Alpha1-antitrypsin deficiency, respiratory infections and airway inflammation: the role of epithelial cells

Alpha-1-Antitrypsin (AAT) is a major inhibitor of neutrophil elastase, and AAT
deficiency (AATD) is associated with early-onset development of COPD. Initially
it was thought that the protease-antiprotease imbalance was the main
explanation for the development of emphysema in AATD. More recently, local
mechanisms operative in the lung were shown to contribute to emphysema and
inflammation in AATD. The most important mutation causing AATD is the
Z-mutation (Z-AATD), and this is accompanied by misfolding and intracellular
accumulation of the polymerized mutant AAT (Z-AAT). This process - extensively
studied in hepatocytes - results in the so-called endoplasmatic reticulum (ER)
stress response, causing cell death and inflammation. ER stress-induced
inflammation and cellular dysfunction may occur in the lung because lung
epithelial cells and macrophages produce AAT. Indeed, studies by other
researchers using cell lines from healthy subjects engineered to overexpress
recombinant Z-AAT have provided preliminary evidence for altered epithelial
cell function. In vivo, epithelial AAT production appears to be driven mainly
by inflammatory cytokines such as oncostatin M. So far, the effect of
intracellular polymerized Z-AAT on epithelial function in patients and its role
in the progression of emphysema has not been studied before.

We hypothesize that ER stress contributes to epithelial dysfunction in Z-AATD,
resulting in enhanced inflammation. In addition, we hypothesize that the
airways of Z-AATD patients are particularly prone to bacterial infection and
colonization through the inflammatory mechanisms outlined above that result in
impaired host defence. We aim to test this hypothesis by:
1. Investigating airway secretions (nasal secretions and induced sputum) from
patients with Z-AATD, non-AATD COPD and healthy controls for the presence of
respiratory pathogens, inflammatory mediators and effector molecules of innate
immunity. Bronchial biopsies will be collected and studied in a subgroup of
Z-AATD and non-Z-AATD COPD patients.
2. Exploring the ER stress response in epithelial cells from Z-AATD patients by
using epithelial cell culture and investigating whether exposure to smoke or
respiratory pathogens further enhances ER stress. Cellular markers of ER
stress, as well as production of antimicrobial peptides will be assessed as
read-out for epithelial polymerization of Z-AAT. In addition, we will explore
whether a deficiency in AAT production itself (induced by siRNA) alters
epithelial cell function in culture.
3. Developing a model system to study the efficacy of compounds that block AAT
polymerization and increase secretion in airway epithelial cells from Z-AATD
subjects. To this end we will generate epithelial cell lines from AATD patients
and non-AATD COPD controls by introducing the catalytic subunit of telomerase
(hTERT) in these cells

We will collect nasal secretions and induced sputum from Z-AATD and non-Z-AATD
patients, as well as healthy controls. These samples will be used to compare
local inflammation, antimicrobial peptides as effector molecules of innate
immunity, and microbial colonization. In addition, bronchial brushes and
biopsies will be collected during bronchoscopy in Z-AATD and non-Z-AATD
patients to study these parameters in the bronchial mucosa (no bronchoscopy
will be performed in healthy subjects for ethical reasons). In addition,
bronchial epithelial cells will be isolated from the brushes to establish cell
culture

There is a 20% chance patients will become desaturated during bronchoscopy,
despite usual oxygen supply of 2 L/min during the bronchoscpy procedure. if
this occurs, the procedure will be stopped.