Elevetad Cysteine-Protease Activity in Bronchiolitis Obliterans Syndrome

Abstract

Purpose Lung transplantation (LTx) is the final treatment option for patients with end-stage lung diseases, however, 50% of LTx patients do not survive the first 5 years due to chronic lung allograft dysfunction (CLAD). Bronchiolitis obliterans syndrome (BOS) is the most common CLAD phenotype, characterized by mononuclear cell infiltration, airway epithelial cell dysfunction, and peribronchial collagen deposition that lead to progressive bronchiolar occlusion and pulmonary function decline. The mechanisms underlying BOS development remain poorly described. Here, we investigated the role of proteases in tissue remodeling in BOS. Methods Bronchoalveolar lavage fluid (BALF) and lung tissue from LTx patients with and without BOS were analyzed by quantitative proteomics, Western blotting (WB), FRET based activity assay, ELISA, and immunostaining. Orthotopic lung transplantation of single mismatched lungs in mice was used as a model of CLAD. BALF and lung tissue were analyzed up to 2 months after LTx by WB and tissue staining. Results Quantitative proteomics of human BALF showed down-regulation of cystatin C (p = 0.0001) and cystatin S (p = 0.0188), members of the cysteine-protease inhibitor superfamily that inhibit cysteine protease activity. WB and protease activity assays further confirmed increased amount and activities of cysteine proteases in human BALF from BOS patients. Histological analysis of lungs from BOS patients demonstrated accumulation of these proteases in the peribronchiolar and perivascular compartment characterized by mononuclear infiltration and fibrosis. Histological analysis of the CLAD mouse model confirmed a significant increase of the expression of cysteine-proteases in the peribronchiolar area starting from seven days after LTx. Conclusion Here, we demonstrate that BOS is associated with an increase in the amount and activity of cysteine-proteases. Their accumulation in the peribronchiolar and perivascular area, which clearly shows signs of fibrosis, justifies further investigation into the role of these proteases in the pathogenesis, early detection, and outcome prediction of BOS.