Skip to main content
Log in

Purification and characterization of defensins from cystic fibrosis sputum

  • Published:
Inflammation Research Aims and scope Submit manuscript

Abstract.

Objective and Design: Cystic fibrosis (CF) sputum contains large numbers of neutrophils whose most abundant granule proteins are defensins. Within phagolysosomes, defensins kill microbes; however, extracellular defensins can be toxic to human cells. To begin to explore the possibility that defensins damage CF airways, this study examines the concentration and properties of defensins in CF sputum. ¶Materials: As a source of biological material in which to assay levels of defensins, purulent sputum was collected from persons with CF hospitalized with exacerbations of bronchitis. Purulent CF sputum was also a source of material for purification of defensins. ¶Methods: Defensin concentration in the cell-free component of CF sputum was measured by immunoassay. Defensins acid-extracted from sputum were purified by Sephacryl S-200 gel filtration chromatography and reverse phase-high pressure liquid chromatography (HPLC). Toxicity of CF defensins was tested by incubating the purified defensins with a line of CF tracheal cells cultured in serum-free medium. ¶Results: In 5 patients with CF, sputum defensin levels ranged from 300 to > 1600 <mu>g/ml, higher than levels previously reported in any human secretion or fluid and greatly exceeding concentrations toxic to mammalian cells in vitro. HPCL-purified CF sputum defensins were pure as judged by acid urea-PAGE and N-terminal sequencing, which revealed a mixture of defensins-1, -2 and -3 at ratios of ∼4 : 2 : 1. At > 10 <mu>g/ml the purified mixture was toxic for a line of CF tracheal cells cultured in serum-free medium, as judged by reductions in cell numbers and increased permeability to trypan blue. ¶Conclusions: This study suggests that defensins in CF sputum are intact and sufficiently abundant that they may damage airway epithelium.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

Author information

Authors and Affiliations

Authors

Additional information

Received 2 July 1996; returned for revision 13 August 1996 and 18 November 1996; accepted by G.W. Carter 13 December 1996

Rights and permissions

Reprints and permissions

About this article

Cite this article

Soong, L., Ganz, T., Ellison, A. et al. Purification and characterization of defensins from cystic fibrosis sputum. Inflamm. res. 46, 98–102 (1997). https://doi.org/10.1007/s000110050114

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1007/s000110050114

Navigation