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.