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S99 A Functional Variant of Elafin with Improved Anti-Inflammatory Activity
  1. Dm Small1,
  2. M Zani2,
  3. D Quinn1,
  4. S Weldon1,
  5. Df McAuley1,
  6. P McNally3,
  7. T Moreau2,
  8. Cc Taggart1
  1. 1Queen's University, Belfast, Northern Ireland
  2. 2Francois-Rabelais University, Tours, France
  3. 3Our Lady's Hospital for Sick Children, Dublin, Ireland

Abstract

Introduction and Objectives Elafin is a well-known serine protease inhibitor produced by epithelial and inflammatory cells with anti-inflammatory properties. Previous work has shown that unregulated protease activity can cause the proteolytic cleavage of elafin, therefore impairing the innate immune function of the protein. Consequently, the aim of this study was to generate a variant of elafin that would demonstrate increased protease resistance whilst retaining many of the beneficial characteristics of the parent molecule.

Methods Two elafin variants (GG-elafin and QQ-elafin) were recombinantly synthesised in a yeast-based expression system and subsequently tested for antiprotease, transglutaminase and protease susceptibility. In addition, the LPS neutralisation activity of the GG-variant was evaluated in in vitro based assays and an in vivo mouse model of LPS-induced acute lung inflammation.

Results GG- and QQ-elafin retained similar antiprotease and transglutaminase activity compared to wild-type elafin (WT-elafin). When incubated with diseased bronchoalveolar lavage fluid (BALF), the elafin variants displayed significantly enhanced resistance to degradation when compared to WT-elafin. Intriguingly, both variants, but particularly GG-elafin, demonstrated improved LPS neutralisation by inhibiting cytokine expression in monocytic cells. Moreover, the GG-elafin showed improved anti-inflammatory properties in a mouse model of LPS-induced acute lung inflammation with significantly decreased inflammatory cell counts, namely neutrophils (p = 0.0362). Furthermore, total BAL protein levels were significantly decreased (p = 0.0336) and a reduction in pro-inflammatory cytokine/chemokine levels was observed in mice treated with GG-elafin compared to those treated with WT-elafin.

Conclusions Site-specific mutants of elafin, particularly GG-elafin, showed increased functionality when compared to WT-elafin and may be of future therapeutic relevance in the treatment of lung diseases, particularly acute lung injury (ALI).

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