Shape matters—the interaction of gold nanoparticles with model lung surfactant monolayers

Abstract

The lung surfactant monolayer (LSM) forms the main biological barrier for any inhaled particles to enter our bloodstream, including gold nanoparticles (AuNPs) present as air pollutants and under investigation for use in biomedical applications. Understanding the interaction of AuNPs with lung surfactant can assist in understanding how AuNPs enter our lungs. In this study, we use coarse-grained molecular dynamics simulations to investigate the effect of four different shape D AuNPs (spherical, box, icosahedron and rod) on the structure and dynamics of a model LSM, with a particular focus on differences resulting from the shape of the AuNP. Monolayer-AuNP systems were simulated in two different states: the compressed state and the expanded state, representing inhalation and exhalation conditions, respectively. Our results indicate that the compressed state is more affected by the presence of the AuNPs than the expanded state. Our results show that in the compressed state, the AuNPs prevent the monolayer from reaching the close to zero surface tension required for normal exhalation. In the compressed state, all four nanoparticles (NPs) reduce the lipid order parameters and cause a thinning of the monolayer where the particles drag surfactant molecules into the water phase. Comparing the different properties shows no trend concerning which shape has the biggest effect on the monolayer, as shape-dependent effects vary among the different properties. Insights from this study might assist future work of how AuNP shapes affect the LSM during inhalation or exhalation conditions.

1. Introduction

Any particles less than approximately 100 nm in size that enter our respiratory system are likely to reach the alveoli [1,2], the tiny air sacs in the bronchioles where gas exchange occurs during the breathing process. The main air/water interface in the alveoli is formed by lung surfactant, also known as pulmonary surfactant, which is secreted from alveolus type II cells and forms a thin but elastic monolayer [3] with a multilayer film associated with it. This lung surfactant monolayer (LSM) forms the main biological barrier for any inhaled particles to enter our bloodstream [4,5]. Examples of such particles include drugs to treat lung disease (e.g. steroids to treat asthma), nano-sized pollutants found in our environment (e.g. diesel particles in exhaust gas fumes) and nanoparticles (NPs) under investigation in various biomedical applications.

The main function of the LSM is to reduce the surface tension at the air/water interface and thus reduce the mechanical work during the breathing process [6–8]. An intact monolayer is critical for normal breathing, while abnormalities of the LSM are associated with lung diseases such as respiratory distress syndrome [9]. During normal breathing, the LSM is repeatedly expanded and compressed. The surface tension oscillates between 0 mN/m at the end of exhalation (maximum compression) and an equilibrium value of approximately 20–25 mN m⁻¹ during inhalation (expansion) [8]. As a result, the LSM undergoes repeated phase transitions between two different states; the liquid condensed (LC) phase and an intermediate phase where the LC and liquid expanded phases (LE) coexist (LC + LE) [10]. These two phases show different structural and dynamic properties, which affect the interactions with any inhaled particles in contact with the LSM. Understanding how particles interact with the LSM during the different stages of breathing contributes to various applications ranging from drug development to minimizing the risk of airborne particles on our health. During the phase change, the surfactant is transferred between the interfacial LSM and associated multilayer or bilayer reservoirs through continuous re-adsorption and reformation of surfactant [11]. Through this process, the LSM and the bilayers are intimately connected and particles interacting with the LSM might also affect the formation of bilayer, adsorption and recycling of surfactant. Nevertheless, the LSM is the first point of contact for inhaled particles.

NPs are increasingly found in our environment with both beneficial and detrimental effects. Potential biomedical applications include the use of NPs to improve the delivery of drugs or probes into cells [12–15], including diagnosing and treating lung diseases such as chronic obstructive pulmonary disease, asthma or cystic fibrosis [16–18]. There are clinical trials of gold nanoparticles (AuNPs) for the photothermal ablation of prostate tumours [19], treatment of other cancers [20] and cancer diagnostics [21]. Surface modification of AuNPs with different functional groups including ligands, surfactant and others stabilizing agents increase the potential of AuNPs to be used in various therapeutic applications [22–24]. The surface modification alters the surface properties of AuNPs and thus, their interactions with the LSM may be different compared to their bare state. Bare AuNPs can also act as harmful pollutants. Several studies reported that gold miners and goldsmiths suffer from various occupational respiratory diseases associated with excessive inhalation of gold dust particles and other NPs [25–27]. In an in vitro study, Bakshi et al. [28] used AuNPs as a model pollutant and reported that the inhalation of AuNPs may hinder the LSM function. Another recent in vivo study postulated that engineered NPs like AuNPs are similar to environmental pollutant NPs [29]. The findings regarding the effects of bare AuNPs on the LSM may help in understanding the effects of other NP pollutants associated with lung diseases. The National Institute of Standards and Technology has suggested the use of AuNPs as a model system for nanotoxicological research after considering multiple lines of evidence on the toxicity of gold particles (see https://ntp.niehs.nih.gov/ntp/noms/final_resconcept/nanoscale_gold_508.pdf (published May 2008; accessed 10 January 2019)). AuNPs are also found in consumer products such as cosmetics [30–32], electronics [33] and food industries [34,35], which increase our daily exposure to potentially harmful particles that can enter the body via inhalation or other means such as ingestion or skin absorption.

While the detailed pathophysiology of different AuNP -related lung diseases is not clear, it is likely related to the particles interfering with the normal function of the LSM. AuNPs can interfere with the ability of the LSM to reduce surface tension or its mechanical properties required for monolayer compressibility and re-expansibility [28,36,37]. Independent of the application, the interaction of NPs with lipid monolayers or bilayers depends on the size, shape and physico-chemical properties of the particle [2,38–40]. For AuNPs, it is known that particle shape affects their diffusion and translocation properties in biological environments, including membranes [41]. For example, in vitro studies reported that mammalian cells preferentially internalize disc-shaped NPs over rod-shaped NPs and that increasing the NP size accelerates intracellular uptake [39]. In the case of the LSM, several studies have shown that the size and shape affect how the NPs translocate across the monolayer or whether they cause monolayer disruption [42–44]. In vitro studies also reported that AuNPs as pollutants could disrupt normal lung function by preventing the LSM from lowering the surface tension values at the air–liquid interface [28,45]. Results from various in silico studies suggest that these effects are directly related to the interaction of the NPs with the LSM [36,46]. For example, a recent in silico study [36] found that AuNPs impede the ability of the LSM to regulate surface tension and create pores in the monolayer when a sufficiently high concentration of NPs is reached. The shape- and size-dependent translocation of NPs can also depend on the state of the LSM itself. A simulation study [42] reporting on the interaction of rod-, barrel- and disc-shaped NPs with a dipalmitoyl-phosphatidylcholine (DPPC) monolayer found that shape affects the ability of a NP to translocate across the monolayer in the contracted state (during exhalation) but not in the expanded form (during inhalation). Also, compared to the barrel- or disc-shaped NPs, rod-like NPs more easily translocate across the monolayer with less disruption to lipid packing [42]. Despite various studies, the detailed mechanism of how the size and shape of AuNPs affect their interactions with the LMS is still poorly understood.

In the current study, we use coarse-grained (CG) molecular dynamics (MD) simulations to investigate the effect of four different shapes (spherical, box, icosahedron and rod) of bare AuNPs on the structure and function of the LSM. To mimic the ratio of the most dominant lipids and protein in the LSM, the monolayer is composed of the following lipids: saturated, neutral (zwitterionic) phospholipid DPPC; unsaturated, negatively charged phospholipid palmitoyl-oleoyl-phosphatidylglycerol (POPG); the sterol lipid cholesterol (CHOL) and the membrane-embedded component of the surfactant protein B (SP-B_1-25). All systems were simulated at surface tensions 0 and 23 mN m⁻¹ to mimic inhalation and exhalation conditions.

2. Methods

2.1. Set-up of simulation systems

For all simulations, the LSM was composed of DPPC : POPG : CHOL : SP-B_1-25 at a ratio of approximately 70 : 30 : 10 : 1 mol %. To construct the simulation system, a monolayer consisting of 666 DPPC, 279 POPG, 90 CHOL and 9 surfactant protein fragments SP-B_1-25 was prepared using the python script insane.py [47]. The structure for the protein fragment SP-B_1-25 was prepared using the coordinates obtained from the protein data bank (PDB ID: 1DFW), which was converted to a CG representation using the python script martinize.py [48]. The nine copies for the proteins were embedded into the monolayer to obtain the final lipid–protein monolayers. The SP-B segment (SP-B_1-25) was added to the lipid monolayers to account for the effect of hydrophobic protein components in the LSM on its the structure and compressibility. The full-length SP-B (79 residues) is a covalently linked homodimer, and the amino-terminal peptide SP-B_1-25 is able to replicate at least some of the interfacial properties of full-length SP-B [49].

To mimic the air–liquid interface inside the lung alveolus, an approximately 21 nm thick water layer was added to separate the two monolayers (figure 1a). Na⁺ and Cl⁻ ions were added to neutralize the system and reach a concentration of 150 mM. The system was placed in the centre of a periodic box (25 × 25 × 60 nm³) with a vacuum below and above the monolayers. The height of the two monolayers enclosed by the water layer was approximately 26 nm, which left a nearly 17 nm vacuum to the hydrophobic side of each monolayer in the box.

Figure 1.

Simulation systems for LSMs interacting with four-different shaped AuNPs. (a) Initial simulation system containing lipids and surfactant proteins. The LSM is composed of DPPC (cyan), POPG (green), CHOL (red) and surfactant protein segments SP-B_1-25 (orange). (b) The four different AuNP shapes investigated in this study. (c) A rod-shaped AuNP at the vacuum–lipid interface of an LSM with APL 0.47 nm² after 3 µs (water has been removed for clarity). An example of an initial configuration is shown in the electronic supplementary material, figure S2.

The system was energy minimized using the steepest descent algorithm. Before introducing AuNPs to the system, it was equilibrated in an NPγT ensemble (constant number of particles, surface tension and temperature) at two surface tensions 0 and 23 mN m⁻¹ to mimic the inhalation and exhalation conditions. Previous studies reported that at high surface tension, the MARTINI model could not reproduce all intrinsic properties of the LSM. However, the model has successfully explored these properties at low surface tension [50,51] as well as at surface tension less of than 30 mN/m that represent the monolayer phase coexistence (LC + LE) [10]. At each surface tension (0 and 23 mN m⁻¹), the system was equilibrated using the area per lipid (APL) to assess equilibration (electronic supplementary material, figure S1). The systems equilibrated to APL values of 0.471 ± 001 nm² and 0.538 ± 0.001 nm² at surface tensions 0 and 23 mN m⁻¹, respectively.

The four shapes of AuNPs used in this study are spherical, icosahedron, box and rod (figure 1b). Particle sizes were chosen such that the four shapes have a similar surface area (table 1). AuNPs were added to the equilibrated system by placing two of the relevant particles (one for each vacuum side) within approximately 0.5–1 nm to the lipid tails (figure 1c; electronic supplementary material, figure S2). Each NP system was simulated at surface tensions of 0 and 23 mN m⁻¹, respectively (table 2).

Table 1.

Dimensions for the four shapes of AuNPs. a = side lengths, r = radius.

NP shapes	dimensions (nm)	surface areas (nm2)
spherical	r = 2.5	78.5
icosahedron	a = 3	77.9
box	a = 3.5	73.5
rod (capsule shape)	a = 5; r = 1.5	75.4

Table 2.

APL and surface tension from simulations of an LSM composed of DPPC : POPG : CHOL : SP-B_1-25 (70 : 30 : 10 : 1) in the absence and presence of AuNPs with different shapes. Data are averaged over the last 1 µs of two independent 3 µs simulations. Uncertainties represent standard deviations across the combined dataset.

system	APL (nm2)		surface tension (mN m−1)
	constant	variable	constant	variable
no AuNP		0.47 ± 0.001	0
		0.54 ± 0.001	23
spherical	0.47			16.12 ± 1.29
	0.54			24.49 ± 2.74
icosahedron	0.47			21.15 ± 0.14
	0.54			28.88 ± 0.17
box	0.47			18.56 ± 1.82
	0.54			28.67 ± 0.09
rod	0.47			14.59 ± 1.20
	0.54			27.10 ± 1.09

2.2. Corse-grained molecular dynamics simulations

All simulations were carried out using GROMACS v. 5.1.4 [52] with the MARTINI CG force field [53] to describe the phospholipids, CHOL, peptides (SP-B_1-25), water and ions. The AuNPs' non-bonded parameters for the Au bead type (C5) were adopted from Song et al. [54] and Lin et al. [22]. The details of this AuNP model are described in our previous paper [36]. Briefly, irrespective of the MARTINI conventional 4 : 1 mapping, a 1 : 1 mapping was used to map the Au atoms to CG gold beads. Au CG beads were connected using harmonic bonds [55,56].

Lennard–Jones (LJ) potentials were used to describe the non-bonded interactions with a cutoff distance of 1.2 nm and shifted from 0.9 nm to the cutoff distance. The electrostatic potential at 0.9 nm was used and shifted from 0 to that cutoff. Both LJ and Coulomb potentials were shifted to zero within their respective cutoffs. Long-range electrostatic interactions were corrected using a relative dielectric constant of 15. The system components (phospholipids and CHOL, water with ions and proteins) were coupled independently using a velocity rescale thermostat at 310 K, with a time constant of 1 ps. The Berendsen pressure barostat [57] was applied for constant surface tension at 0 and 23 mN m⁻¹, with compressibility 4.5 × 10⁻⁵ and 0 bar⁻¹ along the xy plane and z-axis (no compressibility in z-direction for constant box height), respectively. Periodic boundary conditions were applied in all directions of the simulation box, and the neighbour list was updated every 10 steps for non-bonded interactions. The simulations were carried out with a 20 fs time step to complete 500 ns equilibration followed by a 3 µs production run. All simulations of the LSM in the presence of AuNPs (table 2) were simulated in an NVT (constant particle number, volume and temperature) ensemble. The temperature was kept close to the reference temperature 310 K with a velocity rescale thermostat [58] with tau-t 1.0 ps. All production simulations were carried out in duplicate using different starting velocities.

Visual molecular dynamics (VMD) [59] was used to visualize the simulation trajectories and render snapshots for images. Two repeats were performed for each system, and unless otherwise stated, analyses were conducted using all frames from the last 1 µs of each simulation. The results from the two independent runs were averaged. Radial distribution functions (RDFs) were calculated with a cutoff distance of 1.2 nm. The GROMACS tool ‘gmx clustsize’ [52] was used to analyse the cluster size of surfactant peptides over the entire 3 µs of the production simulation, using a cutoff of 1.2 nm. The diffusion coefficients of surfactant phospholipids, peptide and CHOL were calculated from the mean square displacement (MSD) slope over the lipid positions $r(t)$ and $r(t_0)$ at time t and $t_0$, respectively. The following equation is used to calculate the MSD of surfactant constituents.

$$MSD(t)=\mathrm{\Delta }{r}_i(t)=⟨{[r_i(t+t_0)-r_i(t_0)]}^2⟩\hspace{0.17em}\hspace{0.17em},$$

where the angle brackets designate a time-average over t and overall molecules. The two dimensional (d = 2) or lateral diffusion coefficient D is defined by

$$D=\underset{t\to \mathrm{\infty }}{lim}\frac{1}{2dt}MSD(t).$$

3. Results and discussion

3.1. Effect of different-shaped nanoparticles on surface tension

The system without any AuNPs represents the reference system. During exhalation, the LSM compresses and the monolayer reaches approximately 0 mN m⁻¹ surface tension. This compression is reflected in a low APL of 0.470 ± 0.001 nm² (table 2). By contrast, the monolayer expands during inhalation and surface tension reaches approximately 20–25 mN m⁻¹. As a result, the monolayer has a higher APL of 0.538 ± 0.001 nm² in simulations with a surface tension of 23 mN m⁻¹. These values are in excellent agreement with previous simulation studies of LSMs with comparable composition [37,60].

To understand the underlying effect of different-shaped AuNPs on the structure and dynamics of the LSM, simulations of the AuNP–LSM system were carried out at fixed APL representing the compressed and expanded states. From these simulations, the effective surface tension was calculated (table 2). Previous studies [28,45] suggested that NPs impede the LSM's ability to lower the surface tension at the air–liquid interface. Comparison of the surface tension from the LSM in the compressed state (APL = 0.47 nm², exhalation) shows that for all AuNPs, the surface tension is greater than 14 mN m⁻¹, which is significantly higher than the expected surface tension close to 0 mN m⁻¹. This suggests that all four types of AuNPs prevent the LSM from reaching the low surface tension values required for normal exhalation. The four shapes affect the surface tension differently. In order of increasing surface tension: rod < spherical < box < icosahedron. In the expanded state (APL = 0.54 nm², inhalation), the effect of the NPs on the surface tension is much less pronounced. The calculated surface tensions range from approximately 24 to 29 mN m⁻¹ and within the levels of uncertainties, and this is only slightly higher than the expected range of 23–25 mN m⁻¹. There is no statistically significant difference between the four NP shapes, indicating that it is the presence of the particle rather than its specific shape that causes the small increase in surface tension.

Our results are consistent with previous wet-laboratory experiments with lung surfactant and AuNPs [28,45]. For example, Bakshi et al. [28] reported that spherical AuNPs impede the ability of a surfactant to reduce surface tension during monolayer compression. The semisynthetic lung surfactant used was composed of DPPC : POPG : SP-B (70 : 30 : 1) and is thus comparable to the model LSM used in our simulations. Similarly, Zhang et al. [45] showed that AuNPs hinder phase transition and reduce the compressibility of a monolayer composed of DPPC.

3.2. Effect of different-shaped nanoparticles on the phospholipids order parameter

The lipid order parameter is a commonly used parameter to characterize monolayer phases. An increase in the order parameter is indicative of a monolayer phase transition from disordered to ordered. Monolayer compression induces a transition from gas (G) to LE to LC phases, which is accompanied by a reduction of APL. Consequently, the lipid order increases. By contrast, during monolayer expansion, APL increases, which results in the lipid order decreasing. Figure 2 shows the sn-1 and sn-2 order parameters for DPPC and POPG lipids from simulations in the absence and presence of AuNPs in the compressed and expanded states. Comparison of the order parameters for DPPC and POPG shows that for all systems, both the sn-1 and sn-2 tails are more ordered for DPPC than POPG (figure 2a,c). This increased order is more pronounced in the sn-1 tail compared to the sn-2 tail. These results are consistent with the increased order for saturated lipid tails in DPPC, compared to the unsaturated bond in the sn-1 tail of POPG. Comparing order parameters for the different NPs shows that all four shapes reduce the order parameters in both chains sn-1 and sn-2 tails. However, in the compressed state monolayers, rod-shaped AuNP has the least impact on the ordering of the lipid tails (figure 2a,b). A stronger effect is seen for the other shapes (spherical, box and icosahedron).

Figure 2.

Effect of AuNPs of various shapes on the order parameters (S_z) of phospholipids from simulations of the model LSM in the (a,b) compressed state (APL = 0.47 nm²) and (c,d) expanded state (APL = 0.54 nm²). The order parameter, S_z for the sn-1 (a,c) and sn-2 (b,d) chains of DPPC (solid lines) and POPG (dashed lines). The order parameters for the different-shaped NPs are shown in cyan for spherical, orange for rod, green for box and red for icosahedron. Black colour represents the order parameter values of phospholipid tail beads in the monolayer. Error bars in each group/bead have been calculated using the standard deviation across the duplicate runs.

Compared to the compressed state, in the expanded state, the presence of the NPs has a smaller effect on the order of the phospholipids (figure 2c,d). A reduction in the order parameter values can be observed for the sn-1 chain (figure 2c), whereas in the sn-2 chain, there is no significant effect on the order parameters due to the presence of the NPs (figure 2d). The obtained results overall support our previous studies [36], where we showed that the presence of spherical AuNPs (diameter = 3 nm) decreases the order parameters in a concentration-dependent manner, both in the compressed and expanded states.

Overall, all four shapes of AuNPs decrease the lipid order in the monolayer in both expanded and compressed states, with a more pronounced effect on the latter. The results are consistent with surface tension data (table 2) and indicate that the AuNPs affect the monolayer's compressibility.

3.3. The effect of nanoparticles on the packing of lipids in the lung surfactant monolayer

Electronic supplementary material, figure S3 shows the density profiles for the head groups and tails of the phospholipids from simulations of the LSM in the presence of the four AuNPs both in the compressed (electronic supplementary material, figure S3a, c, e, g) and expanded states (electronic supplementary material, figure S3b, d, f, h). The density profiles for water, and the phospholipid head groups and tails, from the LSM in the absence of the AuNP are shown as a reference. In addition, representative snapshots from the simulations of the rod-shaped NP for the compressed-state (top) and expanded-state LSM illustrate the position of the particle in the system relative to the air–lipid interface.

In the absence of AuNPs, surfactant molecules are expected to be packed more densely in the compressed state monolayer than in the expanded state. This is reflected in a higher peak in the density profiles of phospholipids' head and tail groups (electronic supplementary material, figure S3a, c, e, g). Consistent with that, the reduced packing of lipids in the expanded state is reflected in a shift of densities for the lipid tails and heads towards the centre of the system (the water phase; electronic supplementary material, figure S3b, d, f, h). As a result, the water layer thickness between the monolayers is reduced in the expanded state. These changes in the density profiles are consistent with the differences in APL and order parameters between the two states.

When AuNPs are added to the vacuum, the particles adsorb to the phospholipid tails during simulations. To assess the effect on the lipid packing, we compared the density profiles of all four AuNPs in the compressed and expanded states to the reference system. In the compressed state, the presence of the AuNPs causes a slight drop in the peak height for the densities of the phospholipid head groups. The embedded AuNPs also drag some surfactant molecules into the water phase, which is also indicated by the densities of the phospholipids inside the water phase (electronic supplementary material, figure S3). This effect is absent in the simulations without NPs or in the expanded monolayer. In the expanded state, the presence of the AuNPs slightly increases the packing of the heads and tails in the phospholipids, which is indicated by the shift of the densities towards the centre. In both states, the density profiles suggest that there is no difference between the different particle shapes.

3.4. Effect of shape on the interactions of gold nanoparticles and surfactant molecules

The RDF was used to estimate the proximity of AuNPs to the surfactant molecules. The RDFs for the spherical AuNP with the different surfactant lipids (DPPC, POPG and CHOL) and the surfactant peptide SP-B_1-25 for the monolayer at APL = 0.47 nm² are shown in figure 3a. The corresponding plots for the other three shapes are shown in the electronic supplementary material, figure S4. Despite CHOL being present at lower concentrations than DPPC and POPG, the RDF shows that CHOL is much more likely to be found near the AuNP surface than other surfactant molecules (figure 3a; electronic supplementary material, figure S4). A visualization of the trajectories (electronic supplementary material, figure S5) shows an accumulation of CHOL molecules on the NP surface.

Figure 3.

RDF of surfactant molecules to AuNPs in the monolayer at APL = 0.47 nm² is shown. (a) The RDF of the surfactant molecules to spherical AuNPs, and (b) the RDF of CHOL molecules to the four different-shaped AuNPs have been presented. The RDF values of surfactant molecules are calculated over the last 1 µs of simulation and from the centre of mass of the molecules to the centre of mass of each shape AuNPs in the monolayer.

The RDF of CHOL to four different-shaped AuNPs for the monolayer at APL = 0.47 nm² was calculated (figure 3b) to assess the effect of NP shape on this preference for hydrophobic molecules over other phospholipids. The data suggest that CHOL has the highest preference for spherical AuNP surface, as indicated by the high densities of CHOL molecules close to the NPs compared to the other NPs. The fact that the spherical shape lacks edges may facilitate the adsorption of CHOL molecules on its surface. Consistent with this, the rod shape shows closer contact to CHOL than the box- and icosahedral-shaped AuNPs, both of which have edges.

A recent in silico study [61] on model pulmonary surfactant monolayers reported that CHOL increases the order of the hydrophobic tails of lipids and thus decreases the fluidity of the monolayer. The study also reported that hydrophobic carbon NPs preferentially interact with the hydrophobic components of the monolayer. Based on this, we would expect AuNPs to preferentially interact with the lipid tails of the phospholipids (DPPC and POPG). The RDFs comparing the densities for the lipid heads and tails around the AuNPs confirmed this (electronic supplementary material, figure S6).

3.5. Clustering of cholesterol and peptides

To elucidate the potential hydrophobic–hydrophobic interaction between surfactant molecules, we carried out individual cluster analysis of SP-B_1-25 and CHOL in the presence and absence of different-shaped AuNPs in the surfactant monolayer. The same cutoff distance of 1.2 nm was used in all cluster analyses.

Peptide clustering was measured by calculating the number of clusters formed by the nine surfactant peptides SP-B_1-25 in the system and the number of peptides in the largest cluster. Both analyses were carried out for the expanded state (figure 4) and the compressed state (electronic supplementary material, figure S7). In the expanded state and in the absence of AuNPs, the peptides do not show any significant clustering tendencies (figure 4a). All nine peptides are separated by a distance larger than the cutoff of 1.2 nm (i.e. as indicated by the nine individual clusters in the monolayer). In the presence of the AuNPs, however, a tendency for clustering is observed. The extent of the clustering depends on the shape. In the presence of rod and spherical AuNPs, the peptides aggregate into clusters containing up to five peptides per cluster. This aggregation is much less pronounced in the presence of box- and icosahedron-shaped AuNPs. By contrast, in the compressed state monolayer (electronic supplementary material, figure S7), even in the absence of AuNPs in the monolayer, the peptides aggregate into clusters containing up to five peptides. Interestingly, this clustering behaviour is not affected by the presence of any of the AuNPs.

Figure 4.

Clustering of surfactant peptide B (SP-B_1-25) from simulations of the surfactant monolayer in expanded state (APL = 0.54 nm²) with four different AuNPs. On the right are two snapshots showing SP-B_1-25 clustering in simulations in the presence of box (top) and rod (bottom) shaped AuNPs.

Overall, the clustering analysis suggests that the AuNPs do not affect the peptide clustering in the compressed state. Still, in the expanded state, the spherical and rod shapes induced clustering that is not seen in the presence of icosahedron and box AuNPs.

In two independent simulations, it has been noticed that the presence of different-shaped AuNPs in both monolayers (compressed and expanded states) accelerates the CHOL cluster formation (electronic supplementary material, figure S8). Box-shaped AuNP shows strong effects on CHOL cluster numbers and the number of CHOL in the largest clusters, which have not been noticed for the peptide clustering process in the expanded monolayer. One possible reason might be charged residues in the peptide molecules, whereas CHOL has no charges.

3.6. The shape effect of gold nanoparticles on the surfactant molecules' diffusion in the lung surfactant monolayer

The two-dimensional movement of individual surfactant molecules is estimated from the MSD. The MSD of all surfactant molecules in the monolayers, in the absence and presence of AuNP, is calculated for the expanded and compressed states (figures 5 and 6; electronic supplementary material, figure S9).

Figure 5.

MSD for the surfactant monolayer peptide SP-B_1-25 obtained from simulations of the LSM in the (a) expanded and (b) compressed states.

Figure 6.

(a) LSM phospholipids MSD curves with colours representing different AuNPs shapes at approximately 0.1 mol% of AuNPs/lipids applied in the (a) expanded state (APL = 0.54 nm²) and (b) compressed state (APL = 0.47 nm²) monolayers. Black colour represents the phospholipids MSD in the monolayer without any NPs. The time range used for fitting the MSD curves is indicated by two dotted lines.

In the expanded state (figure 5a), the diffusion of surfactant peptide is reduced by all four AuNPs. In particular, in the presence of a box-shaped particle, the peptide essentially lacks any lateral diffusion. In the absence of AuNPs, the peptides in the compressed monolayer laterally diffuse much more slowly than in the expanded state monolayer (electronic supplementary material, table S1). In contrast with the expanded state, in the compressed state monolayer, the presence of AuNPs induces the peptides to diffuse much faster than they do in the absence of AuNPs (figure 5b).

Interestingly, a similar trend in the peptide MSD and cluster formation for different-shaped AuNPs has been identified in the expanded monolayer (box < icosahedron < spherical < rod < no_NPs). By contrast, a different trend is noticed in the compressed monolayer (No_NPs < all shape AuNPs). Therefore, we may infer that the diffusion of peptides might be related to the peptide clustering formation in the monolayer.

The MSD curves of the phospholipids in the presence of the four different AuNPs for the monolayers in expanded and compressed states are shown in figure 6. The corresponding data for CHOL are shown in the electronic supplementary material, figure S9. The diffusion coefficients are calculated from the slope of the corresponding MSD curves and listed in the electronic supplementary material, tables S2 and S3. Similar to the peptide diffusion in the reference systems (figure 5), expanded monolayer is more favourable for phospholipids and CHOL diffusion than compressed state monolayer. In the compressed state, lipids usually show limited to no diffusion as the low APL means an insufficient area for the lipids to move [62,63]. In the expanded state monolayer (figure 6a; electronic supplementary material, figure S9a), the box- and icosahedron-shaped AuNPs have a stronger effect on the diffusion of the phospholipids and CHOL than the spherical- and rod-shaped AuNPs. These results imply that the lipids' translational motion along with the monolayer is slowed by the box and icosahedron AuNPs over the simulation time more than by the other two shapes. The MSD data from the LSM in the presence of the four AuNPs in the compressed state shows that the particles increase the diffusion of the phospholipids and CHOL to a large extent (figure 6b; electronic supplementary material, figure S9b and tables S2 and S3). Thus, analogously to peptides, the presence of AuNPs reduces the diffusion of the lipids in the compressed state monolayer.

These combined results suggest that in the absence of NPs in the monolayer, the surfactant molecules diffuse quicker than in the compressed monolayers. This increased diffusion is not surprising as the diffusion of molecules is facilitated by the larger APL and decreased order in the expanded state compared to the compressed state. The presence of AuNPs in the expanded state monolayer reduces the diffusion of the peptides and phospholipids, while in the compressed state, the diffusion is increased. The diffusion of surfactant lipids and peptides is increased by the NPs in the compressed monolayer due to the introduction of lipid disorder by the NPs, the increase in lateral pressure and the potential compatibility of peptides.

4. Summary and conclusion

In this study, we used CG MD simulations to investigate the effect of AuNPs on the structure and dynamics of a model LSM with a particular focus on differences resulting from the shape of the AuNP. The shapes investigated were spherical, rod, cube and icosahedron. Our findings demonstrate that the presence of AuNPs can significantly affect the structure of the monolayer. Among the most striking effect was that consistent with previous results from wet-laboratory experiments, our simulations showed that in the compressed state, the AuNPs prevent the monolayer from reaching the close to zero surface tension required for normal exhalation. The effect of the AuNPs on the order parameter is consistent with this. All four NPs reduce the order parameters in both chains sn-1 and sn-2 tails, but the effect is much more pronounced in the compressed state than in the expanded state.

Further, all four NPs cause a thinning of the monolayer in the compressed state where the particles drag surfactant molecules into the water phase. This effect is much less pronounced in the expanded state. The difference between compressed and expanded states is also clear from the diffusion data. In the compressed state, all four AuNPs increase the diffusion of phospholipids, CHOL and surfactant peptides. The opposite trend is present for the compressed state, where AuNPs reduce diffusion of all monolayer components. The only exception to this trend appears in the clustering analysis, where the AuNPs change the clustering behaviour of peptides in the expanded monolayer but not in the compressed one. It is important to note that in the compressed state, clustering is already present without AuNPs, while in the expanded state, some of the AuNPs induce clustering.

Concerning the effect of shape, our results indicate that the shape has no apparent effect for some properties, while for other properties, there are significant differences between the four shapes. For APL and membrane thinning, there is no difference between the AuNP shapes. By contrast, for order parameters, diffusion and clustering, the four shapes show significant differences. Comparing the different properties shows no trend concerning which shape has the biggest effect on the monolayer.

Overall, our results indicate that the compressed state is more affected by the presence of the AuNPs than the expanded state. The above insights from this study might assist future studies of how AuNP shapes affect the LSM during inhalation or exhalation conditions.

Data accessibility

All data are presented in the paper in either figure or tabular form. The data are also provided in the electronic supplementary material [64].

Authors' contributions

S.I.H. performed the simulations, analysed data and drafted the manuscript. Z.L. analysed data and edited the manuscript. E.D. analysed data, supervised S.I.H. and edited the manuscript. S.C.S. developed the concept, supervised S.I.H., analysed data and edited the manuscript.

Competing interests

There are no conflicts to declare.

Funding

E.D. was funded by the UTS Chancellor's Postdoctoral Research Fellowship scheme.