Endotoxin Exposure Increases LL-37 - but Not Calprotectin - in Healthy Human Airways

Abstract

Rationale: The antimicrobial peptides (AMPs) LL-37 and calprotectin are important players in the innate immunity of human airways. In patients with diseases characterized by bacterial colonization, the airway concentrations of these AMPs are increased. Less is known about their presence and release patterns in healthy humans. Our aim was to determine whether LL-37 and calprotectin are released after the activation of the innate immune response in the peripheral airways. Methods: Healthy volunteers underwent exposure to endotoxin and vehicle in contralateral segment bronchi. After 12 or 24 h, samples of bronchoalveolar lavage fluid (BALf) were collected bilaterally from exposed segments. Cell and AMP concentrations were assessed, as were the pro-form and active form of LL-37. Results: Both LL-37 and calprotectin were detected in cell-free BALf from both endotoxin- and vehicle-exposed segments. The concentrations of precursor and active LL-37 and neutrophils were significantly higher in endotoxin-exposed segments after 12 and 24 h, and the concentrations of LL-37 and neutrophils correlated positively. The concentrations of calprotectin were not markedly affected by exposure to endotoxin. Conclusions: Local endotoxin exposure elicits the release and activation of LL-37 but not calprotectin in healthy human peripheral airways, suggesting an inducible involvement of LL-37 in the local innate immune response.

Studies on citrullinated LL-37: detection in human airways, antibacterial effects and biophysical properties

“…This has been postulated in previous work, but not formally proven 15,24 . In order to detect citrullinated LL-37, we utilized cell-free BAL fluid obtained from from healthy human volunteers after intra-bronchial exposure to LPS 21,25 . We found that the BAL fluid from LPS-exposed lungs contained low, but clearly detectable, levels of citrullinated LL-37, which was demonstrated by using an in-house polyclonal serum raised against LL-37 Cit5 combined with validation by a sensitive targeted LC-MS/MS analysis.…”

“…Knowing that native LL-37 was present in human BAL fluid 21 , we determined the impact of citrullination on bacterial killing. As expected, synthetic native LL-37 (Table 1) Table 1.…”

“…The cell-free BAL fluid samples were obtained as part of a clinical study performed from 11 th of November 2003 to 3 rd of December 2008 21,25 . This sample collection constituted a part of a larger project studying the innate immune response in healthy human airways with and without prior intra-bronchial LPS exposure to LPS, from which other results have already been presented 21,25 . This clinical study was performed in accordance with the Helsinki declaration after approval by the ethics committee Regional Ethical Review Board in Gothenburg, Sweden (Dnr.…”

“…After either 12 or 24 h after the exposure to vehicle and LPS, respectively, BAL was performed (3 × 50 mL of PBS at 37 °C) in the respective lung. After additional processing as described elsewhere 21,25 , the cell-free BAL fluid samples were stored at −80 °C until further use. For the current study, we utilized BAL samples from LPS-exposed lungs only.…”

Cathelicidin – A Novel Potential Marker of Pediatric Inflammatory Bowel Disease

“…The relationship between cathelicidin and calprotectin has been previously studied in various models. 52–54 Smith et al found that although endotoxin exposure increases the concentration of cathelicidin in bronchoalveolar lavage fluid (BALf), it does not affect the concentration of calprotectin in BALf. 52 That may result from the fact that cathelicidin is mainly expressed in the secretory granules of neutrophils and more easily mobilized than calprotectin which is a cytoplasmic protein.…”