Granulocyte elastase‐mediated proteolysis of alpha₁‐antitrypsin in cystic fibrosis bronchopulmonary secretions

Abstract

Airway secretions of patients with cystic fibrosis (CF) contain large amounts of alpha 1-antitrypsin (alpha 1-AT), yet elastase activity is also often detectable, suggesting that airway alpha 1-AT may not be functional in some CF patients. It is unknown whether in CF sputum alpha 1-AT is inactivated by oxidants, neutrophil metalloproteinases, bacterial elastase, or neutrophil elastase. To investigate the mechanism(s) by which alpha 1-AT may be inactivated in CF airway secretions, sputum samples were obtained from nine patients during respiratory physiotherapy. alpha 1-AT was measured by radial immunodiffusion. Sputum-alpha 1-AT was purified by antibody affinity chromatography. Electrophoresis of alpha 1-AT from seven patients with acute infectious exacerbations revealed two distinct components: a minor band corresponding to an elastase/alpha 1-AT complex and a major band typical of proteolysed alpha 1-AT (Mr = 48 kD). Each patient had large amounts of sputum elastase activity. In contrast, two patients without free sputum elastase activity had intact sputum alpha 1-AT; however, alpha 1-AT was partially truncated by porcine pancreatic elastase suggesting that the alpha 1-AT may have been partially oxidized. Adding alpha 1-AT purified from normal serum to alpha 1-AT-depleted sputum containing elastase activity resulted in a small alpha 1-AT/elastase complex with most alpha 1-AT being truncated. The serine proteinase inhibitor phenylmethylsulfonyl fluoride but not the metalloproteinase inhibitor EDTA prevented alpha 1-AT proteolysis, thus granulocyte elastase can mediate alpha 1-AT degradation in CF. Apparently, the large granulocyte elastase burden in some acutely ill patients with cystic fibrosis can proteolytically inactivate alpha 1-AT.

Plasma biomarkers and cystic fibrosis lung disease

“…Although CF patients express normal to high systemic levels of plasma α1AT, they have a marked de ciency of functional α1AT in their airway secretions [37,38]. Much of this functional deciency in α1AT is related to the molar excess of HLE relative to α1AT in the CF airways [13,14,39,40].…”

Proteases and cystic fibrosis

“…NE sustains proteolytic inflammation by other mechanisms. NE degrades a1AT, decreasing the anti-protease capacity of the lung (Cantin et al, 1989). NE activates pro-MMP-9 and degrades its natural inhibitor, tissue inhibitor of metalloprotease-1 (TIMP-1), further activating MMP-9 in the CF airway.…”

The cell biology of leukocyte-mediated proteolysis

“…Oxidatively and proteolytically inactivated proteinase inhibitors have been detected in vivo in lung secretions from patients with cystic fibrosis [105,110], and in synovial fluid from patients with inflammatory arthritides [111][112][113][114]. Thus it is clear that, during infection and inflammation, oxidants and proteinases released from activated leukocytes and/or bacteria may act synergistically to create microenvironments in which extracellular proteolysis by leukocyte proteinases is facilitated via the inactivation of proteinase inhibitors.…”

“…In vitro studies have demonstrated that reactive oxygen species released by activated leukocytes can inactivate serpins (␣ 1 -proteinase inhibitor and SLPI) as well as ␣ 2 -macroglobulin [98][99][100][101]. Serpins are also susceptible to proteolytic inactivation by several classes of proteinases: (1) several MMPs (MMP-1, MMP-3, MMP-7, MMP-8, MMP-9, and MMP-12) can inactivate ␣ 1 -proteinase inhibitor, ␣ 1 -antichymotrypsin, and also ␣ 2 -macroglobulin [25,26,[102][103][104]; (2) the serine proteinase, HLE, can inactivate its cognate inhibitor (␣ 1 -proteinase inhibitor) [105] and PAI-1 [106]; (3) cathepsin B (a cysteine proteinase) can inactivate ␣ 1 -proteinase inhibitor [107]; and (4) several bacterial proteinases can inactivate ␣ 1 -proteinase inhibitor and SLPI [108]. It is also noteworthy that serine proteinases can proteolytically inactivate TIMPs [109].…”