title: Patterns of neutrophil serine protease-dependent cleavage of surfactant protein D in inflammatory lung disease.
abstract: The manuscript presents definitive studies of surfactant protein D (SP-D) in the context of inflammatory lung fluids. The extent of SP-D depletion in bronchoalveolar lavage fluid (BALF) of children affected with cystic fibrosis (CF) is demonstrated to correlate best with the presence of the active neutrophil serine protease (NSP) elastase. Novel C-terminal SP-D fragments of 27 kDa and 11 kDa were identified in patient lavage fluid in addition to the previously described N-terminal, 35-kDa fragment by the use of isoelectrofocusing, modified blotting conditions, and region-specific antibodies. SP-D cleavage sites were identified. In vitro treatment of recombinant human SP-D dodecamers with NSPs replicated the fragmentation, but unexpectedly, the pattern of SP-D fragments generated by NSPs was dependent on calcium concentration. Whereas the 35- and 11-kDa fragments were generated when incubations were performed in low calcium (200 microM CaCl(2)), incubations in physiological calcium (2 mM) with higher amounts of elastase or proteinase-3 generated C-terminal 27, 21, and 14 kDa fragments, representing cleavage within the collagen and neck regions. Studies in which recombinant SP-D cleavage by individual NSPs was quantitatively evaluated under low and high calcium conditions showed that the most potent NSP for cleaving SP-D is elastase, followed by proteinase-3, followed by cathepsin G. These relative potency findings were considered in the context of other studies that showed that active NSPs in CF BALF are in the order: elastase, followed by cathepsin G, followed by proteinase-3. The findings support a pre-eminent role for neutrophil elastase as the critical protease responsible for SP-D depletion in inflammatory lung disease.
authors.affiliation: Immune Disease Institute, Harvard Medical School, Boston, MA 02115, USA.
authors.forename: Jessica
authors.initials: J
authors.lastname: Cooley
authors.forename: Barbara
authors.initials: B
authors.lastname: McDonald
authors.forename: Frank J
authors.initials: FJ
authors.lastname: Accurso
authors.forename: Erika C
authors.initials: EC
authors.lastname: Crouch
authors.forename: Eileen
authors.initials: E
authors.lastname: Remold-O'Donnell
mesh_terms: D000293:Adolescent; D000328:Adult; D001249:Asthma / Q000097:blood; D001992:Bronchoalveolar Lavage Fluid / Q000737:chemistry / Q000166:cytology; D002648:Child; D002675:Child, Preschool; D003371:Cough / Q000097:blood; D003550:Cystic Fibrosis / Q000097:blood* / Q000201:enzymology; D006801:Humans; D007223:Infant; D019272:Leukocyte Elastase / Q000097:blood*; D008171:Lung Diseases / Q000097:blood* / Q000201:enzymology; D009468:Neuromuscular Diseases / Q000097:blood; D009504:Neutrophils / Q000201:enzymology*; D037663:Pulmonary Surfactant-Associated Protein D / Q000097:blood*; D012016:Reference Values; D012697:Serine Endopeptidases / Q000378:metabolism*
chemical_list: D037663:Pulmonary Surfactant-Associated Protein D; D012697:Serine Endopeptidases; D019272:Leukocyte Elastase