Signal transduction and the regulation of ion transport in airway epithelia
It has recently been shown that a critical component of airways innate defense is the thin (7 µm) liquid layer lining airway surfaces, the periciliary liquid (PCL), that provides a low viscosity solution for ciliary beating and acts a lubricant layer for mucus transport. Normal airways appear to be able to sense the PCL volume and adjust ion channel activity accordingly by unknown mechanisms. A proposed model for ASL volume regulation is shown in Figure 1.
Apical membrane ion channel activity controls the amount of salt (and water) on airway surfaces and hence, PCL volume and mucus hydration levels. It has recently been proposed that the initiating event in CF lung disease is depletion of the PCL due to abnormal ion channel activity (i.e. a lack of CFTR), which causes dehydrated mucus to adhere to airway surfaces, preventing it from being cleared (Figure 2), causing increased bacterial infections.
The effects of new and emerging tobacco products on lung health: Initial studies found that acute tobacco exposure causes a loss of CFTR from the plasma membrane (Figure 5). As part of the UNC Center for Tobacco Regulatory Science and Lung Health, we are currently investigating this phenomenon as well as other markers of how exposure to tobacco smoke affects airway surface liquid homeostasis and innate immunity.
Regulation of epithelial cell function by the extracellular environment: We have hypothesized that nucleotides, proteases, and other molecules contained in the ASL (ATP for example) can regulate airway ion transport. Figure 3 depicts cystic fibrosis airway cells that have lost the ability to regulate ASL volume following infection with GFP-labeled viruses that inhibit intracellular Ca2+ signaling by depleting extracellular ATP. Recently, we identified that the short palate lung and nasal epithelial clone 1 (SPLUNC1), a 256 amino acid secreted protein highly expressed in airway epithelia, is a potent inhibitor of the epithelial sodium channel (ENaC) (Movie 1). Currently, we are working to identify its mechanism of action on ENaC, as well as building and testing peptides based on its ENaC inhibitory domain, and working to elucidate SPLUNC1’s other roles as a surfactant and antimicrobial.
Gender differences in cystic fibrosis lung disease: Females suffer more severely with cystic fibrosis than their male counterparts, resulting in a significantly shorter lifespan. This gender difference becomes apparent after puberty. Accordingly, we are currently investigating whether estrogens affect Ca2+ signaling and ASL homeostasis. shows an example of simultaneous Fura-2 imaging (as a marker of intracellular Ca2+) and a fusion protein of Estrogen Receptor α (ER α) conjugated to orange fluorescent protein (mOr).
Techniques used in our lab:
- Ca2+ imaging
- Confocal microscopy
- Electrophysiology (In vivo nasal potential difference measurements, microelectrodes & Ussing chambers)
- Fluorescence recovery after photobleaching (FRAP)
- Fluorescence resonance energy transfer (FRET)
- Mass Spectrometry
- Molecular Biology & Real Time (q)PCR
- Tissue culture
- Western Blot