Pulmonary surfactant protein A interacts with gel-like regions in monolayers of pulmonary surfactant lipid extract

Epifluorescence microscopy was used to investigate the interaction of pulmo nary surfactant protein A (SP-A) with spread monolayers of porcine surfacta nt lipid extract (PSLE) containing 1 mol % fluorescent probe (NBD-PC) sprea d on a saline subphase (145 mM NaCl, 5 mM Tris-HCl, pH 6.9) containing 0, 0 .13, or 0.16 mug/ml SP-A and 0, 1.64, or 5 mM CaCl2. In the absence of SP-A , no differences were noted in PSLE monolayers in the absence or presence o f Ca2+. Circular probe-excluded (dark) domains were observed against a fluo rescent background at low surface pressures (pi similar to5 mN/m) and the d omains grew in size with increasing pi. Above 25 mN/m, the domain size decr eased with increasing a. The amount of observable dark phase was maximal at 18% of the total film area at pi similar to 25 mN/m, then decreased to sim ilar to3% at pi similar to 40 mN/m. The addition of 0.16 mug/ml SP-A with 0 or 1.64 mM Ca2+ in the subphase caused an aggregation of dark domains into a loose network, and the total amount of dark phase was increased to simil ar to 25% between pi of 10-28 mN/m. Monolayer features in the presence of 5 mM Ca2+ and SP-A were not substantially different from those spread in the absence of SP-A, likely due to a self-association and aggregation of SP-A in the presence of higher concentrations of Ca2+. PSLE films were spread on a subphase containing 0.16 mug/ml SP-A with covalently bound Texas Red (TR -SP-A). In the absence of Ca2+, TR-SP-A associated with the reorganized dar k phase (as seen with the lipid probe). The presence of 5 mM Ca2+ resulted in an appearance of TR-SP-A in the fluid phase and of aggregates at the flu id/gel phase boundaries of the monolayers. This study suggests that SP-A as sociates with PSLE monolayers, particularly with condensed or solid phase l ipid, and results in some reorganization of rigid phase lipid in surfactant monolayers.