CCSP counterbalances airway epithelial-driven neutrophilic chemotaxis

Abstract

Club cell secretory protein (CCSP) knockout mice exhibit increased airway neutrophilia, as found in chronic obstructive pulmonary disease (COPD). We therefore investigated whether treating COPD airway epithelia with recombinant human CCSP (rhCCSP) could dampen exaggerated airway neutrophilia. Control, smoker and COPD air–liquid interface (ALI) cultures exposed to cigarette smoke extract (CSE) were treated with and without rhCCSP. The chemotactic properties of the supernatants were assessed using Dunn chambers. Neutrophil chemotaxis along recombinant human interleukin 8 (rhIL8) gradients (with and without rhCCSP) was also determined. rhCCSP–rhIL8 interactions were tested through co-immunoprecipitation, Biacore surface plasmon resonance (SPR) and in silico modelling. The relationship between CCSP/IL8 concentration ratios in the supernatant of induced sputum from COPD patients versus neutrophilic airway infiltration assessed in lung biopsies was assessed. Increased neutrophilic chemotactic activity of CSE-treated ALI cultures followed IL8 concentrations and returned to normal when supplemented with rhCCSP. rhIL8-induced chemotaxis of neutrophils was reduced by rhCCSP. rhCCSP and rhIL8 co-immunoprecipitated. SPR confirmed this in vitro interaction (equilibrium dissociation constant=8 µM). In silico modelling indicated that this interaction was highly likely. CCSP/IL8 ratios in induced sputum correlated well with the level of small airway neutrophilic infiltration (r2=0.746, p<0.001). CCSP is a biologically relevant counter-balancer of neutrophil chemotactic activity. These different approaches used in this study suggest that, among the possible mechanisms involved, CCSP may directly neutralise IL8.