ACE/ACE2 ratio and MMP-9 activity as potential biomarkers in tuberculous pleural effusions

Abstract

Objective: Pleural effusion is common problem, but the rapid and reliable diagnosis for specific pathogenic effusions are lacking. This study aimed to identify the diagnosis based on clinical variables to differentiate pleural tuberculous exudates from other pleural effusions. We also investigated the role of renin-angiotensin system (RAS) and matrix metalloproteinase (MMPs) in the pathogenesis of pleural exudates.

Experimental design: The major components in RAS and extracellular matrix metabolism, including angiotensin converting enzyme (ACE), ACE2, MMP-2 and MMP-9 activities, were measured and compared in the patients with transudative (n = 45) and exudative (n = 80) effusions. The exudative effusions were come from the patients with tuberculosis (n = 20), pneumonia (n = 32), and adenocarcinoma (n = 28).

Results: Increased ACE and equivalent ACE2 activities, resulting in a significantly increased ACE/ACE2 ratio in exudates, were detected compared to these values in transudates. MMP-9 activity in exudates was significantly higher than that in transudates. The significant correlation between ACE and ACE2 activity that was found in transudates was not found in exudates. Advanced analyses showed significantly increased ACE and MMP-9 activities, and decreased ACE2 activity in tuberculous pleural effusions compared with those in pneumonia and adenocarcinoma effusions. The results indicate that increased ACE and MMP-9 activities found in the exudates were mainly contributed from a higher level of both enzyme activities in the tuberculous pleural effusions.

Conclusion: Interplay between ACE and ACE2, essential functions in the RAS, and abnormal regulation of MMP-9 probably play a pivotal role in the development of exudative effusions. Moreover, the ACE/ACE2 ratio combined with MMP-9 activity in pleural fluid may be potential biomarkers for diagnosing tuberculous pleurisy.

Keywords: angiotensin converting enzyme; angiotensin converting enzyme 2; exudative effusion; matrix metalloproteinase-9; tuberculous effusion..

Figure 1

Enzymatic activity of ACE and ACE2 in pleural transudative and exudative effusions. The specific activities of ACE (A) and ACE2 (B) in pleural transudative (n = 45) and exudative (n = 80) effusions from 125 patients. The ratio of ACE/ACE2 in the pleural effusions (C). Pleural effusion (20 μl) was assayed for the ability to cleave the fluorescent substrate at 37°C for 1 hour with a specific ACE inhibitor or a specific ACE2 inhibitor. Each symbol represents one individual, and horizontal bars represent median values. ** and *** indicate p < 0.01 and p < 0.001, respectively, compared with transudates.

Figure 2

Correlations between ACE and ACE2 activities in pleural effusions. ACE and ACE2 activities were measured in each sample of pleural transudative effusions (A; n = 45) and exudative effusions (B; n = 80). For correlation analysis, Pearson's correlation analysis (SPSS statistics package, Chicago, IL) was applied. Statistically significant differences were established at p < 0.05. ACE and ACE2 positively correlate with each other in the group of transudative effusions (F_(1,43) = 36.052, r² = 0.456, p < 0.001) (A), but not in the group of exudative effusions (F_(1,78) = 1.567, r² = 0.020, p > 0.05) (B).

Figure 3

MMP-2 and MMP-9 activities in pleural transudative and exudative effusions. The activities of MMP-2 (A) and MMP-9 (B) in pleural transudative and exudative effusions from 125 patients were determined with zymography. The gelatinase activities detected in this study were based on pro-MMP-2 (72 kDa) and pro-MMP-9 (95 kDa). Each symbol represents one individual, and horizontal bars represent median values. *** indicates p < 0.001 compared with transudates.

Figure 4

ACE and ACE2 activities in exudative effusions from patients with tuberculosis (TB), pneumonia (Pn), and adenocarcinoma (Ad). The activities of ACE (A), ACE2 (B), and the ratio of ACE/ACE2 (C) in TB (n = 20), Pn (n = 32), and Ad (n = 28) effusions. Each symbol represents one individual, and horizontal bars represent median values. * and *** indicate p < 0.05 and p < 0.001, respectively, compared with values measured in Pn and/or Ad effusions.

Figure 5

MMP-2 and MMP-9 activities in exudative effusions from patients with tuberculosis (TB), pneumonia (Pn), and adenocarcinoma (Ad). The activities of MMP-2 (A) and MMP-9 (B) in TB (n = 20), Pn (n = 32), and Ad (n = 28) effusions. The gelatinase activities detected in this study were based on pro-MMP-2 (72 kDa) and pro-MMP-9 (95 kDa). Each symbol represents one individual, and horizontal bars represent median values. * and ** indicate p < 0.05 and p < 0.01, respectively, compared with values measured in Pn and/or Ad effusions.