Thymosin beta4 sequesters actin in cystic fibrosis sputum and decreases sputum cohesivity in vitro

Filamentous actin (F-actin) forms polymers that contribute to the abnormal biophysical properties of sputum. Thymosin beta4 (Tbeta4) is the major monomeric actin-sequestering peptide in cells and can depolymerize F-actin. Tbeta4 could potentially decrease sputum viscoelasticity and adhesivity and improve sputum clearance. Sputum was collected during pulmonary function testing from 17 subjects during a cystic fibrosis (CF) center visit. Sputum rheology, cough and ciliary transportability, and interfacial tension were measured before and after the addition of dornase alfa at 30 microg/mL; Tbeta4 at 0.3, 3, 30, and 150 microg/mL; and Tbeta4 with dornase alfa at 1.5 microg/mL each. Sputum was separately incubated with Tbeta4 at 30 microg/mL for 0, 10, 20, or 60 min. There was a direct relationship between actin filament length and sputum cohesivity. There was a dose-dependent threshold decrease in cohesivity with Tbeta4 and a time-dependent decrease in cohesivity over 60 min at 30 microg/mL. With the combination of dornase alfa and Tbeta4 at 1.5 microg/mL each, there was a 70% decrease in G*s and a 65% decrease in G' at 1 rad/s (p = 0.013). There was a 44% increase in cough transportability of sputum in vitro (p = 0.037) and a 71% increase in mucociliary transportability of sputum in vitro (p = 0.013) relative to control with the combination of dornase alfa and Tbeta4, but no significant change with dornase alfa or Tbeta4 alone at any concentration. Actin polymer filament length is correlated with sputum cohesivity. Tbeta4 depolymerizes CF sputum actin in both a dose-dependent and a time-dependent manner. An apparent synergy of Tbeta4 on actin and dornase on DNA may be explained by the combined effect of actin depolymerization and DNA filament severing or by virtue of actin depolymerization increasing the effectiveness of dornase alfa