Surfactant protein A (SP-A) inhibits agglomeration and macrophage uptake of toxic amine modified nanoparticles

Abstract

The lung provides the main route for nanomaterial exposure. Surfactant protein A (SP-A) is an important respiratory innate immune molecule with the ability to bind or opsonise pathogens to enhance phagocytic removal from the airways. We hypothesised that SP-A, like surfactant protein D, may interact with inhaled nanoparticulates, and that this interaction will be affected by nanoparticle (NP) surface characteristics. In this study, we characterise the interaction of SP-A with unmodified (U-PS) and amine-modified (A-PS) polystyrene particles of varying size and zeta potential using dynamic light scatter analysis. SP-A associated with both 100 nm U-PS and A-PS in a calcium-independent manner. SP-A induced significant calcium-dependent agglomeration of 100 nm U-PS NPs but resulted in calcium-independent inhibition of A-PS self agglomeration. SP-A enhanced uptake of 100 nm U-PS into macrophage-like RAW264.7 cells in a dose-dependent manner but in contrast inhibited A-PS uptake. Reduced association of A-PS particles in RAW264.7 cells following pre-incubation of SP-A was also observed with coherent anti-Stokes Raman spectroscopy. Consistent with these findings, alveolar macrophages (AMs) from SP-A^−/− mice were more efficient at uptake of 100 nm A-PS compared with wild type C57Bl/6 macrophages. No difference in uptake was observed with 500 nm U-PS or A-PS particles. Pre-incubation with SP-A resulted in a significant decrease in uptake of 100 nm A-PS in macrophages isolated from both groups of mice. In contrast, increased uptake by AMs of U-PS was observed after pre-incubation with SP-A. Thus we have demonstrated that SP-A promotes uptake of non-toxic U-PS particles but inhibits the clearance of potentially toxic A-PS particles by blocking uptake into macrophages.

• Collectin
• mucosal
• nanoparticles
• surface chemistry
• surfactant

Acknowledgements

S. M. acknowledges EPSRC Laser Loan Pool support, which enabled the CARS experiments. M. K., H. W. C. and J. M. developed the concepts in all the detailed experiments. M. K., H. W. C. and J. M. wrote and managed the proposal outlining the experimental design. M. G. and C. M. organised and collected the human BAL. Z. M., H. W., C. E. and P. D. prepared the materials and conducted DLS. Z. M., M. K., H. W., C. E. and P. D. jointly interpreted and synthesised the protein-NP data to form conclusions. Z. M., R-M. M., H. W. C. and J. M. designed the RAW264.7 and alveolar macrophage experiments. Z. M. and R-M. M. conducted the RAW264.7 and alveolar macrophage experiments. S. M. performed the CARS experiments and the analysis Z. M., M. K. R-M. M., H. W. C. and J. M. jointly interpreted and synthesised the cell data to form the protein-NP manuscript. Z. M., M. K., R-M. M., H. W., C. E., P. D., M. G., C. M., S. M., H. W. C and J. M. jointly revised the manuscript critically for important intellectual content. We thank Dr. Tony Willis for the N-terminal sequencing of purified native human SP-A.

Declaration of interest

The authors do not have any competing financial interests with the work in this article.

This work was funded under the Joint Environment and Human Health programme (NERC-EPSRC Project NEE009395-1), funded by agencies of the UK Government: The Natural Environment Research Council (NERC), Department for Environment Food and Rural Affairs (Defra), Environment Agency (EA), Ministry of Defence (MOD) and the Medical Research Council (MRC). We gratefully acknowledge the financial support of the MRC ITTP Toxicology Unit for Z. M. The use of FENAC (Facility for Environmental Nanoscience Analysis and Characterisation) was supported by NERC FENAC access grant 2013/05/004. This work was also supported by the National Institute of Health Research (NIHR) funded Respiratory Biomedical Research Units of University Hospital Southampton and the Royal Brompton and Harefield NHS Foundation Trust.

Supplementary material available online

Supplementary Table S1 and Figures S1-S6.