Cystic Fibrosis Sputum

Abstract

Cystic fibrosis (CF) is characterized by the presence of a viscoelastic mucus layer in the upper airways and bronchi. The underlying problem is a mutation in the gene encoding the cystic fibrosis transmembrane conductance regulator protein. Clinical studies of gene transfer for CF are ongoing. For gene delivery to the airways of CF patients to be effective, the mucus covering the target cells must be overcome. We therefore examined the extent to which CF sputum presents a physical barrier to the transport of nanospheres of a size comparable to that of lipoplexes and other transfection systems currently being clinically evaluated for CF gene therapy. We observed that an extremely low percentage of nanospheres (< 0.3%) moved through a 220-microm-thick CF sputum layer after 150 min. The largest nanospheres studied (560 nm) were almost completely blocked by the sputum, whereas the smaller nanospheres (124 nm) were retarded only by a factor of 1.3 as compared with buffer. Surprisingly, the nanospheres diffused significantly more easily through the more viscoelastic sputum samples. We hypothesize that the structure of the network in sputum becomes more macroporous when the sputum becomes more viscoelastic. Sputum from a patient with chronic obstructive pulmonary disease retarded the transport of nanospheres to the same extent as did CF sputum. When directly mixed with CF sputum, recombinant human deoxyribonuclease I moderately facilitated the transport of nanospheres through CF sputum.

Rapid transport of large polymeric nanoparticles in fresh undiluted human mucus

“…An important role of mucus barriers is to trap foreign particles and then clear them from the body before they reach the underlying epithelia (24,31). The mucus barrier has been cited as a critical bottleneck in the treatment of a variety of diseases (17,24,(31)(32)(33)(34), and it has been widely suggested that nanoparticles are unable to efficiently traverse mucus layers (35)(36)(37)(38)(39), including CV mucus (27). For applications in CV diseases, the need for improved particle transport is further underscored by: (i) the previous observation that polystyrene beads firmly adhere to mucin fibers in human CV secretions, rendering them completely immobile (27); (ii) there is 100-to 1,000-fold reduced D eff for herpes simplex virus (d ϭ 180 nm) in CV mucus compared with water (27); and (iii) there are existing estimates of CV mucus mesh pore size of 10 to at most 200 nm from fluorescence recovery after photobleaching (FRAP) and most electron microscopy studies (27,28).…”

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

“…These entanglements are stabilized by covalent and noncovalent interactions, including hydrogen bonding and electrostatic and hydrophobic interactions (3,4). In the cystic fibrosis (CF) 1 diseased state, the water content of sputum is reduced and the amount of cellular debris is increased, leading to an increase in the concentration of physical entanglements (crosslinking) and a pronounced increase in the viscoelastic nature of sputum (5).…”

“…However, increasing viscoelasticity of CF sputum correlates with an increase in overall transport rates of 100 -200-nm particles (5). Current methods, such as fluorescent recovery after photobleaching and fluorescent imaging, used to determine the rate of diffusion of particles in CF sputum provide only the ensemble-averaged transport of particles through a non-physiologically thick sputum slab (5,7,8).…”

Bronchial Mucus as a Complex Fluid: Molecular Interactions and Influence of Nanostructured Particles on Rheological and Transport Properties

“…Generally, oligosaccharide side chains of mucins are responsible for electrostatic interactions and non-specific hydrogen bonding, while the protein backbone allows for hydrophobic interactions. Such a structure allows them to bind substances with different properties, and therefore to hinder their transport rate through the gel (Bansil and Turner, 2006;Bhat et al, 1995;Sanders et al, 2000;Widdicombe, 1997).…”