Biophysical analysis of gelatin and PLGA nanoparticle interactions with complex biomimetic lung surfactant models

Abstract

Biocompatible materials are increasingly used for pulmonary drug delivery, and it is essential to understand their potential impact on the respiratory system, notably their effect on lung surfactant, a monolayer of lipids and proteins, responsible for preventing alveolar collapse during breathing cycles. We have developed a complex mimic of lung surfactant composed of eight lipids mixed in ratios reported for native lung surfactant. A synthetic peptide based on surfactant protein B was added to better mimic the biological system. This model was used to evaluate the impact of biocompatible gelatin and poly(lactic-co-glycolic acid) nanoparticles. Surface pressure-area isotherms were used to assess lipid packing, film compressibility and stability, whereas the lateral organization was visualized by Brewster angle microscopy. Nanoparticles increased film fluidity and altered the monolayer collapse pressure. Bright protruding clusters formed in their presence indicate a significant impact on the lateral organization of the surfactant film. Altogether, this work indicates that biocompatible materials considered to be safe for drug delivery still need to be assessed for their potential detrimental impact before use in therapeutic applications.

Fig. 1. Pressure–area and compression modulus isotherms of individual lipid systems on an aqueous subphase. (A) Pressure–area isotherm and (B) compression modulus of DPPC, DPPG, and in the presence of SP-B_1–25 at a 10% weight ratio, respectively. All isotherms collected are an average of the replicates (n ≥ 3).

Fig. 2. Pressure–area isotherms and compression moduli of phosphatidylcholine (PC) and phosphatidylglycerol (PG) containing systems on an aqueous subphase. (A) The pressure–area isotherm of PC system, (B) compression moduli of PC system, (C) the pressure–area isotherm of PG system, and (D) compression moduli of PG system. All systems show results for lipid controls (black), presence of lipid system and SP-B_1–25 at 10% wt ratio (blue), presence of PLGA at 10 : 1 wt ratio and SP-B_1–25 at 10% wt ratio (red), presence of PLGA at 1 : 1 wt ratio and SP-B_1–25 at 10% wt ratio (green), and presence of gelatin NPs at 10 : 1 ratio and SP-B_1–25 at 10% wt ratio (yellow). The collapse pressures for the isotherms are highlighted (purple box), while dip corresponding to coexistence phase (arrow) and peak modulus (purple oval) are indicated in the compression moduli. All isotherms collected are average of various replicates (n ≥ 3).

Fig. 3. Pressure–area isotherms and compression modulus of 8 lipid system on an aqueous subphase. (A) The pressure–area isotherm of 8 lipid system, (B) compression moduli of 8 lipid system. All systems show results for lipid controls (black), presence of lipid system and SP-B_1–25 at 10% wt ratio (blue), presence of PLGA at 10 : 1 wt ratio and SP-B_1–25 at 10% wt ratio (red), presence of PLGA at 1 : 1 wt ratio and SP-B_1–25 at 10% wt ratio (green), and presence of gelatin NPs at 10 : 1 ratio and SP-B_1–25 at 10% wt ratio (yellow). The collapse pressures for the isotherms are highlighted (purple box). All isotherms collected are average of various replicates (n ≥ 3).

Fig. 4. BAM images of the lateral organization of PC systems on an aqueous subphase at 15 and 30 mN m⁻¹ surface pressures. Panel (1) control PC system, panel (2) PC system + 10% SP-B_1–25, panel (3) PC system + 10% SP-B_1–25 + PLGA 10 : 1 wt ratio, panel (4) PC system + 10% SP-B_1–25 + PLGA 1 : 1 wt ratio, and panel (5) PC system + 10% SP-B_1–25 + gelatin 10 : 1 wt ratio. Scale bar corresponds to 50 μm. Each image is a representation of at least 3 images.

Fig. 5. BAM images of the lateral organization of PG systems on an aqueous subphase at 15 and 30 mN m⁻¹ surface pressures. Panel (1) control PG system, panel (2) PG system + 10% SP-B_1–25, panel (3) PG system + 10% SP-B_1–25 + PLGA 10 : 1 wt ratio, panel (4) PG system + 10% SP-B_1–25 + PLGA 1 : 1 wt ratio, and panel (5) PG system + 10% SP-B_1–25 + gelatin 10 : 1 wt ratio. Scale bar corresponds to 50 μm. Each image is a representation of at least 3 images.

Fig. 6. Quantitative analysis of area percentage (bar) covered by lipid domains and domain frequency (line) on PC and PG systems using representative BAM images at 30 mN m⁻¹. This analysis was done using ImageJ software.

Fig. 7. BAM images of the lateral organization of 8 lipid systems on an aqueous subphase. Panel (1) control 8 lipid system, panel (2) 8 lipid system in the presence of 10% by weight SP-B_1–25, panel (3) 8 lipid system in the presence of 10% by weight SP-B_1–25 and gelatin NPs at a 10 : 1 lipid to NP weight ratio. Panel (4) 8 lipid system + 10% SP-B_1–25 + PLGA 1 : 1 wt ratio, and panel (5) 8 lipid system + 10% SP-B_1–25 + gelatin 10 : 1 wt ratio. Scale bar corresponds to 50 μm. Each image is a representative sample (n ≥ 3).

Fig. 8. Quantitative analysis of area percentage (bar) covered by lipid domains and domain frequency (line) on 8 lipid system using representative BAM images at 30 mN m⁻¹. This analysis was done using ImageJ software.