New biophysical and immunoregulatory mechanisms in neutrophil extracellular trap mediated lung dysfunction in cystic fibrosis

PROJECT SUMMARY
Mucus acts as a defensive barrier in the airways by trapping inhaled particles within a mucin gel network and
clearing them from the airway via mucociliary transport performed by the underlying airway epithelium. Muco-
obstructive airway diseases including cystic fibrosis (CF), asthma, and chronic obstructive pulmonary disease
are caused by the buildup of thick mucus with an aberrant composition that is not able to be dynamically cleared,
resulting in occluded airways. Mucus accumulation also leads to chronic bacterial infection in the airways,
especially by Pseudomonas aeruginosa in CF patients. One abnormal component found in excess within the
mucus of patients with muco-obstructive diseases, most prominently in CF mucus, is neutrophil extracellular
traps (NETs). NETs are web-like complexes comprised mainly of decondensed chromatin intermixed with
neutrophilic granular proteins that are secreted extracellularly to capture and kill bacteria in a process known as
NETosis. DNase is currently used by CF patients to degrade the chromatin structure of NETs in the mucus but
often does not fully restore mucociliary transport in patients, indicating the granular components of NETs are
also likely involved in mucus dysfunction. In our previous research, we used a synthetic biomaterial model of the
chromatin structure of NETs to evaluate the effects on mucus biophysical properties and mucociliary transport
velocity. Building upon this, we propose to use both biomaterial and human airway tissue culture models to
pursue the following objectives: 1) determine how various granular proteins within NETs differentially affect
mucus biophysical properties and transportability across the airway epithelium, and 2) determine how the
alterations to mucin composition and glycosylation in CF mucus contribute to the increased NETosis observed.
For the first objective, we will employ similar synthetic NET biomaterial models, but incorporate neutrophilic
granular proteins into the formulation to evaluate their specific contributions in enhancing mucus viscosity and
decreasing mucociliary transport in CF airways. We will also account for the effects of inhaled DNase therapy
used by CF patients to determine if granular proteins continue to cause mucociliary transport dysfunction after
degradation of the chromatin scaffold of NETs. In the second objective, we will manipulate the expression of
secreted mucins and mucin glycosylation patterns of human airway tissue to understand how mucins and their
glycans modulate the activation of NETosis in neutrophils. We will account for the effects of P. aeruginosa
bacteria on mucin glycosylation to determine how NETosis is altered during infection in CF patients. This
research will identify novel anti-NET mucosal drug targets to prevent NETosis and neutralize the effects of NETs
in the airway mucus barrier. Ultimately, we believe this will lead to improved treatment of CF and other related
muco-obstructive lung diseases.

Grant Number: 5F31HL176146-02
NIH Institute/Center: NIH
Principal Investigator: Allison Boboltz