Article: article from journal or magazin.
Characterization of the enzyme involved in the processing of big endothelin-1 in human lung epithelial cells.
Pulmonary pharmacology & therapeutics
Publication types: Journal Article ; Research Support, Non-U.S. Gov't ; Research Support, U.S. Gov't, P.H.S. - Publication Status: ppublish
Biosynthesis of active endothelin-1 (ET-1) implies an enzymatic processing of the inactive precursor Big ET-1 (1-39) into the mature, 21 amino acid peptide. The aim of this study was to characterize in airway and alveolar epithelial cells the enzymes responsible for this activation. BEAS-2B and A549 cells, which both produce ET-1, were studied in vitro as models for bronchiolar and alveolar cells, respectively. Both cell lines were able to convert exogenously added Big ET-1 (0.1 microM) into ET-1, suggesting a cell surface or an extracellular processing. The conversion was inhibited by phosphoramidon in both cell lines with an IC50 approximately 1 microM, but not by thiorphan, a specific inhibitor of neutral endopeptidase 24.11 (NEP). The endogenous production of serum-stimulated BEAS-2B and A549 cells was not inhibited by thiorphan, and phosphoramidon showed inhibition only at high concentration (>100 microM). Western blotting following electrophoresis in reducing conditions demonstrated a protein of MR 110 corresponding to the ECE-1 monomer in both BEAS-2B and A549 cells, as well as in whole lung extracts. By RT-PCR we revealed the mRNA encoding for the ECE-1b and/or -1c subtype, but not ECE-1a, in both cell lines. We conclude that BEAS-2B and A549 cells are able to process either endogenous or exogenous Big ET-1 by ECE-1 and that isoforms 1b and 1c could be involved in this processing with no significant role of NEP.
Cell Line, Endothelin-1, Endothelins, Epithelial Cells, Glycopeptides, Humans, Lung, Neprilysin, Protease Inhibitors, Protein Isoforms, Protein Precursors, RNA, Messenger, Reverse Transcriptase Polymerase Chain Reaction, Signal Transduction, Thiorphan
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