Enhanced Viscoelasticity of Human Cystic Fibrotic Sputum Correlates with Increasing Microheterogeneity in Particle Transport

Michelle Dawson,Denis Wirtz,Justin Hanes

doi:10.1074/jbc.m309026200

Michelle Dawson, Denis Wirtz + Show 1 more

Open Access

https://doi.org/10.1074/jbc.m309026200

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Abstract

Current biochemical characterizations of cystic fibrosis (CF) sputum do not address the high degree of microheterogeneity in the rheological properties of the mucosal matrix and only provide bulk-average particle diffusion coefficients. The viscoelasticity of CF sputum greatly reduces the diffusion rates of colloidal particles, limiting the effectiveness of gene delivery to underlying lung cells. We determine diffusion coefficients of hundreds of individual amine-modified and carboxylated polystyrene particles (diameter 100-500 nm) embedded in human CF sputum with 5 nm and 33 ms of spatiotemporal resolution. High resolution multiple particle tracking is used to calculate the effective viscoelastic properties of CF sputum at the micron scale, which we relate to its macroscopic viscoelasticity. CF sputum microviscosity, as probed by 100- and 200-nm particles, is an order of magnitude lower than its macroviscosity, suggesting that nanoparticles dispersed in CF sputum are transported primarily through lower viscosity pores within a highly elastic matrix. Multiple particle tracking provides a non-destructive, highly sensitive method to quantify the high heterogeneity of the mucus pore network. The mean diffusion coefficient becomes dominated by relatively few but fast-moving particles as particle size is reduced from 500 to 100 nm. Neutrally charged particles with a diameter <200 nm undergo more rapid transport in CF sputum than charged particles. Treatment with recombinant human DNase (Pulmozyme) reduces macroviscoelastic properties of CF sputum by up to 50% and dramatically narrows the distribution of individual particle diffusion rates but surprisingly does not significantly alter the ensemble-average particle diffusion rate.

Highlights

Transport of viral particles, bacteria, and gene delivery vehicles to the lung epithelial cells is greatly limited by the respiratory sputum that sits on the apical surface of the cells [1]
Current biochemical characterizations of cystic fibrosis (CF) sputum do not address the high degree of microheterogeneity in the rheological properties of the mucosal matrix and only provide bulk-average particle diffusion coefficients
The elastic modulus of CF sputum was much higher than the viscous modulus over the entire tested range of frequency (Fig. 1A), indicating that CF sputum behaves as a viscoelastic solid when probed by macrorheological techniques

Summary

Microrheology and Nanoparticle Tracking in Cystic Fibrotic Sputum

Slow, an improved understanding of the relationship between CF mucus viscoelastic properties, which can be manipulated by mucolytic agents, and the transport of particles with a variety of surface chemistries may be critical to the design of DNA carriers for delivery in the central airways of the lung [15]. CF sputum was treated with DNase, and the effects on macroviscoelasticity and microviscoelasticity and particle transport were assessed. The viscoelasticity of CF sputum was significantly reduced after the addition of DNase, but the ensembleaveraged diffusion rate remained unchanged. The distribution of transport rates of individual particles was more homogeneous, demonstrating that a reduction in viscoelasticity may correspond to a reduction in heterogeneities in transport rates in CF sputum

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DISCUSSION