse model of pleural injury. NOX1 downregulation blocked THB- and Xa (factor Xa)-mediated MesoMT, as did pharmacologic inhibition of NOX1 with ML-171. NOX1 inhibition also reduced phosphorylation of Akt, p65, and tyrosine 216-GSK-3ß, signaling molecules previously shown to be implicated in MesoMT. Conversely, ML-171 did not reverse established MesoMT. NOX4 downregulation attenuated TGF-ß- and THB-mediated MesoMT. However, NOX1 downregulation did not affect NOX4 expression. NOX1- and NOX4-deficient mice were also protected in our mouse model of Streptococcus pneumoniae-mediated PF. These data show that NOX1 and NOX4 are critical determinants of MesoMT.

Loss of BAP1 Expression in Atypical Mesothelial Proliferations Helps to Predict Malignant Mesothelioma.

Distinguishing reactive mesothelial proliferation from malignant mesothelioma (MM) can be difficult, particularly on small biopsies. In this scenario, a diagnosis of atypical mesothelial proliferation might be rendered. However, the distinction between a reactive process and MM is important for prognosis and treatment. Recently, loss of BRCA1-associated protein 1 (BAP1) expression and/or homozygous deletion of CDKN2A were identified in some MM, but not in reactive mesothelial proliferations. We studied 34 cases of atypical mesothelial proliferation from our institutional files (1993 to 2016) for BAP1 expression, deletion of CDKN2A, and clinical outcome. Fifteen of 34 patients (44%) were subsequently diagnosed with MM. BAP1 expression was lost in 6 of these 15 (40%) patients. Ten of 15 (67%) patients died of disease within a median time of 18.2 months. BAP1 expression was also lost in 1 case of probable MM. In this case atypical mesothelial proliferation was identified in the pleura during a lobectomy procedure for lung adenocarcinoma. Follow-up of 57.0 months was remarkable for visceral and parietal pleural thickening with continued unilateral effusion identified on imaging studies but no subsequent definitive diagnosis of MM. CDKN2A studies by fluorescence in situ hybridization (performed in 31 cases) found no homozygous deletion of that gene in any case. In conclusion, loss of BAP1 expression in atypical mesothelial proliferation helps to predict MM and is a useful adjunct test in these cases. Homozygous deletion of CDKN2A in mesothelial cell proliferations did not prove to be useful to predict MM in cases of atypical mesothelial proliferation.

Heme Oxygenase-1 Deficiency Diminishes Methicillin-Resistant Staphylococcus aureus Clearance Due to Reduced TLR9 Expression in Pleural Mesothelial Cells.

Methicillin Resistant Staphylococcus aureus (MRSA) cause pneumonia and empyema thoraces. TLR9 activation provides protection against bacterial infections and Heme oxygenase-1 (HO-1) is known to enhance host innate immunity against bacterial infections. However, it is still unclear whether HO-1 regulates TLR-9 expression in the pleura and modulates the host innate defenses during MRSA empyema. In order to determine if HO-1 regulates host innate immune functions via modulating TLR expression, in MRSA empyema, HO-1+/+ and HO-1-/- mouse pleural mesothelial cells (PMCs) were infected with MRSA (1:10, MOI) in the presence or absence of Cobalt Protoporphyrin (CoPP) and Zinc Protoporphyrin (ZnPP) or CORM-2 (a Carbon monoxide donor) and the expression of mTLR9 and mBD14 was assessed by RT-PCR. In vivo, HO-1+/+ and HO-1-/- mice were inoculated with MRSA (5x106 CFU) intra-pleurally and host bacterial load was measured by CFU, and TLR9 expression in the pleura was determined by histochemical-immunostaining. We noticed MRSA inducing differential expression of TLR9 in HO-1+/+ and HO-1 -/- PMCs. In MRSA infected HO-1+/+ PMCs, TLR1, TLR4, and TLR9 expression was several fold higher than MRSA infected HO-1-/- PMCs. Particularly TLR9 expression was very low in MRSA infected HO-1-/- PMCs both in vivo and in vitro. Bacterial clearance was significantly higher in HO-1+/+ PMCs than compared to HO-1-/- PMCs in vitro, and blocking TLR9 activation diminished MRSA clearance significantly. In addition, HO-1-/- mice were unable to clear the MRSA bacterial load in vivo. MRSA induced TLR9 and mBD14 expression was significantly high in HO-1+/+ PMCs and it was dependent on HO-1 activity. Our findings suggest that HO-1 by modulating TLR9 expression in PMCs promotes pleural innate immunity in MRSA empyema.

Pleural inhibition of the caspase-1/IL-1ß pathway diminishes profibrotic lung toxicity of bleomycin.

BACKGROUND: Idiopathic and toxic pulmonary fibrosis are severe diseases starting classically in the subpleural area of the lung. It has recently been suggested that pleural mesothelial cells acquire a myofibroblast phenotype under fibrotic conditions induced by TGF-ß1 or bleomycin. The importance and role of inflammation in fibrogenesis are still controversial. In this work, we explored the role of IL-1ß/caspase-1 signaling in bleomycin lung toxicity and in pleural mesothelial cell transformation. METHODS: C57BL/6 mice were intravenously injected with either bleomycin or nigericin or NaCl as control. In vitro, the Met5A cell line was used as a model of human pleural mesothelial cells. RESULTS: Intravenous injections of bleomycin induced lung fibrosis with histologically-proven peripheral distribution, collagen accumulation in the pleural and subpleural area, and overexpression of markers of myofibroblast transformation of pleural cells which migrated into the lung. These events were associated with an inflammatory process with an increase in neutrophil recruitment in pleural lavage fluid and increased caspase-1 activity. TGF-ß1 was also overexpressed in pleural lavage fluid and was produced by pleural cells following intravenous bleomycin. In this model, local pleural inhibition of IL-1ß with the IL-1ß inhibitor anakinra diminished TGF-ß1 and collagen accumulation. In vitro, caspase-1 inhibition interfered with Met5A cell transformation into the myofibroblast-like phenotype induced by bleomycin or TGF-ß1. Moreover, nigericin, a caspase-1 activator, triggered transformation of Met5A cells and its intra-pleural delivery induced fibrogenesis in mice. CONCLUSIONS: We demonstrated, after intravenous bleomycin injection in mice, the role of the pleura and highlighted the key role of IL-1ß/caspase-1 axis in this fibrogenesis process.

Utility of adenosine deaminase (ADA), PCR & thoracoscopy in differentiating tuberculous & non-tuberculous pleural effusion complicating chronic kidney disease.

BACKGROUND & OBJECTIVES: Pleural effusion is a common occurrence in patients with late-stage chronic kidney disease (CKD). In developing countries, many effusions remain undiagnosed after pleural fluid analysis (PFA) and patients are empirically treated with antitubercular therapy. The aim of this study was to evaluate the role of adenosine deaminase (ADA), nucleic acid amplification tests (NAAT) and medical thoracoscopy in distinguishing tubercular and non-tubercular aetiologies in exudative pleural effusions complicating CKD. METHODS: Consecutive stage 4 and 5 CKD patients with pleural effusions underwent PFA including ADA and PCR [65 kDa gene; multiplex (IS6110, protein antigen b, MPB64)]. Patients with exudative pleural effusion undiagnosed after PFA underwent medical thoracoscopy. RESULTS: All 107 patients underwent thoracocentesis with 45 and 62 patients diagnosed as transudative and exudative pleural effusions, respectively. Twenty six of the 62 patients underwent medical thoracoscopy. Tuberculous pleurisy was diagnosed in six while uraemic pleuritis was diagnosed in 20 subjects. The sensitivity and specificity of pleural fluid ADA, 65 kDa gene PCR, and multiplex PCR were 66.7 and 90 per cent, 100 and 50 per cent, and 100 and 100 per cent, respectively. Thoracoscopy was associated with five complications in three patients. INTERPRETATION & CONCLUSIONS: Uraemia remains the most common cause of pleural effusion in CKD even in high TB prevalence country. Multiplex PCR and thoracoscopy are useful investigations in the diagnostic work-up of pleural effusions complicating CKD while the sensitivity and/or specificity of ADA and 65 kDa gene PCR is poor.

Tissue inhibitor of metalloproteinase-1 is responsible for residual pleural thickening in pleural tuberculosis.

Residual pleural thickening (RPT) is the most frequent complication associated with pleural tuberculosis, and may occur even after successful anti-tuberculosis medications. Matrix metalloproteinases (MMPs) are zinc-dependent proteinases capable of degrading all components of the extracellular matrix. The proteolytic action of MMPs may be involved in the pathogenesis of tuberculosis. MMP-9, secreted by monocytes and lymphocyte, may lead to long-term fibrosis. The aim of the present study was to determine whether MMP-2 and/or MMP-9 and their specific inhibitors, tissue inhibitors of metalloproteinase 1 (TIMP-1) and TIMP-2, could be used to predict RPT. This retrospective study enrolled 52 patients diagnosed with pleural tuberculosis. Levels of MMP-2, MMP-9, TIMP-1, and TIM-2 were determined in the pleural fluid by ELISA. The RPT was measured on chest X-ray at the completion of treatment and the final follow-up. The average periods of anti-tuberculosis medication and the follow-up after completion of treatment were 6.7 and 7.6 months, respectively. MMP-2 or MMP-9 levels had no significant correlation to RPT. The patients with RPT > 2 mm at the completion of anti-tuberculosis medication and the final follow-up had higher TIMP-1 levels (p = 0.00 and p = 0.001, respectively). However, patients with RPT > 2 mm at the completion of anti-tuberculosis medication had lower TIMP-2 levels (p = 0.005). In a logistic regression model, elevated TIMP-1 levels at the completion of anti-tuberculosis medications were associated with RPT. In conclusion, higher TIMP-1 levels are responsible for the development of RPT and may be helpful for predicting RPT in pleural tuberculosis.

Pleural fluid adenosine deaminase (pfADA) in the diagnosis of tuberculous effusions in a low incidence population.

INTRODUCTION: Previous studies have assessed the diagnostic ability of pleural fluid adenosine deaminase (pfADA) in detecting tuberculous pleural effusions, with good specificity and sensitivity reported. However, in North Western Europe pfADA is not routinely used in the investigation of a patient with an undiagnosed pleural effusion, mainly due to a lack of evidence as to its utility in populations with low mycobacterium tuberculosis (mTB) incidence. METHODS: Patients presenting with an undiagnosed pleural effusion to a tertiary pleural centre in South-West England over a 3 year period, were prospectively recruited to a pleural biomarker study. Pleural fluid from consecutive patients with robust 12-month follow up data and confirmed diagnosis were sent for pfADA analysis. RESULTS: Of 338 patients enrolled, 7 had confirmed tuberculous pleural effusion (2%). All mTB effusions were lymphocyte predominant with a median pfADA of 72.0 IU/L (range- 26.7 to 91.5) compared to a population median of 12.0 IU/L (range- 0.3 to 568.4). The optimal pfADA cut off was 35 IU/L, which had a negative predictive value (NPV) of 99.7% (95% CI; 98.2-99.9%) for the exclusion of mTB, and sensitivity of 85.7% (95% CI; 42.2-97.6%) with an area under the curve of 0.88 (95% CI; 0.732-1.000). DISCUSSION: This is the first study examining the diagnostic utility of pfADA in a low mTB incidence area. The chance of an effusion with a pfADA under 35 IU/L being of tuberculous aetiology was negligible. A pfADA of over 35 IU/L in lymphocyte-predominant pleural fluid gives a strong suspicion of mTB.

Clinical significance of soluble CD26 in malignant pleural mesothelioma.

There is no established single diagnostic marker for malignant pleural mesothelioma (MPM). CD26 is a 110 kDa, multifunctional, membrane-bound glycoprotein that has dipeptidyl peptidase IV (DPPIV) enzyme activity. The aim of this study was to evaluate the clinical significance of soluble CD26 (sCD26) in patients with MPM. The study included 80 MPM patients, 79 subjects with past asbestos exposure (SPE), and 134 patients with other benign pleural diseases (OPD) that were included as a control group. sCD26 levels and DPPIV activity in serum and/or pleural fluid were determined using an ELISA kit. Serum sCD26 levels and DPPIV enzyme activity in patients with MPM were significantly decreased compared with those in the SPE group (P = 0.000). The level of serum sCD26 was significantly decreased in patients with advanced stages of MPM compared with those with earlier stages (P = 0.047). The median OS of patients with MPM who had higher DPPIV enzyme activity was significantly longer than that of those with lower DPPIV enzyme activity (P = 0.032). The sCD26 levels in the pleural fluid of MPM patients with an epithelioid subtype were significantly increased compared with the OPD cohort (P = 0.012). Moreover, DPPIV enzyme activity in the pleural fluid of patients with MPM with an epithelioid subtype were significantly increased compared with those in the OPD cohort (P = 0.009). Patients with MPM who had lower specific DPPIV activity, determined as DPPIV/sCD26, showed significantly prolonged survival compared with those with higher specific DPPIV activity (P = 0.028). Serum sCD26 and DPPIV enzyme activity appear to be useful biomarkers for differentiating patients with MPM from SPE. The sCD26 levels or DPPIV enzyme activity in pleural fluid appear to be biomarkers in patients with an epithelioid subtype of MPM. DPPIV activity in serum or pleural fluid appears to be predictive for the prognosis of patients with MPM.

Influence of storage time on pleural fluid adenosine deaminase activity.

BACKGROUND: Storing pleural fluid samples for research purposes is a common practice, but whether adenosine deaminase (ADA), an enzyme used for the diagnosis of tuberculous pleuritis, is stable over long periods of time is unknown. METHODS: We evaluated the stability of pleural ADA concentrations in 223 samples frozen at -800C as compared to values obtained immediately following the initial thoracentesis. Sample storage time ranged from several months to slightly more than 10 years. RESULTS: ADA activity was stable for up to 2.6 years. Afterwards, it decreased 6 to 8 U/L, enough to drop 2 (3.3%) tuberculous patients below the diagnostic ADA cutoff. CONCLUSIONS: As far as ADA enzymatic activity is concerned, pleural fluid samples are viable for extended periods of time. However, some caution in interpreting results from specimens stored for > 2.6 years is prudent.

Pleural mesothelium lubrication after hyaluronidase, neuraminidase or pronase treatment.

Coefficient of kinetic friction (µ) of pleural mesothelium has been found to increase markedly after mesothelial blotting and rewetting. This increase disappeared after addition of a solution with hyaluronan or sialomucin, though previous morphological studies showed that only sialomucin occurs in mesothelial glycocalyx. In this research we investigated whether µ of rabbit pleural mesothelium increased after hyaluronidase, neuraminidase or pronase treatment. Hyaluronidase and neuraminidase did not increase µ, though neuraminidase cleaved sialic acid from mesothelial glycocalyx of diaphragm specimens, and removed hystochemical stain of sialic acid from glycocalyx. Sialomucin treated with neuraminidase lowered µ of blotted mesothelium, though less than untreated sialomucin; this feature plus lubrication provided by other molecules could explain why µ did not increase after neuraminidase. Short pronase treatment (in order to affect only glycocalyx proteins) increased µ; this increase was removed by hyaluronan or sialomucin. After pronase treatment µ decreased with increase in sliding velocity, indicating a regime of mixed lubrication, as in blotted mesothelium.

Diagnostic significance of adenosine deaminase in pleural tuberculosis.

BACKGROUND: Tuberculosis (TB) is a major cause of pleural effusion, which in TB usually has lymphocytic and exudative characteristics. Analysis of adenosine deaminase (ADA) activity is a very useful diagnostic approach to achieve a more rapid and precise diagnosis in cases of Pleural TB (pTB). METHODS: Fifty male and fifty female patients presenting with tuberculous pleural effusion was included in the study. The patients were taken from the medical ward of Sir Ganga Ram Hospital between September 2001 and September 2002. Activity of Adenosine Deaminase (ADA) was estimated by the technique of Sodium dodecyl sulphate electrophoresis (SDS-EF) using 10% polyacrylamide gel. RESULTS: Mean age of males was 45.72 +/- 19.22 years and of female was 43.74 +/- 16.09 years. Mean protein level was 3.39 +/- 0.24 g/dl in males, and it was 3.02 +/- 0.26 g/dl in females. Mean specific gravity both in males and females was 1.020 +/- 0.01. The results show an increased level of enzyme ADA in patients as compared to normal subjects. CONCLUSION: Estimation of ADA activity may provide basis for rapid and efficient diagnosis of pleural TB in different clinical settings. However study should be extended to larger number of patients to reach a better conclusion.

Human parietal pleura present electrophysiology variations according to location in pleural cavity.

The aim of the study was to investigate if human pleura from different anatomical locations presents electrophysiology differences. Specimens were stripped over the 2nd-5th rib (cranial), 8th-10th rib (caudal), and mediastinum during open surgery and were mounted between Ussing chambers. Amiloride and ouabain were added towards mesothelial surface and trans-mesothelial potential difference (PD) was measured after 1, 5, 10 and 20 min. Trans-membrane resistance (R) was calculated from Ohm's law. R increased after amiloride addition, for cranial (net increase of 0.40 Omega x cm(2)) and caudal (1.16 Omega x cm(2)) pleural pieces. Mediastinal pleura R remained unchanged (0.09 Omega x cm(2)). R increase was higher for caudal than cranial (P=0.029) or mediastinal tissues (P=0.002). R increased after ouabain addition for caudal (1.35 Omega x cm(2)) and cranial (0.56 Omega x cm(2)) pleural pieces. Mediastinal pleural tissue did not respond (0.20 Omega x cm(2)). Caudally located pleura responded greater than cranial (P=0.043) or mediastinal (P=0.003) pleural tissues. Human pleura shows electrophysiology differences according to the location within the pleural cavity. Surgeons may waste mediastinal pleura when needed but should leave intact caudal parietal pleura, which seems to be electrophysiologically the most important part of the pleural cavity.

Intrapleural activation, processing, efficacy, and duration of protection of single-chain urokinase in evolving tetracycline-induced pleural injury in rabbits.

Intrapleural fibrinolysins have been used to treat pleural loculations. However, the efficacy of clinically available agents has recently been questioned, providing a rationale for investigation of new interventions. Single-chain urokinase plasminogen activator resists inhibition by serpins, and repeated, daily intrapleural administration of this agent prevents intrapleural loculation more effectively than complexes of this proenzyme with its receptor (Idell S, Mazar A, Cines D, Kuo A, Parry G, Gawlak S, Juarez J, Koenig K, Azghani A, Hadden W, McLarty J, Miller E. Am J Respir Crit Care Med 166: 920-926, 2002). Understanding of the protective mechanism and intrapleural processing remains unclear. We speculated that single-chain urokinase could induce sustained local fibrinolysis and protection by selective administration either before, during, or following loculation after pleural injury induced by tetracycline in rabbits. Enzymography, immunoassays, histology, immunohistochemistry, morphology, and morphometry were used to test the efficacy, duration of protective effect, and processing of single-chain urokinase. Intrapleural single chain urokinase prevented loculation at 72 h after injury (P < 0.01) if given either before or during adhesion formation and was converted to two-chain high-molecular-weight urokinase, which remained active for at least 24 h within pleural fluids. The effect was dose dependent, and established loculations at 72 h after tetracycline-induced injury were reversed at 96 h by single-dose treatment. Single-chain urokinase bound and saturated intrapleural plasminogen activator inhibitory (PAI)-1-like activity and urokinase-related immunoreactivity of the mesothelium was comparable in treatment or vehicle-control groups. Adhesions recurred by 2 wk after treatment with recurrence of excess local PAI activity. Single-chain urokinase induces sustained local fibrinolysis and reversibly prevents pleural loculation for up to 48 h after intrapleural administration after tetracycline-induced injury.

Pleural injury and pleurisy-induced progelatinase B/proMMP-9 is associated with markers of neutrophil degranulation.

OBJECTIVE: Progelatinase B/proMMP-9 has recently been identified as an indicator of pleural inflammation, presumably originating from granulocytes. The aim of this study was to verify the origin of progelatinase B by simultaneous estimation of specific markers of neutrophil recruitment and activation in pleural effusions following induced pleurisy and pleural injury. MATERIAL AND METHODS: Sixty-three samples of pleural fluid from patients undergoing therapeutic talc pleurodesis (n = 8) and explorative thoracoscopy (n = 3) collected before and at different time intervals after the intervention were analyzed for progelatinase B and neutrophil gelatinase-associated lipocalin (NGAL)-gelatinase complex by substrate electrophoresis, for myeloperoxidase (MPO) and interleukin-8 (IL-8) by immunoadsorbent sandwich assay, as well as for leukocyte count, C-reactive protein (CRP) and total protein (TP). RESULTS: A significant increase in free and NGAL-complexed progelatinase B, MPO and IL-8 was recorded within 48 h following treatment in all subjects. Progelatinase B was strongly correlated with NGAL-gelatinase complex (r = 0.88, p = 0.001), MPO (r = 0.81, p = 0.001), neutrophil count (r = 0.75, p = 0.01) and IL-8 (r = 0.71, p = 0.001), but not with CRP and TP. CONCLUSIONS: The results support the neutrophil origin of the proenzyme, which confirms progelatinase B as an indicator of a local inflammatory reaction. Quantifying the inflammatory reaction may be helpful in the evaluation of both the technical variants of therapeutic pleurodesis and finer discrimination of paraneoplastic effusions.

Pleural adenosine deaminase in the separation of transudative and exudative pleural effusions.

OBJECTIVE: The purpose of this study was to evaluate the usefulness of a new parameter, pleural adenosine deaminase (PADA), for separating transudative pleural effusion from exudative pleural effusion, and to compare the results with other tests (albumin gradient and protein gradient). METHODS: From November 2001 to January 2003, 359 consecutive patients with pleural effusion who underwent a diagnostic thoracentesis were included in the study. Effusions were individually classified as transudates or exudates after the careful evaluation of all clinical data and biochemical parameters of pleural fluid and serum of patients on the basis of Light's criteria. The means and standard deviations of PADA, pleural/serum ADA (P/S ADA) ratio, albumin gradient and protein gradient were evaluated for transudative and exudative effusions. The best cut-off values for each test were identified by using the receiver operating characteristic (ROC) curve. The optimum cut-off level was determined by selecting points of test values that provided the greatest sum of sensitivity and specificity. RESULTS: There were 113 transudates and 246 exudates. For each test, differences in mean value between the transudate group and the exudate group were statistically significant (t test, P<0.001). The optimum cut-off levels for PADA and P/S ADA were 15.3 U/L and 0.66 U/L, respectively. ROC analysis confirmed previous recommendations for albumin gradient (12 g/L) and protein gradient (31 g/L). For detecting exudates, the PADA test yielded a sensitivity and specificity of 85.8% and 82.3%, respectively. Sensitivity and specificity of the albumin gradient were found to be 88.5% and 79.3%, and of the protein gradient 85% and 83.2%, respectively. The areas under the curve (AUC) data and accuracy demonstrated similar discriminative properties in the examined tests. CONCLUSIONS: The measurement of PADA is suggested as a reliable test in the separation of pleural exudates from transudates with accuracy similar to that of the albumin gradient and protein gradient.

The relation of the pleural thickening in tuberculosis pleurisy with the activity of adenosine deaminase.

BACKGROUND: Residual pleural thickening (RPT) still occurs in most patients with tuberculosis pleurisy despite advances in the treatment of tuberculosis. The aim of this study was to evaluate the significance of RPT in tuberculosis pleurisy with the patients clinical findings, biochemical and microbiological properties of pleural effusion and with the total adenosine deaminase (ADA) and isoenzymes levels. METHODS: 121 tuberculosis pleurisy patients were evaluated retrospectively. According to posteroanterior chest x-rays, the 63 (52%) cases with the thickness 2 mm or more in lower lateral hemithorax were grouped as I and the 58 (48%) cases without pleural thickness were grouped as II. The amount of pleural effusion was classified into small, medium or massive according to their chest x-rays. In both groups; sex, age, symptoms score, bacteriological and biochemical tests and ADA levels were recorded. RESULTS: 81 (67%) male and 40 (33%) female, overall 121 patients were enrolled into the study. RPT was found higher in males (p=0.014) and the increase ran parallel with the amount of cigarette smoking (p=0.014). RPT was found to be lower in small effusions (p=0.001). The group with RPT, the serum albumin was found lower (p=0.002), pleural fluid total protein (p=0.047) and the ratio of pleural fluid protein to serum protein (p=0.002) were found higher. In group I, total ADA: 69.5 +/- 38.9 IU/L and ADA2: 41.3 +/- 31.6 IU/L were higher than the cases without RPT (p=0.032, p=0.017, respectively). CONCLUSIONS: We suggest that the immunological mechanisms are effective in the development of pleural thickening.

Expression of matrix metalloproteinase-9 in pleural effusions of tuberculosis and lung cancer.

BACKGROUND: Matrix metalloproteinase (MMP)-9 has been implicated in the development of pleural effusions. OBJECTIVES: The aim of this study was to assess the expression of MMP-9 in pleural effusions of tuberculosis, lung cancer and transudates. METHODS: Ninety-one patients (37 tuberculous pleural effusions, 42 malignant pleural effusions of lung cancer and 12 transudates) were included. Concentrations of pleural fluid MMP-9 and tissue inhibitors of matrix metalloproteinase (TIMP)-1 were determined by immunoassay. We also investigated the cellular localization of MMP-9 and TIMP-1 by reverse-transcriptase polymerase chain reaction on lymphocytes from pleural effusions and by immunohistochemical analysis of pleural tissues. RESULTS: Pleural fluid MMP-9 levels, MMP-9/total protein and MMP-9/TIMP-1 ratios were significantly higher in tuberculous pleural effusions, whilst TIMP-1 levels were similar in the three groups. MMP-9 levels positively correlated with TIMP-1 and lactate dehydrogenase levels, and negatively with pH and glucose levels in pleural effusions. MMP-9 mRNA expression in lymphocytes tended to be higher in malignant pleural effusions of lung cancer than in the other groups without reaching statistical significance. The strongest immunoreactivity for MMP-9 was observed in epithelioid cells of tuberculous pleural tissues. Much lower levels of MMP-9 expression were found in tumor cells of pleural tissues. CONCLUSIONS: MMP-9 is increased in tuberculous pleural effusions compared with transudates and malignant pleural effusions of lung cancer and is produced predominantly by epithelioid cells in the granulomas of tuberculous pleural tissues.

ADA2 isoform of adenosine deaminase from pleural fluid.

Adenosine deaminase isoenzyme 2 (ADA2) was isolated from human pleural fluid for the first time. Molecular and kinetic properties were characterized. It was shown that the inhibitors of adenosine deaminase isoenzyme 1 (ADA1), adenosine, and erithro-9-(2-hydroxy-3-nonyl)adenine (EHNA) derivatives are poor inhibitors of ADA2. Comparison of the interaction of ADA2 and ADA1 with adenosine and its derivative, 1-deazaadenosine, indicates that the isoenzymes have similar active centers. The absence of ADA2 inhibition by EHNA is evidence of a difference of these active centers in a close environment. The possible role of Zn2+ ions and the participation of acidic amino acids Glu and Asp in adenosine deamination catalyzed by ADA2 were shown.