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Physiological concentration of calcium inhibits elastase-induced cleavage of a functional recombinant fragment of surfactant protein D

Physiological concentration of calcium inhibits elastase-induced cleavage of a functional recombinant fragment of surfactant protein D
Physiological concentration of calcium inhibits elastase-induced cleavage of a functional recombinant fragment of surfactant protein D
Surfactant protein D (SP-D) plays an important role in lung host defence. SP-D levels have been shown to be depleted in cystic fibrosis (CF) patients. A recombinant fragment of the human SP-D (rfhSP-D) which consist of a hydrophobic neck and a CRD has been shown to be active in vivo and partially reverses the symptoms of the SP-D deficiency in the lungs when administered to SP-D knock-out mice. In this paper we studied the in vitro effect of different proteolytic enzymes commonly found in CF patients lungs, such as neutrophil elastase, cathepsin G and protease 3 as well as Pseudomonas elastase, on rfhSP-D. It was also shown that cleavage was inhibited by physiological concentration of calcium. When Western blot was compared with ELISA, we show that an anti-SP-D ELISA is a not a reliable assay of functional SP-D levels since non-functional fragments of SP-D are also detected. Thus, ELISA cannot be used as a reliable "diagnostic" tool for SP-D deficiency. Finally, we observe that SP-D is not cleaved in control patients but is degraded in about half the samples from cystic fibrosis patients, indicating that degradation of endogenous SP-D, by enzymes present in CF bronchioalveolar lavage fluid (BALF), may lead to deficiency of the protein as seen in CF and therefore rfhSP-D may be a useful future therapy.
bronchioalveolar lavage, cathepsin g, cystic fibrosis, neutrophile elastase, protease 3, pseudomonas elastase, surfactant protein d
0171-2985
72-79
Duvoix, Annelyse
f9113c62-9335-4c0b-b34b-294181995343
Mackay, Rose-Marie
19cf1b92-c65d-4baa-a165-ab630bf77ec3
Henderson, Neil
1a283a67-c5f7-48f5-8a7a-dbb023e03708
McGreal, Eamon
3367e432-3b4a-46c8-af8c-1aad0ab3917b
Postle, Anthony
0fa17988-b4a0-4cdc-819a-9ae15c5dad66
Reid, Kenneth
a148830a-ba86-4e98-a418-e1fb9e673c30
Clark, Howard
70550b6d-3bd7-47c6-8c02-4f43f37d5213
Duvoix, Annelyse
f9113c62-9335-4c0b-b34b-294181995343
Mackay, Rose-Marie
19cf1b92-c65d-4baa-a165-ab630bf77ec3
Henderson, Neil
1a283a67-c5f7-48f5-8a7a-dbb023e03708
McGreal, Eamon
3367e432-3b4a-46c8-af8c-1aad0ab3917b
Postle, Anthony
0fa17988-b4a0-4cdc-819a-9ae15c5dad66
Reid, Kenneth
a148830a-ba86-4e98-a418-e1fb9e673c30
Clark, Howard
70550b6d-3bd7-47c6-8c02-4f43f37d5213

Duvoix, Annelyse, Mackay, Rose-Marie, Henderson, Neil, McGreal, Eamon, Postle, Anthony, Reid, Kenneth and Clark, Howard (2010) Physiological concentration of calcium inhibits elastase-induced cleavage of a functional recombinant fragment of surfactant protein D. Immunobiology, 216 (1-2), 72-79. (doi:10.1016/j.imbio.2010.03.006). (PMID:20378199)

Record type: Article

Abstract

Surfactant protein D (SP-D) plays an important role in lung host defence. SP-D levels have been shown to be depleted in cystic fibrosis (CF) patients. A recombinant fragment of the human SP-D (rfhSP-D) which consist of a hydrophobic neck and a CRD has been shown to be active in vivo and partially reverses the symptoms of the SP-D deficiency in the lungs when administered to SP-D knock-out mice. In this paper we studied the in vitro effect of different proteolytic enzymes commonly found in CF patients lungs, such as neutrophil elastase, cathepsin G and protease 3 as well as Pseudomonas elastase, on rfhSP-D. It was also shown that cleavage was inhibited by physiological concentration of calcium. When Western blot was compared with ELISA, we show that an anti-SP-D ELISA is a not a reliable assay of functional SP-D levels since non-functional fragments of SP-D are also detected. Thus, ELISA cannot be used as a reliable "diagnostic" tool for SP-D deficiency. Finally, we observe that SP-D is not cleaved in control patients but is degraded in about half the samples from cystic fibrosis patients, indicating that degradation of endogenous SP-D, by enzymes present in CF bronchioalveolar lavage fluid (BALF), may lead to deficiency of the protein as seen in CF and therefore rfhSP-D may be a useful future therapy.

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More information

Published date: 15 March 2010
Keywords: bronchioalveolar lavage, cathepsin g, cystic fibrosis, neutrophile elastase, protease 3, pseudomonas elastase, surfactant protein d
Organisations: Faculty of Medicine, Medicine

Identifiers

Local EPrints ID: 158125
URI: http://eprints.soton.ac.uk/id/eprint/158125
ISSN: 0171-2985
PURE UUID: 95d9eda7-88d2-4ecc-aecb-fbbc6f889fdc
ORCID for Rose-Marie Mackay: ORCID iD orcid.org/0000-0002-9493-9654
ORCID for Anthony Postle: ORCID iD orcid.org/0000-0001-7361-0756

Catalogue record

Date deposited: 15 Jun 2010 13:28
Last modified: 20 Jul 2019 01:29

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